Table of contents of the topic "Virology. Reproduction of viruses. Genetics of viruses.":
1. Virology. History of virology. Chamberlan. RU. Pasteur. Ivanovsky.
2. Reproduction of viruses. Reproduction of +RNA viruses. Picornaviruses. Reproduction of picornaviruses.
3. Togaviruses. Reproduction of togaviruses. Retroviruses. Reproduction of retroviruses.
4. Reproduction of -RNA viruses. Reproduction of viruses with double-stranded RNA.
5. Reproduction of DNA viruses. Replicative cycle of DNA viruses. Reproduction of papovaviruses. Reproduction of adenoviruses.
6. Reproduction of herpes viruses. Replicative cycle of herpes viruses. Poxviruses. Reproduction of poxviruses.
7. Reproduction of the hepatitis B virus. Replicative cycle of the hepatitis B virus.
8. Genetics of viruses. Characteristics of viral populations. Gene pool of viral populations.

10. Genetic interactions between viruses. Recombination and redistribution of genes by viruses. Exchange of genome fragments by viruses. Antigenic shift.

Nucleic acids viruses are subject to mutations, that is, sudden inherited changes. The essence of these processes lies in violations of the genetic code in the form of changes in nucleotide sequences, their deletions (deletions), insertions or rearrangements of nucleotides or pairs in single- and double-stranded nucleic acid molecules. These disorders can be limited to individual nucleotides or spread over larger areas. Viruses have spontaneous and induced mutations. Their biological significance may be associated with the acquisition or loss of pathogenic properties, as well as with the acquisition of properties that deprive them of sensitivity to the action of the host’s defense mechanisms. Mutations that completely disrupt the synthesis or function of vital proteins lead to loss of reproductive ability and are otherwise known as lethal mutations. They are based on changes that lead to the appearance of meaningless codons (with disruption of the synthesis of the protein chain) or to the appearance of insertions or deletions (with profound violations of the genetic code). Mutations with loss of the ability to synthesize a certain protein or with disruption of its functions, which under certain conditions can lead to loss of the ability to reproduce, are called conditionally lethal.

Spontaneous mutations of viruses

Spontaneous mutations arise under the influence of various natural mutagens and occur with a frequency of l:10-8 viral particles. They can be observed more often in retroviruses, which is associated with a higher frequency of failures in reverse transcription.

Induced mutations of viruses

Induced mutations caused by various chemical agents and UV irradiation (for DNA viruses). There is no fundamental difference in genome rearrangement caused by spontaneous or induced mutations. It is generally accepted that the mutagens used only increase the frequency of spontaneous mutations. When classifying viral mutations, two different approaches are used: they are divided according to the nature of the genotype changes or according to the phenotypic changes that occur as a result of mutations. The study of changes in the genotype of viruses is rarely carried out, since this requires a detailed study of their genomes. The phenotypic manifestations of mutations are studied more often as they are more accessible for research.

Manifestation of virus mutations in phenotype

According to phenotypic manifestations virus mutations can be divided into four groups.

Mutations, which do not have phenotic manifestations, do not change the properties of viruses and are detected only with a special analysis.

Mutations, having a phenotypic manifestation (for example, a change in the size of plaques formed by viruses in cell culture or the thermostability of viruses). Mutations that increase or decrease pathogenicity can be divided into point mutations (localized in individual genes) and gene mutations (affecting larger areas of the genome).

Introduction

Increasing the safety and productivity of farm animals is impossible without further improvement of veterinary services for livestock farming. Among veterinary disciplines, virology plays an important role. A modern veterinarian must know not only the clinical and pathological side of the disease, but also have a clear understanding of viruses, their properties, laboratory diagnostic methods and the characteristics of post-infectious and post-vaccination immunity.

Viruses change their properties both under natural reproduction conditions and in experiments. Hereditary changes in the properties of viruses can be based on two processes: 1) mutation, i.e., a change in the nucleotide sequence in a certain part of the virus genome, leading to a phenotypically expressed change in the property; 2) recombination, i.e. the exchange of genetic material between two viruses that are close but differ in hereditary properties.

Mutation in viruses

Mutation is variability associated with changes in the genes themselves. It can have an intermittent, spasmodic character and lead to persistent changes in the hereditary properties of viruses. All virus mutations are divided into two groups:

· spontaneous;

· induced;

Based on their extent, they are divided into point and aberration (changes affecting a significant portion of the genome). Point mutations are caused by the replacement of a single nucleotide (for RNA viruses). Such mutations can sometimes revert, restoring the original genome structure.

However, mutational changes can also affect larger sections of nucleic acid molecules, i.e., several nucleotides. In this case, deletions, insertions and movements (translocation) of entire sections and even rotations of sections by 180° (so-called inversions), shifts of the reading frame - larger rearrangements in the structure of nucleic acids, and, consequently, violations of genetic information, can also occur.

But point mutations do not always lead to a change in phenotype. There are a number of reasons why such mutations may not appear. One of them is the degeneracy of the genetic code. The protein synthesis code is degenerate, i.e. some amino acids can be encoded by several triplets (codons). For example, the amino acid leucine can be encoded by six triplets. That is why, if in an RNA molecule due to some influences the triplet TsUU is replaced by TsUC, TsUA by TsUG, then the amino acid leucine will still be included in the molecule of the synthesized protein. Therefore, neither the structure of the protein nor its biological properties will be damaged.

Nature uses a unique language of synonyms and, replacing one codon with another, puts into them the same concept (amino acid), thus preserving its natural structure and function in the synthesized protein.

It’s another matter when some amino acid is encoded by only one triplet, for example, the synthesis of tryptophan is encoded by only one UGG triplet and there is no replacement, i.e., a synonym. In this case, some other amino acid is included in the protein, which can lead to the appearance of a mutant trait.

Aberration in phages is caused by deletions (loss) of a different number of nucleotides, from one pair to a sequence that determines one or more functions of the virus. Both spontaneous and induced mutations are also divided into direct and reverse.

Mutations can have different consequences. In some cases, they lead to changes in phenotypic manifestations under normal conditions. For example, the size of plaques under an agar coating increases or decreases; neurovirulence increases or decreases for a certain animal species; the virus becomes more sensitive to the action of a chemotherapeutic agent, etc.

In other cases, the mutation is lethal because it disrupts the synthesis or function of a vital virus-specific protein, such as viral polymerase.

In some cases, mutations are conditionally lethal, since the virus-specific protein retains its functions under certain conditions and loses this ability under non-permissive conditions. A typical example of such mutations are temperature-sensitive - ts-mutations, in which the virus loses the ability to reproduce at elevated temperatures (39 - 42 ° C), while maintaining this ability at normal growing temperatures (36 - 37 ° C).

Morphological or structural mutations may concern the size of the virion, the primary structure of viral proteins, changes in genes that determine early and late virus-specific enzymes that ensure virus reproduction.

According to their mechanism, mutations can also be different. In some cases, a deletion occurs, i.e., the loss of one or more nucleotides, in others, the incorporation of one or more nucleotides occurs, and in some cases, the replacement of one nucleotide with another.

Mutations can be direct or reverse. Direct mutations change the phenotype, and reverse mutations (reversions) restore it. True reversions are possible, when a reverse mutation occurs together with the primary damage, and pseudoreversions, if the mutation occurs in another part of the defective gene (intragenic suppression of mutation) or in another gene (extragenic suppression of mutation). Reversion is not a rare event, since revertants are usually more adapted to a given cellular system. Therefore, when obtaining mutants with specified properties, for example, vaccine strains, one has to take into account their possible reversion to the wild type.

Viruses differ from other representatives of the living world not only in their small size, selective ability to reproduce in living cells, structural features of the hereditary substance, but also in significant variability. Changes may relate to size, shape, pathogenicity, antigenic structure, tissue tropism, resistance to physical and chemical influences and other properties of viruses. The significance of the causes, mechanisms and nature of the change is of great importance in obtaining the necessary vaccine strains of viruses, as well as for the development of effective measures to combat viral epizootics, during which, as is known, the properties of viruses can significantly change one of the reasons for the relatively high ability of viruses to change their properties is that the hereditary substance of these microorganisms is less protected from environmental influences.

Mutation of viruses can occur as a result of chemical changes in cistrons or a violation of the sequence of their location in the structure of the viral nucleic acid molecule.

Depending on the conditions, a distinction is made between natural variability of viruses, observed under normal conditions of reproduction, and artificial, obtained in the process of numerous special passages or by exposing viruses to special physical or chemical factors (mutagens).

Under natural conditions, variability does not manifest itself equally in all viruses. This symptom is most pronounced in the influenza virus. The pangolin virus is subject to significant variability. This is evidenced by the presence of a large number of variants in different types of these viruses, and significant changes in its antigenic properties at the end of almost every epizootic.

The influenza virus is a champion of mutation
Every year, between three and five million people suffer from a severe form of influenza, up to 500 thousand of whom die from the flu itself or its complications (according to according to WHO). Flu shots, of course, significantly reduce the likelihood of getting sick. However

Unlike diseases such as measles or tuberculosis, immunity to which is developed after the first illness or vaccination and remains effective throughout life, many people get the flu almost every year.

The effectiveness of immunity is determined by how successfully the immune system recognizes and neutralizes the source of infection - a virus or bacteria. When first infected or vaccinated, the immune system learns to produce antibodies - molecules that bind to viral particles or bacteria and neutralize them. Once antibodies have been produced, the immune system keeps them “in service” for the rest of life.

Therefore, if a person becomes infected with the same infection again, the immune system is triggered and the infection is quickly neutralized. It is on this principle that vaccinations against measles, tuberculosis and other diseases work. Why does this mechanism fail with the influenza virus and why do you have to get vaccinated against the flu every year again?

This is due to two reasons. The first is the peculiarity of the interaction between our immune system and the virus. The surface of influenza virus particles is coated with molecules of two proteins called hemagglutinin (HA) and neuraminidase (NA) (see figure). Various variants of human influenza are classified by the type of these proteins, for example, H1N1 (hemagglutinin type 1, neuraminidase type 1). The human immune system is able to produce antibodies that successfully bind to these proteins. The problem is that these antibodies are quite finicky. Even small changes in the structure of HA and NA lead to the fact that antibodies lose the ability to bind to them and neutralize the virus.

From the point of view of the immune system, such modified versions of an already known virus look like completely new infections.

Secondly, the virus comes to the aid of an extremely useful property (and harmful to us) - the ability to quickly evolve. Like all other organisms, the influenza virus is subject to random mutations. This means that the genetic information of the descendant viruses is slightly different from the genetic information of the parent viruses. Thus, mutations constantly create new variants of the HA and NA proteins. However, unlike higher living organisms and many other viruses, influenza changes very quickly:

To accumulate as many mutations as mammalian proteins accumulate over millions of years, the influenza virus takes only a few years or even months.

Thus, we can observe the evolution of the influenza virus literally in real time.

Some of the flu mutations lead to the fact that the immune system, “trained” on the old strain, recognizes the mutated virus worse than the unmutated one. While the immune system effectively fights unmutated viruses, mutant viruses multiply and infect more and more people. This is the classic process of natural selection discovered by Charles Darwin.

The selection is carried out by the immune system, which, while protecting us, unwittingly does us a disservice.

After some time—usually two to three years—the old, non-mutated strain (virus variant) completely dies out, and the mutant virus becomes the new dominant strain. Most people's immune systems learn to cope with the new strain, and the cycle repeats. This “arms race” between the virus and the immune system has been going on for decades.

How to fight the flu

How to fight the flu in this case? There are several ways to help our immune system. First, antiviral drugs, such as oseltamivir (known under the brand name Tamiflu) or amantadine, are created to prevent the virus from reproducing inside cells. Unfortunately, viruses develop resistance to such drugs over time through the same process of mutation and natural selection:

Thus, almost the entire H1N1 subtype virus circulating in 2009 turned out to be resistant to oseltamivir (Tamiflu).

Secondly, scientists are trying to teach the immune system to recognize the less volatile parts of the virus (I wrote about this).

Third, scientists are trying to predict which strain of the virus will be most common next year. If we learn to do this, we can “retrain” our immune system as needed, vaccinating in advance against the strain that will dominate next season, and our immunity will get a head start in the arms race with the virus. Actually,

Already today, the World Health Organization updates the composition of the influenza vaccine every six months.

However, sometimes - once every few years - the predominant strain is not the one on the basis of which the vaccine was developed; in this case, the vaccination is less effective. Therefore, accurately predicting the strain that will be most common next year is one of the important tasks in the fight against influenza.

Our group (Jonathan Dushoff, Joshua Plotkin, Georgy Bazykin and Sergey Kryazhimsky) has been studying the evolution of the influenza virus and other organisms for several years. Our collaboration began at Princeton University in the laboratory of Professor Simon Levin, whose graduate students we were over the years. From the very beginning we were interested in both practical questions (how to most effectively predict the next dominant strain) and fundamental questions of evolution, e.g.

whether the evolution of influenza is directed or random.

The goal of our latest collaborative project was to determine the relationship between mutations occurring in different parts of the HA and NA proteins. The point is that the same mutation in, say, the HA protein can have very different consequences for the virus depending on whether mutations have occurred in other parts of the same protein. For example, mutation A allows the virus to become “invisible” to the immune system only when paired with mutation B, while each mutation on its own is useless for the virus. Such pairs of mutations, called epistatic, can be detected by analyzing statistical patterns in the genetic sequences of the virus. That's what we did.

Such analysis has only become possible in recent years, when the cost of “sequencing,” that is, identifying genetic sequences, has dropped sharply.

The number of genetic sequences of the influenza virus registered in the database has grown more than sixfold over the past five years to reach 150 thousand. This amount of data is sufficient to detect epistatic pairs of mutations that have occurred in the influenza virus over the past 100 years.

It turns out that the number of epistatic mutations in influenza is quite large, that is, only very specific variants of the virus that acquire the necessary combinations of mutations can avoid an attack by the immune system or gain immunity to an antiviral drug. For example, immunity to the drug oseltamivir appeared in 2009 only in viruses with at least three specific mutations in the NA protein.

From a practical point of view, the fact that mutations in the influenza virus are epistatic allows us to hope that in the near future we will learn to predict subsequent mutations from previous ones. As long as the virus "assembles" all the necessary mutations for a successful combination, we will be able to develop a new vaccine against a strain with the entire combination, which will spread only after several months or even years.

To determine the success of a particular mutation in combination with others, it is necessary to understand exactly how the interaction between mutations occurs

and how they, together and separately, affect the structure of the HA and NA proteins, as well as understand how the immune system reacts to modified versions of these proteins. These questions are now being actively researched, especially in Joshua Plotkin's group at the University of Pennsylvania, with which we actively collaborate, as well as other groups.

Influenza virus. Why does he mutate?

Every six out of ten sick children and four out of ten adults registered at the clinic suffer from the flu (it is clear that these data are far from complete: not everyone goes to the doctor!). Not only that, the flu “spurs up” cardiovascular and pulmonary diseases. The severe damage to people's health makes the problem extremely acute.

Viruses cause hundreds of diseases in animals, plants, and even bacteria. They account for the majority of infectious diseases of modern humans, and among them such terrible ones as smallpox, rabies, and polio.

The virus is very variable and adapts to its environment. The essence of this variability was deciphered relatively recently. The “outer dress” of the virus—its “outer” or, more precisely, “entrance” suit—is extremely practical. It could also be called a “hunting” suit: it is perfectly adapted for hunting cages. The suit is “sewn” from two main protein materials - hemagglutinins (with their help the virus attaches to the surface of the victim cell) and neuraminidases (whose enzymes remove the guard at the fortress gates when the virus needs to penetrate the cell and then exit it).

But the body also encounters the virus “by its clothes”: it is the protein shell that is the sphere of application of protective forces. As soon as at least some part of the protein coat of the virus is changed, the previously produced antibodies are no longer valid.

So why does the flu virus mutate?
There are two opposing points of view on the nature of the variability of the influenza virus.

Here is the first one.

In laboratory experiments, sensitive cells were infected with influenza virus containing different neuraminidases. As a result, we obtained not only exact copies of the original viruses, but also viruses with rearranged fragments. The mechanism of such rearrangement (recombination) is more or less clear.

The influenza virus nucleic acid strand is made up of eight separate fragments. Each of them is replaced relatively easily... If a fragment of nucleic acid changes, the corresponding protein in the virus envelope immediately changes.

But where do these new fragments come from? It would seem that they have nowhere to come from.

This question puzzled the researchers. It seemed to lead to a dead end. Until we started studying animal and bird flu. It turned out that viruses reminiscent of the human influenza pathogen circulate among domestic and wild animals. Especially many of them were isolated from birds, including migratory ones. Hybrids of influenza viruses of various types have been isolated, for example, from ducks; an influenza virus similar to the human one has been found in whales.

Please note: avian viruses contain all the types of neuraminidases found in humans and other mammals. For example, neuraminidase from viruses that circulated from 1933 to 1957, as well as neuraminidase from the so-called “Asian” influenza that appeared after 1957.

This is how the assumption arose: the mutation of the influenza virus is associated with the relationships between organisms in nature and the exchange of influenza viruses between humans and animals. This hypothesis is also supported by the fact that variants of currently circulating human influenza viruses have been isolated in humans and birds.

Still, this is nothing more than a guess. Although recombinations of human and animal viruses are obtained in laboratory experiments, no one has observed such phenomena in nature. It is unclear how new virus variants, if they arise in animals, can infect humans. It will take a lot of effort to find out.

This hypothesis looks logical, harmonious and therefore very attractive. She has many supporters. However, other scientists believe that it is impossible to look for the reasons for the variability of influenza in interaction with the animal world. Yes, hybrids of human and animal viruses can be found in nature and in laboratory test tubes. But they are not viable and not so aggressive.

Proponents of the second point of view turn to the human body. Everyone searches where they expect to find it. And, what’s most surprising, he finds it! Special studies have confirmed: in the blood of older people there are antibodies against influenza pathogens that have been circulating for a long time or are not yet circulating!

But studies of whales, ducks, pigs and many other representatives of the animal world seem to convince us that the same influenza virus (meaning its nucleic acid - the pathogenic principle) is found in different kingdoms of life?..

In addition to large, noticeable changes in the protein appearance of the virus (they are associated with the replacement of one of the fragments of the hereditary apparatus), less noticeable, but progressive changes in hemagglutinins are also observed from year to year. Scientists' proposed explanations for this protein drift are being tested experimentally.

What about the truth? She, as usual, is somewhere in the middle. As soon as it is possible to erect a harmonious and harmonious building of a well-founded theory of influenza at the crossroads of modern sciences, all observations will acquire the only true meaning in our minds and will take their rightful place among other factors. Most likely, extreme points of view will also converge. This has happened more than once when passionate seekers of truth argued.

Instructions

Among scientists, interest in influenza is caused, first of all, by the fact that, despite all the progressiveness of modern medicine, an absolutely effective cure against this disease has not been found. As many years ago, people during illness use various “grandmother’s” remedies, such as drinking large amounts of liquid, honey, various herbal infusions, etc. Yes, today there are many drugs that can improve the immunity and general well-being of a person infected with the flu, however, they are not an absolute panacea. Even with vaccinations it is not always possible to avoid infection. Surprisingly, influenza is still "uncharted territory" for medical scientists.

Perhaps the most effective medicine has not yet been found due to the constant mutation of the influenza virus. But is this happening? It is impossible to answer this question with accuracy, but the virus, like any other living organism in nature, tries to survive and adapt to new conditions of existence. Most likely, it is this desire that causes the influenza virus to change, to acquire different forms that are more resistant to various influences.

Today, scientists identify two paths that the influenza virus can take in its mutation processes, they are called “antigenic drift” and “antigenic shift.” Any organism that tries to capture the influenza virus begins to provide all possible resistance to it. In this case, special antibodies are produced, their task is to eliminate the influenza virus and free the body. However, the influenza virus begins to resist such an attack; it is able to change its structure in order to resist antibodies. As a result of such a struggle, new, previously unknown forms of influenza are formed. That is why these mutational processes are “antigenic”. After mutation, the antibodies produced by the body no longer pose any threat to the new form of the virus. Thanks to this, the flu easily overcomes the barriers of the immune system and begins its destructive activity in the body.

The first type of influenza mutation, “drift,” does not occur immediately, the virus changes gradually, and therefore does not pose a particular danger to the body; usually the immune system still copes with the disease. However, the second type of mutation – “shift” – is very serious. The virus is able to significantly change its structure in the shortest possible time, forming new genetic combinations. It is because of the second type of mutation that such frightening varieties of influenza as “bird” and “swine” appeared. With such a sharp shift in the structure of the virus, the immune system has practically no chance in the fight, since antibodies simply do not have time to be produced. In this case, the virus is able to spread very quickly, and an epidemic begins that can take many human lives.