History of virology. Principles of classification of viruses Virology is the science that studies the morphology, physiology, genetics, ecology and evolution of viruses
QUESTION No. 1 “HISTORY OF VIRUSOLOGY. ROLE OF VIRUSES IN INFECTIOUS PATHOLOGY OF HUMAN ANIMALS."
In the first period, people did not know the essence of the disease, they only described it. In the 18th century, the doctor Gener developed a vaccine against smallpox, with which it was treated. Further credit goes to Pasteur; rabies existed in his time. He proved that rabies is transmitted by biting. Nothing grew on nutrient media. After Pasteur's work, it was found that infectious diseases are caused by tiny organisms (microbes). Not a single method of bacterial research made it possible to isolate microbes whose presence is associated with smallpox, foot-and-mouth disease, and plague.
In 1931, a method for cultivating chicken embryos was proposed. This method is highly sensitive; infection by spontaneous viruses is excluded. The most rapid development of virology began after 1948. Enders proposed a method of single-layer cell and tissue cultures. This method made it possible to study many viruses and obtain vaccines. The study of viruses was formed into the independent science of virology, which studies viruses and the diseases caused by them. General virology studies the nature and origin of viruses, structure and chemical composition, resistance to physicochemical factors; its subject is also the interaction of virus and cell, genetics of viruses, features of the formation of immunity against viruses, general principles of diagnosis and prevention. She studies the same issues as general virology. Viruses as objects have units of measurement.
QUESTION No. 2 “SUBJECT AND TASKS OF GENERAL AND PRIVATE VETERINARY VIRUSOLOGY. HISTORY OF THE DISCOVERY OF VIRUSES. ACHIEVEMENTS OF DOMESTIC VIRUSOLOGY".
Virology is a science that studies the nature and origin of viruses and the diseases they cause. General virology studies the nature and origin of viruses, structure and chemical composition, resistance to physicochemical factors; its subject is also the interaction of virus and cell, genetics of viruses, features of the formation of immunity against viruses, general principles of diagnosis and prevention. She studies the same issues as general virology. Viruses as objects have units of measurement. Period - people did not know the essence of the disease, they only described it. In the 18th century, the doctor Gener developed a vaccine against smallpox, with which it was treated. Further credit goes to Pasteur; rabies existed in his time. He proved that rabies is transmitted by biting. Nothing grew on nutrient media. After Pasteur's work, it was found that infectious diseases are caused by tiny organisms (microbes). Not a single method of bacterial research made it possible to isolate microbes whose presence is associated with smallpox, foot-and-mouth disease, and plague.
The idea of the existence of a pathogen different in nature from microbes did not occur to Pasteur. The first virus discovered affected tobacco plants (tobacco mosaic). At that time, this virus caused great economic damage. Scientists set out to find out the cause of this disease. This work was entrusted to D.I. Ivanovsky.
As a result of observations, D.I. Ivanovsky and V.V. Polovtsev first suggested that the tobacco disease, described in 1886 by A.D. Mayer in Holland under the name mosaic, is not one, but two completely different diseases of the same plant: one of them is hazel grouse, the causative agent of which is a fungus, and the other is of unknown origin. D.I. Ivanovsky continues his study of tobacco mosaic disease in the Nikitin Botanical Garden (near Yalta) and the botanical laboratory of the Academy of Sciences and comes to the conclusion that tobacco mosaic disease is caused by bacteria passing through Chamberlant filters, which, however, are not able to grow on artificial substrates . The causative agent of mosaic disease is called by Ivanovsky either “filterable” bacteria or microorganisms, since it was very difficult to immediately formulate the existence of a special world of viruses. Emphasizing that the causative agent of tobacco mosaic disease could not be detected in the tissues of diseased plants using a microscope and was not cultivated on artificial nutrient media.
He founded virology. Increased interest in virology was caused by the fact that viral diseases are of leading importance. 75% of diseases are caused by viruses. They cause enormous economic damage. After Ivanovsky’s discovery, the Danish scientist Beyering repeated Ivanovsky’s experiments and confirmed that the mosaic pathogen passes through porcelain filters and proved that it is a liquid living contagium. The virus gave it its name. In 1903, the causative agents of swine fever and infectious anemia were discovered. In 1915-1917, bacterial viruses were bacteriophages; by the end of the 40s, more than 40 viruses were discovered, and over the past 40 years, more than 500 viral diseases have become known. Scientists set out to obtain viral agents.
In 1931, a method for cultivating chicken embryos was proposed. This method is highly sensitive; infection by spontaneous viruses is excluded. The most rapid development of virology began after 1948. Enders proposed a method of single-layer cell and tissue cultures.
QUESTION No. 3 “PRINCIPLES OF MODERN CLASSIFICATION OF VIRUSES, MAIN GROUPS OF VIRUSES.”
The modern classification of viruses is universal for viruses of vertebrates, invertebrates, plants and protozoa. It is based on the fundamental properties of virions, of which the leading ones are those characterizing nucleic acid, morphology, genome strategy, and AG properties. Fundamental properties are placed in 1st place, since viruses with similar AG properties also have a similar type of nucleic acid, similar morphological and biophysical properties. An important feature for classification, which is taken into account with structural characteristics, is the strategy of the viral genome, which is understood as the method of reproduction used by the virus, determined by the characteristics of its genetic material. AG and other biological properties are characteristics that underlie the formation of a species and have significance within the genus. The modern classification is based on the following main criteria: 1) type of nucleic acid (RNA or DNA), its structure (number of strands); 2) the presence of a lipoprotein membrane; 3) viral genome strategy; 4) size and morphology of the virion, type of symmetry, number of capsomeres; 5) phenomena of genetic interactions; 6) range of susceptible hosts; 7) pathogenicity, including pathological changes in cells and the formation of intracellular inclusions; 8) geographical distribution; 9) method of transmission; 10) AG properties. Based on the listed characteristics, viruses are divided into families, subfamilies, genera and types. A number of rules have been developed to organize the names of viruses. The family names end in "viridae" "virinae" "virus". The name may contain the usual Latinized designations, numbers and type designations, abbreviations, letters and their combinations.
QUESTION No. 4 “CHEMICAL COMPOSITION AND PHYSICAL STRUCTURE OF VIRUSES. CONCEPT OF VIRION, CAPSIDS, CAPsomeres. TYPE OF SYMMETRY.
Viruses are made up of a piece of genetic material, either DNA or RNA, that makes up core virus, and a protective protein shell surrounding this core, which is called capsid. A fully formed infectious particle is called virion. Some viruses, such as herpes or influenza viruses, also have an additional lipoprotein shell, which arises from the plasma membrane of the host cell. Unlike all other organisms, viruses do not have a cellular structure. The shell of viruses is often constructed of identical repeating subunits - capsomeres. Capsomeres form structures with a high degree of symmetry that are capable of crystallization. This makes it possible to obtain information about their structure using both crystallographic methods based on the use of X-rays and electron microscopy. As soon as virus subunits appear in the host cell, they immediately exhibit the ability to self-assemble into a whole virus. Self-assembly is also characteristic of many other biological structures and is of fundamental importance in biological phenomena. An essential component of a viral particle is one of two nucleic acids, protein and ash elements. These three components are common to all viruses without exception, while the remaining lipids and carbohydrates are not included in all viruses. Viruses, which, in addition to protein and nucleic acid, also contain lipids and carbohydrates, as a rule, belong to the group of complex viruses. In addition to the proteins that make up the nucleoprotein “core,” virions may also contain a virus—specific proteins that have been built into the plasma membranes of infected cells and cover the viral particle when it leaves the cell or “buds off” from its surface. In addition, some enveloped viruses have a submembrane matrix protein between the envelope and the nucleocapsid. The second large group of virus-specific proteins consists of non-capsid viral proteins. They are mainly related to the synthesis of virion nucleic acids. The fourth component sometimes found in purified viral preparations is carbohydrates (in an amount exceeding the sugar content of the nucleic acid). The elementary bodies of the influenza virus and classical fowl plague contain up to 17% carbohydrates.
According to morphological characteristics, all viruses are divided into:
1) Rod-shaped
2) Globular
3)Cuboidal
4) Club-shaped
5) Thread-like
The main ones are the first 4, filamentous in the intermediate form.
The concept of the type of symmetry.
Depending on the location of capsomeres in the protein shell, all viruses are divided into 3 groups:
1) Spiral type
2)Cubic type
3) Combined
1 – have viruses that are large in size and have high polymorphism. Their capsomeres are arranged in the form of a spiral with different diameters and thus most often have a spherical shell, sometimes they are covered with a second shell (peplos). Nucleic acid is twisted like a spring and arranged in coils in the form of protein molecules.
2 – in such viruses, capsomeres are arranged in the form of a regular polyhedron (icosahedron). It is twisted into a ball and is located in the center.
In most viruses, capsomeres have the shape of 5-6 faceted prisms.
3 – this type of symmetry is characteristic of bacteriophages. All varieties of bacteriophages have a head of cubic symmetry, and a tail with a spiral structure. The surface of the head is covered with a protein shell, which consists of homogeneous protein subunits. One of the nucleic acids is located in the cavity of the head. The tail end consists of a hollow rod. It ends with a hexagonal plate at the end. The tail end is surrounded by a collar, to which is attached a sheath covering the entire shaft.
Chemical composition of viruses.
Methods of purification and concentration of viruses by salting out, adsorption, ultrafiltration, and sedimentation made it possible to study the chemical composition. Viruses contain proteins and one of the nucleic acids. Viruses of large and medium sizes also contain lipids, carbohydrates and some other organic and inorganic compounds.
Most of the protein and associated lipids and carbohydrates are the membrane. The substances that make up viruses have characteristics, both chemically and biologically.
Proteins – the main part (20 AA).
The significance of viral proteins is their protective function (capsid formation).
The virus contains enzymes of a protein nature (adsorption, targeting function) and endowed with immune properties (determine antigenic properties).
Features of viral proteins:
1. They have the property of self-assembly (as they accumulate, viral proteins aggregate).
2. They have selective sensitivity in relation to physical and chemical factors.
3.Do not undergo hydrolysis under the influence of proteolytic enzymes.
Proteins make up 50-75% of the mass of virions.
Cells infected with the viral gene encode the synthesis of 2 protein groups:
Structural===, ===non-structural===
1.Structural – the amount in the virion, depending on the complexity of the virion’s organization. Structural proteins of group 2 are divided: a. capsid b. supercapsid (peplomers).
Complex viruses contain both types of proteins. A number of such viruses contain enzymes in their capsids that carry out transcription and replication.
Supercapsid proteins form spines (up to 7-10 nm). The main function of glycoproteins is interaction with specific cell receptors. Another function is participation in the synthesis of cellular and viral membranes.
“Address function” is developed in the process of evolution; it is a search for a sensitive cell.
It is realized through the presence of special proteins that recognize special receptors on the cell.
Non-structural (temporary) viral proteins are precursors of viral proteins, DNA/RNA polymerase synthesis enzymes, ensure transcription and replication of the viral genome, regulatory proteins, polymerases.
Lipids - in complex viruses are found in the supercapsid (from 15 to 35 percent). The lipid component stabilizes the structure of the viral particle.
Carbohydrates – up to 10-13%. They are part of glycoproteins. Play an essential role in protein structure and function.
Nucleic acids are a constant component. Complex polymer compounds. Isolated by Miescher in 1869 from leukocytes. Unlike bacteria, they contain only 1 amino acid. Structurally, nucleic acids are different.
1.Linear double helix with open ends.
2. Linear double helix with closed ends.
3.Linear single-spiral.
4.Ring single spiral.
1.Linear single-spiral.
2. Linear fragmented.
3.Ring single-spiral.
5.Linear double-helix fragmented.
QUESTION No. 5 “RESISTANCE OF VIRUSES TO PHYSICAL AND CHEMICAL FACTORS. PRACTICAL USE OF THESE PROPERTIES."
Different groups of viruses have different resistance in the external environment. The least resistant viruses are those that have lipoprotein membranes; the most resistant are isometric viruses. Thus, orthomyxoviruses and paramyxoviruses are inactivated on surfaces in a few hours, while polioviruses, adenoviruses, and reoviruses remain infectious for several days. However, there are exceptions to this rule. Thus, the smallpox virus is resistant to desiccation and persists in excreta for many weeks and months. The hepatitis B virus is resistant to adverse external factors and retains its activity in serum even after short-term boiling. The sensitivity of viruses to ultraviolet and x-ray irradiation depends primarily on the size of their genome. The sensitivity of viruses to formaldehyde and other chemicals that inactivate genetic material depends on many conditions, including the density of packaging of the nucleic acid in the protein case, the size of the genome, and the presence or absence of outer shells. Viruses with lipoprotein envelopes are sensitive to ether, chloroform and detergents, while simply constructed isometric and rod-shaped viruses are resistant to their action. An important feature of viruses is sensitivity to pH. There are viruses that are resistant to acidic pH values (2.2-3.0), for example, viruses that cause intestinal infections and enter the body through nutrition. However, most viruses are inactivated at acidic and alkaline pH values.
QUESTION No. 6 “VIRAL NUCLEIC ACIDS. THEIR VARIETIES, STRUCTURES, BASIC PROPERTIES.
Viral DNA molecules can be linear or circular, double-stranded or single-stranded along their entire length, or single-stranded only at the ends. Most nucleotide sequences occur only once in the viral genome, but there may be repeating or redundant regions at the ends. There are also large differences in the size of the genome in the structure of the terminal regions of viral DNA. Animal viruses undergo almost no modifications to their DNA. For example, although the DNA of host cells contains many methylated bases, viruses have, at best, only a few methyl groups per genome. The sizes of RNA virus virions vary greatly - from 7.106 daltons in picornaviruses to >2.108 daltons in retroviruses; however, the size of RNA and, therefore, the amount of information it contains varies to a much lesser extent. The RNA of picornaviruses is probably the smallest known, containing about 7,500 nucleotides, and the RNA of paramyxoviruses is perhaps the largest, almost 15,000 nucleotides. Apparently, all independently replicating. Nucleic acids are a constant component. Complex polymer compounds. Isolated by Miescher in 1869 from leukocytes. Unlike bacteria, they contain only 1 amino acid. Structurally, nucleic acids are different.
1. Linear single-helix. 2. Linear fragmented. 3. Ring single-helix. 5. Linear double-helix fragmented.
QUESTION No. 7 “VIRUS PROTEINS, THEIR FEATURES (CHARACTERISTICS OF THE PROPERTIES OF NEURAMINIDASES AND ANTIGENS OF MIXOVIRUSES).”
They represent an extremely heterogeneous class of biological macromolecules. AKs are essential components of proteins. Alpha-AA are relatively simple organic molecules. The molecular weight of AK lies in the range of 90-250D. The polypeptide can contain from 15 to 2000 AA. The most common polypeptides weighing from 20 to 700 kDa, consisting of 100-400 AA. Viral proteins—proteins encoded by the virus genome—are synthesized in the infected cell. Based on the function of localization, structure and regulation of synthesis, viral proteins are divided into structural and non-structural; enzymes, precursors, histone-like capsid proteins; membrane, transmembrane.
Structural proteins– all proteins that are part of mature extracellular virions. They perform a number of functions in the virion: 1) protection of the NK from external damaging influences; 2) interaction with the membrane of sensitive cells during the first stage of their infection; 3) interaction with viral NK during and after its packaging into the capsid; 4) interaction with each other during capsid self-assembly; 5) organizing the penetration of the virus into a sensitive cell. These 5 functions are inherent in the structural proteins of all viruses without exception. All functions can be realized by one protein. 6) ability to be destroyed during the liberation of the NK; 7) organization of exit from the infected cell during the formation of the virion. 8) organization of “melting” and fusion of cell membranes.
Proteins may also have the ability to catalyze certain biochemical reactions: 9) RNA-dependent RNA polymerase activity. This function is performed by the structural proteins of all viruses whose virions contain RNA, which does not play the role of mRNA; 10) RNA-dependent DNA polymerase activity - this function is performed by special retroviral proteins called reversetases; 11) protection and stabilization of the viral NK after its release from the capsid in the infected cell.
Depending on the location of a particular protein in the virion, groups of proteins are distinguished: A) Capsid proteins - in the virions of complexly organized viruses, these proteins can perform only 2-3 functions - protection of the NK, the ability to self-assemble and destroy during the release of the NK. In the virions of simple viruses, their functions are usually more diverse. B) Proteins of the viral supercapsid shell - their role is reduced mainly to organizing the budding of virions, the ability to self-assemble, interacting with the membrane of sensitive cells, organizing penetration into a sensitive cell. C) Matrix proteins are proteins of the intermediate layer of virions, located immediately under the supercapsid shell of some viruses. Their main functions: organizing budding, stabilizing the structure of the virion due to hydrophobic interactions, mediating the connection of supercapsid proteins with capsid ones. D) Viral core proteins - represented mainly by enzymes. Viruses with multilayer capsids may also have a protective role. E) Proteins associated with NK of the innermost layer of virions.
Non-structural proteins– all proteins encoded by the viral genome, but not included in the virion. They have been studied less well, which is due to the incomparably greater difficulties that arise in their identification and isolation compared to structural proteins. Non-structural proteins, depending on their function, are divided into 5 groups: 1) Regulators of viral genome expression - directly affect the viral NK, preventing the synthesis of other viral proteins, or, conversely, triggering their synthesis. 2) Precursors of viral proteins - are precursors of other viral proteins that are formed from them as a result of complex biochemical processes. 3) Non-functional peptides – are formed in an infected cell. 4) Inhibitors of cellular biosynthesis and inducers of cell destruction - these include proteins that destroy cellular DNA and mRNA, modify cellular enzymes, giving them virus-specific activity. 5) Viral enzymes - enzymes encoded by the viral genome, but not included in the virions.
QUESTION No. 8 “PERIODS AND STAGES OF VIRUS REPRODUCTION. TYPES OF INTERACTION.”
Interaction of viruses with host cells and virus reproduction.
Viruses go through a complex development cycle in a cell. Morphogenesis of viruses is the main stage of this development and consists of formative processes leading to the formation of a virion as the conclusion of the form of virus development. Ontogenesis and reproduction of the development of the virus are regulated by the genome.
In the 50s it was established that the virus propagates through reproduction, i.e. reproduction of nucleic acids and proteins followed by virion assembly. These processes occur in different parts of the cell, for example in the nucleus and cytoplasm (disjunctive mode of reproduction). Viral reproduction is a unique form of expression of a foreign infection in the cells of humans, animals, insects and bacteria.
Morphogenesis is regulated by morphogenetic genes. There is a directly proportional relationship between the complexity of the virion ultrastructure and its morphogenesis. The more complex the organization of the virion, the longer the development path of the virus. This entire process is carried out with the help of special enzymes. Because Viruses do not have their own metabolism and therefore require enzymes. However, over 10 enzymes, different in origin and functional significance, have been found in viruses.
By origin: virion, virus-induced, cellular, modified by viruses. The former are part of many DNA and RNA viruses. DNA-dependent RNA polymerase, protein kinase, ATPase, ribonuclease, RNA-dependent RNA polymerase, exonuclease and others.
Virion forms include: hemoglutinin and neuraminidase, lysozyme.
Virus-inducing enzymes are enzymes whose structure is encoded in the genome, and synthesis occurs on the host ribosome - early virion proteins.
Cellular - include enzymes of the host cell, are not virus-specific, however, when interacting with viruses, the activity can be modified.
According to their functional significance, enzymes are divided into 2 groups:
— Participating in replication and transcription;
— Neuraminidase, lysozyme and ATPase, which contribute to the penetration of the virus into the cell and the exit of mature virions from the cell.
Reproduction of virions is characterized by a change of stages:
According to modern data, there are 3 main periods in the reproduction cycle:
1. Initial (preparatory) 2. Middle (latent) 3. Final (final)
Each period includes a number of stages:
First stage
1.Adsorption of the virus on the cell.
2. Penetration into the cell.
3.Deproteinization (release of nucleic acid).
Second stage
1.Biosynthesis of early viral proteins
2.Biosynthesis of viral components
Third stage
1.Formation of mature virions
2. Exit of mature virions from the cell.
1.Adsorption is a physical and chemical process that is a consequence of the difference in charges. This stage is reversible; its outcome is influenced by the acidity of the environment, temperature and other processes.
The main role in virus adsorption is played by the interaction of the virus with complementary cell receptors. By chemical nature they belong to mucopolyproteins. The rate of adsorption is influenced by hormones acting on the receptors. Adsorption of the virus may not occur, which is due to the different sensitivity of cells to viruses. Sensitivity, in turn, is determined by:
The presence in the cell membrane and cytoplasm of enzymes that can destroy the membrane and release nucleic acid.
The presence of enzymes, material that ensures the synthesis of viral components.
2.Virus penetration into the cell:
The virus penetrates in 3 ways - by direct injection (typical of phages); by destroying the cell membrane (fusion path - typical for plant viruses); by pinocytosis (characteristic of vertebrate viruses).
3. Reproduction of DNA-containing viruses.
4. Exit of the virion from the cell:
1. They leak through the cell membrane and are covered with a supercapsid, which includes cell components: lipids, polysaccharides. In this case, the cell retains its vital activity and then dies. In some cases, during the process of reproduction, processes can occur over several years, but vital activity is maintained. With this method, mature virions leave the cell gradually and over a relatively long period of time. This path is typical for complex viruses that have a double shell.
Anomalous viruses.
During the reproduction process, various abnormal viruses are formed. Through the efforts of Academician Zhdanov, in recent years pseudoviruses have been discovered, consisting of an RNA virus and cell proteins that form a capsid. They have infectious properties, but due to the peculiarity of the capsid, they are not susceptible to the action of antibodies that form a response to this virus.
The formation of such viruses is explained by prolonged virus carriage in the presence of specific antibodies in the body.
The reasons for the formation of such virions are:
1.High multiplicity, as a result of which the cell is not able to provide all its offspring with energy material.
2. The action of interferon - it affects the synthesis of DNA and RNA viruses.
QUESTION No. 9 “PECULIARITIES OF BIOSYNTHESIS OF DNA-CONTAINING VIRUSES. THE CONCEPT OF TRANSCRIPTION AND BROADCASTING.”
Transcription - the rewriting of DNA into RNA - is carried out using the enzyme RNA polymerase, the products of which are the biosynthesis of mRNA. DNA viruses that reproduce in the nucleus use cellular polymerase for transcription. RNA-containing viruses are produced by the genome itself. In some RNA-containing viruses, the transfer of genetic information is carried out according to the RNA-RNA-protein formula. This group of viruses includes picornoviruses and cornoviruses.
Protein synthesis occurs as a result of translation into RNA.
Under the influence of enzymes in DNA-containing viruses, mRNA is synthesized, and the mRNA is sent to the ribosomes of the sensitive cell. The synthesis of early virion proteins begins on the ribosomes of the cell (endowed with the properties of enzymes, blocking cellular metabolism).
Early virion proteins give rise to the formation of early virion acids.
As early virion proteins accumulate, they block themselves and the process is rearranged on the ribosomal apparatus. Virions are assembled and the newly formed virions leave the mother cell.
QUESTION No. 10 “TYPES OF INTERACTION, MAIN OUTCOMES OF VIRUS INTERACTION WITH A CELL.”
1) Productive interaction - when viruses multiplying in a cell form a new generation 2) Abortive - when reproduction cycles are interrupted at some stage. 3) Lytic reaction - when after the formation of the virus the cell dies. 4) Latent reaction - when an infected cell retains its viability for a long time. 5) Integration - when the genomes of viruses and cells are combined. In this case, reproduction of genomes occurs in cells and is subject to general regulation. Reproduction of viruses causes pathological changes in the affected cells, expressed by functional and morphological disorders of the cells. Possible outcomes of the processes of interaction between various viruses and cells can be divided into 5 types: 1) Degeneration of cells - leads to their death. In this case, the cell acquires an irregular round shape, becomes rounded, becomes denser, granularity appears in the cytoplasm, wrinkling and fragmentation of the nuclei. 2.The formation of symplasts is multinucleate. accumulations outside the cell. substances. 3) Cell transformation – i.e. formation of foci of random three-dimensional growth. Cells in these foci acquire new hereditary properties, continuously piling up on each other (tumors). 4. Arr. extracellular inclusions, which are products of the cell reaction to the viral particle. 5) Latent infection is a kind of condition. equilibrium between the virus and the cell., when the infection does not manifest itself in any way. Insignificant production of the virus is observed, without cell damage.
QUESTION No. 11 “PHASES OF INTERACTION OF RNA CONTAINING VIRUS WITH A CELL.”
See question No. 8
QUESTION No. 12 “PATHOGENESIS OF VIRAL INFECTIONS
Tropism is the tendency of a virus to one or another site of infection. For respiratory infections, the virus is localized in the nasopharynx, trachea and lungs; for enteroviruses - in feces; for neurotropic ones - in the GM or SM; with dermotropic - in the skin.
Pathogenesis of viral infections.
Pathogenesis is understood as a set of processes that cause a disease, its development and outcome.
Pathogenesis is determined by:
1.Tropism of the virus
2.Number of infectious particles
3. Cell response to infection.
4. The body's response to changes in cells and tissues.
5. Speed of reproduction.
The tropism of viruses is based on the sensitivity of certain cells to the virus.
Pathogenesis is determined by the main mechanisms of interaction of viruses with cells:
Atrophy or dystrophy (CPD)
Formation of inclusion bodies
Formation of symplasts and syncytia
Transformation
Latent (chronic) infection.
Pathogenesis at the cellular level - this includes CPD (visible morphological changes in cells under the influence of a particular viral agent). The nature of CPP is different and depends on:
1.Type of cell
2.Biochemical properties of the virus
3. Infectious dose
The nature of CPP is assessed using a 4-point cross system and changes are taken into account when cell cultures are used for titration (i.e.).
Pathogenesis at the organismal level.
The state of infection as any biological process is dynamic; the dynamics of interaction are usually called the infectious process. On the one hand, the infectious process includes: the introduction, reproduction and spread of the pathogen in the body, as well as the pathogenic action, and on the other hand, the body’s reaction to this action.
The pathogenic effect of the pathogen may be different. It manifests itself in the form of an infectious disease of varying severity, in others without pronounced clinical signs, in others it manifests itself only by changes identified by virological, biochemical, and immunological methods. It depends on:
The quantity and quality of the pathogen that has penetrated into a susceptible organism, the conditions of the internal and external environment that determine the resistance of the animal and are characterized by the interaction of micro and macroorganisms. Based on the nature of the interaction between the pathogen and the organism, there are 3 forms:
1.an infectious disease is an infectious process characterized by certain clinical signs, as well as disorders, functional disorders and morphological tissue damage.
2. Microbial carriage is an immunological subinfection. A differentiated approach to various forms of infection makes it possible to correctly diagnose the infection and identify infected animals in a dysfunctional herd. The pathogenesis of any infectious disease is determined by the special action of the pathogen and the body's responses, depending on the conditions in which the interaction of micro and macroorganism occurs. In this case, the routes of penetration and distribution of the pathogen are of no small importance. Pathogen gates: skin, mucous membranes, genitourinary system, placenta.
Each type of pathogen has evolutionarily adapted to such routes of introduction, which provide favorable conditions for reproduction and spread - the entrance gate for each infection is characterized by specificity. To carry out prevention, it is necessary to take into account the specificity of the infection gate. For example, with INAN, the pathogen penetrates the skin through an insect bite. In case of foot and mouth disease, the main route is nutritional; in case of rabies, it is through a bite.
Classification of viral infections.
There are autonomous and integrated infections. Autonomous - in this case, the viral genome replicates independently of the cellular genome. Autonomous infection is typical for most viruses.
Integrated infections - the viral genome is included in the cellular genome, i.e. integrated into the cellular genome and replicated with it. In this case, the viral genome replicates and functions as an integral part of the cellular genome. It can integrate both the entire genome and a part. In integrated infections, there is no assembly of viral particles or exit.
Autonomous infection - a cell sometimes acquires the ability to divide unlimitedly as a result of disruption of the regulatory mechanisms that control division. This is more often observed in oncogenic infections.
Productive and abortive infections:
1. Productive – ends with the release of infectious offspring.
2. Abortive – infectious offspring are not formed or there are few of them.
Forms of the course - both productive and abortive - can occur in acute and chronic forms. An acute infection is an infection that results in the cell either recovering or dying. Acute infection at the cellular level can be cytolytic (when cell death occurs).
A chronic infection is an infection in which a cell continues to produce viral particles for a long time and transfers this ability to daughter cells. More often, an abortive infection takes on a chronic form because viral material accumulates and is transmitted to the daughter cell.
Mixed infection - a cell is infected with two or more different viruses, as a result of which two or more infectious processes can be combined in the cell. There are several possible options for virus interaction during a mixed infection:
1. Interference - one virus suppresses the action of another.
2. Complementation (exaltation) - one virus enhances the effect of another.
Classification of viral infections at the organismal level.
The classification is based on:
1. Generalization of the virus
2.Duration of infection
3.Manifestation of clinical symptoms
4. Release of viruses into the environment
One of the forms can transform into another (for example, focal to generalized, acute to chronic).
Focal infection.
The virus acts near the entry gate of infection, due to local reproduction. They have a shorter latent period compared to generalized ones.
Generalized infections.
After a limited period of reproduction in primary foci, generalization of infections occurs - viruses penetrate other systems, for example, foot-and-mouth disease, polio, and smallpox.
Acute infection.
It lasts for a short period and is released into the environment. Ends in death or recovery.
Persistent infection.
With prolonged interaction of the virus with the body. It can be latent, chronic, slow.
Latent infection - is not accompanied by the release of the virus into the environment; under certain conditions it can become acute and chronic.
For influenza, sepsis, AIDS, etc.
Chronic infection.
This is a long-term process. Characterized by periods of remission (adenovirus, herpes).
Slow infections are a kind of interaction between a virus and a phage and are characterized by long incubation periods.
Sources of infection.
When studying any infectious disease, it is important to know the source, place of permanent residence and reproduction, routes of spread, place and timing of preservation, occurrence in the external environment, methods of transmission from sick to healthy.
The natural environment is a living organism, here it finds all the conditions for existence. The duration of stay of viruses varies widely and depends on the biological properties and reactivity of the body. From the conditions of pathogenesis. Sources of infection are only infected organisms. They only play a role in the transmission process. Most animals excrete viruses in excreta, secretions, blood, effluent, and sputum. In most viral infections, the pathogenesis is based on viremia (foot-and-mouth disease, plague, etc.). In these diseases, the virus is released in all possible ways. In chronic cases, viral shedding is less intense, but can be prolonged. For viral diseases, localization is limited to one way: pneumonia - with drops of sputum. The most intense release of the virus into the external environment is observed during the acute period of the disease, but in a number of diseases it also occurs during the incubation period. Asymptomatic infections occur when vaccinated with live vaccines.
QUESTION No. 13 “RULES FOR COLLECTING PATHMATERIAL FROM SICK AND DEAD ANIMALS IN THE EVENT OF SUSPECTED VIRAL DISEASES. TRANSPORTATION AND PREPARATION OF IT FOR VIRUSOLOGICAL STUDIES.
Material for research from sick, dead or forcedly killed animals should be taken as quickly as possible after the appearance of clear signs of the disease or no later than 2-3 hours after clinical death or slaughter. This is due to the fact that immediately after the disease or in the first 1-2 days, the barrier role of the intestine is significantly weakened, which, along with increased permeability of blood vessels, contributes to the dissemination of intestinal flora. In addition, as the infectious process continues and even deepens, the amount of virus may decrease as a result of the influence of the body's defense mechanisms. When taking material for virus isolation, one should proceed from the pathogenesis of the infection being studied (entry gate, routes of spread of the virus in the body, places of its reproduction and routes of excretion). For respiratory infections, nasopharyngeal swabs, nasal and pharyngeal swabs are taken to isolate viruses; for enterovirus - feces; with dermotropic - fresh skin lesions. Various excreta and secretions, pieces of organs, blood, and lymph can serve as materials for isolating the virus. Blood is taken from the jugular vein; in pigs, from the tip of the tail or ear. Washings from the conjunctiva, from the nasal mucosa, from the posterior wall of the pharynx, rectum and cloaca in birds are taken with sterile cotton swabs and immersed in penicillin vials. When taking material from the nasopharynx, you can use the device designed by Thomas and Scott. Saliva flowing from the mouth can be collected directly into a test tube. Urine is collected using a catheter into a sterile container. Feces are removed from the rectum with a spatula or stick and placed in a sterile tube. Vesicular fluid can be collected with a syringe or Pasteur pipette into a sterile tube. The walls of the canker sores and crusts from the surface of the skin are removed with tweezers. After the death of the animal, it is important to take pieces of organs as quickly as possible, because... With many viral infections, the phenomenon of post-mortem autosterilization is observed, as a result of which the virus may not be detected at all or its amount will be very small. Next, the pathological material is placed in low temperatures (dry ice + alcohol; snow + salt) or glycerin on ICH. Patent material must be provided with a reliable and clear label. You need to write down what material was obtained from what animal. A cardboard or plywood tag is hung on the thermos with PM samples, indicating the farm, type of animal, type of material, and date. The thermos must be sealed and delivered by express. It is recommended that samples delivered to the laboratory be used immediately for virus isolation. In the laboratory, the resulting pathological material is freed from preservatives, thawed, washed from glycerin, weighed and measured. Some are taken for research, some in the refrigerator. The preparation of organs and tissues is carried out as follows: the virus is released from the cells of organs and tissues - the material is thoroughly crushed and ground in a mortar with sterile quartz sand. A 10% suspension is usually prepared from the ground material in Hanks or phosphate buffer. The suspension is centrifuged at 1500-3000 rpm, the supernatant is sucked off and freed from microflora by treating with antibiotics (penicillin, nystatin). The suspension is exposed to AB for at least 30-60 minutes at room temperature, then the material is subjected to bacteriological control by inoculation on MPA, MPB, MPPB, Sabouraud's medium. The suspension is stored at minus 20-minus 70 C.
QUESTION No. 14 “METHODS FOR PRESERVATION OF VIRUSES AND THEIR PRACTICAL IMPORTANCE.”
The following methods of virus preservation are used:
1) when storing viral material (pieces of organs or tissues), glycerin (50% solution in ICN) is often used, which has a bacteriostatic effect and at the same time protects viruses. In this case, it can be stored for several months at 4C.
2) viruses are most often stored in refrigerators that provide temperatures of -20, -30, -70C. At this temperature, some viruses lose their infectivity relatively quickly without the addition of protective substances. The addition of inactivated blood serum or skim milk or 0.5-1.5% gelatin has a good protective effect when freezing and storing viruses.
3) Quick freezing to minus 196C with liquid nitrogen. Viruses sensitive to low pH values should be frozen in liquids that do not contain monobasic phosphates.
4) Lyophilization - frozen drying under vacuum conditions - is a very good method of canning. In lyophilized form, viruses can be stored for several years.
QUESTION No. 15 “WORK RULES IN A VIRUSOLOGY LABORATORY. SAFETY PRECAUTIONS WHEN WORKING WITH VIRUS-CONTAINING MATERIAL.”
All laboratory personnel are instructed and trained in safe working methods, provided with overalls, safety footwear, sanitary protection and protective equipment in accordance with current standards. The basic rules of work are as follows: 1) entry of unauthorized persons into the production premises, as well as entry of employees into the laboratory without a gown and spare shoes is strictly prohibited; 2) it is prohibited to leave the laboratory in gowns and special shoes or to put on outerwear over a gown, to smoke, to eat and to store food in the laboratory. In boxing they wear a sterile gown, mask, cap, and if necessary, wear rubber gloves and goggles. Be sure to change your shoes. 3) all material entering the laboratory for testing must be considered infected. It must be handled very carefully; when unpacking, the jars should be wiped on the outside with a disinfectant solution and placed on a tray or in cuvettes. The work area on the table is covered with several layers of gauze moistened with a 5% chloramine solution. When working with pipettes, use rubber bulbs. Pipettes, slides and cover glasses and other used glassware are disinfected by immersing them in 5% chloramine, phenol, Lysol, and sulfuric acid. 4) upon completion of work, the workplace is tidied up and thoroughly disinfected. Virus-containing material necessary for further work is stored in a refrigerator and sealed. 5) hands are thoroughly washed with 5% chloramine, gloves are removed, disinfected a second time, disinfected and washed. When working in a virology laboratory, employees must strictly adhere to the methods and rules of asepsis and antisepsis. Asepsis is a system of measures and work methods that prevent the entry of microorganisms and viruses from the environment into the human body, as well as the material being studied. It involves the use of sterile instruments and materials, disinfection of employees’ hands, and compliance with special sanitary and hygienic rules and work practices. Antiseptics are a set of measures aimed at destroying microorganisms and viruses that can cause an infectious process when they come into contact with damaged or intact areas of the skin and mucous membranes. Ethyl alcohol (70%), alcohol solution of iodine, brilliant green and others are used as antiseptics. Disinfection is the disinfection of environmental objects by destroying pathogenic microorganisms and viruses for humans and animals by physical means and with the help of chemicals. Sterilization – sterilization, complete destruction of microorganisms and viruses in various materials. It is carried out using physical and chemical methods.
QUESTION No. 16 “SCHEME FOR LABORATORY DIAGNOSTICS OF VIRAL INFECTIONS.”
Laboratory diagnostics is a system of measures to detect and indicate the virus. It includes: receipt of sent pathological material, examination of pathological material using a rapid diagnostic method, research using long-term methods (retrospective diagnosis, examination of paired sera in seroreactions).
Laboratory research. I. Indication of the virus in pathological material. 1. Detection – light microscopy of large viruses (Poxviridae), electron microscopy. 2. Detection of inclusion bodies. (Babes-Chenegri bodies in rabies) 3. Detection of viral antigens: serological reactions. 4. Detection of viral NK (DNA probes and PCR - polymerase chain reaction). 5. Detection of the active form of the virus by bioassay (laboratory animals, chicken embryos, cell culture). 6. Detection of hemagglutinins in hemagglutinating viruses (currently practically not used due to the availability of more accurate methods). II. Isolation (isolation) of the virus from pathological material. At least three blind passages are carried out, and a bioassay is performed. A) Laboratory animals (clinical, death, pathological changes) B) Chicken embryos (death, pathological changes, RGA) C) Cell culture (CPD, RGAd, plaque method) III. Identification of the isolated virus - serological reactions. IV. Evidence of etiological role. Sometimes it is necessary to prove the etiological role of the isolated virus. For this purpose, paired blood sera are used in serological reactions. An isolated virus is used as an AG, and paired sera are used as an AT. An increase in antibody titer in the second serum by 4 or more times indicates the etiological role of the isolated virus.
QUESTION No. 17 “CLINICAL-EPIZOOTOLOGICAL DIAGNOSTICS OF VIRAL DISEASES OF ANIMALS, ESSENCE, SIGNIFICANCE.”
Clinical-epidemiological or pre-laboratory diagnostics - carried out on farms and allows only a preliminary diagnosis to be made; recognition is carried out based on collection, comparison of analysis of sick animals (clinical symptoms of the disease, pathological changes in organs). Collecting epidemiological data is very important; it allows us to obtain data on how the disease progresses and information about farms. If the farms are not prosperous, then this once again confirms the diagnosis. A clinical examination focuses the veterinarian on only a few types of diseases. Laboratory diagnostics are still of primary importance.
QUESTION No. 18 “METHODS FOR DETECTING VIRUSES IN PATTERN MATERIAL.”
I. Indication of the virus in pathological material. 1. Detection – light microscopy of large viruses (Poxviridae), electron microscopy. 2. Detection of inclusion bodies (Babes-Chenegri bodies in rabies) 3. Detection of viral antigens: serological reactions. 4. Detection of viral NK (DNA probes and PCR - polymerase chain reaction). 5. Detection of the active form of the virus by bioassay (laboratory animals, chicken embryos, cell culture). 6. Detection of hemagglutinins in hemagglutinating viruses (currently practically not used due to the availability of more accurate methods). Serological tests are used to identify the isolated virus. 1.RIF – immunofluorescence reaction. AG + AT labeled with fluorochrome. Contact is allowed for 30 minutes at 37 C, then a thorough wash is carried out in the laboratory. Detection method: fluorescent light under a microscope. 2.ELISA – enzyme-linked immunosorbent assay. AG + AT with enzyme. Contact, wash, then add a substrate, which upon contact with the AT-enzyme complex gives a color reaction. 3.RSK – complement fixation reaction. AG + AT + complement. Contact. Then the heme system (hemolysin + sheep red blood cells) is added. Contact. If hemolysis does not occur, then AG and AT have fixed complement. Delayed hemolysis is a positive reaction. If hemolysis occurs, then complement is bound by the heme system - the reaction is negative. 4.RDP – diffuse precipitation reaction. AG + AT (diffusion in agar gel). The detection method is the formation of a precipitation contour. 5.RNHA – indirect hemagglutination reaction. Erythrocytes are loaded with antigen and when the antigen-antigen complex is formed, agglutination of erythrocytes occurs. 6.RTGA – hamagglutination inhibition reaction 7.RTGAd – hemadsorption inhibition reaction 8.RN – neutralization reaction. Virus + AT. Contact. Entering a virus-sensitive system. The detection method is to neutralize the infectious activity of the virus.
QUESTION No. 19 “THE PRINCIPLE OF RETROSPECTIVE DIAGNOSTICS, ITS PROS AND CONS.”
Retrospective diagnostics - the goal is to detect the dynamics of the increase in AT, based on the study of paired sera, which are taken twice, at the beginning of the disease and at the end. They are tested in one of the seroreactions. If the increase in AT is 4-5 times greater, the diagnosis is 100%.
Role - the method allows you to reliably make a diagnosis in most cases.
Role – duration of retrospective diagnosis.
QUESTION No. 20 “AUJESKY’S DISEASE VIRUS.”
Aujeszky's disease (pseudorabies, pruritic plague, rabid scabies, infectious bulbar palsy) is an acute disease of all types of farm animals, fur-bearing animals and rodents. It is characterized by signs of damage to the brain and spinal cord, severe itching and scratching.
AD causes particular damage in pig farming and fur farming. This is an acute food infection in fur-bearing animals. The cause is food, which is often slaughterhouse waste and offal obtained from sick animals or virus-carrying animals.
Clinic. The incubation period is 1.5 days - 10-12 days, depending on the method of infection, the virulence of the virus and the resistance of the animal. The virus is pantropic.
In pigs the clinical course proceeds without signs of itching. Suckers and weanlings are seriously ill. The disease is septic in nature. Piglets usually die within 4-12 hours. In piglets from 10 days to 3 months, the first signs of the disease are fever (40-42), depression, mucous discharge from the nose. Later, signs of central nervous system damage appear: restlessness, maneuvering movements, loss of orientation, convulsions, arching of the back, paralysis of the pharynx, larynx, limbs, pulmonary edema, salivation. The illness lasts from several hours to 3 days. Mortality: 70-100%
In sows it manifests itself as a flu-like syndrome with recovery after 3-4 days.
In cattle, the temperature rises to 42 C, chewing stops, severe itching in the nostrils, lips, cheeks, refusal to feed, lethargy, anxiety, fear, rapid breathing, sweating, cramps of the chewing and neck muscles. Death occurs with increasing lethargy after 1-2 days. Recovery is extremely rare.
Carnivorous animals experience food refusal, fearfulness, restlessness, and severe itching. Sometimes dogs and cats show signs of rabies. Then paralysis of the pharynx occurs. Death in 2-3 days. Animals are not the source of the virus and do not excrete it, being an ecological dead end.
Aujeszky's disease can be suspected based on characteristic clinical symptoms and pathological changes (clinical, epizootological and pathological diagnostics).
Material for research: swabs from the nasal cavity and blood (preferably paired serum), from corpses - pieces of the brain, lungs, liver, spleen.
Express method - detection of viral antigen in RIF. Virological method: a) isolation of the virus on a culture of piglet kidney cells: b) bioassay on rabbits (characteristic itching and scratching at the site of infection).
Identification: RIF, RN.
Retrospective diagnosis: based on the increase in antibody titer in paired serum samples.
It is necessary to differentiate Aujeszky's disease from rabies, swine fever, influenza, erysipelas, and table salt poisoning.
Live VGNKI virus vaccine and inactivated cultural vaccine are used - immunity for 6-10 months. Subunit and recombinant vaccines are used abroad.
QUESTION No. 21 “SIGNIFICANCE AND FEATURES OF VIRAL PROTEINS.”
See question number 7
QUESTION No. 22 “GENERAL PRINCIPLES OF SEROLOGICAL REACTIONS AND THEIR USE IN THE DIAGNOSIS OF VIRAL DISEASES.”
In order to determine the type of a given virus, serological methods are used when studying protective processes in the body of a sick person or infected animal. Serology (from the Latin Serum - serum, liquid component of blood) is a branch of immunology that studies antigen reactions with specific protective substances, antibodies, that are found in blood serum. Antibodies neutralize the effect of the virus. They bind to certain antigenic substances located on the surface of viral particles. As a result of the binding of antibody molecules to the surface structure of the virus, the latter loses its pathogenic properties. To establish the level (quantity) of antibodies in the serum or determine the type of a given virus, a virus neutralization reaction is performed. It can be carried out both in animals and in cell culture.
The minimum concentration of serum containing antibodies sufficient to neutralize the virus and prevent it from exhibiting CPE is called the titer of serum neutralizing the virus. This concentration can also be detected using the plaque method.
To detect antibodies, the method of inhibiting hemagglutination (the gluing of red blood cells under the influence of a virus) and the method of complement fixation are used. Of the methods used in virology for various research purposes, we can also mention the methods by which virological material is prepared for physical and chemical analyzes that facilitate the study of the fine structure and composition of viruses. These tests require large quantities of completely pure virus. Virus purification is a process in which all foreign particles that contaminate it are eliminated from a suspension with a virus. These are mainly pieces and “fragments” of host cells. Simultaneously with purification, thickening of the suspension usually occurs, increasing the concentration of the virus. This provides the source material for many studies.
Using a serological reaction, you can: determine the antibody titer to the hemagglutinating virus in serum; identify an unknown hemagglutinating virus from known sera; establish the degree of antigen relatedness of 2 viruses, determine the titer of virus-neutralizing antibodies in serum, or the neutralization index, identify an unknown virus by testing it with various known sera.
Serological reactions.
1. RIF – immunofluorescence reaction.
AG + AT labeled with fluorochrome. Contact is allowed for 30 minutes at 37 C, then thoroughly washed in saline solution. Detection method: fluorescent light under a microscope.
2. ELISA – enzyme-linked immunosorbent assay.
AG + AT with enzyme. Contact, wash, then add a substrate, which upon contact with the AT-enzyme complex gives a color reaction.
3. RSK – complement fixation reaction.
AG + AT + complement. Contact. Then the heme system (hemolysin + sheep red blood cells) is added. Contact. If hemolysis does not occur, then AG and AT have fixed complement. Delayed hemolysis is a positive reaction. If hemolysis occurs, then complement is bound by the heme system - the reaction is negative.
4. RDP – diffuse precipitation reaction.
AG + AT (diffusion in agar gel). The detection method is the formation of a precipitation contour.
5. RNHA – indirect hemagglutination reaction.
Erythrocytes are loaded with antigen and when the antigen-antigen complex is formed, agglutination of erythrocytes occurs.
6. RTGA – hamagglutination inhibition reaction
7. RTGAd – hemadsorption inhibition reaction
8. RN – neutralization reaction.
Virus + AT. Contact. Entering a virus-sensitive system. The detection method is to neutralize the infectious activity of the virus.
QUESTION No. 23, 25 “RTGA AND ITS USE IN VIRUSOLOGY. ADVANTAGES AND DISADVANTAGES."
One of the simplest serological reactions is the hemagglutination inhibition reaction. It is based on the fact that antibodies, when encountering a homologous antigen, neutralize not only its infectious, but also hemagglutinating activity, because block virion receptors responsible for hemagglutination, forming an “AG + AT” complex with them. The principle of RTGA is that equal volumes of blood serum and virus suspension are mixed in a test tube and, after exposure, it is determined whether the virus is preserved in the mixture by adding a suspension of red blood cells. Agglutination of erythrocytes indicates the presence, and the absence of hemagglutination indicates the absence of virus in the mixture. The disappearance of the virus from the virus + serum mixture is regarded as a sign of interaction between serum and virus ATs. RTGA allows you to solve the following problems: determine the titer of antibodies to the hemagglutinating virus in serum; identify an unknown hemagglutinating virus from known sera; establish the degree of AG relationship between the two viruses. Advantages of RTGA: simplicity of technique, speed, no sterile work required, specificity, low cost. Disadvantage of RTGA: only possible with hemagglutinating viruses.
The principle of AT titration in RTGA is as follows: prepare a series of successive (usually 2-fold) dilutions of the test serum in equal volumes (usually 0.25 or 0.2 ml); to each dilution add the same volumes of homologous virus in a titer of 4 HAE; the mixtures are kept for a certain time at a certain temperature, equal volumes of a 1% suspension of washed erythrocytes are added to all mixtures; After exposure, hemagglutination in each mixture is assessed in crosses.
QUESTION No. 26 “RDP. IMMUNOLOGICAL BASIS OF THE METHOD, STATEMENT AND ACCOUNTING OF RESULTS. ADVANTAGES AND DISADVANTAGES."
RDP in the gel is based on the ability to diffusion in gels of AT and soluble AG and the absence of such ability in the “AG + AT” complex. This complex is formed upon contact of homologous AG and AT diffusing towards each other. It is deposited at the site of formation in the thickness of the gel in the form of a precipitation band. Starch, gelatin, agar-agar and more are used as gels. Agar gel is often used in laboratory practice. Serum antibodies are Ig molecules, which, despite their rather large size. Able to diffuse in agar gel. Viral Ags are viral proteins. They can be found in virions, representing so-called corpuscular AGs. Large sizes that do not allow them to diffuse in the agar gel. But viral proteins can also be in the form of free molecules formed as a result of the destruction of virions and (or) the destruction of the cells in which they were formed. These are soluble antigens. They are capable of diffusion in an agar gel. The technique for setting up RDP in a gel is to make several depressions in a layer of agar gel and pour AG and serum into them. So that AG and serum are in adjacent wells. From the wells, AG and serum begin to diffuse into the gel layer. Diffusion is directed in all directions from each hole. In the space between the wells containing AG and serum, the latter diffuse towards each other. If they turn out to be homologous, then an “AG + AT” complex is formed, which is not capable of diffusion due to its larger size. It settles at the site of formation in the form of a whitish precipitation streak. RDP solves the following problems: 1) detection of antibodies homologous to antigens in blood serum; 2) detection in the material of an antigen homologous to known serum antibodies; 3) identification of an unknown virus; 4) titration of serum AT. Here, the highest serum dilution, which still gives precipitation with homologous antigen, serves as an indicator of the antibody titer in the serum. RDP is often used to diagnose bovine leukemia and equine infectious anemia. The reaction can be carried out in Petri dishes, on glass slides, or capillaries (rarely). To carry out RDP on glass slides, you need: defatted glass slides, graduated pipettes (2-5 ml), Pasteur pipettes; a tube with a diameter of 5 mm or a stamp, a wet chamber, a tool for extracting gel, agar, AG, serum from the wells. Setting up the RDP: The glass slides are placed on a cold surface. Pour agar from a pipette (layer 1.5-2 mm), allow to cool for 5-10 minutes. Holes are cut out and soldered. The RDP components are poured into the wells and placed in a humid chamber (where they are left at room temperature or placed in a thermostat). The RDP preparation on glass slides can be dried after 48-72 hours and stained with amide black solution. This allows the preparation to be stored indefinitely and improves the ability to photograph precipitation bands. Advantages of RDP: simplicity of the technique, quick response, undemanding to the purity of the components, no sterile work required, minimal need for components, suitability for working with any soluble antigens, the ability to document the result by photographing. Disadvantages of RDP: low sensitivity. The reaction is used to detect rabies viruses, infectious bovine rhinotracheitis, African swine fever, canine fever, and others in pathological material; And also for the identification of equine infectious anemia viruses, adenoviruses, respiratory syncytial virus, bovine diarrhea virus, for the detection in blood serum of antibodies to equine infectious anemia viruses, bovine respiratory syncytial virus and in many other cases.
QUESTION No. 27 “RSK. IMMUNOLOGICAL BASIS AND CHARACTERISTICS OF REACTION COMPONENTS.”
The complement fixation test (FFR) is one of the traditional serological tests used to diagnose many viral diseases. The name itself largely reflects the essence of the method, which consists of two separate stages. The first stage involves an antigen and an antibody (one of these ingredients is known in advance), as well as a certain amount of pre-titrated complement. If the antigen and antibody match, their complex binds complement, which is detected at the second stage using an indicator system (a mixture of sheep red blood cells and antiserum to them - hemolysin). If complement is bound by the interaction of antigen and antibody, then lysis of red blood cells does not occur (positive RBC). With negative RSC, unbound complement promotes hemolysis of erythrocytes (Fig. 80).
RSCs are often used in diagnostic practice for the detection and identification of viruses, detection and titration of antibodies in blood serum.
The main components of RSC are antigens (known or detectable), antibodies (known antisera or test sera), complement, hemolytic serum and sheep red blood cells; Isotonic sodium chloride solution (pH 7.2-7.4) or various buffer solutions are used as a diluent. Antigens and serums may have anti-complementarity, i.e. the ability to adsorb complement, which delays hemolysis and distorts the results of the reaction. To get rid of anticomplementarity, antigens are purified by various methods: acetone, freon, ether, chloroform, etc., depending on the type of tissue used as the antigen and the virus. Serums are freed from anticomplementarity by heating, treating complement and other methods.
Antigens for CSC are prepared from the organs of infected animals, from the allantoic or amniotic fluid of infected chicken embryos, as well as from the liquid medium of infected cell cultures.
differs significantly from its preparation for bacterial infections. This is due to a number of specific properties of viruses.
First, to release viral antigen from a cell, it is often necessary to further process the infectious material in order to destroy the cells and release the antigen.
Secondly, the greater thermolability of viral antigens compared to bacterial ones. In most viruses, the complement-fixing antigen is associated with the infectious particle, and its destruction occurs in parallel with the loss of the infectious one. Therefore, materials for obtaining the antigen must be taken from dead animals only in the first hours after their death, or better yet, during life. Preservation of virus-containing material with various disinfectants often does not give positive results, since many of them cause the destruction of the viral antigen.
Thirdly, the unevenness of complement fixation when they are worn differently; with an excess of antibodies, complement fixation sharply decreases, since the active antigen + antibody complex is presented mainly in the form of antibodies and the active complement surface is insignificant. The same is observed in the zone of excess antigen, where the suppression of complement fixation occurs even faster. Therefore, to establish the optimal zone of complement fixation, preliminary titration of antigen and antibodies is necessary.
Fourthly, the volume of the antigen + antibody complex is insignificant. The size of the viral particles entering the complex is very small, and therefore the area of complement fixation is insignificant. With an increase in the volume of the antigen + antibody complex by lengthening the period of complement fixation (up to 18 hours at 4 °C), the sensitivity of the reaction increases, but its specificity decreases, since with a long period of fixation, the fixation of complement by nonspecific antigens (tissue) increases.
And finally, fifthly, the high complementary activity of the viral antigen. To exclude nonspecific complement fixation, more complete purification of the viral antigen from tissue fragments is necessary.
A big obstacle to the use of RSC in the diagnosis of viral diseases of animals and humans is the uneven accumulation of viral antigen during different periods of the disease and especially during different infections.
RSC is used to determine the types and subtypes (variants) of the foot-and-mouth disease virus that cause disease in animals, to test production strains of the foot-and-mouth disease virus in the manufacture of vaccines and laboratory strains in research work.
QUESTION No. 28 “TITER OF VIRUSES AND PRINCIPLES OF ITS DETERMINATION IN UNITS OF 50% INFECTIOUS ACTION.”
The titer is the amount of virus contained in a unit volume of material. Of the local damage caused by viruses, the best known are plaques and pockmarks on the XAO EC. If there is evidence to the contrary, the infectious activity of the virus can be measured in plaque-forming units (PFU) or pox-forming units (PFU) 1 PFU = dose of virus capable of causing the formation of one plaque, and one PFU - one pock. Methods: several CCs or ECs are infected at the HAO. The arithmetic mean number of pockmarks or plaques is calculated. It = PFU or OFU of the virus. Calculate how many PFU or PFU are per unit volume of virus-containing material. This is the title. T=n/Va, where n is the arithmetic mean of plaques or pockmarks, and is the dilution of the material, V is the administered dose. Method of 50% infectious action. One unit of the amount of virus is a dose that can cause an infectious effect in 50% of those infected. The number of such doses per unit of material will express the titer of the virus in this material. A 10-fold dilution of the test material is prepared, then equal groups of living test objects are infected with equal doses. They take into account the result of the action and find in what dilution the virus showed its effect by 50%. If such a dilution is not immediately found, then it is calculated using the formula T=lgB – (b-50)/(b-a) *lgd, where B is the dilution giving an infectious effect of more than 50%, b is the percentage giving an infectious effect of more than 50%, a – less than 50% d – dilution factor. 1HAE is taken to be a dose of the virus that is capable of agglutinating approximately 50% of the red blood cells contained in the same volume as the virus, 1% of a suspension of washed red blood cells. A series of successive multiple dilutions of the material are prepared and a 1% suspension is added to each dilution. The reaction is scored in crosses. The 2-cross reaction contains 1GAE, which is multiplied by the dilution factor.
QUESTION No. 29 “BIOLOGICAL CHARACTERISTICS OF FOOT-AND-MOUTH VIRUS. DIAGNOSTIC PRINCIPLE"
Foot and mouth disease is an acute, highly contagious disease of artiodactyls, manifested by fever, vesicular lesions of the mucous membranes of the mouth, skin of the corolla and udder, in young animals, damage to the mucous membranes of the mouth, skin of the corolla and udder, and in young animals, damage to the myocardium and skeletal muscles. Foot and mouth disease is recorded in many countries around the world. The incubation period lasts 1-3 days. Sometimes up to 7-10 days. The most characteristic sign of this disease in animals is vesicular lesions of the mucous membranes of the mouth and the skin of the corolla and udder. In cattle - it is acute, benign in adults. Initially, a deterioration in appetite, increased salivation, and an increase in body temperature are noted. On the 2-3 day, aphthae appear on the inner surface of the lips and tongue (in some, in the area of the interhoof gap, on the udder). After a day, erosions form. After 2-3 weeks, the erosions heal and the animal recovers. The virus belongs to the family Picornaviridae, genus Aphthovirus, RNA-containing, does not have a supercapsid shell. Virions are small particles of icosahedral shape. The virus is quite resistant to environmental influences. Domestic and wild artiodactyls are susceptible. The virus can be isolated already during the incubation period. Relapse may be accompanied by prolonged viral carriage. About 50% of recovered cattle can shed the virus for 8 months, and some for up to 2 years. The virus is cultivated on naturally susceptible and laboratory animals: newborn mice, rabbits and guinea pigs. Proliferates well in bud cells. It does not have hemagglutinating properties. There are 7 known types of FMD: A, O, C, Sat-1, Sat-2, Sat-3, Asia-1. In the body of naturally susceptible animals, the virus induces the formation of virus-neutralizing, complement-binding and precipitating antibodies.
Foot and mouth disease virus is usually determined in the RSC. The main components of RSC are AG, AT, complement, hemolytic serum and sheep erythrocytes; ICN or various buffer solutions are used as a diluent. AG and serum may have anticomplementarity - the ability to adsorb complement, which delays hemolysis and distorts the results of the reaction. To get rid of anticomplementarity, AG is purified using various methods: acetone, freon, ether, chloroform, depending on the type of tissue used as AG and virus. AG for RSC is prepared from the organs of infected animals, from the allantoic and amniotic fluid of infected EC, as well as from the liquid medium of infected EC. RSC is used to determine the types and subtypes of foot-and-mouth disease virus that cause disease in animals, to test production strains of foot-and-mouth disease virus in the manufacture of vaccines and laboratory strains in research work.
QUESTION No. 30 “LUMINESTENCE MICROSCOPY. BASICS OF IMMUNOFLUORESCENCE".
The method is based on the phenomenon of luminescence, the essence of which is that by absorbing various types of energy (light, electrical), atoms of certain substances go into an excited state, and then, returning to their original state, release the absorbed energy in the form of light radiation. Luminescence is observed in the form of fluorescence - a glow that occurs at the moment of irradiation with exciting light and stops immediately after its completion. Phosphorescence is a glow that continues for a long time even after the end of the excitation process.
QUESTION No. 31 “RABIS VIRUS, ITS PROPERTIES. PATHOGENICITY. PRINCIPLES OF DIAGNOSTICS".
Rabies is an acute infectious disease that occurs with severe damage to the nervous system, usually with a fatal outcome. Humans and all mammals are susceptible. Rabies is widespread. The pathogen is transmitted by dogs, cats, wild rodents and predators, as well as blood-sucking vampire bats. The duration of the incubation period depends on the location, strength of the bite, the amount and virulence of the virus that has entered the wound, and the resistance of the bitten animal. The incubation period lasts from 1-3 weeks to a year or more. The disease is acute. Clinical signs of an atypical course are loss of appetite, rumen atony, pharyngeal paralysis, drooling. There can also be a violent and quiet course of the disease. The rabies virus (RV) has pronounced neuroprobasia. Penetrating from the periphery along the nerve trunks into the central nervous system centripetally, it spreads in the body centrifugally along the peripheral nerves and enters various organs, including the salivary glands.
The virus belongs to the family Rhabdoviridae, genus Lyssavirus. Virions have the shape of a rod with a chopped end. The virus virion is RNA-containing with a helical type of symmetry and has a lipoprotein envelope. Low temperatures preserve the virus. The VB virion contains glycoprotein and nucleocapsid Ag. The first induces the formation of virus-neutralizing antibodies, and the second – complement-fixing and precipitating antibodies. In the body, the virus is localized mainly in the central nervous system, in the salivary glands, and saliva. Cultivated in mice, rabbits, guinea pigs, and in primary cell cultures. Reproduction of the virus in CC does not always manifest itself as CPD. The sources of infection are sick animals. They transmit the virus through a bite. The diagnosis of rabies is made on the basis of epidemiological, clinical data and laboratory test results, which are of critical importance. For research, fresh corpses of small animals are sent to the laboratory as a whole, and from large and medium-sized animals - the head with 2 cervical vertebrae. The corpses of small animals are treated with insecticides before being sent for research. Laboratory diagnostics include: detection of viral hypertension in RIF and RDP, Babes-Negri bodies and bioassays on white mice. RIF - for this reaction, the bioindustry produces fluorescent anti-rabies gamma globulin. Principle – 1) Make prints or smears from various sections of the left and right side of the GM on glass slides (at least 2 preparations from each section); 2) They are dried and fixed in chilled acetone; 3) Dry, apply fluorescent gamma globulin; 4) Place in a humid chamber; 5) I thoroughly wash the ICN, rinse it with water, dry it in air, apply non-fluorescent immersion oil and view it under a fluorescent microscope. In preparations containing antigen WB, yellow-green fluorescent granules of different sizes and shapes are observed in neurons, but more often outside cells. RDP – 1) Agar gel is poured onto glass slides 2) Wells are made (D = 4-5 mm); 3) The wells are filled with a paste-like mass from the GM sections. 4) Controls with “+” and “-“ AG are placed on a separate glass using the same stencil; 5) After filling the wells, the preparations are placed in a humid chamber and placed in a thermostat at 37C for 6 hours, then at room temperature for 18 hours. The reaction is considered positive when one or 2-3 lines of precipitation of any intensity appear between the wells containing the brain suspension and rabies gamma globulin. DETECTION OF BODIES - thin smears or imprints are made on glass slides from all parts of the GM and stained according to Sellers or Muromtsev or Mann or Lenz. BIOPEST - white mice (16-20 grams) are selected, the nervous tissue from all parts of the GM is ground in a mortar with sterile sand, ICN is added to a 10% suspension, left for 30-40 minutes and the supernatant is used for infection to infect the pups. Infect 10-12 pieces: half intracerebrally with 0.03 ml, half subcutaneously in the area of the nose or in the upper lip with 0.1-0.2 ml. Observed for 30 days. In the presence of VB in the pathological material, from 7-10 days after infection, symptoms are observed in mice: ruffled fur, a peculiar hunchback of the back, impaired coordination of movements, paralysis of the hind, then forelimbs and death. In dead mice, the GM is examined in the RIF for the detection of Babes-Negri bodies and a RDP is placed. A bioassay for rabies is considered positive if Babes-Negri bodies are detected in preparations from the brains of infected mice or if AG is detected using RIF or RDP methods. A negative diagnosis is the absence of death of mice within 30 days.
QUESTION No. 32 “MODERN CLASSIFICATION OF IMMUNITY. AT STRUCTURE CHARACTERISTICS OF DIFFERENT CLASSES OF IMMUNOGLOBULINS AND THEIR STRUCTURE.”
Immunity is a state of the body's immunity to the effects of pathogenic microbes, their toxins and other foreign substances of biological nature.
The body's immune system is a system of organs and cells that responds against foreign substances.
Innate immunity is immunity to infectious agents, located in the genome and manifested by the number and order of arrangement of gangliosides of a certain type on the surface of cell membranes. It is very durable, but not absolute.
Acquired immunity is the body’s resistance only to a specific pathogen. This immunity is divided into natural and artificial. Natural is divided into 1. active - it is formed after the animal has naturally recovered from the disease, sometimes after exposure to repeated small doses of the pathogen (immunizing subinfection). 2.passive – immunity of newborns acquired due to the receipt of antibodies to the fetus from the mother through the placenta or after birth through the intestines with colostrum. There are natural and artificial colostral immunity; in the first case, immunity arises due to antibodies naturally produced in the mother’s body under the influence of various environmental antigens. In the second case, through targeted immunization of the mother’s body. Naturally acquired active immunity can last 2 years, sometimes for life; artificially acquired immunity can provide a state of immunity from several weeks to several months.
Artificially acquired immunity is also divided into 1.active - occurs as a result of immunization of animals with vaccines (develops after 7-14 days and lasts up to several months to 1 year or more) and passive - is created when immune serum containing specific antibodies against a specific pathogen.
There are also types of immunities: 1. Antibacterial immunity - protective mechanisms are directed against the pathogenic microbe. 2. Antiviral - the body produces antiviral antibodies. 3. Antitoxic immunity - during the formation of which bacteria are not destroyed, but antibodies are produced that effectively neutralize toxins in the patient’s body.
4.Local immunity. 5. Sterile immunity - if after an illness the body is freed from the pathogen, while maintaining a state of immunity. 6. Non-sterile - when immunity is maintained only as long as the pathogen is in the body. 7. Humoral immunity - the production of specific antibodies in the infected body. 8. Cellular – ensured by the formation of T-lymphocytes that specifically react with the pathogen.
Nonspecific factors of the body's defense.
They act as the first protective barrier and do not need to be rebuilt.
The skin is a powerful barrier to the penetration of microorganisms, and mechanical factors are important.
Mucous membranes - in the respiratory tract with the help of ciliated epithelium (moves a film of mucus along with microorganisms towards natural openings), in the mouth to the nasal passages (coughing and sneezing). These membranes secrete secretions that have bactericidal properties, in particular due to lysozyme and IgA. The secretions of the digestive tract have the ability to neutralize many pathogenic microbes. Saliva contains lysozyme, amylase, and phosphatase. Bile causes the death of pasteurella. The intestinal mucosa contains powerful antimicrobial factors.
Lymph nodes - inflammation develops in them, in its zone microbes are fixed by fibrin threads. The complement system and endogenous mediators are involved in inflammation.
Phagocytosis is the process of active absorption by the cells of the body of pathogenic living or dead microbes and other foreign particles entering it, followed by digestion with the help of enzymes.
Antibodies can exist in millions of varieties, each with its own unique antigen binding site. Collectively called immunoglobulin (Ig), AT proteins form one of the major classes of blood proteins, accounting for approximately 20% of total plasma protein by weight. When Ag binds to the membrane antigen-specific receptors of the B cell, cell proliferation and differentiation occurs to form cells that secrete Ab. ATs have 2 identical Ag-binding sites. The simplest AT molecules are schematically shaped like the letter gamma with two identical Ag-binding sites, one at the end of each of the two “branches.” Since there are 2 such sites, these ATs are called bivalent. The protective effect of antigens is not simply explained by their ability to bind antigens. They also perform a number of other functions in which the “tail” is involved; they are called effector functions and are determined not by the participation of the “tail” in them, but by the structure of the Fc fragment. This region of the molecule determines what will happen to the AG if it is bound. Antibodies with the same antigen-binding regions can have very different “tail” regions, and therefore different functional properties. The Ig G, D, E and serum IgA molecule consists of 4 polypeptide chains - 2 light and 2 heavy. In higher vertebrates, there are 5 different classes of antibodies - IgA, IgD, IgE, IgG, IgM, each with its own class of heavy chains. IgG antibodies constitute the main class of Ig found in the blood. They are produced in large quantities during the secondary response and are the only antibodies that can pass from mother to fetus. This is the predominant class of antibodies produced in most secondary immune responses; in the early stages of the primary immune response, mainly IgM antibodies enter the blood - they are also the first class of antibodies produced by developing B cells. IgA is the main class of antibodies in milk secretions, saliva, tears, secretions of the respiratory tract and intestinal tract. ATs protect vertebrates from infections by inactivating viruses, mobilizing complement and various cells that kill and engulf invading MOs.
QUESTION No. 33 “PECULIARITIES OF ANTI-VIRAL IMMUNITY.”
1. Antiviral immunity is associated with unique protective mechanisms, because Viruses are unable to develop and reproduce in a non-living cell. The body's protective adaptation is aimed at 2 forms of existence of the virus. On the extracellular viral nonspecific and specific immunity factors, on the intracellular form - the process of phagocytosis. During viral infections, it is always incomplete, interferon has an exogenous effect on the extracellular form, viruses lose their ability to adsorb, endogenous interferon is synthesized in cells in response to viral Ag.
2.Means and methods of influencing viruses can be effective only at certain stages of the virus’s existence, which is most clearly manifested when treating patients with immune drugs, because Abs are not able to penetrate into cells.
3. Antiviral immunity is longer lasting than bacterial immunity, and in some viral infections it is lifelong (rinder, canine, bluetongue, smallpox).
QUESTION No. 34 “ROLE OF LYMPHOID CELLS IN ANTI-VIRAL IMMUNITY (CHARACTERISTICS OF T AND B LYMPHOCYTES).”
T lymphocytes. Thymus-dependent lymphocytes are formed from stem cells of hematopoietic tissue. The precursors of T-lymphocytes enter the thymus, undergo differentiation in it and emerge as cells with various functions, bearing characteristic markers. There are several subpopulations of T lymphocytes depending on their biological properties.
T helper cells (helpers) belong to the category of regulatory support cells. Stimulate the proliferation of B lymphocytes and differentiation into antibody-forming cells (plasma cells). It has been established that the response of B lymphocytes to the influence of most protein antigens is completely dependent on the help of T helper cells, which is carried out in two ways. The first requires direct action of a helper T cell and a responding B cell. It is believed that the T cell recognizes the determinants of the antigenic molecule that is already fixed on the B cell by cell receptors: In the second case, the helper function of T cells in activating B lymphocytes can also be carried out through the formation of soluble nonspecific helper factors - lymphokines (cytokines).
T-killers (killers) perform effector functions, carrying out cellular forms of the immune response. They recognize and lyse cells on the surface of which there are antigens foreign to a given organism (tumor, viral and histocompatibility). Proliferation and differentiation of T-killers occurs with the participation of T-helpers, whose action is carried out mainly with the help of soluble factors, in particular interleikin. It has been established that killer T cells carry out a delayed-type hypersensitivity reaction.
T-y s i l and t e l and activate the immune response within the T-subsystem of immunity, and T-helpers provide the opportunity for its development in the B-link of immunity in response to thymus-dependent antigens.
T-suppressors (suppressors) provide internal self-regulation of the immune system in two ways: suppressor cells limit the immune response to antigens; prevent the development of autoimmune reactions. T-suppressors inhibit the production of antibodies and the development of delayed-type hypersensitivity; the formation of T-killers ensures the formation and maintenance of immunological tolerance.
Immune memory T cells provide a secondary type of immune response in the event of repeated contact of the body with this antigen. Antigen-binding receptors and Fe receptors, IgA or IgM are found on the membranes of T cells. Null lymphocytes do not have distinctive markers of T and B lymphocytes. They are capable of carrying out antibody-dependent, complement-free, lysis of target cells in the presence of antibodies specific against these cells. K lymphocytes are a type of null lymphocyte. For them, the target cells are tumor cells, T- and B-lymphocytes modified by viruses, monocytes, fibroblasts, and erythrocytes.
B lymphocytes. Like T-lymphocytes, they are formed from stem cells of hematopoietic tissue. The precursors of B lymphocytes in the bursa of Fabricius undergo differentiation and then migrate to the lymph nodes and spleen, where they perform their specific functions.
The presence of two classes of B cells has been established: B-effectors and B-regulators. The effector cells of B-lymphocytes are antibody-forming cells (plasma), synthesizing antibodies of one specificity, i.e., against one antigenic determinant. B-regulators, in turn, are divided into suppressors and amplifiers (amplifiers). The function of the regulators is to release mediators that inhibit DNA production in T and B lymphocytes only within the bone marrow, as well as to enhance B effectors. B lymphocytes are larger than T lymphocytes (8 and 5 µm, respectively). Thanks to electron microscopy, it was found that the surface of B-lymphocytes is covered with numerous villi and folded, and the surface of T-lymphocytes is smooth.
QUESTION No. 35 “ROLE OF CELLULAR FACTORS IN ANTI-VIRAL IMMUNITY.”
It differs from humoral in that the effector elements of cellular immunity are T-lymphocytes, and humoral - plasma cells. It is of particular importance for infections caused by many viruses, bacteria, and fungi.
Formation of cytotoxic T cells (TCC) - among cell surface Ags that can cause the formation of the TTC cycle - MHC products (mononuclear system), viruses, tumor-specific Ags. TCA cycles have receptors through which antigen binds and processes that trigger cell lysis are triggered. The lytic activity of T cells begins with a close interaction between the killer cell and the target cell, a change in the membrane permeability of the target cell occurs, ending with the rupture of the cell membrane.
PCs have the ability to directly lyse a wide range of target cells, especially tumor cells; they can lyse cells regardless of MHC products (interferon and IL-2 enhance the lytic activity of PCs).
HRT is a T-cell dependent immunological response that manifests itself as inflammation at the site of Ag entry into the body, usually the skin. Lymphocytes that can tolerate HRT are T-cells and are called TGRT lymphocytes (they can be activated and respond to protein Ags, alloantigens, tumor antigens, to Ags of viruses, bacteria, fungi, protozoa.
Macrophages play a major role in cellular immunity. When pathogens multiply inside phagocytes, intracellular destruction occurs only after macrophages receive a stimulus from specially sensitized T lymphocytes. T lymphocytes activate macrophages by releasing lymphokines.
QUESTION No. 36 “ROLE OF HUMORAL FACTORS IN ANTI-VIRAL IMMUNITY”
In addition to AT - a specific factor of antiviral immunity - the body produces special virus-tropic substances - inhibitors that can interact with viruses and suppress their activity. Serum inhibitors have a wide range of action: some suppress the hemagglutinating properties of viruses, others suppress their cytopathogenic effect, and others suppress their infectious activity. Heat-labile inhibitors are found in normal human and animal sera. They have a wide range of virus-neutralizing effects, are able to block the hemagglutinating activity of influenza viruses, New Castle disease, measles, arboviruses and others and neutralize the infectious and immunogenic properties of inhibitor-sensitive viruses. Heat-stable gamma inhibitors are highly active against modern variants of the influenza virus. Heat-stable alpha inhibitors block the hemagglutinating, but not the infectious, activity of the virus.
QUESTION No. 37 “ANTIVIRAL AT, THEIR PROPERTIES, BIOLOGICAL ROLE, DETECTION AND TITRATION METHODS.”
AT are proteins formed in the body upon parenteral administration of high-molecular substances with signs of genetic foreignness for the given organism. Antibodies are capable of interacting with antigen in response to which it was formed and neutralizing its biological activity. The usual source of AT is blood serum. When encountering an antigen, AT neutralizes not only its infectious, but also its hemagglutinating activity, because blocks virion receptors responsible for hemagglutination, resulting in the formation of the “AG + AT” complex.
Antibodies can exist in millions of varieties, each with its own unique antigen binding site. Collectively called immunoglobulin (Ig), AT proteins form one of the major classes of blood proteins, accounting by weight for approximately 20% of total plasma protein. When Ag binds to the membrane antigen-specific receptors of the B cell, cell proliferation and differentiation occurs to form cells that secrete Ab. ATs have 2 identical Ag-binding sites. The simplest AT molecules are schematically shaped like the letter gamma with two identical Ag-binding sites, one at the end of each of the two “branches.” Since there are 2 such sites, these ATs are called bivalent. The protective effect of antigens is not simply explained by their ability to bind antigens. They also perform a number of other functions in which the “tail” is involved; they are called effector functions and are determined not by the participation of the “tail” in them, but by the structure of the Fc fragment. This region of the molecule determines what will happen to the AG if it is bound. Antibodies with the same antigen-binding regions can have very different “tail” regions, and therefore different functional properties. The Ig G, D, E and serum IgA molecule consists of 4 polypeptide chains - 2 light and 2 heavy. In higher vertebrates, there are 5 different classes of antibodies - IgA, IgD, IgE, IgG, IgM, each with its own class of heavy chains. IgG antibodies constitute the main class of Ig found in the blood. They are produced in large quantities during the secondary response and are the only antibodies that can pass from mother to fetus. This is the predominant class of antibodies produced in most secondary immune responses; in the early stages of the primary immune response, mainly IgM antibodies enter the blood - they are also the first class of antibodies produced by developing B cells. IgA is the main class of antibodies in milk secretions, saliva, tears, secretions of the respiratory tract and intestinal tract. ATs protect vertebrates from infections by inactivating viruses, mobilizing complement and various cells that kill and engulf
implemented MOs.
QUESTION No. 38 “INTERFERON AND ITS ROLE IN ANTI-VIRAL IMMUNITY.”
Human cells have 27 genetic loci for interferons (hereinafter referred to as I) – 14 are functional. And encoded in the genetic apparatus of the cell. There are alpha, beta, gamma - I. Its system does not have a central organ, all cells have the ability to synthesize it. For its formation, inducers are needed (viruses, bacterial toxins, extracts from bacteria and fungi, double-stranded RNA (the most effective) and others). Virus-infected I – alpha and beta; Gamma-I is formed under the influence of phytohemagglutinin with SEA. During induction of I, 2 or more of its types are synthesized. The most actively inducing viruses are myxo- and arboviruses. The interferonogenicity of viruses increases with a decrease in their virulence for the body. Inducers of a non-viral nature stimulate a more rapid and short-term accumulation of “heavy” I (with a high molecular weight) in the body. And can be obtained 4 hours after intravenous administration of Ig. And it does not affect adsorption, viropexis, deproteinization of virions, it suppresses the production of the virus. It does not act on any specific virus, but on many types in general. And it is able to enhance phagocytic activity (macrophages, when exposed to I, have significantly more vacuoles, attach more quickly to glass, and more actively capture bacteria). Interferon drugs inhibit cell growth and suppress the growth of tumor cells. And it inhibits AT formation and has a direct effect on B-lymphocytes. And it helps to increase the killer activity of T cells. Pre-treatment of cells or animals with small doses of And leads to an increase in the production of And in response to the last induction of its synthesis (priming). When processing producers, And increases in quantities of And, blocking is observed (the opposite effect). The production of I is influenced by external conditions (weather, air temperature). Ionizing radiation reduces I production. As the body grows, the amount of I inhibitors decreases. And young animals exhibit a reduced antiviral effect compared to And an adult animal, because the production of mononuclear phagocytes is reduced. During the formation of I in the cells of newborns, cathepsin D is activated and released from lysosomes, which leads to proteolytic degradation of I. As growth occurs, the components that contribute to the release of cathepsin D from lysosomes decrease. The most sensitive to I are viruses that have an outer shell and contain lipids (myxoviruses, smallpox group, arboviruses). For medical and veterinary purposes, inducers of endogenous I are mainly used, but exogenous I is also used. Like hormones, I-ins are secreted by some cells and transmit a specific signal through the intercellular space to other cells. And - “protein factor”, which does not have virus specificity and its antiviral activity is carried out with the participation of cellular metabolism, involving the synthesis of RNA and protein.
QUESTION No. 39 “PRINCIPLE OF OBTAINING BACTERIOPHAGES. DETERMINATION OF ACTIVITY AND PRACTICAL USE OF PHAGES."
The phage is obtained by adding a special phage to the MO culture, kept for 24 hours at a temperature of 37 degrees, filtered through bacterial filters, and the filtrate is checked for purity by inoculation; harmlessness and activity, phage titer.
Determination of phage activity.
Use qualitative and quantitative methods. The amount of phage is determined by titration on liquid or solid nutrient media. To do this, the phage is diluted tenfold. The same amount of daily bacterial broth culture is added to each dilution. Then they are placed in a thermostat and the result is taken into account. The titer is determined after separating the mixture on 1 day in a thermostat.
The phage titer is taken to be its highest dilution, which is capable of delaying the growth of MO. Expressed by the degree of its dilution. Only virulent phages cause complete destruction of the cell and the formation of phage particles.
QUESTION No. 40 “PASSIVE SPECIFIC PREVENTION OF VIRAL DISEASES. THE PRINCIPLE OF RECEIVING."
Preparations for passive IP– for parenteral and oral administration of AT or Ig. For the purpose of carrying out IP, immune, hyperimmune sera, convalescent and allogeneic sera are used.
Convalescent serum– serum from donors of recovered or infected animals. It is used when there are no more effective agents at a dose of 1 ml/kg body weight.
Hyperimmune serums– donor serum, which is obtained as a result of a single administration of massive doses of antigen according to a certain scheme. A healthy donor who has not previously suffered from this disease is selected. He is vaccinated and after 2-3 weeks they begin to administer it according to a certain scheme in increasing doses, bringing it to the peak of the increase in AT. The peak is determined by setting a serological reaction to the AT titer (serum is checked for sterility, activity and harmlessness. Dose 2 ml/kg (therapeutic), 1-1.5 ml/kg (prevention). Administered fractionally. First, a sensitized dose is administered, and after 2-3 hours is the permissive dose to avoid anaphylactic shock.
Allogeneic whey– combined whey, which is obtained from different animals in the same farm. It contains a large set of ATs and various AGs.
QUESTION No. 41 “SPECIFIC PREVENTION OF VIRAL DISEASES. TYPES OF VACCINES AND METHODS OF THEIR ADMINISTRATION".
1. In the practice of epizootology, an increase in the size and density of animal populations increases the risk of epizootics. The main principle in the fight against them is to break the infectious chain in all areas or stop the transition of the epizootic process to a latent state. One of the main tools for breaking the chain is timely prevention. For livestock farming, developing on an industrial basis, the fight against all factors, incl. with pathogenic MOs and viruses is one of the most important conditions for a healthy livestock. IP (immunoprevention), when properly included in the strategy to combat infectious diseases, significantly reduces the danger.
The goal of IP is not only the eradication of infectious diseases, but also the preservation of productivity, therefore it is necessary to strive to create vaccines that can provide a high degree of protection for the entire livestock immediately after vaccination, regardless of the age of the animals.
IP has a number of advantages:
1. The principle of action of IP is based on a specific change in the animal’s body in the direction of maximally reducing the possibility for the pathogen to cause an infectious disease.
2.IP acts continuously and for a long time, sometimes throughout life.
3.IP not only changes the reactivity of the animal’s body, but also increases the ability to immune defense in the entire livestock.
4.The effect of IP on the epizootic process can be accurately calculated.
5. With appropriate selection of vaccination moments, PIs provide maximum protection during the most dangerous periods of life for infection.
6.IP can be linked to the technological process in animal husbandry.
7. The drugs used for IP can be dosed, used in different combinations and standardized.
8. Unlike AB and chemical drugs, PI does not cause resistance in MO.
9. IP requires lower economic costs and costs of raw materials.
10. IP does not have any impact on the quality of animal products.
Negatives:
1.Evaluation of the individual entrepreneur’s capabilities. The owner of the animal is often convinced that everything has already been done for protection with vaccination, which leads to a weakening of sanitary and hygienic measures.
2.Too large increase in the final cost of production.
3. Post-vaccination reactions, which for a certain time reduce productivity if an insufficiently tested vaccine is used.
4. Too frequent disturbance of animals, leading to a decrease in productivity.
5. The emergence of diagnostic problems and increasing difficulties in the fight against diseases if vaccine and pathogenic strains under normal conditions do not differ or are distinguished with great difficulty.
The inappropriate use of vaccines can cause harm, therefore, for each specific infectious disease and epizootic situation, it is necessary to carefully select a vaccine and the option for its use, taking into account economic costs and effectiveness, in order to ensure the highest result of mass vaccinations.
Immunoprophylaxis was developed on the basis of the long-standing experience of mankind, according to which people who had infectious diseases did not become ill with them a second time. Back when there was human plague in Athens. Thucydides reported that the sick were left without help if they were not cared for by people who were recovering. In China in the 16th century, when dealing with human smallpox, there was a custom: to inhale dried crushed smallpox crusts through the nose. Jenner invented the smallpox vaccine. Pasteur proposed a method of vaccination against rabies.
Prevention of viral diseases is based on the same principles as the prevention of other infectious diseases:
1. Carrying out organizational activities.
3. Chemoprophylaxis.
Specific prevention of viral diseases is ensured by the use of live, inactivated, poly- and monovalent sera.
Classification and characteristics of immunopreparations:
Biological products are products of biological origin used for active and passive IP.
Preparations for passive IP – for parenteral and oral administration of AT or Ig. For the purpose of carrying out IP, immune, hyperimmune sera, convalescent and allogeneic sera are used.
Convalescent serum is serum from donors of recovered or infected animals. It is used when there are no more effective agents at a dose of 1 ml/kg body weight.
Hyperimmune sera are donor sera that are obtained as a result of a single injection of massive doses of antigen according to a certain scheme. A healthy donor who has not previously suffered from this disease is selected. He is vaccinated and after 2-3 weeks they begin to administer it according to a certain scheme in increasing doses, bringing it to the peak of the increase in AT. The peak is determined by setting a serological reaction to the AT titer (serum is checked for sterility, activity and harmlessness. Dose 2 ml/kg (therapeutic), 1-1.5 ml/kg (prevention). Administered fractionally. First, a sensitized dose is administered, and after 2-3 hours is the permissive dose to avoid anaphylactic shock.
Gamma globulins are obtained from hyperimmune sera by releasing ballast proteins. They are administered subcutaneously or intramuscularly at a dose of 0.5-2 ml/kg. First, sensitization is administered, then a resolving dose.
Allogeneic whey is a combined whey that is obtained from different animals in the same farm. It contains a large set of ATs and various AGs.
Preparations for active immunization - vaccines. There are live and inactivated vaccines.
Vaccines are also classified according to: 1) Initial virus-containing material - tissue, embryo-virus vaccines, cultured virus vaccines; 2) by the attenuation method - lapinized (against foot-and-mouth disease, rinderpest and others, use rabbits), caprinized (through the body of a goat, against sheep pox by passing through several goats, against cattle), ovinized (through sheep - against rinderpest, foot-and-mouth disease).
Vaccine administration methods:
1.Subcutaneously
2. Intramuscular
3. Aerosol
4.Rectal method
5.Intranasal
QUESTION No. 42 “INACTIVATED ANTI-VIRAL VACCINES, THEIR OBTAINING, PROPERTIES, APPLICATION, DIFFERENCE FROM LIVE VACCINES.”
Inactivated vaccines are complex preparations. Their production requires a large amount of virus. The main requirement for killed vaccines is complete and irreversible inactivation of the genome with maximum preservation of the antigen determinant and immune protection of vaccinated animals. To obtain inactivated vaccines, formalin, chloroform, thiomersal, hydroxylamine, ethanol, beta-propiolactone, ethyleneimine, UV and gamma irradiation, and temperature are widely used as inactivants. Inactivated vaccines are used only parenterally. They necessarily include adjuvants - nonspecific stimulators of immunogenesis. A larger dosage is required and, as a rule, repeated administration. They create less intense, short-lasting immunity than with live vaccines.
QUESTION No. 43 “FACTORS OF ANTI-VIRAL IMMUNITY, THEIR CHARACTERISTICS.”
Specific
1) Associated with qualitatively unique protective mechanisms, because viruses are not able to develop outside a living cell 2) Protection is aimed at 2 forms of nouns. virus: outside and inside cells. The resting form is affected by specific and nonspecific factors, humoral and cellular protective factors. Vegetative forms – interferon, which prevents the synthesis of viral mRNA. 3) Virus neutralizing antibody does not react with viral information NK. 4)Methods and means of neutralizing the virus are effective only at a certain stage. 5) Special protective factors: the formation of extracellular oxyphilic and basophilic granules and the presence of antiviral inhibitors. 6) This immunity is long-lasting, and sometimes lifelong.
Nonspecific cellular and general physiological reactions.
Temperature
Hormones reduce resistance, but somatotropic hormones increase resistance and enhance the inflammatory response.
A pregnant animal gets sick faster and the disease is more severe.
The physiological state of the excretory system is the rate of virus release from the body.
Humoral factors - the presence of serum inhibitors (heat-stable or heat-labile). Each species has its own predominant type.
QUESTION No. 44 “LIVE ANTI-VIRAL VACCINES, THEIR PROPERTIES, APPLICATION AND DIFFERENCES FROM INACTIVATED VACCINES.”
Live antiviral vaccines are lyophilized suspensions of vaccine strains of viruses grown in various biological systems (EC, CC, laboratory animals) or use naturally weakened strains of the pathogen that are created during a long-term epizootic. The main property is the persistent loss of the ability to cause a typical infectious disease in the body of a vaccinated animal; they also have the ability to “take root” in the animal’s body, that is, to multiply. The residence and reproduction of the vaccine strain usually lasts 5-10 days. up to several weeks and are not accompanied by clinical manifestations characteristic of this disease, lead to the formation of immunity against the infectious disease. Advantages: high intensity and duration of immunity they create, approaching post-infectious immunity. Possibility for most single administration. Administration is not only subcutaneous, but also orally and internally. Disadvantages: sensitivity to adverse factors. Strict storage and transportation limits - temperature - 4-8C. It is unacceptable to break the vacuum in vaccine ampoules. Strict adherence to asepsis rules. Quality control: 1) comprehensive examination of donors. 2) assessment of the quality of the nutrient medium and QC for sterility. 3)Supervision over the quality of production virus strains. 4) Creation of optimal conditions for the preservation of biomaterials.
Inactivated vaccines create less intense and long-lasting immunity; they must be administered repeatedly.
QUESTION No. 45 “BACTERIOPHAGES, THEIR IMPORTANCE AND BASIC PROPERTIES.”
Bacteriophages (from Lat. Bacteriophaga) – destroying bacteria. These are viruses that have the ability to penetrate bacterial cells, reproduce in them and cause their death.
The history of the discovery of the bacteriophage is associated with Academician Gamaley, who observed the accidental lysis of anthrax bacteria.
Tvort - described the degeneration of staphylococci (1915). D'Herelle (1917) studied in detail the interaction of the phage and bacteria of the dysentery bacillus and gave the agent the name “bacteriophage”. Subsequently, viruses of fungi, mycoplasmas and other microorganisms were isolated. Therefore, to designate these viruses, the term “phage” is used - eater.
Phage structure and morphology.
Phages consist of DNA/RNA nucleic acid surrounded by a capsid containing strictly oriented capsomers. Large phages have a tadpole-like structure, have a head, a collar and a tail process ending in a 6-gonal basal plate to which fibrils are attached. The head has 2 shells: an outer membrane and an inner membrane in which the AK is enclosed. The average size of the head is 60-100 nm, the tail is 100-200 nm. According to morphology, phages are divided into 6 groups:
Phages with a long process, the sheath of which contracts - T-even phages.
Phages with a long process, the sheath of which does not contract.
Phages with a process analogue.
Phages with a short process.
Filamentous phages.
Phages without a process.
Chemical composition of the phage.
The phage head contains one of the nucleic acids. The shell also contains lipids and carbohydrates. Phages can withstand pressure up to 6 thousand atmospheres. They are resistant to environmental influences and retain their activity in spare ampoules for up to 13 years.
They quickly die under the influence of boiling, UV light, and certain chemicals (1% phenol, alcohol, chloroform ether do not change the phage). Some substances: thymol, chloroform, dinitrophenol have no effect on phages, but kill bacteria.
A 1% formaldehyde solution inactivates the phage. Phages are distinguished: polyphages (lyse related bacteria), monophages (lyse related bacteria), monophages (lyse bacteria of the same species), phages that cause lysis of a specific serotype of the 1st species. Based on their type-specific properties, phages are divided into serotypes. Special phages can be easily adapted to related bacteria by riding on bacteria of the same species. The phenomenon of bacteriophagy can be easily observed in both liquid and solid nutrient media. If a culture is inoculated into a dish with a nutrient medium and a few drops of high concentration phage are applied, then there will be no growth at this place - sterile spots. According to the mechanism of interaction with cells, phages are divided into virulent and moderate.
The phenomenon of bacteriophagy caused by temperate phages manifests itself only in the form of phases of adsorption, penetration into cells, reproduction and release of the phage. The entire reproduction process follows the pattern of DNA viruses. Virulent phages ensure the formation of new phages and lysis of bacterial cells. It has been established that 7-8 phage particles appear in phage-infected bacteria within 1 minute.
Reproduction diagram.
1.Adsorption of phage on the MO shell and its dissolution. Phages are adsorbed by their flagella, these flagella firmly connect to the receptors of the cell wall, as a result of which the phage particle contracts and the end of the process penetrates the bacterial cell membrane and at the same time the phage secretes a lysozyme-like enzyme that dissolves the cell membrane.
2. Injection of nucleic acid into the microbial cell. All the nucleic acid and part of the proteins are injected into the microbial cell; the sheath remains on the surface of the bacterial cell.
3.Latent phase – eclipse phase. The phase promotes the development of DNA viruses. At the beginning, mRNA is synthesized, it gives rise to the synthesis of early viral proteins, which stop cellular metabolism and give rise to the formation of daughter nucleic acids.
4. Formation of new phage particles. The connection of two main phage particles by filling the phage protein shell with nucleic phage particles.
5. Dissolution of the bacterial cell membrane and the release of newly formed particles outside the cell. The rupture of the cell wall is facilitated by: a strong increase in intracellular pressure, and on the other hand, the action of enzymatic processes caused by phages. The number of perceived phages varies and ranges from 1 to 1000 or more.
The entire reproduction process takes from 3 to 10 hours.
Lysogeny - along with virulent phages, there are also temperate phages that differ in the nature of their interaction with the bacterial cell. Their main feature is that they are able to transform from a vegetative state into a non-infectious form called a prophage, which is unable to cause bacterial lysis. Bacterial cells containing a prophage on the chromosome are called lysogenic, and the phenomenon is lysogeny. In this phenomenon, bacteria infected with the phage are not lysed. But artificial lysis can release a phage that can infect bacteria of a given species. The transition of a prophage to a vegetative phage does not occur often. When infected with temperate phages, one part of the cells is lysed to form a vegetative phage, and the other part survives and becomes lysogenic.
In lysogenic bacteria, the phage DNA is integrated into the DNA of the cell and the temperate phage is converted into a prophage that does not have lytic properties.
Lysogenic bacteria formed as a result of lysogenization become carriers of the phage and acquire immunity for a long time. This connection is strong and is disrupted when the bacterium is exposed to inducing agents. These are UV rays, ionizing radiation, chemical mutagens. Under the influence of these factors, the prophage is transferred to an autonomous state and disintegration occurs.
Lysogenization of bacteria is accompanied by a change in their properties (morphological, cultural and biological properties). Non-toxic strains become toxigenic. Changing the properties of bacteria - phage conversion. Lysogenic bacteria are the most convenient models for studying the interaction of viruses and cells.
Currently, temperate phages are widely used to study questions of genetics, with the help of which it is possible to more accurately differentiate the processes of variability. Under the influence of radiation, the number of phage particles produced by the cells of lysogenic bacteria increases.
Practical use of phages - phages are used to titrate bacteria, treat and prevent a number of infectious diseases, and are used to determine the dose of radiation on spacecraft.
QUESTION No. 46 “LABORATORY ANIMALS, OBJECTIVES AND METHODS OF THEIR USE IN VIRUSOLOGY.”
Due to the fact that viruses can reproduce only in living cells, at the earliest stages of the development of virology, the cultivation of viruses in the body of laboratory animals, specially raised for research on them, was widely used.
Used: 1) to detect the virus in the PM 2) primary isolation of the virus from the PM 3) accumulation of the viral mass 4) maintaining the virus in the laboratory in an active state. 5) titration of the virus 6) as a test object in pH 6) obtaining hyperimmune sera. Animals used: white mice (rabies, foot and mouth disease), white rats (swine flu, Aujeszky's disease), guinea pigs (rabies, foot and mouth disease, canine distemper). Rabbits (rabies, rabbit myxomas).
Requirements for laboratory animals - the animal must be sensitive to this virus; its age is of great importance for the cultivation of many viruses. Most viruses reproduce better in the body of young and even newborn animals; standard sensitivity is achieved by selecting animals of a certain age and the same weight. In terms of sensitivity, the so-called linear animals obtained as a result of inbreeding over a number of generations have the greatest standard; laboratory animals must be healthy. Animals entering the vivarium of the virology laboratory must be brought from a farm free of infectious diseases. They are kept in quarantine and undergo clinical observation. If there is a disease, they are destroyed.
Animals are placed in such a way that, on the one hand, the functioning of all body systems within physiological norms is ensured, and on the other hand, mutual reinfection and the spread of infection beyond the vivarium are excluded. Different methods of individual marking are used for animals of different species. For large animals and chickens, metal tags with a stamped number are used. When using a small group of animals in an experiment and for a short period of time, the hair can be trimmed with marks on the back and hips. Marking of white mice and white rats can be carried out by amputation of individual fingers on the front or hind limbs. The method of applying colored spots to unpigmented wool is often used. Infection of laboratory animals.
1. subcutaneously - back.
2. Intradermal – heel
3. Intramuscular – thigh
4. Intravenously - into the tail (preliminarily rubbed with hot water and squeezing)
5. Intranasally - a drop in the nose (a weak ether anesthesia is first given to prevent sneezing)
6. Interocerebral - the skull is carefully drilled with a needle, do not press, the drop goes away on its own.
All surfaces are pre-lubricated with iodized alcohol.
Preparation lab. animals (using the example of a white mouse)
The skin is lubricated with a disinfectant.
An incision is made along linea alba.
Opening the sternum - the lungs are taken and placed in tube No. 1
Opening the abdominal cavity - the liver, spleen, kidney are taken and placed in tube No. 2.
The skull is opened. The brain is taken, sections of 4 layers are made, the pieces are placed on filter paper and prints are made on glass.
QUESTION No. 47 “STRUCTURE OF A DEVELOPING CHICKEN EMBRYO. MAIN PROBLEMS SOLVED BY THE METHOD OF INFECTION BY TBE AND ITS ADVANTAGES OVER CULTIVATION OF VIRUSES IN LABORATORY ANIMALS.
CE is used in virology mainly for the same purposes as LV: detection of active virus by bioassay in pathological material; primary isolation of the virus; maintaining viruses in the laboratory; virus titration; accumulation of the virus for laboratory research and obtaining vaccines; as a test object in the neutralization reaction.
Structure: 1. Shell 2. Subshell membrane 3. Air chamber 4. Allantoic cavity 5. Yolk sac 6. Albumin sac 7. CHAO - chorion-allantoic membrane 8. Amniotic cavity 9. Embryo 10. Cord (connection of the yolk sac with the umbilical cord) . From 5-12 days EC can be used for infection
1) The shell and subshell membrane serve as good protection from environmental factors. 2) EC contain a substrate for growing the virus. 3) CE are resistant to impacts associated with the release of the material under study. 4) ECs are easily accessible, environmentally friendly, do not require care or feeding, and do not form AT.
6 methods of infection with TBE: 1) Infection in the allantoic cavity (influenza, Newcastle disease). The EC is fixed vertically with the blunt end up, and a 1mm hole is made on the side of the embryo 5-6mm above the border of the air chamber. The needle is inserted parallel to the longitudinal axis to a depth of 10-12 mm. 2) for CAO (smallpox, canine plague): a) B/w natural air chamber. FE into a tripod with the blunt end up, a 15-20mm window in the shell against the center of the air chamber. Remove the shell membrane. 0.2 mm of suspension is applied to the XAO. Hole adhesive plaster. b) B/w artificial air chamber. The tripod is horizontal with the bud up. Make 2 holes: above the center of the air chamber, another 0.2-0.5 cm on the side, on the side of the embryo. The air is sucked out of the first embryo, an artificial air chamber is formed, the bottom of which is CAO, an infectious liquid is applied to it, and it is sealed with an adhesive plaster. 3) In the yolk sac (chlamydia, Marek's b.): a) EC is placed vertically in a stand. A hole above the center of the air chamber, a needle 3.5-4 cm at an angle of 45, opposite to the location of the embryo. b) a similar route of infection is carried out on a horizontally reinforced CE stand; in this case, the embryo is at the bottom, and the yolk is above it. 4) Into the amniotic cavity (flu, Newcastle disease): the method is closed - the embryo. up. The needle is inserted with a blunt end towards the embryo of the open. method - a hole of 1.5-2.5 cm above the air cavity. The subshell membrane is removed. Tweezers push the XAO towards the embryo. Then the amniotic membrane along with the CAO is pulled to the window, and the suspension is introduced there. They let you go. Band-Aid. 5) Infection into the body of the embryo. 6) into blood vessels.
QUESTION No. 48 “TYPES OF CELL CULTURES AND THEIR USE IN VIRUSOLOGY. BRIEF CHARACTERISTICS OF EACH SPECIES.”
Cell culture (CC) is the cells of a multicellular organism that live and multiply in artificial conditions outside the body. The cultivation technique began to develop especially successfully after the 40s. This was facilitated by the following events: the discovery of antibiotics that prevent bacterial infection of CC, the discovery by Hang and Enders of the ability of viruses to cause specific cell destruction. Dulbecco and Vogt (1952) proposed a method for trypsinizing tissues and obtaining single-layer CCs. The following CCs are used: 1) PTCC – cells obtained directly from organs or tissues of the body, growing in vitro in one layer. CC can be obtained from almost any human or animal organ or tissue. It is better to do this from embryonic organs, because embryonic cells have a higher growth potential. Most often, they are obtained from the kidneys, lungs, skin, thymus, and testicles. To obtain primary cells from a healthy animal, no later than 2-3 hours after slaughter, the corresponding organs or tissues are taken, crushed, and treated with trypsin, pancreatin, and collagenase. Enzymes destroy intercellular substances, and the resulting individual cells are suspended in a nutrient medium and cultured on the inner surface of test tubes or mattresses in a thermostat at 37C. The cells attach to the glass and begin to divide. A layer one cell thick forms on the glass, usually after 3-5 days. The nutrient medium is changed as it becomes contaminated with cell waste products. The monolayer will remain viable for 7-21 days. When cultivating viruses in CC, it is possible to obtain preparations with a high titer of the virus, which is important when obtaining antigens and vaccines. 2) Subcultures - they are often used and obtained from primary cells grown in mattresses by removing them from the glass with a solution of versene or trypsin, resuspending them in a new nutrient medium and reseeding them on new mattresses or test tubes. After 2-3 days, a monolayer is formed. They are not inferior in sensitivity to PTCA and are more economical. 3) Transplantable CCs are cells capable of multiplying outside the body for an indefinitely long time. They are maintained in the laboratory by subculture from one vessel to another (subject to replacement of the nutrient medium). They are obtained from primary CCs with increased growth activity through long-term subcultures in a certain cultivation mode. The cells of transplanted cultures have the same shape, a heteroploid set of chromosomes, are stable under in vitro growth conditions, and some of them have oncogenic activity. “+” before the primary ones - it’s easier to prepare, you can check in advance for the presence of latent viruses and microflora; clonal lines provide more standard conditions for virus propagation than primary lines. Most transplanted cells have a wider spectrum of sensitivity to viruses than the corresponding primary cultures. But they are prone to malignant degeneration. 4) Diploid CCs are a morphologically homogeneous population of cells, stabilized during in vitro cultivation, having a limited lifespan, characterized by 3 growth phases, preserving the karyotype characteristic of the original tissue during passaging, free from contaminants and not having tumorigenic activity when transplanted into hamsters. They are also obtained from primary cells. In contrast, they have limited passaging capabilities. The maximum number of passages is 50 -\+ 10, then the number of dividing cells sharply decreases and they die. Advantages over transplantable CCs - they can be in a viable state for 10-12 days without changing the nutrient medium; when changing the medium once a week, they remain viable for 4 weeks; They are especially suitable for long-term cultivation of viruses; they retain the sensitivity of the original tissue to viruses. 5) Suspension CCs – continuous cell cultures in suspension.
QUESTION No. 49 “PRIMARY TRYPSINIZED CELL CULTURES. THEIR ADVANTAGES AND DISADVANTAGES. APPLICATION IN VIROLOGICAL RESEARCH".
PTCA are cells obtained directly from organs or tissues of the body, growing in vitro in one layer. CC can be obtained from almost any human or animal organ or tissue. It is better to do this from embryonic organs, because embryonic cells have a higher growth potential. Most often, they are obtained from the kidneys, lungs, skin, thymus, and testicles. To obtain primary cells from a healthy animal, no later than 2-3 hours after slaughter, the corresponding organs or tissues are taken, crushed, and treated with trypsin, pancreatin, and collagenase. Enzymes destroy intercellular substances, and the resulting individual cells are suspended in a nutrient medium and cultured on the inner surface of test tubes or mattresses in a thermostat at 37C. The cells attach to the glass and begin to divide. A layer one cell thick forms on the glass, usually after 3-5 days. The nutrient medium is changed as it becomes contaminated with cell waste products. The monolayer will remain viable for 7-21 days. When cultivating viruses in CC, it is possible to obtain preparations with a high titer of the virus, which is important when obtaining antigens and vaccines. Using the QC method, some theoretical questions were resolved - about the interaction of the virus with the cell, the place of virus reproduction, the mechanism of antiviral immunization. Currently, QC is used for isolating viruses from pathogenic material, their indication, identification, for setting up a neutralization reaction, determining the titer of viruses, for preparing diagnostic Ags and vaccines, and as test objects in a neutralization reaction.
QUESTION No. 50 “NUTRIENT MEDIA AND SOLUTIONS USED IN VIRUSOLOGY. REQUIREMENTS FOR Utensils for CC CULTIVATION, ITS PROCESSING.”
The most widely used solutions when working with CC are Hanks and Earle solutions, which are prepared using bidistilled water with the addition of various salts and glucose. These balanced salt solutions are used to prepare all culture media because... they ensure the preservation of pH, osmotic pressure in cells and the appropriate concentration of necessary inorganic substances. They are also used for washing away growth media, virus dilutions, etc. When culturing cells, dispersing solutions of trypsin and versene are used. Trypsin solution is used to separate pieces of tissue into individual cells and to remove the layer of cells from the glass. Versene solution - used to remove cells from glass. Nutrient media (hereinafter referred to as NS) - distinguish: 1) natural media, which consist of a mixture of saline solution, blood serum, tissue extract, cow amniotic fluid, etc. The number of components varies. They are rarely used. 2) artificial PS - enzymatic hydrolysates of various protein products: lactalbumin hydrolysate, muscle enzymatic hydrolysate, etc. Of the synthetic media, the most widely used are medium 199 and Eagle’s medium. The indicator phenol red is added to all culture media and some saline solutions to determine the concentration of hydrogen ions. To destroy microflora before use, add AB to the media: penicillin and streptomycin, 100 units per ml. All PS are divided into 2 groups: growth – ensure the life and reproduction of cells; supporting - ensuring the vital activity of cells, but not their reproduction (they do not contain blood serum). Glassware – The quality of the glassware is important for the successful cultivation of cells outside the body. It should be sterile, fat-free, and have no toxic effect. For cell cultivation, test tubes, mattresses of 50, 100, 250, 500, 1000, 1500 ml, roller flasks of 500, 1000, 2000 ml, various pipettes, bottles for PS and solutions, and flasks of various capacities are used. The processing of glassware consists of several stages: 1) the infected glassware is immersed in a 2-3% NaOH solution for 5-6 hours; 2) rinse in 3-4 changes of tap water; 3) soak in a 0.3-0.5% powder solution; 4) wash thoroughly with a brush in a warm powder solution; 5) rinse in several changes of tap water; 6) rinse in distilled water containing 0.5% HCl; 7) rinse 4-5 times with tap water and 3 changes of distilled water; 8) dried in a drying cabinet; 9) mounted and sterilized in an oven or autoclaved.
QUESTION No. 51 “THE PRINCIPLE OF INFECTION OF CELL CULTURES BY VIRUS-CONTAINING MATERIAL. INDICATION OF VIRUSES IN CELL CULTURE.”
For infection, test tubes (mattresses) with a continuous cell monolayer are selected, viewing them under a low magnification microscope. The growth nutrient medium is drained, the cells are washed 1-2 times with Hanks solution to remove serum antibodies and inhibitors. 0.1-0.2 ml of virus-containing material is added to each tube and distributed evenly over the cell layer by shaking. Leave for 1-2 hours at 22-37C for virus adsorption on the cell surface. The virus-containing material is removed from the containers and the supporting medium is poured. For indication, there are the following main methods for indicating the virus in CC: by cytopathic effect or cytopathic effect; by a positive hemadsorption reaction; by plaque formation; for detection of intracellular inclusions; to detect viruses in immunofluorescence reactions; to detect virus interference; to suppress cell metabolism (color test); electron microscopy. Detection of specific cell degeneration (by CPD) - a simple sign is degenerative changes in cells (manifestation of CPD). Visible changes in the cell are called cytopathic changes. These changes in infected cells depend on the dose and biological properties of the virus being studied; the manifestation of CPE and its features sometimes allows identification of the isolated viruses. When KK is infected with medium doses of the virus, the nature of these changes is specific and can be classified into groups: focal fine-grained degeneration, fine-grained degeneration throughout the entire monolayer, focal cluster-shaped accumulation of round cells, uniform granularity, association of cells into giant multinuclear symplasts and syncytia. The degree of degeneration is assessed using a 4-point system.
Sometimes the absence of CPD is observed, but this is not considered to be the absence of a virus, and therefore 2-3 blind passages are carried out and at the 2-3 passage the viruses can exhibit the desired properties.
QUESTION No. 52 “METHODS FOR DETECTING VIRIONS AND VIRAL INCLUSION BODIES, THEIR PRACTICAL IMPORTANCE.”
It is usually possible to examine virus virions and establish their structure using electron microscopy, which allows one to distinguish objects up to 0.2-0.4 nm in size. Detection of virions in material from sick animals using electron microscopy can serve as evidence of the presence of viruses in this material and in some cases is used to diagnose viral diseases. But this method is technically complex and expensive, and does not allow accurately identifying the detected virus. With a light microscope it is possible to see only virions of smallpox viruses at the limit of visibility. The ability to be stained with certain dyes, the size, shape, structure, location in the cell of inclusion bodies formed by different viruses are not the same, but are specific to each virus. Therefore, the detection of intracellular inclusion bodies with certain characteristics in material from sick animals allows us to judge what kind of virus they were formed, and therefore the presence of this virus in the material under study. To detect inclusion bodies, smears or prints are prepared (posthumously or intravitally), which are subjected to special staining methods followed by microscopy. For inclusion bodies formed by different viruses, staining methods are different. Many coloring recipes have been developed. Among them there are also universal ones, which include hematoxylin-eosin staining.
Virology.
Other mycoplasmas pathogenic for humans.
Mycoplasma pneumonia.
Mycoplasma pneumoniae.
M. pneumoniae differs from other species by serological methods, as well as by characteristics such as b-hemolysis of sheep red blood cells, aerobic reduction of tetrazolium, and the ability to grow in the presence of methylene blue.
M. pneumoniae is the most common cause of nonbacterial pneumonia. Infection with this mycoplasma may also take the form of bronchitis or mild respiratory fever.
Asymptomatic infections are common. Familial outbreaks are common, and large outbreaks have occurred in military training centers. The incubation period is approximately two weeks.
M. pneumoniae can be isolated by culture of sputum and throat swabs, but diagnosis is more easily made by serology, usually the complement fixation test. The diagnosis of mycoplasma pneumonia is helped by the empirical finding that many patients develop cold agglutinins to human red blood cells of group 0.
Mycoplasmas are normally inhabitants of the reproductive tract of men and women. The most commonly encountered species is M. hominis, which is responsible for some cases of vaginal discharge, urethritis, salpingitis and pelvic sepsis. It is the most common cause of postpartum sepsis.
The microorganism can enter the mother's blood during childbirth and be localized in the joints. A group of mycoplasmas (ureaplasmas) that form tiny colonies are considered a possible cause of nongonococcal urethritis in both sexes. Other species are normal commensals of the oral cavity and nasopharynx.
Prevention. It comes down to maintaining a high level of general resistance of the human body. A vaccine made from killed mycoplasmas for the specific prevention of atypical pneumonia has been obtained in the USA
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Timakov V.D., Levashev V.S., Borisov L.B. Microbiology. - M: Medicine, 1983. - 312 p.
2. Borisov L.B., Kozmin-Sokolov B.N., Freidlin I.S. Guide to laboratory classes in medical microbiology, virology and immunology / ed. Borisova L.B. – G.: Medicine, 1993. – 232 p.
3. Medical microbiology, virology and immunology: Textbook, ed. A.A. Vorobyova. – M.: Medical Information Agency, 2004. - 691 p.
4. Medical microbiology, virology, immunology / ed. L.B.Borisov, A.M.Smirnova. - M: Medicine, 1994. - 528 p.
Odessa-2009
Lecture No. 21. Subject and tasks of medical virology. General characteristics of viruses
We are starting to study a new science - virology, the science of viruses. Virology is an independent science of modern natural science, occupying a vanguard position in biology and medicine, and the role and importance of virology is steadily increasing. This is due to a number of circumstances:
1. Viral diseases occupy a leading place in human infectious pathology. The use of antibiotics makes it possible to effectively solve the treatment of most bacterial diseases, while there are still no sufficiently effective and harmless drugs for the treatment of viral diseases. As the incidence of bacterial infections decreases, the proportion of viral diseases is steadily increasing. The problem of mass viral infections - respiratory and intestinal - is acute. For example, the well-known flu often takes the form of massive epidemics and even pandemics, in which a significant percentage of the world's population falls ill.
2. The viral-genetic theory of the origin of tumors and leukemia has gained recognition and is increasingly being confirmed. Therefore, we expect that the development of virology will lead to a solution to the most important problem of human pathology - the problem of carcinogenesis.
3. Currently, new viral diseases are emerging or previously known viral diseases are becoming acute, which constantly poses new challenges for virology. An example is HIV infection.
4. Viruses have become a classic model for molecular biology and molecular genetic research. Many issues of fundamental research in biology are solved using viruses; viruses are widely used in biotechnology.
5. Virology is a fundamental science of modern natural science, not only because it enriches other sciences with new methods and new ideas, but also because the subject of the study of virology is a qualitatively special form of organization of living matter - viruses, which are radically different from all other living beings on Earth .
2. HISTORICAL SKETCH OF THE DEVELOPMENT OF VIRUSOLOGY
The credit for the discovery of viruses and the description of their main characteristics belongs to the Russian scientist Dmitry Iosifovich Ivanovsky (1864-1920). It is interesting that Ivanovsky began his research as a 3rd year student at St. Petersburg University, when he was doing course work in Ukraine and Bessarabia. He studied tobacco mosaic disease and found out that it was an infectious plant disease, but its causative agent did not belong to any of the then known groups of microorganisms. Later, already a certified specialist, Ivanovsky continues his research at the Nikitsky Botanical Garden (Crimea) and performs a classic experiment: he filters the juice of the leaves of the affected plant through a bacterial filter and proves that the infectious activity of the juice does not disappear.
Subsequently, the main groups of viruses were discovered. In 1898, F. Leffler and P. Frosch proved the filterability of the causative agent of foot-and-mouth disease (the foot-and-mouth disease virus affects animals and humans), in 1911, P. Raus proved the filterability of the causative agent of the tumor disease - chicken sarcoma, in 1915, F. Twort and in 1917 Mr. D'Herelle discovered phages - bacterial viruses.
This is how the main groups of viruses were discovered. Currently, more than 500 types of viruses are known.
Further progress in the development of virology is associated with the development of methods for cultivating viruses. At first, viruses were studied only when they infected sensitive organisms. A significant step forward was the development of a method for cultivating viruses in chicken embryos by Woodruff and Goodpasture in 1931. A revolution in virology was the development of a method for cultivating viruses in single-layer cell cultures by J. Enders, T. Weller, F. Robbins, and in 1948. Not without reason in 1952 This discovery was awarded the Nobel Prize.
Already in the 30s the first virological laboratories were created. Currently, Ukraine has the Odessa Research Institute of Epidemiology and Virology named after. I.I. Mechnikov, there are virological laboratories in a number of research institutes of epidemiology, microbiology, and infectious diseases. There are virological laboratories for practical health care, which are primarily engaged in the diagnosis of viral diseases.
3. Compose the ultrastructure of viruses
First of all, it must be said that the term “virus” was introduced into scientific terminology by L. Pasteur. L. Pasteur received his vaccine to prevent rabies in 1885, although he did not discover the causative agent of this disease - there were still 7 years left before the discovery of viruses. L. Pasteur called the hypothetical pathogen the rabies virus, which translated means “rabies poison.”
The term “virus” is used to refer to any stage of virus development - both extracellularly located infectious particles and intracellularly reproducing virus. To designate a viral particle, the term “ virion».
By chemical composition Viruses are basically similar to other microorganisms; they have nucleic acids, proteins, and some also have lipids and carbohydrates.
Viruses contain only one type of nucleic acid - either DNA or RNA. Accordingly, DNA genomic and RNA genomic viruses are isolated. Nucleic acid in the virion can contain from 1 to 40%. Typically, the virion contains only one nucleic acid molecule, often closed in a ring. Viral nucleic acids are not much different from eukaryotic nucleic acids; they consist of the same nucleotides and have the same structure. True, viruses can contain not only double-stranded, but also single-stranded DNA. Some RNA viruses may contain double-stranded RNA, although most contain single-stranded RNA. It should be noted that viruses may contain plus-strand RNA, which can act as messenger RNA, but they may also contain minus-strand RNA. Such RNA can perform its genetic function only after the complementary plus strand is synthesized in the cell. Another feature of viral nucleic acids is that in some viruses the nucleic acid is infectious. This means that if RNA without protein admixture is isolated from a virus, for example the polio virus, and introduced into a cell, a viral infection will develop with the formation of new viral particles.
Proteins are contained in viruses in an amount of 50-90%; they have antigenic properties. Proteins are part of the envelope structures of the virion. In addition, there are internal proteins associated with the nucleic acid. Some viral proteins are enzymes. But these are not enzymes that ensure the metabolism of viruses. Viral enzymes are involved in the penetration of the virus into the cell, the exit of the virus from the cell, some of them are necessary for the replication of viral nucleic acids.
Lipoids can be from 0 to 50%, carbohydrates - 0 - 22%. Lipids and carbohydrates are part of the secondary shell of complex viruses and are not virus-specific. They are borrowed by the virus from the cell and are therefore cellular.
Let us note a fundamental difference in the chemical composition of viruses - the presence of only one type of nucleic acid, DNA or RNA.
Ultrastructure of viruses- this is the structure of virions. The sizes of virions vary and are measured in nanometers. 1 nm is a thousandth of a micrometer. The smallest typical viruses (poliomyelitis virus) have a diameter of about 20 nm, the largest (variola virus) - 200-250 nm. Average viruses have sizes of 60 - 120 nm. Small viruses can only be seen in an electron microscope; large ones are at the limit of the resolution of a light microscope and are visible in a dark field of view or with special staining that increases the size of the particles. Individual viral particles visible under a light microscope are usually called elementary Paschen-Morozov bodies. E. Paschen discovered the variola virus using a special stain, and Morozov proposed a silvering method that made it possible to see even medium-sized viruses in a light microscope.
The shape of virions can be different - spherical, cuboidal, rod-shaped, sperm-like.
Each virion consists of a nucleic acid, which in viruses constitutes a “nucleon.” Compare - nucleus in eukaryotes, nucleoid - in prokaryotes. The nucleon is necessarily associated with the primary protein shell - the capsid, consisting of protein capsomers. As a result, a nucleoprotein is formed - a nucleocapsid. Simple viruses consist only of a nucleocapsid (poliomyelitis viruses, tobacco mosaic disease virus). Complex viruses also have a secondary shell - a supercapsid, which in addition to proteins also contains lipids and carbohydrates.
The combination of structural elements in the virion may be different. There are three types of symmetry of viruses - helical, cubic and mixed. Speaking about symmetry, the symmetry of the viral particles relative to the axis is emphasized.
At spiral type of symmetry individual capsomeres, visible in an electron microscope, are arranged along the nucleic acid helix so that the thread passes between two capsomeres, covering it on all sides. The result is a rod-shaped structure, such as the rod-shaped tobacco mosaic virus. But viruses with a helical type of symmetry do not necessarily have to be rod-shaped. For example, although the influenza virus has a helical type of symmetry, its nucleocapsid is folded in a certain way and is covered with a supercapsid. As a result, influenza virions are usually spherical in shape.
At cubic type symmetry, the nucleic acid folds in a certain way in the center of the virion, and capsomers cover the outside of the nucleic acid, forming a three-dimensional geometric figure. Most often, the figure of an icosahedron, a polyhedron with a certain ratio of the number of vertices and faces, is formed. For example, polio viruses have this form. In profile, the virion has the shape of a hexagon. A more complex form of adenovirus, also of cubic type of symmetry. Long threads and fibers extend from the vertices of the polyhedron, ending in a thickening.
With a mixed type of symmetry, for example, in bacteriophages, the head with a cubic type of symmetry has the shape of an icosahedron, and the process contains a spirally twisted contractile fibril.
Some viruses have a more complex structure. For example, the variola virus contains a large nucleocapsid with a helical type of symmetry, and the supercapsid is complex, containing a system of tubular structures.
Thus, viruses are quite complex. But we must note that viruses do not have a cellular organization. Viruses are non-cellular creatures, and this is one of their fundamental differences from other organisms.
A few words about the stability of viruses. Most viruses are inactivated at 56 - 60 °C for 5 - 30 minutes. Viruses tolerate refrigeration well; at room temperature, most viruses are quickly inactivated. The virus is more resistant to ultraviolet radiation and ionizing radiation than bacteria. Viruses are resistant to glycerol. Antibiotics have no effect on viruses at all. Of the disinfectants, the most effective is 5% Lysol; most viruses die within 1 - 5 minutes.
4. VIRUS REPRODUCTION
Usually we do not use the term “reproduction of viruses”, but rather say “reproduction”, reproduction of viruses, since the method of reproduction of viruses is fundamentally different from the method of reproduction of all organisms known to us.
To better study the mechanism of virus reproduction, we offer you a table that is not included in textbooks, but helps to understand this complex process.
stages of virus reproduction
The first, preparatory period, begins with the stage of virus adsorption on the cell. The adsorption process is carried out due to the complementary interaction of virus attachment proteins with cellular receptors. Cellular receptors can be of glycoprotein, glycolipid, protein and lipid nature. Each virus requires specific cellular receptors.
Viral attachment proteins located on the surface of the capsid or supercapsid act as viral receptors.
The interaction between virus and cell begins with nonspecific adsorption of the virion on the cell membrane, and then specific interaction between viral and cellular receptors occurs according to the principle of complementarity. Therefore, the process of virus adsorption on a cell is a specific process. If the body does not have cells with receptors for a particular virus, then infection with this type of virus in such an organism is impossible - there is species resistance. On the other hand, if we managed to block this first stage of interaction between the virus and the cell, then we could prevent the development of a viral infection at a very early stage.
Stage 2 - penetration of the virus into the cell - can occur in two main ways. The first one, which was described earlier, is called viropexis. This pathway closely resembles phagocytosis and is a variant of receptor endocytosis. The viral particle is adsorbed on the cell membrane; as a result of the interaction of receptors, the state of the membrane changes, and it invaginates, as if flowing around the viral particle. A vacuole is formed, delimited by a cell membrane, in the center of which the viral particle is located.
When a virus enters through membrane fusion mutual penetration of the elements of the virus shell and the cell membrane occurs. As a result, the “core” of the virion ends up in the cytoplasm of the infected cell. This process occurs quite quickly, so it was difficult to register it on electron diffraction patterns.
Deproteinization - liberation of the viral genome from the supercapsid and capsid. This process is sometimes called “undressing” of virions.
Release from the envelopes often begins immediately after the virion attaches to cellular receptors and continues inside the cell cytoplasm. Lysosomal enzymes take part in this. In any case, for further reproduction to occur, deproteinization of the viral nucleic acid is necessary, since without this the viral genome is not able to induce the reproduction of new virions in the infected cell.
Average reproduction period called latent, hidden, since after deproteinization the virus seems to “disappear” from the cell, it cannot be detected on electron diffraction patterns. During this period, the presence of the virus is detected only by changes in the metabolism of the host cell. The cell is rebuilt under the influence of the viral genome on the biosynthesis of the components of the virion - its nucleic acid and proteins.
First stage of the middle period, t transcription viral nucleic acids, rewriting genetic information through the synthesis of messenger RNA is a necessary process to begin the synthesis of viral components. It occurs differently depending on the type of nucleic acid.
Viral double-stranded DNA is transcribed in the same way as cellular DNA by DNA-dependent RNA polymerase. If this process is carried out in the cell nucleus (in adenoviruses), then cellular polymerase is used. If in the cytoplasm (smallpox virus), then with the help of RNA polymerase, which penetrates the cell as part of the virus.
If the RNA is minus-strand (in influenza, measles, rabies viruses), first the messenger RNA must be synthesized on the viral RNA matrix using a special enzyme - RNA-dependent RNA polymerase, which is part of the virions and penetrates the cell along with the viral virus. RNA. The same enzyme is also found in viruses containing double-stranded RNA (reoviruses).
Regulation of the transcription process is carried out by sequential rewriting of information from “early” and “late” genes. “Early” genes contain information about the synthesis of enzymes necessary for gene transcription and their subsequent replication. In the “late” ones there is information for the synthesis of virus envelope proteins.
Broadcast- synthesis of viral proteins. This process is completely analogous to the known scheme of protein biosynthesis. Virus-specific messenger RNA, cellular transfer RNA, ribosomes, mitochondria, and amino acids are involved. First, enzyme proteins necessary for the transcription process are synthesized, as well as for partial or complete suppression of the metabolism of the infected cell. Some virus-specific proteins are structural and are included in the virion (for example, RNA polymerase), others are non-structural, which are found only in the infected cell and are necessary for one of the processes of virion reproduction.
Later, the synthesis of viral structural proteins - components of the capsid and supercapsid - begins.
After the synthesis of viral proteins on ribosomes, their post-translational modification can occur, as a result of which the viral proteins “mature” and become functionally active. Cellular enzymes can carry out phosphorylation, sulfonation, methylation, acylation and other biochemical transformations of viral proteins. The process of proteolytic cutting of viral proteins from large molecular precursor proteins is essential.
Replication viral genome - synthesis of viral nucleic acid molecules, reproduction of viral genetic information.
Replication of viral double-stranded DNA occurs with the help of cellular DNA polymerase in a semi-conservative manner in the same way as cellular DNA replication. Single-stranded DNA replicates through an intermediate double-stranded replicative form.
There are no enzymes in the cell that can carry out RNA replication. Therefore, this process is always carried out by virus-specific enzymes, information about the synthesis of which is encoded in the viral genome. During the replication of single-stranded RNA genomes, an RNA strand complementary to the viral one is first synthesized, and then this newly formed RNA strand becomes the template for the synthesis of genome copies. Moreover, in contrast to the process of transcription, in which often only relatively short chains of RNA are synthesized, during replication a complete strand of RNA is immediately formed. Double-stranded RNA replicates similarly to double-stranded DNA, but with the help of the corresponding enzyme - RNA polymerase of viral origin.
As a result of the process of viral genome replication, funds of viral nucleic acid molecules necessary for the formation of mature virions accumulate in the cell.
Thus, the synthesis of individual components of the virion is separated in time and space, occurring in different cellular structures and at different times.
IN final period During reproduction, virions are assembled and the virus leaves the cell.
Virion assembly may occur in different ways, but it is based on the process of self-assembly of viral components transported from the sites of their synthesis to the site of assembly. The primary structure of viral nucleic acids and proteins determines the order of conformation of the molecules and their connection with each other. First, a nucleocapsid is formed due to the strictly oriented connection of protein molecules into capsomers and capsomers with nucleic acid. For simple viruses, this is where the assembly ends. The assembly of complex viruses with a supercapsid is multi-stage and usually ends during the process of virions leaving the cell. In this case, elements of the cell membrane are included in the supercapsid of the virus.
Exit of the virus from the cell can happen in two ways. Some viruses that lack a supercapsid (adenoviruses, picornaviruses) exit the cell in an “explosive” manner. In this case, the cell is lysed, and the virions exit the destroyed cell into the intercellular space. Other viruses that have a lipoprotein secondary envelope, for example influenza viruses, leave the cell by budding from its envelope. The cell can remain viable for a long time.
The entire virus reproduction cycle usually takes several hours. In the 4 to 5 hours that pass from the moment one molecule of viral nucleic acid enters a cell, from several tens to several hundred new virions can be formed, capable of infecting neighboring cells. Thus, the spread of viral infection in cells occurs very quickly.
Thus, the way viruses reproduce is fundamentally different from the way all other living things reproduce. All cellular organisms reproduce by division. When viruses multiply, individual components are synthesized in different places in the virus-infected cell and at different times. This method of reproduction is called “disconnected” or “disjunctive”.
It should be said that the interaction of the virus and the cell may not necessarily lead to the described result - early or delayed death of the infected cell with the production of a mass of new mature viral particles. There are three possible types of viral infection in a cell.
The first option, which we have already discussed, occurs when productive or virulent infections.
Second option - persistent infection of a virus in a cell, when there is a very slow production of new virions with their release from the cell, but the infected cell remains viable for a long time.
Finally, the third option is integrative type interaction between a virus and a cell, during which the integration of viral nucleic acid into the cellular genome occurs. This involves the physical inclusion of a viral nucleic acid molecule into the host cell chromosome. For DNA genomic viruses, this process is quite understandable; RNA genomic viruses can integrate their genome only in the form of a “provirus” - a DNA copy of viral RNA synthesized using reverse transcriptase - RNA-dependent DNA polymerase. In the case of integration of the viral genome into the cellular genome, the viral nucleic acid replicates together with the cellular one during cell division. A virus in the form of a provirus can persist in a cell for a long time due to constant replication. This process is called " virogeny».
5. CARDINAL FEATURES OF VIRUSES
However, the size of large viruses is comparable to the size of chlamydia and small rickettsia, and filterable forms of bacteria have been described. Currently, the term “filterable viruses”, which for a long time was common to refer to viruses, is practically not used. Therefore, small size is not a fundamental difference between viruses and other living beings.
Therefore, at present, the fundamental differences between viruses and other microorganisms are based on more significant biological properties, which we discussed in this lecture.
Based on the knowledge of the properties of viruses we have analyzed, we can formulate the following 5 fundamental differences between viruses from other living beings on Earth:
1. Lack of cellular organization.
2. The presence of only one type of nucleic acid (DNA or RNA).
3. Lack of independent metabolism. Metabolism in viruses is mediated through the metabolism of cells and organisms.
4. The presence of a unique, disjunctive method of reproduction.
Thus, we can give the following definition to viruses.
History of virology, nature and origin of viruses
Virus discovery
Virology is a young science, its history goes back a little over 100 years. Having begun its journey as the science of viruses that cause diseases in humans, animals and plants, virology is currently developing in the direction of studying the basic laws of modern biology at the molecular level, based on the fact that viruses are part of the biosphere and an important factor in the evolution of the organic world.
The history of virology is unusual in that one of its subjects - viral diseases - began to be studied long before viruses themselves were discovered. The beginning of the history of virology is the fight against infectious diseases and only subsequently the gradual disclosure of the sources of these diseases. This is confirmed by the work of Edward Jenner (1749-1823) on the prevention of smallpox and the work of Louis Pasteur (1822-1895) with the causative agent of rabies.
Since time immemorial, smallpox has been the scourge of humanity, claiming thousands of lives. Descriptions of smallpox infection are found in the manuscripts of ancient Chinese and Indian texts. The first mention of smallpox epidemics on the European continent dates back to the 6th century AD (an epidemic among the soldiers of the Ethiopian army besieging Mecca), after which there was an inexplicable period of time when there were no mentions of smallpox epidemics. Smallpox began to spread across continents again in the 17th century. For example, in North America (1617-1619) in the state of Massachusetts, 9/10 of the population died, in Iceland (1707) after a smallpox epidemic, only 17 thousand remained from 57 thousand people, in the city of Eastham (1763) ) from 1331 inhabitants there are 4 people left. In this regard, the problem of combating smallpox was very acute.
A technique for preventing smallpox through vaccination, called variolation, has been known since ancient times. Mentions of the use of variolation in Europe date back to the mid-17th century, with references to earlier experience in China, the Far East, and Turkey. The essence of variolation was that the contents of pustules from patients suffering from a mild form of smallpox were introduced into a small wound on the human skin, which caused a mild disease and prevented an acute form. However, there remained a high risk of contracting a severe form of smallpox and the mortality rate among vaccinated people reached 10%. Jenner revolutionized smallpox prevention. He was the first to notice that people who had cowpox, which was mild, never subsequently suffered from smallpox. On May 14, 1796, Jenner introduced liquid from the pustules of milkmaid Sarah Selmes, who had cowpox, into the wound of James Phipps, who had never suffered from smallpox. At the site of the artificial infection, the boy developed typical pustules, which disappeared after 14 days. Then Jenner introduced highly infectious material from the pustules of a smallpox patient into the boy’s wound. The boy did not get sick. This is how the idea of vaccination was born and confirmed (from the Latin word vacca - cow). In Jenner's time, vaccination was understood as the introduction of infectious cowpox material into the human body in order to prevent smallpox. The term vaccine was applied to a substance that protected against smallpox. Since 1840, smallpox vaccine began to be obtained by infecting calves. The human smallpox virus was discovered only in 1904. Thus, smallpox is the first infection against which a vaccine was used, i.e., the first vaccine-preventable infection. Advances in vaccine prevention of smallpox have led to its worldwide eradication.
Nowadays, vaccination and vaccine are used as general terms denoting vaccination and vaccination material.
Pasteur, who essentially did not know anything specific about the causes of rabies, except for the indisputable fact of its infectious nature, used the principle of weakening (attenuation) of the pathogen. In order to weaken the pathogenic properties of the rabies pathogen, a rabbit was used, into whose brain the brain tissue of a dog that died of rabies was injected. After the death of the rabbit, its brain tissue was injected into the next rabbit, and so on. About 100 passages were carried out before the pathogen adapted to the rabbit's brain tissue. When injected subcutaneously into the dog's body, it exhibited only moderate pathogenicity properties. Pasteur called such a “re-educated” pathogen “fixed”, in contrast to the “wild” one, which is characterized by high pathogenicity. Pasteur later developed a method of creating immunity, consisting of a series of injections with gradually increasing amounts of a fixed pathogen. The dog that completed the full course of injections turned out to be completely resistant to infection. Pasteur came to the conclusion that the process of development of an infectious disease is essentially a struggle between microbes and the body's defenses. “Every disease must have its own pathogen, and we must promote the development of immunity to this disease in the patient’s body,” said Pasteur. Not yet understanding how the body produces immunity, Pasteur was able to use its principles and direct the mechanisms of this process to the benefit of humans. In July 1885, Pasteur had the opportunity to test the properties of a “fixed” rabies pathogen on a child bitten by a rabid dog. The boy was given a series of injections of an increasingly toxic substance, with the last injection containing a completely pathogenic form of the pathogen. The boy remained healthy. The rabies virus was discovered by Remlanger in 1903.
It should be noted that neither the smallpox virus nor the rabies virus were the first viruses discovered to infect animals and humans. The first place rightfully belongs to the foot-and-mouth disease virus, discovered by Leffler and Frosch in 1898. These researchers, using multiple dilutions of the filterable agent, showed its toxicity and made a conclusion about its corpuscular nature.
By the end of the 19th century, it became clear that a number of human diseases, such as rabies, smallpox, influenza, and yellow fever, are infectious, but their causative agents were not detected by bacteriological methods. Thanks to the work of Robert Koch (1843-1910), who pioneered the use of pure bacterial culture techniques, it became possible to distinguish between bacterial and non-bacterial diseases. In 1890, at the X Congress of Hygienists, Koch was forced to declare that “... with the diseases listed, we are not dealing with bacteria, but with organized pathogens that belong to a completely different group of microorganisms.” This statement by Koch indicates that the discovery of viruses was not a random event. Not only the experience of working with pathogens of unknown nature, but also an understanding of the essence of what was happening contributed to the formulation of the idea of the existence of an original group of pathogens of infectious diseases of a non-bacterial nature. It remained to experimentally prove its existence.
The first experimental evidence of the existence of a new group of pathogens of infectious diseases was obtained by our compatriot - plant physiologist Dmitry Iosifovich Ivanovsky (1864-1920) while studying mosaic diseases of tobacco. This is not surprising, since infectious diseases of an epidemic nature were often observed in plants. Back in 1883-84. The Dutch botanist and geneticist De Vries observed an epidemic of greening of flowers and suggested the infectious nature of the disease. In 1886, the German scientist Mayer, working in Holland, showed that the sap of plants suffering from mosaic disease, when inoculated, causes the same disease in plants. Mayer was sure that the culprit of the disease was a microorganism, and searched for it without success. In the 19th century, tobacco diseases caused enormous harm to agriculture in our country. In this regard, a group of researchers was sent to Ukraine to study tobacco diseases, which, as a student at St. Petersburg University, included D.I. Ivanovsky. As a result of studying the disease described in 1886 by Mayer as mosaic disease of tobacco, D.I. Ivanovsky and V.V. Polovtsev came to the conclusion that it represents two different diseases. One of them - "grouse" - is caused by a fungus, and the other is of unknown origin. The study of tobacco mosaic disease was continued by Ivanovsky at the Nikitsky Botanical Garden under the leadership of Academician A.S. Famytsina. Using the juice of a diseased tobacco leaf, filtered through a Chamberlant candle, which retains the smallest bacteria, Ivanovsky caused a disease of tobacco leaves. Cultivation of the infected juice on artificial nutrient media did not produce results and Ivanovsky comes to the conclusion that the causative agent of the disease is of an unusual nature - it is filtered through bacterial filters and is not able to grow on artificial nutrient media. Warming the juice at a temperature from 60 °C to 70 °C deprived it of infectivity, which indicated the living nature of the pathogen. Ivanovsky first named the new type of pathogen “filterable bacteria” (Figure 1). Results of the work of D.I. Ivanovsky were used as the basis for his dissertation, presented in 1888, and published in the book “On Two Diseases of Tobacco” in 1892. This year is considered the year of the discovery of viruses.
A – Electron micrograph after oblique deposition with carbon and platinum; 65,000´. (Photo by N. Frank.) B – Model. (Karlson, Kurzes Lehrbuch der Biochemie, Stuttgart, Thieme, 1980).
Figure 1 – Tobacco mosaic virus
For a certain period of time, in foreign publications, the discovery of viruses was associated with the name of the Dutch scientist Beijerinck (1851-1931), who also studied tobacco mosaic disease and published his experiments in 1898. Beijerinck placed the filtered juice of an infected plant on the surface of an agar, incubated and obtained bacterial colonies on its surface. After this, the top layer of agar with bacterial colonies was removed, and the inner layer was used to infect a healthy plant. The plant is sick. From this, Beijerinck concluded that the cause of the disease was not bacteria, but some liquid substance that could penetrate inside the agar, and called the pathogen “liquid living contagion.” Due to the fact that Ivanovsky only described his experiments in detail, but did not pay due attention to the nonbacterial nature of the pathogen, a misunderstanding of the situation arose. Ivanovsky’s work became famous only after Beijerinck repeated and expanded his experiments and emphasized that Ivanovsky was the first to prove the non-bacterial nature of the causative agent of the most typical viral disease of tobacco. Beijerinck himself recognized the primacy of Ivanovsky and the current priority of the discovery of viruses by D.I. Ivanovsky is recognized throughout the world.
Word VIRUS means poison. This term was also used by Pasteur to denote an infectious principle. It should be noted that at the beginning of the 19th century, all pathogenic agents were called the word virus. Only after the nature of bacteria, poisons and toxins became clear, the terms “ultravirus” and then simply “virus” began to mean “a new type of filterable pathogen.” The term “virus” took root widely in the 30s of our century.
It is now clear that viruses are characterized by ubiquity, that is, ubiquity of distribution. Viruses infect representatives of all living kingdoms: humans, vertebrates and invertebrates, plants, fungi, bacteria.
The first report related to bacterial viruses was made by Hankin in 1896. In the Chronicle of the Pasteur Institute, he stated that “... the water of some rivers of India has a bactericidal effect...”, which is no doubt related to bacterial viruses. In 1915, Twort in London, while studying the causes of lysis of bacterial colonies, described the principle of transmission of “lysis” to new cultures over a series of generations. His work, as often happens, was virtually unnoticed, and two years later, in 1917, the Canadian de Hérelle rediscovered the phenomenon of bacterial lysis associated with a filtering agent. He called this agent a bacteriophage. De Herelle assumed that there was only one bacteriophage. However, research by Barnett, who worked in Melbourne in 1924-34, showed a wide variety of bacterial viruses in physical and biological properties. The discovery of the diversity of bacteriophages has generated great scientific interest. At the end of the 30s, three researchers - physicist Delbrück, bacteriologists Luria and Hershey, working in the USA, created the so-called “Phage Group”, whose research in the field of genetics of bacteriophages ultimately led to the birth of a new science - molecular biology.
The study of insect viruses has lagged significantly behind the virology of vertebrates and humans. It is now clear that viruses that infect insects can be divided into 3 groups: insect viruses themselves, animal and human viruses for which insects are intermediate hosts, and plant viruses that also infect insects.
The first insect virus to be identified was the silkworm jaundice virus (silkworm polyhedrosis virus, called Bollea stilpotiae). As early as 1907, Provacek showed that a filtered homogenate of diseased larvae was infectious for healthy silkworm larvae, but it was not until 1947 that the German scientist Bergold discovered rod-shaped viral particles.
One of the most fruitful studies in the field of virology is Reed's study of the nature of yellow fever on US Army volunteers in 1900-1901. It has been convincingly demonstrated that yellow fever is caused by a filterable virus that is transmitted by mosquitoes and mosquitoes. It was also found that mosquitoes remained non-infectious for two weeks after absorbing infectious blood. Thus, the external incubation period of the disease (the time required for virus reproduction in an insect) was determined and the basic principles of the epidemiology of arbovirus infections (viral infections transmitted by blood-sucking arthropods) were established.
The ability of plant viruses to reproduce in their vector, an insect, was demonstrated in 1952 by Maramorosh. The researcher, using insect injection techniques, convincingly demonstrated the ability of the aster jaundice virus to multiply in its vector, the six-spotted cicada.
Stages of development of virology
The history of achievements in virology is directly related to the success of the development of the methodological base of research.
The end of the 19th - the beginning of the 20th century. The main method of identifying viruses during this period was the method of filtration through bacteriological filters (Chamberlan candles), which were used as a means of separating pathogens into bacteria and non-bacteria. Using filterability through bacteriological filters, the following viruses were discovered:
– 1892 – tobacco mosaic virus;
– 1898 – foot and mouth disease virus;
– 1899 – rinderpest virus;
– 1900 – yellow fever virus;
– 1902 – fowl and sheep pox virus;
– 1903 – rabies virus and swine fever virus;
– 1904 – human smallpox virus;
– 1905 – canine distemper virus and vaccine virus;
– 1907 – dengue virus;
– 1908 – smallpox and trachoma virus;
– 1909 – polio virus;
– 1911 – Rous sarcoma virus;
– 1915 – bacteriophages;
– 1916 – measles virus;
– 1917 – herpes virus;
– 1926 – vesicular stomatitis virus.
30s– the main virological method used for the isolation of viruses and their further identification are laboratory animals (white mice - for influenza viruses, newborn mice - for Coxsackie viruses, chimpanzees - for hepatitis B virus, chickens, pigeons - for oncogenic viruses, gnotobiont piglets – for intestinal viruses, etc.). The first to systematically use laboratory animals in the study of viruses was Pasteur, who, back in 1881, conducted research on inoculating material from rabies patients into the brain of a rabbit. Another milestone was work on the study of yellow fever, which resulted in the use of newborn mice in virological practice. The culmination of this cycle of work was the isolation by Cycles in 1948 of a group of epidemic myalgia viruses using suckling mice.
1931 - chicken embryos, which are highly sensitive to influenza, smallpox, leukemia, chicken sarcoma and some other viruses, began to be used as an experimental model for isolating viruses. And currently, chicken embryos are widely used to isolate influenza viruses.
1932 - English chemist Alford creates artificial finely porous colloidal membranes - the basis for the ultrafiltration method, with the help of which it became possible to determine the size of viral particles and differentiate viruses on this basis.
1935 - the use of the centrifugation method made it possible to crystallize the tobacco mosaic virus. Currently, centrifugation and ultracentrifugation methods (acceleration at the bottom of the tube exceeds 200,000 g) are widely used for the isolation and purification of viruses.
In 1939, an electron microscope with a resolution of 0.2 to 0.3 nm was used for the first time to study viruses. The use of ultrathin tissue sections and the method of negative contrasting of aqueous suspensions made it possible to study the interaction of viruses with cells and to study the structure (architecture) of virions. The information obtained using the electron microscope was significantly expanded by X-ray diffraction analysis of crystals and pseudocrystals of viruses. The improvement of electron microscopes culminated in the creation of scanning microscopes that make it possible to obtain three-dimensional images. Using electron microscopy, the architecture of virions and the features of their penetration into the host cell were studied.
During this period, the bulk of viruses were discovered. Examples include the following:
– 1931 – swine influenza virus and equine western encephalomyelitis virus;
– 1933 – human influenza virus and eastern equine encephalomyelitis virus;
– 1934 – mumps virus;
– 1936 – mouse mammary cancer virus;
– 1937 – tick-borne encephalitis virus.
40s. In 1940, Hoagland and his colleagues discovered that the vaccinia virus contains DNA but not RNA. It became obvious that viruses differ from bacteria not only in size and inability to grow without cells, but also in that they contain only one type of nucleic acid - DNA or RNA.
1941 - American scientist Hurst discovered the phenomenon of hemagglutination (erythrocyte gluing) using a model of the influenza virus. This discovery formed the basis for the development of methods for detecting and identifying viruses and contributed to the study of virus-cell interactions. The principle of hemagglutination is the basis of a number of methods:
HRA - hemagglutination reaction - used to detect and titrate viruses;
HAI - hemagglutination inhibition reaction - is used to identify and titrate viruses.
1942 - Hearst discovers the presence of an enzyme in the influenza virus, which is later identified as neuraminidase.
1949 – discovery of the possibility of culturing animal tissue cells under artificial conditions. In 1952, Enders, Weller and Robbins received the Nobel Prize for developing the cell culture method.
The introduction of the cell culture method into virology was an important event that made it possible to obtain cultured vaccines. Of the currently widely used cultural live and killed vaccines created on the basis of attenuated strains of viruses, vaccines against polio, mumps, measles and rubella should be noted.
The creators of polio vaccines are American virologists Sabin (a trivalent live vaccine based on attenuated strains of polioviruses of three serotypes) and Salk (a killed trivalent vaccine). In our country, Soviet virologists M.P. Chumakov and A.A. Smorodintsev developed a technology for the production of live and killed polio vaccines. In 1988, the World Health Assembly set WHO the goal of eradicating polio worldwide by completely stopping the circulation of wild poliovirus. To date, enormous progress has been made in this direction. The use of global vaccination against polio using “round” vaccination schemes made it possible not only to radically reduce the incidence, but also to create areas free from the circulation of wild poliovirus.
Viruses discovered:
– 1945 – Crimean hemorrhagic fever virus;
– 1948 – Coxsackie viruses.
50s. In 1952, Dulbecco developed a method for titrating plaques in a monolayer of chicken embryo cells, which introduced a quantitative aspect to virology. 1956-62 Watson, Caspar (USA) and Klug (Great Britain) develop a general theory of the symmetry of viral particles. The structure of the viral particle has become one of the criteria in the virus classification system.
This period was characterized by significant advances in the field of bacteriophages:
– induction of the prophage of lysogenizing phages was established (Lvov et al., 1950);
– it has been proven that infectivity is inherent in phage DNA, and not in the protein shell (Hershey, Chase, 1952);
– the phenomenon of general transduction was discovered (Zinder, Lederberg, 1952).
The infectious tobacco mosaic virus was reconstructed (Frenkel-Conrad, Williams, Singer, 1955-1957), and in 1955 the polio virus was obtained in crystalline form (Shaffer, Shwerd, 1955).
Viruses discovered:
– 1951 – murine leukemia viruses and ECHO;
– 1953 – adenoviruses;
– 1954 – rubella virus;
– 1956 – parainfluenza viruses, cytomegalovirus, respiratory syncytial virus;
– 1957 – polyoma virus;
– 1959 – Argentine hemorrhagic fever virus.
60s characterized by the flourishing of molecular biological research methods. Advances in the field of chemistry, physics, molecular biology and genetics formed the basis of the methodological base of scientific research, which began to be used not only at the level of techniques, but also entire technologies, where viruses act not only as an object of research, but also as a tool. Not a single discovery in molecular biology is complete without a viral model.
1967 – Cates and McAuslan demonstrate the presence of a DNA-dependent RNA polymerase in the vaccinia virion. The following year, RNA-dependent RNA polymerase was discovered in reoviruses, and then in paramyxo- and rhabdoviruses. In 1968, Jacobson and Baltimore established that polioviruses have a genomic protein connected to RNA; Baltimore and Boston established that the poliovirus genomic RNA is translated into a polyprotein.
Viruses discovered:
– 1960 – rhinoviruses;
– 1963 – Australian antigen (HBsAg).
70s. Baltimore, simultaneously with Temin and Mizutani, reported the discovery of the enzyme reverse transcriptase (revertase) in RNA-containing oncogenic viruses. It is becoming possible to study the genome of RNA viruses.
The study of gene expression in eukaryotic viruses provided fundamental information about the molecular biology of eukaryotes themselves - the existence of the cap structure of mRNA and its role in RNA translation, the presence of a polyadenylate sequence at the 3" end of mRNA, splicing and the role of enhancers in transcription were first identified in the study of animal viruses.
1972 - Berg publishes a report on the creation of a recombinant DNA molecule. A new branch of molecular biology is emerging - genetic engineering. The use of recombinant DNA technology makes it possible to obtain proteins that are important in medicine (insulin, interferon, vaccines). 1975 – Köhler and Milstein produce the first lines of hybrids producing monoclonal antibodies (MAbs). The most specific test systems for diagnosing viral infections are being developed based on mAbs. 1976 - Blumberg receives the Nobel Prize for the discovery of HBsAg. It has been established that hepatitis A and hepatitis B are caused by different viruses.
Viruses discovered:
– 1970 – hepatitis B virus;
– 1973 – rotaviruses, hepatitis A virus;
– 1977 – hepatitis delta virus.
80s. Development of the ideas laid down by domestic scientist L.A. Zilber's idea that the occurrence of tumors may be associated with viruses. The components of viruses responsible for the development of tumors are called oncogenes. Viral oncogenes have proven to be among the best model systems that help study the mechanisms of oncogenetic transformation of mammalian cells.
– 1985 – Mullis receives the Nobel Prize for the discovery of the polymerase chain reaction (PCR). This is a molecular genetic diagnostic method, which has also made it possible to improve the technology for obtaining recombinant DNA and discover new viruses.
Viruses discovered:
– 1983 – human immunodeficiency virus;
– 1989 – hepatitis C virus;
– 1995 – the hepatitis G virus was discovered using PCR.
Related information.
The achievements of modern virology are enormous. Scientists are increasingly and more deeply and successfully understanding the subtle structure, biochemical composition and physiological properties of these ultramicroscopic living beings, their role in nature, human life, animals, and plants. Oncovirology persistently and successfully studies the role of viruses in the occurrence of tumors (cancer), trying to solve this problem of the century.
By the beginning of the 21st century, more than 6 thousand viruses belonging to more than 2,000 species, 287 genera, 73 families and 3 orders. For many viruses, their structure, biology, chemical composition and replication mechanisms have been studied. The discovery and research of new viruses continues, and they continue to amaze with their diversity. So in 2003, the largest known virus, mimivirus, was discovered.
The discovery of a large number of viruses required creating their collections and museums. The largest among them are in Russia (state collection of viruses at the D.I. Ivanovsky Institute of Virology in Moscow), USA (Washington), Czech Republic (Prague), Japan (Tokyo), Great Britain (London), Switzerland (Lausanne) and Germany (Brunschweig). The results of scientific research in the field of virology are published in scientific journals and discussed at international congresses organized every 3 years (first held in 1968). In 1966, at the 9th International Congress of Microbiology, the International Committee on Taxonomy of Viruses (ICTV) was elected for the first time.
Within the framework of general, that is, molecular virology, the study of the fundamental principles of interaction between viruses and cells continues. Advances in molecular biology, virology, genetics, biochemistry and bioinformatics have shown that the importance of viruses is not limited to the fact that they cause infectious diseases.
It has been shown that the replication features of some viruses lead to the virus capturing cellular genes and transferring them into the genome of another cell - horizontal transfer of genetic information, which can have consequences both in evolutionary terms and in terms of malignant degeneration of cells.
When sequencing the genome of humans and other mammals, a large number of repeating nucleotide sequences were identified, which are defective viral sequences - retrotransposons (endogenous retroviruses), which may contain regulatory sequences that affect the expression of neighboring genes. Their discovery and study led to active discussion and research into the role of viruses in the evolution of all organisms, in particular in the evolution of humans.
A new direction in virology is ecology of viruses. Detecting viruses in nature, identifying them and estimating their abundance is a very difficult task. At present, some methodological techniques have been developed that make it possible to estimate the amount of certain groups of viruses, in particular bacteriophages, in natural samples and to trace their fate. Preliminary data have been obtained indicating that viruses have a significant impact on numerous biogeochemical processes and effectively regulate the abundance and species diversity of bacteria and phytoplankton. However, the study of viruses in this aspect has just begun, and there are still many unresolved problems in this area of science.
The achievements of general virology have given a powerful impetus to the development of its applied areas. Virology has developed into a vast field of knowledge important for biology, medicine and agriculture.
Virologists diagnose viral infections in humans and animals, study their spread, and develop methods of prevention and treatment. The greatest achievement was the creation of vaccines against polio, smallpox, rabies, hepatitis B, measles, yellow fever, encephalitis, influenza, mumps, and rubella. A vaccine has been created against the papilloma virus, which is associated with the development of one type of cancer. Thanks to vaccination, smallpox has been completely eradicated. International programs for the complete eradication of polio and measles are being implemented. Methods are being developed for the prevention and treatment of hepatitis and human immunodeficiency (AIDS). Data on substances with antiviral activity are accumulating. Based on them, a number of drugs have been created for the treatment of AIDS, viral hepatitis, influenza, and diseases caused by the herpes virus.
The study of plant viruses and the characteristics of their distribution throughout the plant has led to the creation of a new direction in agriculture - the production of virus-free planting material. Meristem technologies, which make it possible to grow plants free of viruses, are currently used for potatoes and a number of fruit and flower crops.
Of exceptional importance at this stage is the knowledge accumulated about the structure of viruses and their genomes for the development of genetic engineering. A notable example of this is the use of bacteriophage lambda to produce libraries of cloned sequences. In addition, based on the genomes of various viruses, a large number of genetically engineered vectors have been created and continue to be created to deliver foreign genetic information into cells. These vectors are used for scientific research, for the accumulation of foreign proteins, especially in bacteria and plants, and for gene therapy. Genetic engineering uses some viral enzymes that are now produced commercially.
Small sizes and the ability to form regular structures have opened up the prospect of using viruses in nanotechnology to produce new bioinorganic materials: nanotubes, nanoconductors, nanoelectrodes, nanocontainers, for encapsidation of inorganic compounds, magnetic nanoparticles and inorganic nanocrystals of strictly controlled sizes. New materials can be created through the interaction of regularly organized viral protein structures with metal-containing inorganic compounds. “Spherical” viruses can serve as nanocontainers for storing and delivering drugs and therapeutic genes into cells. Surface modified infectious virions and viral substructures can be used as nanotools (for example, for the purposes of biocatalysis or the production of safe vaccines).
17. Bacteriophage titer, methods for its determination. Identification of animal and plant viruses.
The bacteriophage titer is the number of active phage particles per unit volume of the material being studied. To determine the bacteriophage titer, the agar layer method is most widely used when working with bacteriophages. , proposed by A. Grazia in 1936. This method is distinguished by its ease of implementation and accuracy of the results obtained and is also successfully used for the isolation of bacteriophages.
The essence of the method is that a bacteriophage suspension is mixed with a culture of sensitive bacteria, added to a low concentration agar (“soft agar”) and layered on the surface of a previously prepared 1.5% nutrient agar in a Petri dish. Water (“hungry”) 0.6% was used as the top layer in the classical Grazia method. - nd agar. Currently, 0.7% nutrient agar is most often used for these purposes. When incubated for 6-18 hours, the bacteria multiply within the upper "soft" layer of agar in the form of many colonies, receiving nutrition from the lower layer of 1.5% nutrient agar, which is used as a substrate. The low concentration of agar in the upper layer creates a reduced viscosity, which promotes good diffusion of phage particles and their infection of bacterial cells. Infected bacteria undergo lysis, resulting in progeny phage that again infects bacteria in close proximity to them. The formation of a negative colony for T-group phages is caused by only one bacteriophage particle, and, therefore, the number of negative colonies serves as a quantitative indicator of the content of plaque-forming units in the test sample.
The culture of phage-sensitive bacteria is used in the logarithmic growth phase in a minimal amount to ensure a continuous lawn of bacteria. The ratio of the number of phage particles and bacterial cells (multiplicity of infection) for each phage-bacterium system is selected experimentally so that 50-100 negative colonies are formed on one plate.
To titrate a bacteriophage, a single-layer method can also be used, which consists of adding suspensions of bacteria and bacteriophage to the surface of a plate with nutrient agar, after which the mixture is distributed with a glass spatula. However, this method is inferior in accuracy to the agar layer method and therefore is not widely used.
Technique for titration and cultivation of bacteriophages. To determine the bacteriophage titer, the initial phage suspension is sequentially diluted in a buffer solution or in broth (dilution step 10 -1). For each dilution, use a separate pipette, and the mixture is stirred vigorously. From each dilution of the suspension, the phage is “seeded” onto a lawn of sensitive bacteria E. coli B. To do this, 1 ml of the diluted phage is added to a test tube with 3 ml of “soft agar” melted and cooled to 48-50°C, after which it is added to each test tube 0.1 ml of a culture of a sensitive microorganism (E. coli B) in the logarithmic growth phase. The contents are mixed by rotating the test tube between the palms and avoiding the formation of bubbles. Then quickly pour it onto the surface of the agar (1.5%) nutrient medium in a Petri dish and distribute it evenly over it, gently shaking the dish. When titrating using the agar layer method, at least two plates of the same phage dilution should be inoculated in parallel. After the top layer has hardened, the cups are turned upside down and placed in a thermostat with a temperature of 37°C, optimal for the development of sensitive bacteria. The results are recorded after 18-20 hours of incubation.
The number of negative colonies is counted in the same way as counting bacterial colonies, and the phage titer is determined using the formula:
Where N is the number of phage particles in 1 ml of the test material; n is the average number of negative colonies per plate; D - dilution number; V is the volume of the inoculated sample, ml.
In the case when it is necessary to determine the multiplicity of infection, the titer of viable cells of E. coli B bacteria in 1 ml of nutrient broth is determined in parallel. To do this, dilute the initial suspension of bacterial cells to 10 -6 and inoculate it (0.1 ml) in parallel onto 2 cups. After incubation at 37 °C for 24 hours, the number of colonies formed on a Petri dish is counted and the cell titer is determined.
To isolate viruses from humans, animals and plants, the material under study is introduced into the body of experimental animals and plants sensitive to viruses or infects cell (tissue) cultures and organ cultures. The presence of the virus is proven by characteristic damage in experimental animals (or plants), and in tissue cultures - by damage to cells, the so-called cytopathic effect, which is recognized by microscopic or cytochemical examination. With V. and. The “plaque method” is used - observation of defects in the cell layer caused by destruction or damage to cells in areas of virus accumulation. Virions, which have a characteristic structure among different viruses, can be identified by electron microscopy. Further identification of viruses is based on the integrated use of physical, chemical and immunological methods. Thus, viruses differ in sensitivity to ether, which is associated with the presence or absence of lipids in their shells. The type of virus nucleic acid (RNA and DNA) can be determined by chemical or cytochemical methods. To identify viral proteins, serological reactions are used with sera obtained by immunizing animals with the corresponding viruses. These reactions make it possible to recognize not only types of viruses, but also their varieties. Serological research methods make it possible to diagnose a viral infection in humans and higher animals by the presence of antibodies in the blood and to study the circulation of viruses among them. To identify latent (hidden) viruses of humans, animals, plants and bacteria, special research methods are used.
Municipal state educational institution
"Secondary school No. 3"
Stavropol region, Stepnovsky district,
Bogdanovka village
MKOU secondary school No. 3, 10th grade student
Scientific supervisor:
Toboeva Natalya Konstantinovna
teacher of geography, biology, MKOU secondary school No. 3
I .Introduction
II.Main part:
1. Discovery of viruses
2.Origin of viruses
3. Structure
4.Penetration into the cell
5.Flu
6. Chicken pox 7. Tick-borne encephalitis 8. The future of virology
III.Conclusion
IV. References
V.Appendix
Object of study:
Non-cellular life forms are viruses.
Subject of research:
The present and future of virology.
Purpose of the work:
Find out the significance of virology at the present time and determine its future. The set goal could be achieved as a result of solving the following tasks:
1) study of literature covering the structure of viruses as non-cellular life forms;
2) research into the causes of viral diseases, as well as their prevention.
This determined the topic of my research.
I. Introduction.
The action-packed and fascinating history of virology is characterized by triumphant victories, but, unfortunately, also defeats. The development of virology is associated with the brilliant successes of molecular genetics.
The study of viruses has led to an understanding of the fine structure of genes, decoding of the genetic code, and the identification of mutation mechanisms.
Viruses are widely used in genetic engineering and research.
But their cunning and ability to adapt know no bounds, their behavior in each case is unpredictable. The victims of viruses are millions of people who died from smallpox, yellow fever, AIDS and other diseases. Much remains to be discovered and learned. And yet, the main successes in virology have been achieved in the fight against specific diseases. That is why scientists say that virology will take a leading place in the third millennium.
What has virology given to humanity in the fight against its formidable enemy - the virus? What is its structure, where and how does it live, how does it reproduce, what other “surprises” does it prepare? I considered these questions in my work.
II.Main part:
1. Discovery of viruses.
The discoverer of the world of viruses was the Russian botanist D.I. Ivanovsky. In 1891-1892 he persistently searched for the causative agent of tobacco mosaic disease. The scientist examined the liquid obtained by rubbing diseased tobacco leaves. I filtered it through filters that were not supposed to let a single bacteria through. Patiently, he pumped liters of juice taken from mosaic tobacco leaves into hollow bacterial filters made of fine porcelain, reminiscent of long candles. The walls of the filter sweated with transparent droplets that flowed into a pre-sterilized vessel. By lightly rubbing, the scientist applied a drop of this filtered juice to the surface of the tobacco leaf. After 7-10 days, previously healthy plants showed undoubted signs of mosaic disease. A drop of filtered juice from an infected plant affected any other tobacco bush with a mosaic disease. The infestation could pass from plant to plant endlessly, like a flame of fire from one thatched roof to another.
Subsequently, it was possible to establish that many other viral pathogens of infectious diseases in humans, animals and plants are capable of passing through, which could be seen through the most advanced light microscopes. Particles of various viruses could only be seen through the window of an all-seeing device - an electron microscope, which provides a magnification of hundreds of thousands of times.
D.I. himself Ivanovsky did not attach much importance to this fact, although he described his experience in detail.
His work gained fame after the Dutch botanist and microbiologist Martin Beijerinck confirmed the results of D. I. Ivanovsky’s research in 1899. M. Beyerinck proved that the mosaic of tobacco can be transferred from one plant to another using filtrates. These studies marked the beginning of the study of viruses and the emergence of virology as a science.
2. Origin of viruses.
3. Structure.
Being completely primitive creatures, viruses have all the basic properties of living organisms. They reproduce offspring similar to the original parental forms, although their method of reproduction is peculiar and differs in many respects from what is known about the reproduction of other creatures. Their metabolism is closely related to the metabolism of host cells. They have heredity characteristic of all living organisms. Finally, they, like all other living beings, are characterized by variability and adaptability to changing environmental conditions.
The largest viruses (for example, smallpox viruses) reach a size of 400-700 nm and are close in size to small bacteria, the smallest (causative agents of polio, encephalitis, foot-and-mouth disease) measure only tens of nanometers, i.e. are close to large protein molecules, in particular blood hemoglobin molecules.
Viruses come in a variety of shapes, from spherical to filamentous. Electron microscopy allows not only to see viruses, determine their shapes and sizes, but also to study their spatial structure - molecular architectonics.
Viruses have a relatively simple composition: nucleic acid (RNA or DNA), protein; more complex structures contain carbohydrates and lipids, and sometimes have a number of their own enzymes.
As a rule, the nucleic acid is located in the center of the viral particle and is protected from adverse effects by a protein shell - capsomers. Electron microscope observations showed that the virus particle
(or virions) come in several basic types in shape.
Some viruses (usually the simplest ones) resemble regular geometric bodies. Their protein shell almost always approaches the shape of an icosahedron (regular twenty-sided structure) with faces of equilateral triangles. These virions are called cubic (such as the polio virus). The nucleic acid of such a virus is often twisted into a ball. Particles of other viruses are shaped like oblong rods. In this case, their nucleic acid is surrounded by a cylindrical capsid. Such virions are called helical virions (for example, tobacco mosaic virus).
Viruses of a more complex structure, in addition to the icosahedral or helical capsid, also have an outer shell, which consists of a variety of proteins (many of them enzymes), as well as lipids and carbons.
The physical structure of the outer shell is very varied and is not as compact as that of the capsid. For example, the herpes virus is an enveloped helical virion. There are viruses with an even more complex structure. Thus, the smallpox virus does not have a visible capsid (protein shell), but its nucleic acid is surrounded by several shells.
4.Penetration into the cell.
As a rule, the penetration of the virus into the cytoplasm of the cell is preceded by its binding to a special receptor protein located on the cell surface. Binding to the receptor occurs due to the presence of special proteins on the surface of the viral cell. The area of the cell surface to which the virus has attached is immersed in the cytoplasm and turns into a vacuole. A vacuole is a wall that consists of a cytoplasmic membrane that can merge with other vacuoles or the nucleus. This way the virus is delivered to any part of the cell.
The receptor mechanism for virus penetration into the cell ensures the specificity of the infectious process. The infectious process begins when viruses that have entered the cell begin to multiply, i.e. The viral genome is reduplicated and the capsid self-assembles. For reduplication to occur, the nucleic acid must be freed from the capsid. After the synthesis of a new nucleic acid molecule, it is dressed with viral proteins synthesized in the cytoplasm of the host cell - a capsid is formed.
The accumulation of viral particles leads to elimination from the cell. For some viruses, this occurs through an “explosion”, in which the integrity of the cell is disrupted and it dies. Other viruses are released in a manner reminiscent of budding. In this case, the cells can maintain their viability.
Bacteriophage viruses have a different way of entering cells. The bacteriophage inserts a full rod into the cell and pushes out the DNA (or RNA) found in its head through it. The bacteriophage genome enters
cytoplasm, and the capsid remains outside. In the bacterial cytoplasm, the reduplication of the bacteriophage genome, the synthesis of its proteins and the formation of the capsid begin. After a certain period of time, the bacterial cell dies and mature particles enter the environment.
5.Flu.
Influenza is an acute infectious disease, the causative agent of which is a filter virus, causing general intoxication and damage to the mucous membrane of the upper respiratory tract.
It has now been established that the influenza virus has several serological types, differing in their antigenic structure.
There are the following types of influenza virus: A, B, C, D. Virus A has 2 subtypes, designated:A 1 and A2.
The influenza virus outside the human body is unstable and dies quickly. The virus dried in a vacuum can persist for a long time.
Disinfectants quickly destroy the virus; ultraviolet irradiation and heat also have a detrimental effect on the virus.
Allow the possibility of infection from a virus carrier. The virus is transmitted from a sick person to a healthy person through airborne droplets. Coughing and sneezing contribute to the spread of infection.
Viral influenza epidemics most often occur during the cold season.
A person with the flu is contagious for 5-7 days. All people who have not had the flu are susceptible to this disease. After suffering from the flu, immunity remains for 2-3 years.
The incubation period is short - from several hours to 3 days. Most often 1-2 days.
Usually there are no prodromes, and a sudden onset is typical. Chills, headache, general weakness appear, and the temperature rises to 39-40 degrees. Patients complain of pain when rotating the eyes, aching muscle joints, disturbed sleep, and sweating. All this indicates general intoxication with the involvement of the nervous system in the process.
The central nervous system is especially sensitive to the toxic effects of the influenza virus, which is clinically expressed in severe adynamia, irritability, and decreased sense of smell and taste.
On the part of the digestive tract, the phenomena of influenza intoxication also differ: decreased appetite, stool retention, and sometimes, more often in young children, diarrhea.
The tongue is coated and slightly swollen, which leads to the appearance of teeth marks along the edges. The temperature remains elevated for 3-5 days and, in the absence of complications, drops to normal gradually or drops critically.
After 1-2 days, a runny nose, laryngitis, and bronchitis may appear. Bleeding from the nose is common. The cough is dry at first and turns into a cough with sputum. Vascular disorders are expressed in the form of low blood pressure, pulse instability and disturbances in its rhythm.
Uncomplicated flu usually ends within 3-5 days, however, full recovery takes 1-2 weeks.
Like any infection, influenza can occur in mild, severe, hypertoxic and fulminant forms.
Along with this, viral flu can be extremely mild and spread on the legs, ending within 1-2 days. These forms of influenza are called erased.
Influenza infection can cause complications in various organ systems. Most often in children, the flu is complicated by pneumonia, otitis media, which is accompanied by fever, anxiety, and sleep disturbances.
Complications from the peripheral nervous system are expressed in the form of neuralgia, neuritis, radiculitis.
Treatment:
The patient must be provided with bed rest and rest. Bed rest must be maintained for some time, even after the temperature drops. Systematic ventilation of the room, daily warm or hot baths, good nutrition - all this increases the body's resistance to fighting the flu.
Specific treatment of viral influenza is carried out using the anti-influenza polyvalent serum proposed by A.A. Smorodintsev.
Among the symptomatic remedies for headache, muscle and joint pain, as well as neurological pain, pyramidon, phenacetin, and aspirin with caffeine are prescribed.
In case of severe toxicosis, intravenous glucose is prescribed. For uncomplicated influenza, antibiotics are not used, because They no longer work on the virus. For a dry cough, hot milk with soda or Borjomi is useful.
Prevention:
Patients should be isolated at home or in hospitals. If the patient is left at home, it is necessary to place him in a separate room or separate his bed with a screen or sheet. Caregivers should wear a gauze mask covering the nose and mouth.
6. Chicken pox.
Chickenpox is an acute infectious disease caused by a virus and characterized by a macular vesicular rash on the skin and mucous membranes.
The causative agent of chickenpox is a filter virus and is found in chickenpox vesicles and in the blood. The virus is unstable and exposed to various environmental influences and dies quickly.
The source of infection is the patient, who is contagious during the period of rash and at the end of incubation. The infection is spread by airborne droplets. The disease is not transmitted through objects.
Immunity after chickenpox remains for life. The incubation period lasts from 11 to 21 days, with an average of 14 days.
In most cases, the disease begins immediately, and only sometimes there are precursors in the form of a moderate increase in temperature with symptoms of general malaise. Prodromes may be accompanied by a rash resembling scarlet fever or measles.
With a moderate rise in temperature, a spotted rash of varying sizes appears on different parts of the body - from a pinhead to a lentil. Over the next few hours, a bubble with transparent contents, surrounded by a red rim, forms at the site of the spots. Chickenpox blisters (vesicles) are located on unchanged skin, tender and soft to the touch. The contents of the bubble soon become cloudy, and the bubble itself bursts (2-3 days) and turns into a crust, which disappears after 2-3 weeks, usually leaving no scar. The rash and subsequent formation of blisters can be abundant, affecting the entire scalp, trunk, and limbs, while on the face and distal parts of the limbs they are less abundant.
The course of chickenpox is usually accompanied by a slight disturbance in the general condition of the patient. Each new rash causes an increase in temperature to 38° and above. At the same time, appetite decreases.
In addition to the skin, chicken rash can affect the mucous membranes of the oral cavity, conjunctiva, genitals, larynx, etc.
Treatment:
Bed linen must always be clean. Take warm baths (35°-37°) from weak solutions of potassium permanganate. The patient's hands should be clean with short-cut nails.
Individual bubbles are lubricated with iodine or potassium solution, 1% alcohol solution of brilliant green.
For purulent complications caused by secondary infection, treatment is carried out with antibiotics (penicillin, streptomycin, biomycin)
Prevention:
A person infected with chickenpox must be isolated at home. Disinfection is not carried out, the room is ventilated and subjected to wet cleaning.
7. Tick-borne encephalitis.
An acute viral disease characterized by damage to the gray matter of the brain and spinal cord. The reservoir for sources of infection are wild animals (mainly rodents) and ixodid ticks. Infection is possible not only by sucking on a tick, but also by consuming the milk of infected goats. The causative agent is an arbovirus. The gateway of infection is the skin (if ticks are sucked on) or the mucous membrane of the digestive tract (if there is alimentary infection). The virus hematogenously penetrates the central nervous system and causes the most pronounced changes in the nerve cells of the anterior horns of the cervical spinal cord and in the nuclei of the medulla oblongata.
The incubation period is from 8 to 23 days (usually 7-14 days). The disease begins acutely: chills, severe headache, and weakness appear. After encephalitis, lasting consequences may remain in the form of flaccid paralysis of the muscles of the neck and shoulder girdle.
Treatment:
Strict bed rest:
for mild forms - 7-10 days,
for moderate cases - 2-3 weeks,
for severe ones - even longer.
Prevention:
When a tick bites in an area unfavorable for encephalitis, it is necessary to administer anti-encephalitis gamma globulin. According to indications, preventive vaccination is carried out.
8.The future of virology.
What are the prospects for the development of virology in the 21st century? In the second half of the 20th century, progress in virology was associated with classical discoveries in biochemistry, genetics and molecular biology. Modern virology is intertwined with the successes of fundamental applied sciences, so its further development will follow the path of in-depth study of the molecular basis of the pathogenicity of viruses of new previously unknown pathogens (prions and virions), the nature and mechanisms of persistence of viruses, their ecology, the development of new and improvement of existing diagnostic methods and specific prevention of viral diseases.
There is currently no more important aspect in virology than the prevention of infections. Over the 100 years of the existence of the science of viruses and viral diseases, vaccines have undergone great changes, going from attenuated and killed vaccines from the time of Pasteur to modern genetically engineered and synthetic vaccine preparations. This direction will continue to develop, based on physicochemical genetic engineering and synthetic experiments with the goal of creating polyvalent vaccines that require minimal vaccinations as early as possible after birth. Chemotherapy will develop, an approach relatively new to virology. These drugs are so far useful only in isolated cases.
III. Conclusion.
Humanity faces many complex unsolved virological problems: hidden viral infections, viruses and tumors, etc. The level of development of virology today, however, is such that means of combating infections will definitely be found. It is very important to understand that viruses are not an element alien to living nature; they are a necessary component of the biosphere, without which adaptation, evolution, immune defense and other interactions of living objects with their environment would probably be impossible. Understanding viral diseases as pathologies of adaptation, the fight against them should be aimed at improving the status of the immune system, and not at destroying viruses.
Analysis of various literary sources and statistical data allowed us to draw the following conclusions:
viruses are autonomous genetic compounds of structure that are unable to develop outside the cell;
3) come in a variety of shapes and simple composition.
References:
1. Great Soviet Encyclopedia: T.8 / Ed. B.A. Vvedensky.
2. Denisov I.N., Ulumbaev E.G. Directory - a guide for a practicing physician. - M.: Medicine, 1999.
3. Zverev I.D. A book for reading on human anatomy, physiology and hygiene. - M.: Education, 1983.
4. Luria S. et al. General virology. - M.: Mir, 1981.
6. Pokrovsky V.I. Popular medical encyclopedia. - M.: Onyx, 1998.
7.Tokarik E.N. Virology: present and future // Biology at school. - 2000. - No. 2-3.
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