What is the peculiarity of the nature of viruses? General characteristics of viruses. Pathogenesis of viral diseases The nature of viruses and their study
At the annual conference of the American Society of Clinical Oncology, just held in San Francisco, there was a sensational report by scientists who discovered a virus in the tissues of prostate cancer patients that had previously been found only in mice with cancer. And since the virus is known, it is much clearer how to create antitumor drugs. By the way, the viral theory of cancer was substantiated by the Soviet scientist Lev Zilber back in the 1940s.
Moreover, this virus was found only in the tissues of those patients who have a certain genetic defect.
"We're not saying that the virus is the direct cause of the disease," said study co-author Dr. Eric Klein of the Cleveland Clinic. "But this exciting discovery points to entirely new avenues for studying prostate cancer."
Researchers do not yet know how the mouse virus enters the human body, but they suggest that it may be inherited genetically. Dr. Klein and his colleague Dr. Joe Derisi from the University of California used "genetic chip" technology. A similar diagnosticum was created by Russian scientists from the Institute of Molecular Biology of the Russian Academy of Sciences. On a special plate, Derisi placed characteristic segments of the genetic material of 20 thousand known viruses. Klein provided him with 86 tissue samples from his prostate cancer patients. DNA samples were extracted from them and placed on a chip. The DNA of those 20 patients in whom the mutant gene was detected coincided with the DNA of the mouse oncovirus. A mutation is a duplication of a gene that encodes the production of enzymes that destroy viruses that invade the body.
It turned out that men with a double gene produce significantly less of these enzymes. Among 66 patients with a normal gene, the virus was detected in only one. Scientists plan to examine hundreds of sick and healthy people to clarify the connection between the presence of the virus and prostate cancer.
The discovery of American scientists becomes another practical confirmation of the virusogenetic theory of cancer, which was formulated by Lev Zilber back in the 40s of the twentieth century.
Lev Zilber created the theory of the origin of cancer in prison, ahead of the scientific world by half a century
Academician Lev Kiselev, son of Lev Zilber, answers Izvestia’s questions.
It is known that Lev Aleksandrovich created his theory in the 40s in the camp. But the viral hypothesis has been expressed before?
The first hypotheses about the viral nature of cancer were expressed at the beginning of the twentieth century, including by our compatriot Ilya Mechnikov. But Lev Zilber formulated a holistic virogenetic theory that was far ahead of its time.
- But contemporaries did not accept the new theory?
Yes, for 20 years he single-handedly proved that he was right. Only in the 60s did the first experimental confirmation of the theory appear. Great support was provided by the work of the Czech Jan Svoboda from the Institute of Genetics of Czechoslovakia, he is still alive.
- Today there are no doubts about the correctness of Lev Alexandrovich?
Today it is believed that up to 25% of all cancers arise with the participation of oncoviruses. This has been proven, in particular, for liver cancer, which is caused by chronic hepatitis B and C viruses, and for cervical cancer (human papillomavirus). There are suggestions that diseases such as breast cancer, stomach cancer and some others arise not without the participation of viruses. It has also been proven that all tumors in animals are caused by viruses. My father’s words “cancer is a disease of the genome” turned out to be prophetic, because at that time deciphering the genome was incredibly far away.
Great scientist, bright man
Lev Zilber was born in 1894 in Pskov. After graduating from the Faculty of Medicine, he worked in Moscow and Baku, participated in the eradication of the plague in the USSR, and developed the theory of tick-borne encephalitis, suggesting that the disease virus was transmitted by ticks. He was imprisoned twice on absurd charges (in 1937-39 and 1940-44). His brother, the writer Veniamin Kaverin, and his ex-wife Zinaida Ermolyeva, known as the creator of “Soviet penicillin,” fought selflessly for his release. It is known that the brother served as the prototype for Sanya Grigoriev from Kaverin’s story “Two Captains,” beloved by millions of people. The novel "Open Book" is dedicated to Ermolyeva.
Zilber created his theory of the origin of cancer in prison, conducting experiments in a scientific “sharashka”. Rats and mice were caught for him by prisoners, with whom he paid with tobacco. Studying the mechanisms of tumor development, Zilber came to the conclusion that when a virus enters a healthy cell, it changes its genetic basis, so the cell goes beyond the control of the body and begins to divide unhindered - this is how a tumor arises. Lev Aleksandrovich published the first article about his theory in our newspaper in 1945. In the same year, his monograph on this topic appeared.
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Viruses were discovered by the Russian botanist D.I. Ivanovsky (1864 – 1920) in 1892 while studying mosaic disease of tobacco leaves. The term “virus” was first proposed in 1898 by the Dutch scientist M. Beijerinck (1851 – 1931).
Currently, about 3,000 different types of viruses are known.
The sizes of viruses range from 15 to 350 nm (the length of some filamentous ones reaches 3,000 nm; 1 nm = 1·10–9 m), i.e. most of them are not visible in a light microscope (submicroscopic) and their study became possible only after the invention of the electron microscope.
Unlike all other organisms, viruses do not have a cellular structure!
Mature viral particle (i.e. extracellular, resting – virion) structure is very simple: it consists of one or more molecules of nucleic acids that make up core virus and protein shell (capsid)- these are the so-called simple viruses.
Complex viruses(For example, herpes or flu) in addition, capsid proteins and nucleic acid contain additional lipoprotein membrane(envelope, supercapsid formed from the plasma membrane of the host cell), various carbohydrates And enzymes(Fig. 3.1).
Enzymes facilitate the penetration of viral NK into the cell and the release of the resulting virions into the environment ( neuraminidase myxoviruses, ATPase And lysozyme some phages, etc.), and also participate in the processes of transcription and replication of viral NK (various transcriptases And replicase).
Protein shell protects nucleic acid from various physical and chemical influences, and also prevents the penetration of cellular enzymes into it, thereby preventing its breakdown (protective function). Also, the capsid contains a receptor that is complementary to the receptor of the infected cell - viruses infect a strictly defined range of hosts (defining function).
Virions many plant viruses and a number of phages have spiral a capsid in which the protein subunits (capsomeres) are arranged in a spiral around an axis. For example, VTM ( tobacco mosaic virus) has the shape of rods with a diameter of 15–17 nm and a length of up to 300 nm (Fig. 3.2.). Inside its capsid there is a hollow channel with a diameter of 4 nm. The genetic material of TMV is Rice. 3.2.
The structure of the tobacco mosaic virus.
There is single-stranded RNA tightly packed in the groove of the helical capsid. For virions with a helical capsid is characterized by a high protein content (90 – 98%) in relation to
The capsids of virions of many viruses (for example, adenovirus, virus herpes, yellow virus turnip mosaics– VZhMT) have the shape of a symmetrical polyhedron, most often an icosahedron (a polyhedron with 12 vertices, 20 triangular faces and 30 edges). Such capsids are called isometric(Fig. 3.3.). In such virions, the protein content is about 50% relative to NK.
A virus always contains one type of nucleic acid (either DNA or RNA), therefore all viruses are divided into DNA-containing and RNA-containing. The nucleic acid molecules in the virion can be linear (RNA, DNA) or ring-shaped (DNA). Moreover, these nucleic acids can consist of one chain or two. Viral NK has from 3 to 200 genes.
The nucleic acid of the virus combines the functions of both acids (DNA and RNA) - storage and transmission of hereditary information, as well as control of protein synthesis.
Unlike viruses, all cellular organisms contain both types of nucleic acids.
Bacterial viruses have a more complex structure – bacteriophages(Fig. 3.4.). They consist of a head and tail (rod and sheath, basal plate and process filaments). A long NA molecule (RNA or DNA) is folded into a spiral inside the bacteriophage head (protein shell).
Viruses also include viroids– infectious agents that are low molecular weight (short) single-stranded circular RNAs that do not encode their own proteins (lacking a capsid). They are causative agents of a number of diseases.
TO
IN
Rice. 3.4.
The structure of a bacteriophage. Threads of the process
Scientists believe that viruses arose about 3 billion years ago from the nucleic acids of organisms (prokaryotes) as a result of the isolation of free fragments from the genome that have acquired the ability to synthesize a protein shell and divide (double, replicate) inside cells. It is believed that new types of viruses are still being formed from the genome of bacteria and eukaryotes(nuclei, plastids, mitochondria) .
In nature, viruses are of great importance, since they are ubiquitous and affect all groups of living organisms, often causing various diseases.
More than 1000 diseases are known plants caused by viruses (RNA-containing). The most common are various necrosis(areas of dead tissue), mosaics(spots, specks, stripes on plant organs), in which parenchyma tissue is damaged, the number of chloroplasts decreases, phloem is destroyed, etc.; wrinkled or dwarfed leaves are observed. Viruses cause stunted plant growth, which leads to reduced yields.
VZhMT – turnip yellow mosaic virus, VTM – tobacco mosaic virus, VKKT – tomato dwarf bushiness virus.
The appearance of stripes on the flowers of some varieties of tulips (variegated) is also caused by a virus, but flower growers sell these tulips, passing them off as a special variety.
U animals viruses (DNA and RNA containing) cause diseases such as: foot and mouth disease(in cattle) rabies(in dogs, foxes, wolves), myxomatosis(in rats) sarcoma, leukemia And plague(in chickens), etc. Very often people become infected with these diseases (through contact with infected animals).
U person Viruses cause diseases such as: smallpox(variola virus) piggy(paramyxovirus), flu(myxovirus), respiratory diseases(ARI; rhinoviruses RNA-), infectious hepatitis, polio(infantile paralysis; picornavirus), rabies, herpes, AIDS(human immunodeficiency virus - HIV).
Flu - the only infectious disease that manifests itself in the form of periodic global epidemics that are dangerous to human life. The infectious properties of the influenza virus (affects the mucous membranes of the respiratory tract), like other viruses, depend on specific proteins of the viral envelope, which constantly change as a result of recombinations or mutations. Therefore, new strains of the influenza virus cause new epidemics, since humans have not yet developed immunity to them.
Thus, in the winter of 1968/69, 50 million cases were registered in the USA hong kong flu while 70,000 people died. The epidemic of 1918/19 swept the entire globe, occurred in three waves and claimed 20 million human lives.
Viral diseases are difficult to treat because viruses are not sensitive to antibiotics. Fortunately, in many cases the immune system limits the infection from spreading further.
Numerous viral diseases of humans and animals can be prevented by immunization– carrying out preventive vaccinations that allow you to develop immunity against viruses.
Viruses are widely used by humans in microbiological research (biotechnology, genetic engineering). It is possible to use viruses to control crop pests.
In the USA with cotton boll effectively fight with the help of the virus. This method of control is practically harmless - the virus, as a rule, is species-specific (that is, it affects only a certain type of organism).
It was also found that, for example, rice necrotic mosaic virus inhibits the growth of rice. But other plants, for example, jute(a source of coarse fibers for bags and ropes), grow better when affected by this virus than when healthy. Scientists cannot yet explain this phenomenon.
Bacteriophages infect bacteria (penetrate and actively destroy them), including pathogenic ones. Therefore, it is possible to use them to prevent and treat many infectious diseases and to combat pathogenic bacteria: plague, typhoid fever, cholera and etc.
Lecture 1
Nature, origin and structure of viruses.
Module 1
Integrated purpose of the module
The comprehensive goal of the module is to give students an understanding of the basic properties of viruses: their nature, origin, architecture and morphology of viral particles, types of symmetry, and chemical properties of viruses. This information should become a theoretical basis for further understanding of the biological essence of viruses, which is expressed in the processes of reproduction and viral pathogenesis. The module consists of three lectures, the material of which allows you to solve the set goal.
NATURE OF VIRUSES
From the time of the discovery of viruses to the present, ideas about the nature of viruses have undergone significant changes.
D.I. Ivanovsky and other researchers of that time emphasized two properties of viruses that made it possible to isolate them from the general mass of microorganisms: filterability and inability to reproduce in all artificial nutrient media. Later it turned out that these properties are not absolute, since filterable forms of bacteria and mycoplasma were discovered growing on artificial nutrient media, approaching in size the largest viruses (human and animal smallpox viruses).
However, even in the case when integration does not occur and the viral genome is in an autonomous state, the occurrence of infection is due to competition between the viral and cellular genomes.
The unique properties of the virus include its method of reproduction, which differs sharply from the methods of reproduction of all other cells and organisms (binary fission, budding, spore formation). Viruses do not grow, and their reproduction is referred to as disjunctive (separated) reproduction, which emphasizes the disunity in space (on the territory of the cell) and time of synthesis of viral components (nucleic acids and proteins) with subsequent assembly and formation of virions.
In connection with the above, discussions have arisen more than once about what viruses are - living or non-living, organisms or non-organisms. Of course, viruses have the basic properties of all other forms of life - the ability to reproduce, heredity, variability, adaptability to environmental conditions; they occupy a certain ecological niche, they are subject to the laws of evolution of the organic world on earth. Therefore, by the mid-40s, the idea of viruses as the simplest microorganisms had developed. The logical development of these views was the introduction of the term “virion”, which denoted an extracellular viral individual. However, with the development of research on the molecular biology of viruses, facts began to accumulate that contradict the idea of viruses as organisms.
The absence of their own protein-synthesizing systems, a disjunctive mode of reproduction, integration with the cellular genome, the existence of satellite viruses and defective viruses, the phenomena of multiple reactivation and complementation - all this does not fit well into the idea of viruses as organisms. This idea loses even more meaning when we turn to virus-like structures - plasmids, viroids and agents such as the scrapie pathogen.
Plasmids (other names - episomes, epiviruses) are double-stranded circular DNA with a molecular weight of several million, replicated by the cell. They were first discovered in prokaryotes, and their existence is associated with various properties of bacteria, such as resistance to antibiotics. Because plasmids are not usually associated with the bacterial chromosome (although many are capable of integration), they are considered extrachromosomal factors of heredity.
Plasmids have also been discovered in eukaryotes (yeast and other fungi); moreover, ordinary viruses of higher animals can also exist in the form of plasmids, i.e. circular DNA, devoid of their own proteins and replicated by cellular DNA synthesis enzymes. In particular, bovine papillomaviruses and simian virus 40 (SV-40) can exist in the form of plasmids. When the herpes virus persists in cell culture, plasmids can be formed - circular DNA, which constitute only part of the genome of this virus.
Related to viruses are viroids, agents discovered by T. O. Diner in 1972, which cause diseases of some plants and can be transmitted like ordinary infectious viruses. When studying them, it turned out that these are relatively small molecules of circular supercoiled RNA, consisting of a few, 300-400 nucleotides. The mechanism of viroid replication is not entirely clear.
Finally, mention should be made of the agent scrapie, the causative agent of subacute transmissible spongiform encephalopathy in sheep. Probably, similar agents cause other forms of spongiform encephalopathy in animals and humans, which are based on the progressive destruction of nerve cells, as a result of which the brain acquires a spongiform (spongioform) structure. The scrapie agent has a protein nature and even received a special name - prion (from the words proteinaceous infectious particle - protein infectious particle). It is assumed that this protein is both an inducer and a product of some cellular gene that has become autonomous and escaped regulation (“gone crazy”).
All viruses, including satellite and defective viruses, plasmids, viroids and even scrapie agents (their genes), have something in common that unites them. All of them are autonomous genetic structures capable of functioning and reproducing in cells of animals, plants, protozoa, fungi, and bacteria susceptible to them. Apparently, this is the most general definition that allows us to outline the kingdom of viruses. Based on the formulated definition, viruses, while not being organisms, are nevertheless a unique form of life and therefore obey the laws of evolution of the organic world on earth.
ORIGIN OF VIRUSES
Various assumptions have been made regarding the origin of viruses. Some authors believed that viruses are the result of an extreme manifestation of the regressive evolution of bacteria or other single-celled organisms. The hypothesis of regressive evolution cannot explain the diversity of genetic material in viruses, their non-cellular organization, disjunctive mode of reproduction and the absence of protein synthesizing systems. Therefore, at present, this hypothesis has rather historical significance and is not shared by the majority of virologists.
According to the second hypothesis, viruses are the descendants of ancient, precellular life forms - protobionts, which preceded the appearance of cellular life forms, from which biological evolution began. This hypothesis is also not currently shared by the majority of virologists, since it does not explain the same issues that the first hypothesis was powerless to resolve.
A third hypothesis suggests that viruses evolved from genetic elements of cells that became autonomous, although it is not clear which of these elements gave rise to such a wide variety of genetic material in viruses. This hypothesis, which was ironically called the “running genes” hypothesis, finds the largest number of supporters, but not in the original form in which it was expressed, since it does not explain the presence of forms of genetic material in viruses (single-stranded DNA, double-stranded RNA), absent in cells, capsid formation, the existence of two forms of symmetry, etc.
It is likely that viruses are indeed derivatives of the genetic elements of cells, but they arose and evolved along with the emergence and evolution of cellular life forms. Nature, as it were, tried all possible forms of genetic material (different types of RNA and DNA) on viruses before finally choosing its canonical form - double-stranded DNA, common to all cellular forms of organisms, from bacteria to humans. Being, on the one hand, autonomous genetic structures, on the other hand, incapable of developing outside cells, viruses over the course of billions of years of biological evolution have followed such diverse development paths that their individual groups do not have a successive connection with each other. Apparently, different groups of viruses arose at historically different times from different genetic elements of cells and therefore the currently existing different groups of viruses have a polyphyletic origin, that is, they do not have a single common ancestor. However, the universality of the genetic code also applies to viruses, thereby indicating that they too are a product of the organic world of the earth.
ROLE OF VIRUSES IN EVOLUTION
Since viruses, being special forms of life, are not microorganisms, virology is not a branch of microbiology, but an independent scientific discipline that has its own object of study and its own research methods.
Nature and origin of viruses
Antigenic variability of the influenza virus and aspects of its study.
The solution to obtaining effective attenuated variants of the influenza virus is hampered by the unique plasticity and variability of its antigenic properties. Almost annual influenza epidemics at various intervals become global. In recent years, the infection causing pandemics is the influenza A virus. Analysis of antigenic shifts within each of its three types shows that the change in the antigenic composition of virus strains of type AO to type A occurred gradually, and the transition from type A1 to A2 was sharp.
After it was recorded in 1957 that a new serological type A2 had appeared in nature, it seemed stable for some time, although there were slight changes. But already in 1964 they became significant, and after the epidemic in Hong Kong, the viruses differed so dramatically that it was assumed that a new antigenic type A had emerged. During the evolution of the virus, not only the antigenic properties of surface proteins changed, but also other characteristics. The influenza virus strain isolated during the 1971-1972 epidemic, in contrast to previously circulating strains, significantly increased its reproductive and neuraminidase activity, sharply increased the thermostability of viruses and acquired the ability to regularly cause viremia in mice (Zakstelskaya et al., 1969; Sokolov, Podchernyaeva, 1975).
If previously type B viruses were relatively stable, then since 1967 there has been a continuous change (Seihachiro, Mitsuo, 1974). In April–May 1974, new strains of the influenza virus were isolated, of which B/Hong Kong 15/72 are considered as a new variant, and others as intermediate between the old and new strains, which allows us to revise the data on the antigenic stability of the influenza B virus and suggest emergence of a new serotype.
Thus, a picture emerges of significant antigenic changes within types A and B. This, naturally, attracts the close attention of scientists working on the problem of influenza. Since even the presence of intense immunity in the population cannot cause such large antigenic changes in the influenza virus, a hypothesis was put forward about the periodicity of recombinations that occur between human and animal influenza viruses, as well as between human and avian viruses. When developing this hypothesis, it turned out that influenza naturally affects pigs, horses, turkeys, chickens, ducks, terns, and this list continues to grow. They have antibodies to human influenza viruses in their blood serum. This is not surprising, since the fragmentation of the influenza virus genome determines the possibility of not only intraspecific, but interspecific recombination.
The preparative separation of neuraminidase and hemagglutinin opens up prospects for an in-depth analysis of the relationship between individual characteristics. Some researchers (Webster a. o., 1973; Gorev et al., 1974) note that the recombinant virus simultaneously with hemagglutinin acquires virulence. There is a set of recombinants with specified hemagglutinin and neuraminidase.
Currently, many virology laboratories around the world are studying influenza epizootics and analyzing antigenic relationships with human influenza. Work in this direction is coordinated and stimulated by WHO. The complexity of this problem dictates the need for an ambiguous approach to its solution. Parallel searches for other approaches to this issue should not be considered as alternatives.
In the 40-50s, the emergence of antigenic variants during passage of the virus in the body of immunized animals was experimentally proven (Archetti, Horsfoll, 1960). These changes were quite persistent, the viruses maintaining their new antigenic specificity in serial passages in ovo and in the absence of immunoserum. Moreover, long-term passages of the influenza virus through the bodies of non-immunized healthy animals change its biological properties. For example, K. Paucker (1960), during passages of the PR8 strain, received a virus for a long time that was antigenically different from the original and not similar to other types of influenza virus. The author believes that between passages 103 and 107 a mutant was formed, which subsequently replaced the original virus. Similar data are provided by K. Zgozelska et al. (1973).
Here we see a manifestation of the basic law of development of any population, including a viral one - the gene pool of a population changes over time: on the one hand, it becomes depleted as a result of the extinction of organisms containing individual genes, and on the other hand, it becomes enriched due to mutations giving rise to new genes .
The work of S. Fazekas de Sent Groth, C. Hannoun (1973) on the selection of spontaneous antigenic mutants of the influenza A virus under “immunopress” (i.e., in the presence of immunoserum) made it possible to reproduce the hierarchical order of viruses within each type. Moreover, in all his conclusions he was based on the indicators of the cross-sectional RZGA. In experiments on the selection of late mutants obtained with the help of antibodies, he was able to reproduce the natural process of selection of epidemic strains. He also proposed a simple model of the interaction of an antibody with an antigen. The author presented the antigenic zone of the protein shell of the virus in the form of a small number of amino acid protein chains protruding beyond the surface of the virus. Schematically, it looks like a fork with teeth of different lengths and widths, and the corresponding antibodies are cavities complementary to some or all of the teeth. Thus, contact of the antiserum with a related antigen leads to the elimination of homologous antigens, and antigens with non-complementary regions, i.e., mutants, remain in the population.
This diagram represents the logical development of the basic principles of immunology that emerged in the 40s, about the interaction of antigen and antibody and the theory of antibody biosynthesis. According to these works, the active group of antibodies has a configuration additional to the configuration of the determining group of the antigen. It was assumed that these groups relate to each other as an object relates to its mirror image. K. Landsteiner (1946) conducted experiments with an artificial antigen obtained by complexing protein molecules with various low-molecular compounds, which showed that the specificity of this antigen can be determined only by a small group attached to the protein. Antibodies do not “recognize” an antigen if it differs only in the position of the methyl group in the aromatic ring from the one that stimulated the formation of these antibodies, or in the spatial position of the hydroxyl (Boyd 1969).
Thus, returning to the issue of antigenic variability of the virus, we can state the selection role of antibodies in this process. How mutant particles arise in a viral population is one of the questions that must be answered to understand the evolution of influenza viruses.
Any viral population contains spontaneous mutants that arise as a result of the action of external or internal factors. Depending on the acquired properties, a mutant may have an advantage in reproduction and predominate in the population. In some cases, it is possible to identify the factor that played a decisive role in the emergence of the mutant. The 1918 pandemic is of greatest interest to researchers dealing with the problem of influenza, since its virus was extremely pathogenic for humans. Retrospective analysis of this virus has led some researchers to believe that the pandemic was caused by a swine influenza virus isolated in 1930 because the strains have antigens related to those of swine viruses. According to another point of view, the increase in virus activity is caused by the appearance of mutant particles under the influence of mustard gas, which was used during the First World War, i.e., before the pandemic wave of influenza (Blashkovich 1966). Indeed, mustard gas is an extremely strong biologically active chemical agent. Its mutagenic activity was first shown by C. Auerbach and T. M. Robson (1946). At the same time, it was found that mustard gas has a direct mutagenic effect on chromosomes. It was later discovered that mustard gas can cause mutations in viruses and bacteria. Consequently, its possible role as a mutagenic agent cannot be excluded, if we take into account that chemical and physical factors can cause genetic changes in biological objects at all stages of development, and viruses, apparently, are no exception.
Factors that can act as mutagens under natural conditions include pharmacological drugs. There are works that analyze the relationship between teratogenic activity and the chemical structure of drug molecules; In microorganisms, a similar phenomenon of the widespread emergence of drug-resistant mutant forms is observed. At the height of the flu, when the virus reproduces in the body, patients take medications that are synthetic chemical compounds.
It is known that antiviral agents are quite effective only if they are able to selectively suppress the synthesis of nucleic acids, that is, they come into direct contact with the genetic apparatus. Apparently, due to the peculiarities of the influenza virus genome, the line between the purely antiviral and mutagenic effects of chemical compounds is easily crossed.
Our experiments studying the effect of chemical compounds on the antigenic specificity of influenza viruses belonging to the AO serotype showed that some compounds from the class of supermutagens can cause changes that do not extend beyond the homologous serotype. In particular, the first two representatives of nitrosoalkyl ureas induced mutations for this trait (Chulanova, 1968; Akhmatullina et al. 1974). We used our proposed modification of the RZGA, which made it possible to establish the coefficient Ap and, based on it, to determine the degree of difference in the antigenic specificity of wild and mutant viruses.
Experiments with a large set of chemical compounds revealed another agent among them - 1,4-bis-diazoacetylbutane, which is active in mutation based on antigen specificity. We also used the immunopress method; after exposure to the mutagen, the virus was passaged in the presence of homologous serum. Unmodified viral particles were neutralized with complementary antibodies, and selective conditions were created for induced mutants. The resulting antigenic mutants were studied in a cross-reaction test with serum to wild and mutant viruses and in a precipitation reaction and indicated significant antigenic shifts.
Thus, further experimental study of induced mutants using a large set of chemical compounds will provide information on the problem under study.
Flu. Treatment and prevention.
Influenza is an acute infectious disease of the upper respiratory tract. Dangerous in itself, the flu aggravates the course of other chronic diseases and causes serious complications in the cardiovascular and central nervous systems, digestive organs, kidneys, etc. The flu is most dangerous for children and the elderly. The speed of spread of influenza, the severity of the disease, the frequency of complications, and sometimes death - all this makes its prevention especially important. People involved in sports and gymnastics are much less likely to be exposed to the influenza virus. There are several types of influenza virus known - A, B, C, etc.; under the influence of environmental factors, their number may increase. Due to the fact that immunity to influenza is short-term and specific, repeated illness is possible in one season. According to statistics, an average of 20-35% of the population suffers from influenza every year.
The source of infection is a sick person; Patients with a mild form of the virus are the most dangerous as spreaders of the virus, since they do not isolate themselves in a timely manner - they go to work, use public transport, and visit places of entertainment.
The infection is transmitted from sick to healthy through airborne droplets when talking, sneezing, coughing or through household items.
The latent period of influenza lasts from 1 – 12 hours to 3 days. The disease begins acutely: a sharp increase in temperature to 38-400, chills, headache, pain in the bones and muscles, general weakness; pain and sore throat, disturbances of taste and smell occur; After 12-24 hours, nasal discharge appears.
The temperature lasts 1-3 days, sometimes up to 6-7 days. As a rule, by the end of the first week the temperature returns to normal. With proper treatment and care, recovery occurs in 7-9 days.
If influenza is suspected, the patient should be isolated and put to bed. This must be done before the doctor arrives. Considering that influenza pathogens are very unstable in the external environment and are easily destroyed under the influence of oxygen and disinfectants, the room must be regularly ventilated. At least once a day, wet clean the room using bleach, formaldehyde, soda, chloramine, and laundry soap.
The patient must have individual dishes. The patient's tableware and teaware should be washed with boiling water and baking soda or treated with a 5% chloramine solution. Systematic disinfection of the patient's underwear and bed linen by boiling in a soap solution is mandatory.
All medications prescribed by a doctor should be stored in a specially designated place. In addition to medications, it is advisable to drink plenty of fluids during all periods of the disease: tea with honey or lemon, cranberry juice, warm milk, fruit and vegetable juices. Food must be high in calories. The doctor's instructions must be strictly followed. Self-medication is unacceptable. Medicines cannot be taken without a doctor's prescription. Particular caution should be given to antibiotics and sulfonamides - they do not act on the influenza virus, and if taken without permission and inaccurate dosages, they can cause allergic reactions. But what you can use painlessly are foot baths, mustard plasters, onions, garlic. Released through the lungs, the essential oils contained in onion and garlic increase the secretion of mucus and thereby facilitate easier expectoration in case of respiratory diseases.
Healthcare also has a number of specific anti-influenza agents, which include primarily a live vaccine and a special serum containing protective proteins. The drugs introduced into practice are interferon and oxolinic ointment.
Hardening, a balanced diet, fresh air, and timely treatment of chronic diseases will help you prevent colds, in particular the flu.
It is believed that viruses (from Lat. virus- poison) - something nasty that brings nothing but trouble. But this is a grave mistake. Viruses are the key creators of living nature and the engines of its evolution.
One of the main arguments against the hypothesis that viruses “escaped” from cells is the fact that viral genetic systems are much more diverse than cellular ones. As is known, cellular organisms have only double-stranded - linear or circular - DNA genomes. And the genome of the virus can be represented by both single- and double-stranded RNA or DNA molecules, linear or circular. There are also systems that use reverse transcription. Thus, in retroviruses (for example, some oncoviruses, HIV) and pararetroviruses (hepatitis B viruses, cauliflower mosaic, etc.), one of the genomic DNA chains is synthesized on an RNA template. Viruses, unlike cellular organisms, implement all theoretically possible ways of storing and expressing genetic information.
The second important argument against considering viruses as originating from cells is that there are many viral genes that do not exist in cellular organisms. Cellular organisms not only evolved from viruses, but also inherited (and continue to inherit) a significant part of their genetic material from them. Of particular interest in this regard are endogenous viruses (parts of the genome of RNA or DNA viruses integrated into the genome of a cell), among which genes derived from retroviruses predominate. It is believed that mammals have inherited more than half of their genome from viruses and their closest relatives - “selfish” genetic elements, for example, plasmids and transposons. Thus, viruses are co-parents of humans. Often the gene sequences of endogenous viruses, which are found in large numbers in the human genome, are changed and no longer code for proteins. There is good reason to believe that such sequences are involved in the regulation of cellular genes, although their specific biological functions are often unknown. However, we do know something important: for example, the protein syncytin, which is encoded by the envelope gene of one endogenous retrovirus, is necessary for cell fusion during the formation of the placenta. This means that neither humans nor placental animals could be born without this endogenous virus. There is another important example. It turned out that a component of the genome of one of the endogenous viruses controls the expression of proline dehydrogenase in some regions of the central nervous system. This enzyme may have played an important role in the evolution of the human brain. If the expression of this enzyme is disrupted as a result of mutations, mental illnesses occur, including schizophrenia. Viruses and their relatives also play an important role in the horizontal transfer of cellular genes - from one organism to another.
However, despite their key role in evolution, viruses are best known as pathogens of humans, animals and plants (by the way, this is why they were first discovered). And then we will talk about the nature of viral pathogenicity. Viruses (especially eukaryotic viruses) have no special “desire” to harm the host, much less kill him. And in many cases, viruses coexist quite peacefully and friendly with cells. Why are so many viruses so harmful? The usual explanation is that the pathology of an infected cell is caused by the “plunder” of its resources (material and structural), which the virus directs to its own needs of reproduction. However, the greatest harm can occur from injudicious protective actions of hosts and the protective activity of viruses, which is not directly related to their reproduction.
Mechanisms of protection and counter-defense
What are the main defense mechanisms of an infected cell? These are components of innate immunity: degradation of RNA (viral as well as cellular), inhibition of protein synthesis (both viral and cellular), self-destruction (apoptosis and other types of programmed death) and, finally, inflammation. Actually, many viruses discovered their existence in this way - due to the inflammation they cause (encephalitis, pneumonia, etc.). The cell fights the virus by disrupting its own metabolism and/or structure, and its defense mechanisms are usually self-damaging. You could say that a person who dies from polio (and less than 1% does) killed himself while fighting the infection.
In response to cellular defenses, viruses evolve to develop defenses, and there is an arms race between the virus and the cell. These drugs are directed primarily against the general metabolic processes underlying the cell's defense reactions. This again is inhibition of the synthesis of cellular RNA and proteins, disruption of intracellular infrastructure and cell transport, suppression or, conversely, the launch of apoptosis and other mechanisms that cause programmed cell death. Thus, the defense strategy of a virus is in many ways similar to the defense behavior of a cell. Figuratively speaking, wrestlers use the same techniques and hit the same goals. For example, a cell, suppressing the synthesis of viral proteins, uses interferon, and to inhibit its formation, the virus, in turn, inhibits protein synthesis in the cell. Depending on the circumstances, one side or the other benefits. It turns out that the main contribution to pathology is not the reproduction of the virus as such, but the confrontation between cellular defense and viral defense. In plant pathology, the concept of “tolerance” has long existed: a pathogenic virus can actively multiply in an infected plant without causing painful symptoms.
Below we will mainly talk about RNA viruses (this is a simpler example). How does an RNA virus, once it enters a cell, reveal its presence? And how does a cell know that a virus has entered it? The main feature due to which the cell “understands” this is viral double-stranded RNA, which, in principle, can be formed in an uninfected cell, but not in such quantities and places. In some cases, the cell also recognizes viral single-stranded RNA, and sometimes (much less often) viral proteins. It is important that recognition of viral RNA is nonspecific: having “sensed” double-stranded RNA, the cell may “think” that a virus has entered it, but it does not know which one. RNA is captured by two types of sensors: toll-like (from the English. toll-like and from him. toll- remarkable) receptors and specialized RNA helicases. They involve a number of protective mechanisms at the transcriptional level, including the formation of interferon. In addition, viral RNAs are recognized by the “executors” - the double-stranded RNA-dependent protein kinase PKR, which phosphorylates some translation initiation factors, thereby inhibiting protein synthesis; oligoadenylate synthetase (OAS), which activates RNase L, which cleaves RNA; RNA interference system, leading to RNA degradation and disruption of its translation.
Because the virus is recognized as something nonspecific, the cell cannot know its “intentions.” And in general, it would be impossible to come up with an individual innate defense system for any possible virus. This means that the cell can fight the virus only with standard techniques. And therefore, its defensive actions are often disproportionate to the existing threat. However, if the cell's defense reactions are so nonspecific, why do different viruses still cause different diseases? Firstly, each virus can only infect a certain type of cell in a specific organism. This is due to the fact that in order to penetrate the cell it must interact with cellular receptors that are “suitable” for it. In addition, for the reproduction of viruses, a certain intracellular environment is required (specific cellular proteins are often needed). Secondly, while the cell's defense reactions are standard, the virus's defenses are largely individual, although they are directed against standard cellular mechanisms.
In plants, RNA interference plays a very important role as an antiviral mechanism. Double-stranded RNA is formed from viral RNA (an important factor by which the cell learns about the presence of a virus). With the participation of components of the RNA interference system - the Dicer enzyme, which cuts this double-stranded RNA into fragments of 21-25 nucleotide pairs in length, and then the RNA-protein complex RISC - eventually single-stranded short RNA fragments are formed. Hybridizing with viral RNA, they cause either its degradation or inhibition of its translation. This protective mechanism is effective, but can damage the cell itself, as is clearly seen in the example of viroids. These are plant pathogens, short (several hundred nucleotides) circular single-stranded RNA molecules not covered with a protein shell. Viroids do not code for proteins but can cause severe symptoms in an infected plant. This happens because the cell is protecting itself. The resulting viroid double-stranded RNA is exposed to all components of the RNA interference system, resulting in the formation of single-stranded RNA fragments that hybridize not with viral RNA, but with cellular RNA. This leads to its degradation and the development of disease symptoms. However, many plant viruses encode a variety of proteins that interfere with RNA interference ( viral suppressors of RNA silencing - VSR). They either inhibit the recognition and cleavage of viral RNAs, or suppress the formation and functioning of the RISC complex. Therefore, these VSR proteins can disrupt physiologically important (non-viral) RNA interference mechanisms, causing pathological symptoms.
Security proteins
The defense of viruses, in particular picornaviruses, small RNA-containing pathogens, largely depends on the work of such proteins. This large group includes, in particular, the pathogens of poliomyelitis, hepatitis A, foot-and-mouth disease, etc. The peculiarity of these viruses is that, with rare exceptions, their proteins are synthesized in the form of a single polyprotein, from which individual mature proteins are then formed. Among them, three groups can be distinguished. The first consists of key proteins - vital, with fixed functions, directly ensuring the reproduction of the virus: RNA-dependent RNA polymerases, necessary for the replication of the viral genome; capsid proteins that form the protein shell of the virus; proteases involved in the process of converting polyproteins into mature proteins; VPg protein ( viral protein genome linked- a viral protein connected to the genome) that serves as a primer for the synthesis of RNA molecules; Helicase is a very valuable enzyme that all picornaviruses have, but plays a role that is not very clear. The second group also includes vital proteins, but those that perform “auxiliary” work - hydrophobic “guide” proteins 2B and 3A. They direct key proteins to their destinations and help create an optimal intracellular environment for virus reproduction. The third group includes the leader protein L, discovered in our laboratory 30 years ago, and protein 2A; we called them “security” proteins ( security- security). This is a specialized anti-defensive “weapon” of picornaviruses. In general, all three of these classes of proteins can fight the cell's defense mechanisms. But key proteins and guide proteins do this job part-time because they have other important responsibilities that their structure and function must accommodate. Consequently, their defensive capabilities are limited by the need to perform basic work. But “security” squirrels work full-time in their specialty - evolution “hired” them specifically for “security” (later some of them “learned” to do something else). They can have any necessary structure to carry out their duties.
One of the most important functions of security proteins is that they take part in determining the fate of an infected cell. There are many different options for its death, but the two main, most well-known mechanisms are necrosis and apoptosis, which differ in morphological and biochemical characteristics. During necrosis, the cell is lysed, and its contents are poured out into the intercellular space. During apoptosis, clearly visible protrusions are formed on its surface, its DNA is degraded to nucleosomal fragments, and ultimately the cell is fragmented into individual apoptotic bodies limited to the plasma membrane. It is very important how exactly the cell dies. With necrosis, protective inflammation develops, but the virus leaves the cell and spreads. With apoptosis, the spread of the virus is limited and there is usually no inflammatory reaction. The death of an infected cell, as a rule, is an act of self-sacrifice that limits the reproduction of the virus.
We found that infection with picornaviruses, in particular poliovirus (poliovirus), triggers a cell apoptotic program. This occurs along one of the classical pathways, when cytochrome is released from mitochondria c and a cascade of proteolytic enzymes, caspases, is activated. But, on the other hand, it turned out that viruses have an anti-apoptotic mechanism - the ability to suppress the apoptotic reaction of the cell. Thus, HeLa cells infected with poliovirus or encephalomyocarditis virus (also picornavirus) die with signs of necrosis. But if you turn off the anti-apoptotic “weapon” (suppress the synthesis of viral proteins), the cell dies from apoptosis (self-sacrifice). In both viruses, “security” proteins serve as such weapons. However, in the encephalomyocarditis virus this role is played by the L protein, and in the poliovirus it is the 2A protein. The leader protein has no enzymatic activity, while the 2A protein is a protease. They have nothing in common either structurally or biochemically, but they both have antiapoptotic effects based on different molecular mechanisms.
Another anti-protective mechanism of picornavirus security proteins is disruption of nuclear-cytoplasmic transport [10–12]. We have shown that when infected with these viruses, the permeability of the nuclear membrane increases and the active exchange of macromolecules between the cytoplasm and the nucleus is disrupted. And if the cell structure is damaged, then it cannot turn on its regulatory mechanisms to fight the virus. In poliovirus, security protein 2A disrupts nuclear-cytoplasmic transport by hydrolyzing nucleoporins, components of nuclear pores. And the leader protein of the encephalomyocarditis virus works - it affects the cellular cascade of phosphorylation of nucleoporins [,].
The anti-protective function of “security” proteins can manifest itself in other ways. Thus, L-proteins of cardioviruses (including encephalomyocarditis virus) and 2A-proteins of enteroviruses (including poliovirus) inhibit the formation of interferon. And its action is inhibited by the L-protein of the foot-and-mouth disease virus and the 2A protein of the poliovirus. However, the “security” proteins of picornaviruses are not vital. Both guardians can be removed or large deletions can be caused in them (as in the case of the L protein of cardioviruses, 2A proteins of the hepatitis A virus and cardioviruses) without the virus being rendered viable.
Mutual disarmament
What will be the consequences of inactivation of viral “security” proteins for the cell? On the one hand, the sensitivity of viruses to the protective mechanisms of innate cellular immunity will increase. But, on the other hand, his self-harming, suicidal activity will also increase. What happens if you simultaneously turn off the defense mechanisms of the cell and the virus? We studied this situation using the example of interaction between mengovirus (a strain of encephalomyocarditis virus) and HeLa cells. Infected with a wild-type virus, they quickly die from necrosis. And if the virus is partially disarmed (the leader protein is inactivated), HeLa cells live a little longer and die not from necrosis, but from apoptosis. When mutual defense is reduced (apoptosis is turned off in the cell by a chemical compound that inhibits caspases, and the virus’s leader protein is inactivated), even after a twice as long period of time, the cells feel significantly better than those that were not disarmed. And the reproduction of the virus (both dynamics and harvest) proceeded in exactly the same way, regardless of whether only its defenses were turned off or whether the cellular defenses were also removed at the same time. It turns out that in cells that do not yet have serious pathological damage (the so-called cytopathic effect), a lot of viral particles can already form. Thus, cell damage is not necessary for the virus to reproduce. Therefore, an effective strategy for antiviral therapy aimed at alleviating the symptoms of the disease may be the simultaneous suppression of both viral and cellular defenses.
Programmed death
This series of our experiments also provided an opportunity to penetrate deeper into the nature of necrotic death caused by the virus. What is it - the killing of a cell by a virus or its suicide (self-sacrifice), when it decides that for the sake of the common good it is more expedient to die? The following are subject to necrotic lesions:
- plasma membrane (its permeability increases, “blisters” form),
- cytoplasm (microtubules and microfilaments change),
- nucleus (shrinks, deforms, chromatin condenses),
- metabolic activity (NADH-dependent reduction reactions, viability change).
When apoptosis is turned off (the addition of a chemical caspase inhibitor), various necrotic changes depend on whether the viral leader protein is functioning or not. For example, if it is inactivated, the cell’s membrane permeability does not change, “blisters” associated with an imbalance in osmotic balance do not appear, and a number of other necrotic lesions do not occur. One possible explanation for this effect is that L protein acts on multiple targets in different cellular compartments. But because the protein is small and has no enzymatic activity, it is more likely that its direct targets are much smaller. We hypothesize that the leader protein acts on one or more key cellular elements that control the fate of the cell, and as a result, its necrotic program is launched, which is responsible for most of the listed pathological changes. Consequently, it is not the virus that kills the cell necrotically, but the cell itself commits suicide (carries out an act of self-sacrifice). This point of view is consistent with new ideas, according to which, in addition to apoptosis, there are a number of other physiologically important types of programmed (encoded in the cellular genome) cell death, including necroptosis, which is similar to necrosis.
Thus, cell self-sacrifice during viral infection can manifest itself as necroptosis or apoptosis. Necroptosis may be a protective reaction of a cell to a viral infection, and not only in the case of picornaviruses. Which mechanism is more beneficial for the virus depends on the conditions. We see that its anti-protective effect can manifest itself in the form of “rerouting” of mechanisms encoded in the cell genome. This is an important (although not the only) method of defense and one of the main mechanisms of pathogenicity of viruses. Virus-induced apoptosis and necrosis programs compete with each other. We have shown that when HeLa cells are infected with poliovirus, apoptosis is first activated, and then it is suppressed and the necrotic pathway is launched. Thus, infection of a cell with a virus activates a number of protective actions in it, among which there are two suicidal mechanisms of programmed death - apoptotic and necrotic. And then competition occurs between these pathways: suppression of one of them activates the other, and vice versa. And all this is regulated by cellular proteins, viral proteins (primarily “security” proteins), as well as external factors.
Arms race
Since cells have defense mechanisms, and viruses have defense mechanisms, naturally, there is an arms race between them. The non-conservative nature of security proteins suggests that they are adapted to counteract the defense mechanisms of a particular host. And therefore, its change may be accompanied by a loss of the “security” protein function and, as a consequence, an increase in the host’s defensive reactions. This can explain the special pathogenicity of the “new” ( newly emerging- emerging) viruses. Thus, the influenza virus is a low-pathogenic, almost harmless intestinal virus of wild birds. When it infects a person, Spanish flu, bird flu or swine flu can occur. The SARS virus is relatively safe for bats, but in humans it causes severe acute respiratory syndrome, accompanied by high mortality. Finally, HIV (more precisely, its ancestor) is practically harmless to monkeys, but in humans it causes AIDS. It is very important that these viruses do not develop new pathogenicity factors when moving to a new host (simply, as a result of several mutations that ensure penetration into the cell, they acquire the ability to infect humans). Another possible mechanism for the imbalance between the virus and the host and the emergence of new pathogens may be a change in viral defense weapons, for example, the loss of an old one or the acquisition of a new “security” protein.
However, long-term co-evolution of the host and the virus should lead to a decrease in the pathogenicity of the latter (mutually beneficial disarmament). The classic example is the myxoma/fibroma virus. In the middle of the 19th century. European rabbits were brought to Australia, they quickly multiplied and became a serious threat to agriculture. 100 years later, the pathogenic fibroma/myxoma virus (from the poxvirus family, which includes the smallpox virus) began to be used to control their population. Different rabbits react differently to this virus. In Brazilian rabbits, three weeks after infection, it causes a benign tumor - a fibroma (localized nodule on the skin). But European rabbits sensitive to this virus develop a generalized fatal disease within 10 days after infection.
Introduced to Australia, the virus caused mosquito-borne summer epidemics in which more than 99% of infected rabbits died in less than two weeks. Less virulent variants of the virus were more likely to survive the winter, and this led to the selection of weakened (attenuated) strains. And after about 10 years, the mortality rate of European rabbits from the evolved virus was halved. At the same time, resistant rabbits were selected: their mortality from the original virus decreased by approximately four times. In just a decade (a negligible period in evolutionary terms), the relationship between pathogen and host has improved approximately 10-fold. This is, of course, a somewhat simplified scheme, since the arms race does not stop: in response to increased resistance in rabbits, the virulence of the virus may also increase. However, this is a striking example of the role of the interaction of viruses and cellular organisms in the evolution of both. Viruses and cells “teach” each other, and the acquired “knowledge” is inherited. In 2013, two graduates of the Department of Virology of Moscow State University, Evgeny Kunin and Valeryan Dolya, published an article about the “virocentric” view of evolution, according to which the resistance and cooperation of viruses and cellular organisms is the main factor in their evolution.
My story does not exhaust the topic: much more is known about the nature of the pathogenicity of viruses. Much of what we now know has been learned in recent years, and there is every reason to expect new surprises. We can and should blame viruses for serious illnesses and it is necessary to fight them, but we should be grateful to viruses for the existence and diversity of living nature, including the existence of humans.
The author is grateful to his colleagues in scientific cooperation - the staff of the Institute of Poliomyelitis and Viral Encephalitis named after. M. P. Chumakov RAMS, Moscow State University. M.V. Lomonosov, Institute of Protein RAS (Pushchino, Moscow Region), University of Basel (Switzerland), University of Wisconsin (USA), University of Nijmegen. Radboda (Netherlands).
The article is based on a lecture given at the school “Modern biology and biotechnologies of the future” (Zvenigorod, January 26 - February 1, 2014).
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