Bacteriological study for dysentery. Laboratory diagnosis of dysentery, amoebiasis and balantidiasis

Dysentery.

Dysentery - infection, characterized by general intoxication of the body, loose stools and a peculiar lesion of the mucous membrane of the large intestine. It is one of the most common acute intestinal diseases in the world. The disease has been known since ancient times under the name “bloody diarrhea”, but its nature turned out to be different. In 1875 Russian scientist Lesh isolated an amoeba from a patient with bloody diarrhea Entamoeba histolytica, over the next 15 years, the independence of this disease was established, for which the name amoebiasis was retained. The causative agents of dysentery proper are a large group of biologically similar bacteria, united in the genus Shigelta. The pathogen was first discovered in 1888. A. Chantemes and Vidal; in 1891 it was described by A.V. Grigoriev, and in 1898. K. Shiga, using serum obtained from the patient, identified the pathogen in 34 patients with dysentery, finally proving the etiological role of this bacterium. However, in subsequent years, other pathogens of dysentery were discovered: in 1900. - S. Flexner, in 1915 - K. Sonne, in 1917 - K. Stutzer and K. Schmitz, in 1932. - J. Boyd, in 1934 - D. Large, in 1943 - A. Sax.

Currently genus Shigella includes more than 40 serotypes. All of them are short, immobile gram-negative rods that do not form spores or capsules, which (grow well on ordinary nutrient media, do not grow on a medium with citrate as the only carbon source; do not form H2S, do not have urease; Voges-Proskauer reaction is negative; glucose and some other carbohydrates are fermented to form acid without gas (except for some biotypes Shigella flexneri: S.manchester And ewcastle); As a rule, they do not ferment lactose (with the exception of Shigella Sonne), adonitol, inositol, do not liquefy gelatin, usually form catalase, and do not have lysine decarboxylase and phenylalanine deaminase. The G+C content in DNA is 49-53 mol%. Shigella are facultative anaerobes, the optimum temperature for growth is 37 ° C, they do not grow above 45 ° C, the optimal pH of the environment is 6.7-7.2. Colonies on dense media are round, convex, translucent; in case of association, rough R-shaped colonies are formed. Growth on MPB in the form of uniform turbidity, rough forms form a sediment. Freshly isolated cultures of Shigella Sonne J4HO form colonies of two types: small round convex (phase I), large flat (phase 2). The nature of the colony depends on the presence (phase I) or absence (phase II) of a plasmid with mm 120 MD, which also determines the virulence of Shigella Sonne.

In Shigella, O-antigens of different specificity were found: common to the family Enterobacteriaceae generic, species, group and type-specific, as well as K-antigens; They do not have N-antigens.

The classification takes into account only group and type-specific O-antigens. In accordance with these characteristics, the genus Shigella is divided into 4 subgroups, or 4 species, and includes 44 serotypes. In subgroup A (type Shigella dysenteriae) included Shigella, which does not ferment mannitol. The species includes 12 serotypes (1-12). Each stereotype has its own specific type antigen; antigenic connections between serotypes, as well as with other Shigella species, are weakly expressed. To subgroup B (type Shigella flexneri) include Shigella, which usually ferments mannitol. Shigella of this species are serologically related to each other: they contain type-specific antigens (I-VI), by which they are divided into serotypes (1-6), and group antigens, which are found in different compositions in each serotype and by which the serotypes are divided into subserotypes. In addition, this species includes two antigenic variants - X and Y, which do not have typical antigens; they differ in sets of group antigens. Serotype S.flexneri 6 has no subserotypes, but is divided into 3 biochemical types based on the fermentation characteristics of glucose, mannitol and dulcitol.

To subgroup C (type Shlgella boydll) include Shigella, which usually ferments mannitol. Members of the group are serologically different from each other. Antigenic connections within the species are weakly expressed. The species includes 18 serotypes (1-18), each of which has its own main type antigen.

In subgroup D (type Shlgella sonnel) included Shigella, which usually ferments mannitol and is capable of slowly (after 24 hours of incubation and later) fermenting lactose and sucrose. View S. sonnei includes one serotype, but colonies of phases I and II have their own type-specific antigens. For the intraspecific classification of Shigella Sonne, two methods have been proposed:

1) dividing them into 14 biochemical types and subtypes according to their ability to ferment maltose, rhamnose and xylose;

2) division into phage types according to sensitivity to a set of corresponding phages.

These typing methods have mainly epidemiological significance. In addition, Shigella Sonne and Shigella Flexner are typed for the same purpose based on their ability to synthesize specific colicins (colicinogenotyping) and sensitivity to known colicins (colicinotyping). To determine the type of colicins produced by Shigella, J. Abbott and R. Shenon proposed sets of standard and indicator strains of Shigella, and to determine the sensitivity of Shigella to known types colicins use a set of reference colicinogenic strains of P. Frederick.

Resistance. Shigella has a fairly high resistance to environmental factors. They survive on cotton fabric and paper for up to 30-36 days, in dried feces - up to 4-5 months, in soil - up to 3-4 months, in water - from 0.5 to 3 months, on fruits and vegetables - up to 2 food, in milk and dairy products - up to several weeks; at 60 °C they die in 15-20 minutes.

Sensitive to chloramine solutions, active chlorine and other disinfectants.

Pathogenicity factors. The most important biological property Shigella, which determines their pathogenicity, is the ability to invade epithelial cells, multiply in them and cause their death. This effect can be detected using a keratoconjunctival test (introduction of one loop of Shigella culture (2-3 billion bacteria) under the lower eyelid of a guinea pig causes the development of serous-purulent keratoconjunctivitis), as well as by infection of cell cultures (cytotoxic effect), or chicken embryos ( their death), or intranasally in white mice (development of pneumonia). The main pathogenicity factors of Shigella can be divided into three groups:

1) factors determining interaction with the epithelium of the mucous membrane;

2) factors that ensure resistance to humoral and cellular defense mechanisms of the macroorganism and the ability of Shigella to reproduce in its cells;

3) the ability to produce toxins and toxic products that determine the development of the pathological process itself.

The first group includes adhesion and colonization factors: their role is played by pili, outer membrane proteins and LPS. Adhesion and colonization are promoted by enzymes that destroy mucus - neuraminidase, hyaluronidase, mucinase. The second group includes invasion factors that promote the penetration of Shigella into enterocytes and their reproduction in them and in macrophages with the simultaneous manifestation of a cytotoxic and (or) enterotoxic effect. These properties are controlled by the genes of the plasmid with m.m. 140 MD (it encodes the synthesis of outer membrane proteins that cause invasion) and the chromosomal genes of Shigella: KSR A (causes keratoconjunctivitis), cyt (responsible for cell destruction), as well as other genes not yet identified. Protection of Shigella from phagocytosis is provided by the surface K-antigen, antigens 3, 4 and lipopolysaccharide. In addition, lipid A of Shigella endotoxin has an immunosuppressive effect - it suppresses the activity of immune memory cells.

The third group of pathogenicity factors includes endotoxin and two types of exotoxins found in Shigella - Shiga and Shiga-like exotoxins (SLT-I and SLT-II), the cytotoxic properties of which are most pronounced in S. dysenteriae 1. Shiga and Shiga-like toxins have also been found in other serotypes S. dysenteriae, they are also formed S.flexneri, S.sonnei, S.boydii, ETEC and some salmonella. The synthesis of these toxins is controlled by the tox genes of converting phages. Type LT enterotoxins are found in Shigella Flexner, Sonne and Boyd. Their LT synthesis is controlled by plasmid genes. Enterotoxin stimulates the activity of adenylate cyclase and is responsible for the development of diarrhea. Shiga toxin, or neurotoxin, does not react with the adenylate cyclase system, but has a direct cytotoxic effect. Shiga and Shiga-like toxins (SLT-I and SLT-II) have m.m. -70 kDa and consist of subunits A and B (the latter of 5 identical small subunits). The receptor for toxins is a glycolipid of the cell membrane.

The virulence of Shigella Sonne also depends on the plasmid with m.m. 120 MD. It controls the synthesis of about 40 outer membrane polypeptides, seven of them are associated with virulence. Shigella Sonne, having this plasmid, form phase I colonies and are virulent. Cultures that have lost the plasmid form phase II colonies and lack virulence. Plasmids with m.m. 120-140 MD were found in Shigella Flexner and Boyd. Shigella lipopolysaccharide is a strong endotoxin.

Features of epidemiology. The source of infection is only humans. No animals in nature suffer from dysentery. Under experimental conditions, dysentery can only be reproduced in monkeys. The method of infection is fecal-oral. Routes of transmission: water (predominant for Shigella Flexnera), food, especially milk and dairy products (predominant route of infection for Shigella Sonne), and household contact, especially for the species S. dysenteriae.

A feature of the epidemiology of dysentery is a change in the species composition of pathogens, as well as Sonne biotypes and Flexner serotypes in certain regions. For example, until the end of the 30s of the 20th century, the share S.dysenteriae 1 accounted for up to 30-40% of all cases of dysentery, and then this serotype began to occur less and less often and almost disappeared. However, in the 60-80s S.dysenteriae reappeared on the historical arena and caused a series of epidemics that led to the formation of three hyperendemic foci of it - in Central America, Central Africa and South Asia (India, Pakistan, Bangladesh and other countries). The reasons for the change in the species composition of dysentery pathogens are probably associated with changes herd immunity and with changes in the properties of dysentery bacteria. In particular, the return S.dysenteriae 1 and its widespread distribution, which caused the formation of hyperendemic foci of dysentery, is associated with its acquisition of plasmids that caused multidrug resistance and increased virulence.

Features of pathogenesis and clinic. The incubation period for dysentery is 2-5 days, sometimes less than a day. Formation infectious focus in the mucous membrane of the descending part of the large intestine (sigmoid and rectum), where the dysentery pathogen penetrates, it is cyclical in nature: adhesion, colonization, introduction of Shigella into the cytoplasm of enterocytes, their intracellular reproduction, destruction and rejection of epithelial cells, release of pathogens into the intestinal lumen; after this, the next cycle begins - adhesion, colonization, etc. The intensity of the cycles depends on the concentration of pathogens in the parietal layer of the mucous membrane. As a result of repeated cycles, the inflammatory focus grows, the resulting ulcers, connecting, increase exposure intestinal wall, as a result of which blood, mucopurulent lumps, and polymorphonuclear leukocytes appear in the stool. Cytotoxins (SLT-I and SLT-II) cause cell destruction, enterotoxin - diarrhea, endotoxins - general intoxication. The clinical picture of dysentery is largely determined by what type of exotoxins is produced to a greater extent by the pathogen, the degree of its allergenic effect and immune status body. However, many issues of the pathogenesis of dysentery remain unclear, in particular: the features of the course of dysentery in children of the first two years of life, the reasons for the transition of acute dysentery to chronic, the significance of sensitization, the mechanism of local immunity of the intestinal mucosa, etc. The most typical clinical manifestations dysentery is caused by diarrhea, frequent urge- in severe cases, up to 50 or more times a day, tenesmus (painful spasms of the rectum) and general intoxication. The nature of the stool is determined by the degree of damage to the large intestine. The most severe dysentery is caused by S.dysenteriae 1, most easily - Sonne dysentery.

Post-infectious immunity. As observations of monkeys have shown, after suffering from dysentery, strong and fairly long-lasting immunity remains. It is caused by antimicrobial antibodies, antitoxins, increased activity of macrophages and T-lymphocytes. Plays a significant role local immunity intestinal mucosa, mediated by IgAs. However, immunity is type-specific; strong cross-immunity does not occur.

Laboratory diagnostics . The main method is bacteriological. The material for research is feces. Pathogen isolation scheme: inoculation on differential diagnostic media Endo and Ploskirev (in parallel on enrichment medium followed by inoculation on Endo and Ploskirev media) to isolate isolated colonies, obtain pure culture, the study of its biochemical properties and, taking into account the latter, identification using polyvalent and monovalent diagnostic agglutinating sera. The following commercial serums are produced:

1. To Shigella, which does not ferment mannitol: to S.dysenteriae 1 to 2 S.dysenteriae 3-7(polyvalent and monovalent), to S.dysenteriae 8-12(polyvalent and monovalent).

2. To Shigella fermenting mannitol:

to typical antigens S.flexneri I, II, III, IV, V, VI,

to group antigens S.flexneri 3, 4, 6,7,8- polyvalent,

to antigens S.boydii 1-18(polyvalent and monovalent),

to antigens S. sonnei I phase, II phase,

to antigens S.flexneri I-VI+ S.sonnei- polyvalent.

To detect antigens in blood (including as part of the CEC), urine and feces can be used following methods: RPHA, RSK, coagglutination reaction (in urine and feces), IFM, RPGA (in blood serum). These methods are highly effective, specific and suitable for early diagnosis.

For serological diagnosis the following can be used: RPGA with appropriate erythrocyte diagnosticums, immunofluorescence method (indirect modification), Coombs method (determining the titer of incomplete antibodies). Diagnostic value also has an allergy test with dysenterin (solution of protein fractions of Shigella Flexner and Sonne). The reaction is taken into account after 24 hours. It is considered positive in the presence of hyperemia and infiltrate with a diameter of 10-20 mm.

Treatment. The focus is on restoring normalcy water-salt metabolism, rational nutrition, detoxification, rational antibiotic therapy (taking into account the sensitivity of the pathogen to antibiotics). Good effect gives early use of polyvalent dysentery bacteriophage, especially tableted with pectin coating, which protects the phage from the action of HC1 gastric juice; V small intestine pectin dissolves, phages are released and exert their effect. For preventive purposes, the phage should be given at least once every three days (the period of its survival in the intestine).

The problem of specific prevention. For creating artificial immunity Various vaccines were used against dysentery: from killed bacteria, chemical, alcohol, but all of them turned out to be ineffective and were discontinued. Vaccines against Flexner's dysentery have been created from live (mutant, streptomycin-dependent) Shigella Flexner; ribosomal vaccines, but they also did not find wide application. Therefore, the problem of specific prevention of dysentery remains unresolved. The main way to combat dysentery is to improve the water supply and sewerage system, ensure strict sanitary and hygienic regimes in food enterprises, especially the dairy industry, in child care institutions, public places and in maintaining personal hygiene.

Microbiology of cholera

According to WHO, cholera is a disease characterized by acute, severe, dehydrating diarrhea with rice-water-like stool, resulting from infection with Vibrio cholerae. Due to the fact that it is characterized by a pronounced ability for wide epidemic spread, severe course and high mortality, cholera is one of the most dangerous infections.

The historical homeland of cholera is India, more precisely, the delta of the Ganges and Brahmaputra rivers (now Eastern India and Bangladesh), where it has existed since time immemorial (cholera epidemics in this area have been observed since 500 years BC). The long existence of an endemic source of cholera here is explained by many reasons. Vibrio cholerae can not only survive in water for a long time, but also multiply in it under favorable conditions - temperatures above +12 ° C, and the presence of organic substances. All these conditions are available in India - tropical climate (average annual temperature from +25 up to +29 °C), abundance of precipitation and swampiness, high density population, especially in the Ganges Delta, a large number of organic substances in water, continuous year-round water pollution wastewater and excrement, low material standard of living and peculiar religious and cult rituals of the population.

The causative agent of cholera Vibrio cholerae was opened in 1883. during the fifth pandemic by R. Koch, however, vibrio was first discovered in the feces of patients with diarrhea back in 1854. F. Pacini.

V.cholerae belongs to the family Vibrionaceae which includes several genera (Vibrio, Aeromonas, Plesiomonas, Photobacterium). Genus Vibrio since 1985 has more than 25 species, of which highest value for a person have V.cholerae, V.parahaemolyticus, V.alginolyticus, dnificus And V. fluvialis.

Key characteristics of the genus Vibrio : short, not forming spores and capsules, curved or straight gram-negative rods, 0.5 µm in diameter, 1.5-3.0 µm in length, mobile ( V.cholerae- monotrich, some species have two or more polarly located flagella); grow well and quickly on ordinary media, chemoorganotrophs, ferment carbohydrates with the formation of acid without gas (glucose is fermented via the Embden-Meyerhof pathway). Oxidase positive, form indole, reduce nitrates to nitrites (V.cholerae gives a positive nitroso-indole reaction), break down gelatin, often give a positive Voges-Proskauer reaction (i.e., they form acetylmethylcarbinol), do not have urease, do not form H S. have lysine and ornithine decarboxylases, but do not have arginine dihydrolase.

Vibrio cholerae is very unpretentious to nutrient media. It multiplies well and quickly in 1% alkaline (pH 8.6-9.0) peptone water (PV) containing 0.5-1.0% NaCl, outpacing the growth of other bacteria. To suppress the growth of Proteus, it is recommended to add potassium tellurite 4 to 1% (PV) (final dilution 1:100,000). 1% PV is the best enrichment medium for Vibrio cholerae. As it grows, after 6-8 hours on the surface of the PV, it forms a delicate, loose, grayish film, which, when shaken, easily breaks down and falls to the bottom in the form of flakes; the PV becomes moderately cloudy. Various selective media have been proposed for the isolation of Vibrio cholerae: alkaline agar, yolk-salt agar, alkaline albuminate, alkaline blood agar, lactose-sucrose and other media. The best medium is TCBS (thiosulfate citrate-bromothymol sucrose agar) and its modifications. However, most often they use alkaline MPA, on which Vibrio cholerae forms smooth, glassy-transparent disc-shaped colonies with a viscous consistency with a bluish tint.

When sown by injection into a column of gelatin, vibrio after 2 days at 22-23 ° C causes liquefaction from the surface in the form of a bubble, then funnel-shaped and, finally, layer-by-layer.

In milk, vibrio multiplies quickly, causing coagulation after 24-48 hours, and then peptonization of the milk occurs, and after 3-4 days the vibrio dies due to a shift in the pH of milk to the acidic side.

B. Heiberg, based on the ability to ferment mannose, sucrose and arabinose, divided all vibrios (cholera and cholera-like) into a number of groups, the number of which is now 8. Vibrio cholerae belongs to Heiberg’s first group.

Vibrios, similar in morphological, cultural and biochemical characteristics to cholera, were and are called differently: paracholera, cholera-like, NAG-vibrios (non-agglutinating vibrios); vibrios not belonging to group 01. The last name most accurately emphasizes their relationship to Vibrio cholerae. As was established by A. Gardner and K. Venkatraman, cholera and cholera-like vibrios have a common H-antigen, but differ in O-antigens. According to the O-antigen, cholera and cholera-like vibrios are currently divided into 139 O-serogroups, but their number is constantly growing. Vibrio cholerae belongs to group 01. It has a common A-antigen and two type-specific antigens - B and C, which distinguish three serotypes V.cholerae- serotype Ogawa (AB), serotype Inaba (AS) and serotype Gikoshima (ABC). Vibrio cholerae in the dissociation stage has an OR antigen. In this regard, for identification V.cholerae O-serum, OR-serum and type-specific sera Inaba and Ogawa are used.

Pathogenicity factors V.cholerae :

1. Mobility.

2. Chemotaxis. With the help of these properties, the vibrio overcomes the mucous layer and interacts with epithelial cells. In Che" mutants (which have lost the ability to chemotaxis), virulence is sharply reduced. Virulence in Mot" mutants (which have lost mobility) either completely disappears or is reduced by 100-1000 times.

3. Adhesion and colonization factors, with the help of which vibrio adheres to microvilli and colonizes the mucous membrane of the small intestine.

4. Enzymes: mucinase, proteases, neuraminidase, lecithinase, etc.

They promote adhesion and colonization, as they destroy substances that make up the mucus. Neuraminidase, by cleaving sialic acid from epithelial glycoproteins, creates a “landing” site for vibrios. In addition, it increases the number of receptors for choleragen by modifying tri- and disialogangliosides into monosialoganglioside Gm b which serves as a receptor for choleragen.

5. The main factor of pathogenicity V.cholerae is an exotoxin-cholerogen, which determines the pathogenesis of cholera. The cholerogens molecule has a m.m. 84 kDa and consists of two fragments - A and B. Fragment A consists of two peptides - A1 and A2 - and has the specific property of cholera toxin. Fragment B consists of 5 identical subunits and performs two functions: 1) recognizes the receptor (monosialoganglioside) of the enterocyte and binds to it;

2) forms an intramembrane hydrophobic channel for the passage of subunit A. Peptide A 2 Cl is used to connect fragments A and B. The actual toxic function is performed by the peptide A t. It interacts with NAD, causes its hydrolysis, and the resulting ADP-ribose binds to the regulatory subunit of adenylate cyclase. This leads to inhibition of GTP hydrolysis. The resulting GTP + adenylate cyclase complex causes the hydrolysis of ATP with the formation of cAMP. (Another way of accumulating cAMP is the suppression of the enzyme that hydrolyzes cAMP to 5-AMP by cholerogens).

6. In addition to choleragen, Vibrio cholerae synthesizes and secretes a factor that increases capillary permeability.

7. Other exotoxins have also been found in Vibrio cholerae, in particular types LT, ST and SLT.

8. Endotoxin. Lipopolysaccharide V.cholerae has strong endotoxic properties. It is responsible for general intoxication of the body and vomiting. Antibodies formed against endotoxin have a pronounced vibriocidal effect (dissolve vibrios in the presence of complement) and are an important component of post-infectious and post-vaccination immunity.

The ability of vibrios not belonging to group 01 to cause sporadic or group diarrheal diseases in humans is associated with the presence of enterotoxins of the LT or ST type, which stimulate either the adenylate or guanylate cyclase systems, respectively.

Cholerogen synthesis - most important property V.cholerae. The genes that control the synthesis of the A- and B-fragments of cholerogens are combined into the vctAB or ctxB operon; they are located on the vibrio chromosome. Some strains of Vibrio cholerae have two such non-tandem operons. The function of the operon is controlled by two regulatory genes. The toxR gene provides positive control; mutations of this gene lead to a 1000-fold reduction in toxin production. The htx gene exerts negative control; mutations in this gene increase toxin production by 3-7 times.

The following methods can be used to detect cholerogens:

1. Biological tests on rabbits. When cholera vibrios are injected intraintestinal into suckling rabbits (no more than 2 weeks old), they develop a typical cholerogenic syndrome: diarrhea, dehydration and death of the rabbit. At the autopsy - a sharp injection of the vessels of the stomach and small
intestines, sometimes clear liquid accumulates in it. But the changes in the large intestine are especially characteristic - it is enlarged and filled with a completely transparent, straw-colored liquid with flakes and gas bubbles. When cholera vibrios are introduced into the ligated area of ​​the small intestine into adult rabbits, they experience the same changes in the large intestine as when suckling rabbits are infected.

2. Direct detection of cholerogens using immunofluorescent or enzyme-linked immunosorbent methods or a passive immune hemolysis reaction (cholerogen binds to Gm1 of erythrocytes, and they are lysed with the addition of antitoxic antibodies and complement).

3. Stimulation of cellular adenylate cyclase in cell cultures.

4. Using a chromosome fragment as a DNA probe V.cholerae, carrying an operoncholerogen.

During the seventh pandemic, strains were isolated V.cholerae With varying degrees virulence: cholerogenic (virulent), weakly cholerogenic (low virulent) and non-cholerogenic (non-virulent). Non-cholerogenic V.cholerae, as a rule, they have hemolytic activity, are not lysed by cholera diagnostic phage 5 (CDF-5) and do not cause human disease.

For phage typing V.cholerae(including V.eltor) S. Mukherjee proposed corresponding sets of phages, which were then supplemented with other phages in Russia. The set of such phages (1-7) makes it possible to distinguish among V.cholerae 16 phagotypes. HDF-3 selectively lyses Vibrio cholerae classic type, HDF-4 - El Tor vibrios, and HDF-5 lyses only cholerogenic (virulent) vibrios of both types and does not lyse non-cholerogenic vibrios.

Cholerogenic vibrios, as a rule, do not have hemolytic activity, are lysed by HDF-5 and cause cholera in humans.

Resistance of cholera pathogens. Vibrios cholerae survive well at low temperatures: in ice they remain viable for up to 1 month; in sea water - up to 47 days, in river water - from 3-5 days to several weeks, in boiled water mineral water persist for more than 1 year, in soil - from 8 days to 3 months, in fresh feces - up to 3 days, on cooked foods (rice, noodles, meat, cereals, etc.) survive for 2-5 days, on raw vegetables - 2- 4 days, on fruits - 1-2 days, in milk and dairy products - 5 days; when stored in the cold, the survival period increases by 1-3 days: on linen contaminated with feces, they last up to 2 days, and on damp material - a week. Cholera vibrios die at 80 °C in 5 minutes, at 100 °C - instantly; highly sensitive to acids; under the influence of chloramine and other disinfectants they die within 5-15 minutes. They are sensitive to drying and direct sun rays, but they persist well and for a long time and even multiply in open reservoirs and wastewater, rich in organic substances, having an alkaline pH and a temperature above 10-12 ° C. Highly sensitive to chlorine: a dose of active chlorine of 0.3-0.4 mg/l of water in 30 minutes causes reliable disinfection from Vibrio cholerae.

Features of epidemiology. The main source of infection is only a person - a person with cholera or a vibrio carrier, as well as water contaminated by them. No animals in nature suffer from cholera. The method of infection is fecal-oral. Routes of infection: a) main - through water used for drinking, bathing and household needs; b) contact and household and c) through food. All major cholera epidemics and pandemics were waterborne in nature. Vibrios cholerae have such adaptive mechanisms that ensure the existence of their populations both in the human body and in certain ecosystems of open water bodies. The profuse diarrhea caused by Vibrio cholerae leads to the cleansing of the intestines from competing bacteria and contributes to the widespread distribution of the pathogen in the environment, primarily in wastewater and in open water bodies where they are discharged. A person with cholera secretes the pathogen into a huge number- from 100 million to 1 billion per 1 ml of feces, the vibrio carrier releases 100-100,000 vibrios per 1 ml, the infectious dose is about 1 million vibrios. The duration of excretion of cholera vibrio in healthy carriers ranges from 7 to 42 days, and 7-10 days in those who have recovered from the disease. Longer discharge is extremely rare.

The peculiarity of cholera is that after it, as a rule, there is no long-term carriage and persistent endemic foci do not form. However, as already mentioned above, due to the pollution of open water bodies with wastewater containing large quantities organic matter, detergents and table salt, in the summer, the cholera vibrio not only survives in them for a long time, but even multiplies.

Of great epidemiological significance is the fact that cholera vibrios of group 01, both non-toxigenic and toxigenic, can persist for a long time in various aquatic ecosystems in the form of uncultivated forms. Using the polymerase chain reaction, with negative bacteriological studies, veterinary genes of uncultivated forms were discovered in a number of endemic areas of the CIS in various water bodies V.cholerae.

When cholera diseases occur, a set of anti-epidemic measures is carried out, among which the leading and decisive one is active timely detection and isolation (hospitalization, treatment) of patients in acute and atypical form and healthy vibrio carriers; measures are being taken to suppress possible ways of spreading the infection; special attention is paid to water supply (chlorination drinking water), compliance with the sanitary and hygienic regime at food enterprises, in child care institutions, and public places; Strict control, including bacteriological control, is carried out over open water bodies, immunization of the population is carried out, etc.

Features of pathogenesis and clinic. The incubation period for cholera varies from a few hours to 6 days, most often 2-3 days. Once in the lumen of the small intestine, cholera vibrios are directed to the mucus due to motility and chemotaxis to the mucous membrane. To penetrate through it, vibrios produce a number of enzymes: neuraminidase, mucinase, proteases, lecithinase, some destroy substances contained in mucus and facilitate the movement of vibrios to epithelial cells. By adhesion, vibrios attach to the glycocalyx of the epithelium and, losing mobility, begin to multiply intensively, colonizing the microvilli of the small intestine, and at the same time producing large amounts of exotoxin-cholerogen. Cholerogen molecules bind to monosialoganglioside Gm1 and penetrate the cell membrane, activate the adenylate cyclase system, and the accumulating cAMP causes hypersecretion of fluid, cations and anions Na +, HCO 3 ~, K +, SG from enterocytes, which leads to cholera diarrhea, dehydration and desalination body. There are three types of disease:

1. violent, severe dehydrating diarrheal disease, leading to the death of the patient within a few hours;

2. less severe course, or diarrhea without dehydration;

3. asymptomatic course of the disease (vibrio carriage).

In severe forms of cholera, patients develop diarrhea, stools become more frequent, bowel movements become more abundant, become watery, lose their fecal odor and look like rice water (a cloudy liquid with mucus residues and epithelial cells floating in it). Then comes debilitating vomiting, first with intestinal contents, and then the vomit takes on the appearance of rice water. The patient's temperature drops below normal, the skin becomes bluish, wrinkled and cold - cholera algid. As a result of dehydration, blood thickens and cyanosis develops. oxygen starvation, kidney function sharply suffers, convulsions appear, the patient loses consciousness and death occurs. The case fatality rate for cholera during the seventh pandemic ranged from 1.5% in developed countries to 50% in developing countries.

Post-infectious immunity durable, long-lasting, recurrent diseases are rare. Antitoxic and antimicrobial immunity is caused by antibodies (antitoxins last longer than antimicrobial antibodies), immune memory cells and phagocytes.

Laboratory diagnostics. Main and decisive method Diagnosis of cholera is bacteriological. The material for research from the patient is feces and vomit; feces are examined for vibrio carriage; from persons who died from cholera, a ligated segment of the small intestine and gall bladder are taken for research; Among environmental objects, water from open reservoirs and wastewater are most often studied.

When conducting a bacteriological study, the following three conditions must be observed:

1) inoculate material from the patient as quickly as possible (vibrio cholera remains in the feces short term);

2) the container in which the material is taken should not be disinfected with chemicals and should not contain traces of them, since Vibrio cholerae is very sensitive to them;

3) exclude the possibility of contamination and infection of others.

In cases where there are V.cholerae not 01-groups, they must be typed using appropriate agglutinating sera of other serogroups. Discharge from a patient with diarrhea (including cholera-like) V.cholerae not 01-group requires the same anti-epidemic measures as in the case of isolation V.cholerae 01-groups. If necessary, the ability to synthesize choleragen or the presence of choleragen genes in isolated cholera vibrios is determined using a DNA probe using one of the methods.

Serological diagnosis of cholera is auxiliary. For this purpose, an agglutination reaction can be used, but it is better to determine the titer of vibriocidal antibodies or antitoxins (cholerogen antibodies are determined by enzyme-linked immunosorbent or immunofluorescent methods).

Treatment for patients with cholera should consist primarily of rehydration and restoration of normal water-salt metabolism. For this purpose, it is recommended to use saline solutions, for example, of the following composition: NaCl - 3.5; NaHCO 3 - 2.5; KS1 - 1.5 and glucose - 20.0 g per 1 liter of water. Such pathogenetically based treatment in combination with rational antibiotic therapy can reduce the mortality rate in cholera to 1% or less.

Specific prevention. Various vaccines have been proposed to create artificial immunity, including killed Inaba and Ogawa strains; Cholerogen toxoid for subcutaneous use and enteral chemical bivalent vaccine, sos

LABORATORY DIAGNOSTICS OF DYSENTERY, AMOEBIASE AND BALANTIDIASIS

In modern conditions, in isolated cases or in small focal outbreaks of intestinal diseases, often occurring with unclear symptoms, laboratory research methods acquire important practical importance.

Studies carried out for diagnostic purposes must be carried out in strict compliance with established recommendations and as early as possible.

Collection feces for research, produce in clean dishes (night vases, bedpans) that do not contain residual disinfectants; material for research is taken from the rectal mucosa with swabs during sigmoidoscopy.

To bacteriologically confirm the diagnosis, it is better to take stool from patients with dysentery before treatment with antibiotics and sulfonamides, and to determine bacterial carriage - after treatment with these drugs.

Sowing on Petri dishes must be done immediately after taking the material.

First of all, a macroscopic examination of stool is carried out, which can detect: food debris - pieces of meat, fat residues, plant food and pathological impurities - mucus of a viscous consistency in the form of lumps (not transparent in dysentery and transparent in amoebiasis); blood not changed by dysentery and ulcerative lesion the lower part of the colon of a different etiology, and a changed color (“raspberry jelly”) with amebiasis, balantidiasis; pus is detected in severe protracted forms of dysentery.

Microscopic examination of feces is used to detect cellular elements of the blood, amoebae, balantidia and their cysts. The native preparation is prepared as follows: a lump of feces is placed on a glass slide with a loop and a drop of isotonic sodium chloride solution next to it, mixed and covered with a coverslip. Lugol's solution is used to stain protozoa.

To differentiate the cellular elements of the blood, the preparations are treated with Romanovsky-Giemsa dye or azure-eosin. In dysentery, in a preparation prepared from mucus, many neutrophilic granulocytes (over 90% of all cellular elements), single eosinophilic granulocytes (eosinophils) and different quantity red blood cells; in amebiasis, there are few cellular elements, the main mass of which consists of altered cells with a pyknotic nucleus and a narrow rim of protoplasm. Eosinophilic granulocytes and Charcot-Leyden crystals are found.

Tissue forms of amoeba (Entamoeba histolytica) in an unstained preparation are colorless, mobile (with the help of pseudopodia), in an elongated state they reach 50-60 microns, they are often found in the endoplasm with erythrocytes and towards the periphery - the nucleus. The presence of red blood cells in the cell makes it possible to distinguish Entamoeba histolutica from non-pathogenic forms (E. hartmani, E. coli.).

The luminal form of amoeba is smaller in size (up to 20 microns), inactive and does not contain red blood cells. Cysts are even smaller in size (10-12 microns), round shape, motionless; in the early stages of development they contain 2 nuclei, and mature ones - 4. In preparations stained with Lugol's solution, the nuclei of amoebas and their cysts are light brown (Fig. 6).

Balantidia(Balantidium coli) are large ciliates, sometimes reaching 200 microns in length and 50-70 microns in diameter, mobile, due to the presence of cilia, and have oral (peristome) and anal (cytopygous) openings. In the endoplasm, large (macronucleos) and small (micronucleos) nuclei, vacuoles, and trapped erythrocytes are visible. Balantidia cysts are immobile, round in shape, 50-60 µm in diameter, have a double-circuited shell, and contain macronucleoses and vacuoles inside (Fig. 7).

Bacteriological examination of stool for bacterial dysentery is best done soon after defecation, and material (mucus and pus) must be taken from the last portions of stool. The test material is inoculated with a loop onto Petri dishes with elective media (Ploskireva, Ploskireva + chloramphenicol, Levin) and placed in a thermostat for 18-24 hours at a temperature of +37° C. The next day, suspicious (colorless) colonies are subcultured on Ressel’s medium and placed test tubes in a thermostat for a day at a temperature of +370 C. On the third day, having received a pure culture, smears are prepared for microscopy and study of motility (Shigella is immobile). An agglutination reaction is performed on glass with type-specific sera, first an indicative one with sera against the types prevailing in the area, and then expanded and inoculated on the “variegated” row to determine biochemical properties selected culture.

The causative agents of dysentery do not ferment lactose and sucrose (except Sonne), decompose glucose (to acid), and do not form hydrogen sulfide.

The final answer during bacteriological examination is given on the 5th day. Sometimes atypical strains of the pathogen are isolated, cultures that have lost agglutinability and with other characteristics. In such cases, research continues over longer periods.

There are also accelerated bacteriological methods - reseeding suspicious colonies from Petri dishes 18-20 hours from the start of the study into 2 tubes with Ressel's medium (slanted agar with 1% lactose and 0.1% glucose - in one and 1% sucrose and 0.1 % mannitol - in another). After 4 hours, the growth of colonies may already appear, from which smears are prepared, stained with Gram, motility is studied and an approximate agglutination reaction is performed with sera against the most common pathogens in the area. Thus, already on the second day a preliminary answer can be given. The final answer is given on the third day after taking into account the results of sowing on the “variegated” row and a detailed agglutination reaction.

The inoculation rate of dysentery pathogens is not always the same; it depends on many factors - the method of collecting material for research, the quality of media and other reasons, one of which is the number of pathogens per unit volume of feces. It has been proven that dysentery pathogens are sown in cases where one gram of feces contains at least hundreds of millions of microbial bodies. IN in rare cases It is possible to isolate the causative agent of dysentery from the blood.

In the presence of a fluorescent microscope and specific sera with fluorochromes, students are shown the method of direct fluorescence of antibodies.

It is also possible to perform an agglutination reaction with the patient’s blood serum and diagnostic tests, however, antibody titers in patients with dysentery are low and, in addition, para-agglutination phenomena are common, which makes it difficult to obtain reliable results. More sensitive is the indirect hemagglutination reaction (IRHA) with standard erythrocyte diagnostics. By auxiliary method The study is an intradermal allergy test with dysenterin according to D. A. Tsuverkalov, which is taken into account after 24 hours according to the size of the resulting papule.

Guidelines for students for practical lesson No. 28.

Lesson topic:

Target: Studying methods of microbiological diagnosis, etiotropic therapy and prevention of shigellosis.

Module 2 . Special, clinical and environmental microbiology.

Topic 5: Methods for microbiological diagnosis of dysentery.

Relevance of the topic:Shigellosis is widespread and poses a serious problem in countries with a low sanitary cultural level and a high incidence of insufficient and poor-quality nutrition. In developing countries, the spread of infection is facilitated by poor sanitation, poor personal hygiene, overcrowding and a large proportion of children among the population. In Ukraine, outbreaks of shigellosis are more common in closed groups against the background low level sanitation and hygiene, for example in nurseries and kindergartens, on tourist boats, in psychiatric clinics or shelters for the disabled. Shigella has been the cause of diarrhea in travelers and tourists.

The cause of group diseases can be considered the consumption of food products contaminated due to the negligence of sales workers who are carriers of Shigella. There are outbreaks associated with the consumption of drinking water; swimming in contaminated water bodies has also led to infection. However, food and water transmission routes appear to play a lesser role in the spread of shigellosis compared to cholera and typhoid fever, which usually require large doses pathogens. In developing countries, where disease spread is predominantly person-to-person, carriers may be an important reservoir of the infectious agent. In patients who have not taken antibacterial drugs, the excretion of Shigella in feces usually continues for 1×4 weeks, but in a small proportion of cases it continues significantly longer.

Shigellosis is an acute bacterial infection of the intestines caused by one of four types of Shigella. The spectrum of clinical forms of infection includes mild, watery diarrhea, and severe dysentery, which is characterized by cramping pain in the abdomen, tenesmus, fever and signs of general intoxication.

Etiology.

The genus Shigella (named after K. Shiga, who in 1898 studied and described in detail the isolated causative agent of bacterial dysentery by A.V. Grigoriev) of the family Enterobacteriaceae consists of a group of closely related species of bacteria with following properties:

I. Morphological: Shigella - small sticks with rounded ends. They differ from other representatives of the family Enterobacteriaceae in the absence of flagella (immobile), do not have spores or capsules, and are gram-negative.

II. Cultural: Shigella are aerobes or facultative anaerobes; optimal cultivation conditions: temperature 37°C, pH 7.27.4. They grow on simple nutrient media (MPA, MPB) in the form of small, shiny, translucent, grayish, round colonies, 1.5 x 2 mm in size. S form. The exception is Shigella Sonne, which often dissociates, forming large, flat, cloudy colonies with jagged edges R shapes (colonies have the appearance of a “grape leaf”). In liquid nutrient media, Shigella produces uniform turbidity, R forms form a precipitate. The liquid enrichment medium is selenite broth.

III. Enzymatic: the main biochemical characteristics necessary for identifying Shigella when isolating a pure culture are the following:

1. absence of gas formation during glucose fermentation;
2. lack of hydrogen sulfide production;
3. no lactose fermentation for 48 hours.

Overall, the four species are further divided into approximately 40 serotypes. According to the characteristics of the main somatic (O) antigens and biochemical properties, the following four species or groups are distinguished: S. dysenteriae (group A, includes: Grigoriev-Shigi, Stutzer-Schmitz, Large-Sachs), S. flexneri (group B), S. boydii (group C) and S. sonnei (group D).

In relation to mannitol, all Shigella are divided into splitting (Shigella Flexner, Boyd, Sonne) and non-splitting (Shigella Grigoriev-Shiga, Stutzer-Schmitz, Large-Sachs) mannitol.

IV. Pathogenicity factors:

1. Invasion plasmidprovides the ability of Shigella to cause invasion with subsequent intercellular spread and reproduction in the epithelium of the colon mucosa;
2. Toxin formation: Shigella has a lipopolysaccharide endotoxin, which is chemically and biochemically similar to the endotoxins of other members of the family Enterobacteriaceae. In addition, S. dysenteriae type I (Shiga bacillus) produces an exotoxin. Since the discovery of the latter, it has been found that it has enterotoxin activity and can cause intestinal secretion, as well as have a cytotoxic effect directed against intestinal epithelial cells; provides neurotoxic effect which is observed in children with shigellosis. Shiga toxin, entering the blood, along with damage to the submucosal endothelium, also affects the glomeruli of the kidney, as a result of which, in addition to bloody diarrhea, hemolytic uremic syndrome develops with the development of renal failure.

V. Antigenic structure:All Shigella have a somatic O-antigen, depending on the structure of which they are divided into serovars.

VI. Resistance: Temperature 100 0 C kills Shigella instantly. Shigella is resistant to low temperatures in river water they last up to 3 months, on vegetables and fruits - up to 15 months.Under favorable conditions, Shigella is capable of reproducing in food products (salads, vinaigrettes, boiled meat, minced meat, boiled fish, milk and dairy products, compotes and jelly), especially Shigella Sonne.

Epidemiology.

1. Source of infection:A person suffering from acute and chronic forms of shigellosis; bacteria carrier.

2. Transmission routes:

• Food grade (mainly for S. sonnei)
• Aquatic (mainly for S. flexneri)
• Contact household (mainly for S. dysenteriae)

3. Entrance gateinfection serves gastrointestinal tract.

Pathogenesis and pathological changes.

After ingestion, Shigella colonizes upper sections small intestine and multiply there, possibly causing increased secretion in early stage infections. Shigella then penetrates through M cells into the submucosa, where it is absorbed by macrophages. This leads to the death of some Shigella, resulting in the release of inflammatory mediators, which initiate inflammation in the submucosa. Apoptosis of phagocytes allows another part of Shigella to survive and penetrate into the epithelial cells of the mucosa through basement membrane. Shigella multiply and spread intercellularly within enterocytes, resulting in the development of erosions. When Shigella dies, Shiga and Shiga-like toxins are released, the action of which leads to intoxication. Damage to the mucous membrane is accompanied by swelling, necrosis and hemorrhage, which causes the appearance of blood in the stool. In addition, the toxin affects the central nervous system, which leads to trophic disorders.

Clinical manifestations.

The range of clinical manifestations of shigellosis is very wide, from mild diarrhea to severe dysentery with cramping pain in the abdomen, tenesmus, fever and general intoxication.

Incubation periodranges from several hours to 7 days, most often 2-3 days.Initially, patients experience watery stool, fever (up to 41°C), diffuse abdominal pain, nausea and vomiting. Along with this, patients complain of myalgia, chills, lower back pain and headache. In the coming days from the onset of the disease, signs of dysentery appear: tenesmus, frequent, scanty, bloody-mucous stools. Body temperature gradually decreases, pain can be localized in the lower quadrants of the abdomen. The intensity of diarrhea reaches its maximum around the end of the 1st week of illness. Dysentery with bloody stools is more common and appears earlier in the disease caused by S. dysenteriae type I than in other forms of shigellosis.

For shigellosis Sonne A milder course of the disease is characteristic (gastroenteric or gastroenterocolitic variant). The febrile period is shorter, the symptoms of intoxication are short-lived, and destructive changes in the intestinal mucosa are not typical.

Flexner's shigellosisBasically, two variants of the clinical course are characteristic - gastroenterocolitic and colitic.

Extraintestinal complications in shigellosisrare:

1. A complication of shigellosis can be the development of intestinal dysbiosis.
2. Along with headaches, signs of meningitis and seizures may occur.
3. Peripheral neuropathy has been reported in S. dysenteriae type I infections, and Guillain-Barré syndrome (polyneuritis) has been reported during an outbreak of S. boydii gastroenteritis.
4. With the exception of children suffering from dystrophy, hematogenous dissemination of the pathogen is relatively rare; cases of shigellosis abscesses and meningitis have also been described.
5. With shigellosis, the development of Reiter's syndrome with arthritis, sterile conjunctivitis and urethritis is possible; this usually occurs 1-4 weeks after the onset of diarrhea in patients.
6. In children, shigellosis is accompanied by hemolytic-uremic syndrome, often in combination with leukemia-like reactions, severe colitis and endotoxin circulation, but bacteremia is usually not detected.
7. Quite rarely, purulent keratoconjunctivitis is caused by Shigella, which has entered the eyes as a result of self-infection with contaminated fingers.
8. Hypovolemic shock and disseminated intravascular coagulation syndrome.
9. Peritonitis, intestinal gangrene, intestinal bleeding.

Immunity: Humans have natural resistance to shigella infection. After past illness immunity is not stable, and after shigellosis Sonne is practically absent. In case of a disease caused by Shigella Grigoriev Shiga, a more stable antitoxic immunity is developed. In protection against infection, the main role belongs to secretory IgA , preventing adhesion, and cytotoxic antibody-dependent activity of intraepithelial lymphocytes, which, together with secretory IgA destroy Shigella.

Diagnostics and laboratory tests.

Purpose of the study: detection and identification of Shigella for diagnosis; identification of bacteria carriers; detection of Shigella in food products.

Material for research: feces, sectional material, food products.

Diagnostic methods:microbiological (bacteriological, microscopic (luminescent); serological; biological; allergy test.

Progress of the study:

1 day of study:Cultures should be done from freshly excreted feces or using rectal swabs (rectal tube); If suitable conditions are not available, the material must be placed in a transport environment. To do this, use intestinal agar (MacConkey or Shigella-Salmonella medium), moderately selective xylose-lysine deoxycholate agar, KLD) and nutrient broth (selenite broth). If the time between collection and inoculation exceeds 2 hours, then preservative solutions should be used: 20% bile broth, combined Kauffmann's medium.

• The feces in the glycerin mixture are emulsified, a drop of the emulsion is applied to the medium and rubbed in with a spatula. Differential media for Shigella are Ploskirev, Endo and EMS (eosin methylene blue agar). Ploskirev's medium (medium includes: MPA, lactose, salts bile acids and indicator brilliant green) is also an elective environment for Shigella, because inhibits the growth of Escherichia coli.
• In parallel with direct sowing collected material inoculated on enrichment medium selenite broth.
• All crops are placed in a thermostat.

Day 2 of the study:

• The dishes are removed from the thermostat, suspicious colonies are screened out on Ressel's medium (nutrient medium which includes: agar-agar, Andrede indicator, 1% lactose, 0.1% glucose) and mannitol. Inoculation is carried out with strokes on a beveled surface and an injection into an agar column. The inoculated Ressel medium is placed in a thermostat for 18-24 hours (at the same time, reseeding from the selenite medium is done into differential diagnostic media).
• Smears are made (Gram stain) and examined under a microscope.
• Preparations are prepared as a “hanging” or “crushed” drop.
• Establishment of tentative RA with polyvalent diagnostic shigella sera.
• Sowing suspicious colonies onto agar slants.

Day 3 of the study:

• Microscopy of material from agar slants.
• Cultures that have not fermented lactose on Ressel's medium are subjected to further study: smears are made (Gram staining), and the purity of the culture is checked. In the presence of gram-negative rods, inoculation is carried out on Hiss media, broth with indicator papers (to detect indole and hydrogen sulfide) and litmus milk.
• The inoculated media are placed in a thermostat for 18-24 hours.

Day 4 of the study:

• Accounting for a short “variegated row”.
• Cultures suspicious for their enzymatic and cultural properties in relation to Shigella are subjected to serological identification. Statement of RA on glass (typical and group diagnostic sera). Setting up a deployed RA.

As accelerated methods used for shigellosisfluorescence microscopy And biological sample(introduction of virulent strains of Shigella into the conjunctival sac (under the lower eyelid) of guinea pigs; conjunctivitis develops by the end of the 1st day).

Tsuverkalov allergy testintradermal allergy test with dysenterine (injection of 0.1 ml of dysenterine into the forearm positive reaction in case of infiltration and hyperemia). Allergy diagnostics are currently practically not used. Tsurvekalov's test is not specific; positive reactions are recorded not only for shigellosis, but also for salmonellosis, escherichiosis, yersiniosis, etc. OKI, and sometimes in healthy individuals.

Treatment and prevention.For treatment and prevention according to epidemiological indications, bacteriophage is used oral administration, antibiotics after determining the antibiogram; in case of dysbacteriosis probiotic preparations to correct microflora. To replenish the loss of fluids and electrolytes, administer a glucose-electrolyte solution inside.

Specific goals:

Interpret the biological properties of shigellosis pathogens.

Familiarize yourself with the classification of Shigella.

Learn to interpret the pathogenetic patterns of the infectious process caused by Shigella.

Determine methods of microbiological diagnosis, etiotropic therapy and prevention of shigellosis.

Be able to:

• Inoculate the test material on nutrient media.
  • Prepare smears and Gram stain.
  • Conduct microscopy of preparations using an immersion microscope.
  • Analyze the morphological, cultural, enzymatic characteristics of Shigella.

Theoretical questions:

1. Characteristics of shigellosis pathogens. Biological properties.

2. Classification of Shigella. The principles underlying it.

3. Epidemiology, pathogenesis and clinical features Shigellosis

4. Laboratory diagnostics.

5. Principles of treatment and prevention of shigellosis.

Practical tasks performed in class:

1. Microscopy of demonstration preparations from pure cultures of shigellosis pathogens.

2. Work on the bacteriological diagnosis of shigellosis: study of fecal cultures on Ploskirev’s medium.

3. Subculture of suspicious colonies on Ressel’s medium and MPB to determine indole formation and H 2 S.

4. Sketching demonstration preparations and microbiological diagnostic diagrams of shigellosis into the lesson protocol.

5. Drawing up the protocol.

Literature:

1. Korotyaev A.I., Babichev S.A., Medical microbiology, immunology and virology / Textbook for medical universities, St. Petersburg “Special literature”, 1998. - 592 p.

2. Timakov V.D., Levashev V.S., Borisov L.B. Microbiology / Textbook.-2nd ed., revised. And additional - M.: Medicine, 1983, - 512 p.

3. Pyatkin K.D. Krivoshein Yu.S. Microbiology with virology and immunology.- Kyiv: V i scha school, 1992. - 431 p.

4. Medical microbiology / Edited by V.I. Pokrovsky.-M.:GEOTAR-MED, 2001.-768p.

5. Guide to practical classes in microbiology, immunology and virology. Ed. M.P. Zykova. M. "Medicine". 1977. 288 p.

6. Cherkes F.K., Bogoyavlenskaya L.B., Belskan N.A. Microbiology. /Ed. F.K. Circassian. M.: Medicine, 1986. 512 p.

7. Lecture notes.

Additional literature:

1. Makiyarov K.A. Microbiology, virology and immunology. Alma-Ata, “Kazakhstan”, 1974. 372 p.

2. Titov M.V. Infectious illnesses. - K., 1995. 321 p.

3. Shuvalova E.P. Infectious diseases. - M.: Medicine, 1990. - 559 p.

4. BME, T. 1, 2, 7.

5. Pavlovich S.A. Medical microbiology in graphs: Textbook. allowance for medical Inst. Mn.: Higher. school, 1986. 255 p.

Brief guidelines to work in a practical lesson.

At the beginning of the lesson, the students' level of preparation for the lesson is checked.

Independent work consists of studying the classification of Shigella, analyzing the scheme of pathogenetic and clinical signs Shigellosis Study of methods for laboratory diagnosis of shigellosis. Students inoculate biomaterial on nutrient media. Then microslides are prepared, stained with Gram, microscopy is performed, microslides are sketched and the necessary explanations are given. Independent work also includes microscopy of demonstration preparations and their sketching in the lesson protocol.

At the end of the lesson, a test control and analysis of the final results of each student’s independent work are carried out.

Technological map for conducting a practical lesson.

p/p	Stages	Time in minutes	Ways of learning	Equipment	Location
	Checking and correcting the initial level of preparation for the lesson		Test tasks baseline	Tables, atlas	Study room
	Independent work		Logical structure graph	Immersion microscope, dyes, glass slides, bacteriological loops, nutrient media, Ploskirev’s medium, Ressel’s medium, “Hiss variegated series”
	Self check and correction of material mastery		Targeted learning tasks
	Test control		Tests
	Analysis of work results

Target training tasks:

1. Feces containing mucus and pus were obtained from a child with ACI (collection of stool was carried out using a rectal tube). What express diagnostic method should be used?

A. ELISA.

B. REEF.

C. RA.

D. RSK.

E. RIA.

1. The causative agent of dysentery was isolated from a sick child with an acute intestinal infection. Which morphological characteristics characteristic of the pathogen?

A . Gram-negative non-motile rod.

B . Gram-positive motile rod.

C . Forms a capsule on a nutrient medium.

D . Forms spores in the external environment.

E . Gram-positive streptobacilli.

3. A patient who fell ill three days ago and complains of a temperature of 38°C, abdominal pain, frequent loose stool, the presence of blood in the stool was clinically diagnosed by the doctor bacillary dysentery. What microbiological diagnostic method is advisable to use in this case and what material should be taken from the patient to confirm the diagnosis?

A. Bacterioscopic cal.

B. Bacteriological cal.

C. Bacterioscopic blood.

D. Bacteriological urine.

E. Serological blood.

4. Shigella Sonne was isolated from the patient’s feces. What needs to be done additional research to determine the source of infection?

A . Carry out phage typing of the isolated pure culture.

B . Determine the antibiogram.

C . Set up a precipitation reaction.

D . Perform a complement fixation reaction.

E . Set up a neutralization reaction.

5. Among a group of tourists (27 people) who used lake water for drinking, after two days, 7 people developed symptoms acute diarrhea. What material is needed to establish the etiology? of this disease need to be sent to a bacteriological laboratory?

A. Water, feces of patients.

B. Water, the blood of patients.

C. Food products.

D. I'm peeing.

E. Sputum.

6. A significant drawback of the microscopic diagnostic method for acute intestinal infections is its lack of information content due to the morphological identity of bacteria of the family Enterobacteriaceae . What makes this method more informative?

A . Radioimmunoassay.

B . Coombs reaction.

C . Linked immunosorbent assay.

D . Opsonization reaction.

E . Immunofluorescence reaction.

7. A 29-year-old patient was hospitalized with attacks of vomiting, diarrhea, and tenesmus. Feces with pieces of mucus and some blood. A bacteriological study of bacteria from colonies on Ploskirev's medium revealed immobile, gram-negative rods that do not ferment lactose. Name the causative agent of the infectious process.

A. Shigella flexneri.

B. Vibrio eltor.

C. E. Coli.

D. Proteus mirabilis.

E. Salmonella enteritidis.

8. Lettuce was delivered to the microbiological laboratory, which is presumably the cause of acute intestinal infection. What nutrient media are used for primary sowing?

A . Yolk salt agar, MPB.

B. MPA, MPB.

C . Selenite broth, Endo, Ploskireva.

D . Liver broth, Roux medium.

E . Blood agar, alkaline agar.

9. During a microbiological study of minced meat, bacteria belonging to the genus Shigella were isolated. The study of what properties of microbes led to this conclusion?

A . Cultural, tinctorial.

B . Antigenic, cultural.

C . Saccharolytic, proteolytic.

D . Antigenic, immunogenic.

E . Morphological, antigenic.

10. When microscopic examination vomit taken from a patient with symptoms of acute intestinal infection, motionless sticks were found. In what smear or preparation could the mobility of bacteria be studied?

A . In a Gram-stained smear.

B . In a smear stained according to Ziehl-Neelsen.

C . The preparation contains a “thick drop”.

D . In a smear stained according to Neisser.

E . The preparation contains a “crushed drop”.

Algorithm laboratory work:

1. Study of the biological properties of Shigella.

2. Familiarization with the classification of Shigella.

3. Analysis of the scheme of pathogenetic and clinical manifestations of shigellosis.

4. Study of methods for laboratory diagnosis of shigellosis.

5. Study of the basic principles of therapy and prevention of shigellosis.

1. Preparation of fixed preparations from bacterial culture.
2. Coloring microslides according to Gram.
3. Microscopy of microslides With using an immersion microscope, their analysis and recording in the lesson protocol.
4. Mi croscopy and analysis of demonstration preparations from pure cultures of Shigella.
5. Drawing of demonstration preparations and laboratory diagnostic diagrams of shigellosis into the protocol.
6. Drawing up the protocol.

Dysentery is a painful infection accompanied by diarrhea with the release of blood, pus and mucus, abdominal pain and symptoms of general intoxication, occurring with a predominant lesion of the colon, caused by different types sort of Shigella(dysentery bacteria).

Pathogens of dysentery belong to the department Gracilicutes, family Enterobacteriaceae, family Shigella.
Dysentery , called Shigella dysenteriae, is more severe than diseases caused by other Shigella, since in addition to endotoxin, which causes intestinal inflammation, this type of bacteria produces a strong exotoxin that acts as a neurotoxin

Bacterial dysentery , or shigellosis, is an infectious disease caused by bacteria of the genus Shigella,

Dysentery.Morphology and tinctorial properties.
Shigella are gram-negative rods with rounded ends, 2-3 microns long, 0.5-7 microns thick, do not form spores, do not have flagella, and are immobile. In many strains, villi of the general type and sex pili are found. Some Shigella have a microcapsule.

Dysentery. Cultivation.
Dysentery bacilli are facultative anaerobes. They are undemanding to nutrient media and grow well at a temperature of 37 °C and a pH of 7.2-7.4. On dense media they form small transparent colonies, in liquid media - diffuse turbidity. Selenite broth is most often used as an enrichment medium for the cultivation of Shigella.

Dysentery.Enzyme activity.
Shigella has less enzymatic activity than other enterobacteria. They ferment carbohydrates to form acid. An important feature that allows Shigella to be differentiated is their relationship to mannitol: S. dysenteriae does not ferment mannitol, representatives of groups B, C, D are mannitol-positive. The most biochemically active are S. sonnei, which can slowly (within 2 days) ferment lactose. Based on the relationship of S. sonnei to rhamnose, xylose and maltose, 7 biochemical variants are distinguished.

Dysentery.Antigenic structure.
Shigella has an O-antigen, its heterogeneity makes it possible to distinguish serovars and subserovars within groups; Some members of the genus exhibit K-antigen.

Dysentery.Pathogenicity factors.
All dysentery bacilli form endotoxin, which has an enterotropic, neurotropic, and pyrogenic effect. In addition, S. dysenteriae - Shigella Grigoriev-Shiga - secrete an exotoxin that has an enterotoxic, neurotoxic, cytotoxic and nephrotoxic effect on the body, which accordingly disrupts water-salt metabolism and the activity of the central nervous system, leading to the death of epithelial cells of the colon intestines, damage to the renal tubules.

The formation of an exotoxin is associated with a more severe course of dysentery caused by this pathogen. Exotoxin can also be produced by other species of Shigella. The RF permeability factor has been discovered, which causes damage to blood vessels. Pathogenicity factors also include invasive protein, facilitating their penetration into epithelial cells, as well as pili and outer membrane proteins responsible for adhesion, and a microcapsule.

Dysentery.Resistance.
Shigella has low resistance to various factors. S. sonnei, which in tap water last up to 2.5 months; in open water they survive up to 1.5 months. S. sonnei can not only survive for quite a long time, but also reproduce in products, especially dairy products.

Dysentery.Epidemiology.
Dysentery is an anthroponotic infection: the source is sick people and carriers. The mechanism of transmission of infections is fecal-oral. The routes of transmission can be different - with Sonne's dysentery the food route predominates, with Flexner's dysentery - water, for Grigoriev-Shiga dysentery the contact and household route is typical.

Dysentery found in many countries of the world. IN last years There has been a sharp rise in the incidence of this infection. People of all ages are affected, but children aged 1 to 3 years are most susceptible to dysentery. The number of patients increases in July - September. Different types of Shigella are distributed unevenly in individual regions.

Dysentery.Pathogenesis.
Shigella enters the gastrointestinal tract through the mouth and reaches the colon. Possessing tropism for its epithelium, pathogens attach to cells with the help of pili and proteins of the outer membrane. Thanks to the invasive factor, they penetrate inside the cells, multiply there, as a result of which the cells die.

Ulcerations form in the intestinal wall, in place of which scars then form. Endotoxin, released when bacteria are destroyed, causes general intoxication, increased intestinal motility, and diarrhea. Blood from the resulting ulcers enters the feces. As a result of the action of the exotoxin, a more pronounced disturbance of water-salt metabolism, central nervous system activity, and kidney damage is observed.

Dysentery.Clinical picture.
The incubation period lasts from 1 to 5 days. The disease begins acutely with an increase in body temperature to 38-39 ° C, abdominal pain and diarrhea appear. There is an admixture of blood and mucus in the stool. Grigoriev-Shiga dysentery is the most severe.

Dysentery.Immunity.
After an illness, immunity is species-specific and variant-specific. It is short-lived and fragile. Often the disease becomes chronic. Repeated diseases have been observed even within one season.

Dysentery.Laboratory diagnostics.
The patient’s feces are taken as the test material. The basis of diagnosis is the bacteriological method, which allows one to identify the pathogen, determine its sensitivity to antibiotics, and carry out intraspecific identification (determine the biochemical variant, serovar or colicinogenovar). In case of prolonged course of dysentery, it can be used as an auxiliary serological method, which consists of diagnosing RA, RNGA (the diagnosis can be confirmed by the increase in antibody titer when the reaction is repeated).

Dysentery.Treatment.
Patients with severe forms of Grigoriev-Shish and Flexner dysentery are treated with antibiotics wide range actions with mandatory consideration of the antibiogram, since among Shigella there are often not only antibiotic-resistant, but also antibiotic-dependent forms. For mild forms of dysentery, antibiotics are not used, since their use leads to dysbacteriosis, which makes the disease worse. pathological process, and disruption of regenerative processes in the mucous membrane of the colon.

Dysentery.Prevention.
The only drug that can be used in foci of infection for prophylactic purposes is dysentery bacteriophage. Non-specific prevention plays the main role.

Nonspecific prevention involves proper sanitary and hygienic arrangement of people's lives, supplying them with quality water and food.

In the patient's environment, measures must be taken to prevent the spread of the pathogen.

Microbiology of dysentery

Dysentery is an infectious disease characterized by general intoxication of the body, diarrhea and a peculiar lesion of the mucous membrane of the large intestine. It is one of the most common acute intestinal diseases in the world. The disease has been known since ancient times under the name “bloody diarrhea”, but its nature turned out to be different. In 1875, the Russian scientist F.A. Lesh isolated an amoeba from a patient with bloody diarrhea Entamoeba histolytica, in the next 15 years, the independence of this disease was established, for which the name amoebiasis was retained.

The causative agents of dysentery proper are a large group of biologically similar bacteria, united in the genus Shigella. The pathogen was first discovered in 1888 by A. Chantemes and F. Vidal; in 1891 it was described by A.V. Grigoriev, and in 1898 K. Shiga, using serum obtained from a patient, identified the pathogen in 34 patients with dysentery, finally proving the etiological role of this bacterium. However, in subsequent years, other causative agents of dysentery were discovered: in 1900 - by S. Flexner, in 1915 - by K. Sonne, in 1917 - by K. Stutzer and K. Schmitz, in 1932 - by J. Boyd , in 1934 - D. Large, in 1943 - A. Sax. Currently genus Shigella includes more than 40 serotypes. All of them are short, non-motile gram-negative rods that do not form spores or capsules, which grow well on regular nutrient media and do not grow on starvation media with citrate or malonate as the sole carbon source; do not form H 2 S, do not have urease; the Voges–Proskauer reaction is negative; glucose and some other carbohydrates are fermented to form acid without gas (except for some biotypes Shigella flexneri: S. manchester And S. newcastle); As a rule, they do not ferment lactose (with the exception of Shigella Sonne), adonitol, salicin and inositol, do not liquefy gelatin, usually form catalase, and do not have lysine decarboxylase and phenylalanine deaminase. The content of G + C in DNA is 49 – 53 mol%. Shigella are facultative anaerobes, the optimum temperature for growth is 37 °C, they do not grow at temperatures above 45 °C, the optimal pH of the environment is 6.7 - 7.2. Colonies on dense media are round, convex, translucent; in case of dissociation, rough R-form colonies are formed. Growth on MPB in the form of uniform turbidity, rough forms form a sediment. Freshly isolated cultures of Shigella Sonne usually form colonies of two types: small round convex (phase I), large flat (phase II). The nature of the colony depends on the presence (phase I) or absence (phase II) of a plasmid with a molecular weight of 120 MD, which also determines the virulence of Shigella Sonne.

The international classification of Shigella is based on their biochemical characteristics (mannitol-non-fermenting, mannitol-fermenting, slowly lactose-fermenting Shigella) and features of the antigenic structure (Table 37).

In Shigella, O-antigens of different specificity were found: common to the family Enterobacteriaceae, generic, species, group and type-specific, as well as K-antigens; They do not have N-antigens.

Table 37

Classification of bacteria genus Shigella

The classification takes into account only group and type-specific O-antigens. In accordance with these characteristics, the genus Shigella is divided into 4 subgroups, or 4 species, and includes 44 serotypes. In subgroup A (type Shigella dysenteriae) included Shigella, which does not ferment mannitol. The species includes 12 serotypes (1 – 12). Each serotype has its own specific type antigen; antigenic connections between serotypes, as well as with other Shigella species, are weakly expressed. To subgroup B (type Shigella flexneri) include Shigella, which usually ferments mannitol. Shigella of this species are serologically related to each other: they contain type-specific antigens (I – VI), by which they are divided into serotypes (1 – 6), and group antigens, which are found in different compositions in each serotype and by which the serotypes are divided into subserotypes. In addition, this species includes two antigenic variants - X and Y, which do not have typical antigens; they differ in sets of group antigens. Serotype S. flexneri 6 has no subserotypes, but it is divided into 3 biochemical types according to the characteristics of the fermentation of glucose, mannitol and dulcitol (Table 38).

Table 38

Biotypes S. flexneri 6

Note. K – fermentation with the formation of only acid; CG – fermentation with the formation of acid and gas; (–) – no fermentation.

Lipopolysaccharide antigen O in all Shigella Flexner contains group antigen 3, 4 as the main primary structure, its synthesis is controlled by a chromosomal gene localized near the his-locus. Type-specific antigens I, II, IV, V and group antigens 6, 7, 8 are the result of modification of antigens 3, 4 (glycosylation or acetylation) and are determined by the genes of the corresponding converting prophages, the site of integration of which is located in the lac - pro region of the Shigella chromosome.

Appeared in the country in the 80s. XX century and a new subserotype that has become widespread S. flexneri 4(IV:7, 8) differs from subserotype 4a (IV:3, 4) and 4b (IV:3, 4, 6), arose from a variant S. flexneri Y(IV:3, 4) due to lysogenization by its converting prophages IV and 7, 8.

To subgroup C (type Shigella boydii) include Shigella, which usually ferments mannitol. Members of the group are serologically different from each other. Antigenic connections within the species are weakly expressed. The species includes 18 serotypes (1 – 18), each of which has its own main type antigen.

In subgroup D (type Shigella sonnei) included Shigella, which usually ferment mannitol and are capable of slowly (after 24 hours of incubation and later) fermenting lactose and sucrose. View S. sonnei includes one serotype, but colonies of phases I and II have their own type-specific antigens. For the intraspecific classification of Shigella Sonne, two methods have been proposed:

1) dividing them into 14 biochemical types and subtypes according to their ability to ferment maltose, rhamnose and xylose; 2) division into phage types according to sensitivity to a set of corresponding phages.

These typing methods have mainly epidemiological significance. In addition, Shigella Sonne and Shigella Flexner are typed for the same purpose based on their ability to synthesize specific colicins (colicinogenotyping) and sensitivity to known colicins (colicinotyping). To determine the type of colicins produced by Shigella, J. Abbott and R. Chenon proposed sets of standard and indicator strains of Shigella, and to determine the sensitivity of Shigella to known types of colicins, a set of standard colicinogenic strains of P. Frederick is used.

Resistance. Shigella has a fairly high resistance to environmental factors. They survive on cotton fabric and paper for up to 30 - 36 days, in dried feces - up to 4 - 5 months, in soil - up to 3 - 4 months, in water - from 0.5 to 3 months, on fruits and in vegetables – up to 2 weeks, in milk and dairy products – up to several weeks; at a temperature of 60 °C they die in 15 – 20 minutes. Sensitive to chloramine solutions, active chlorine and other disinfectants.

Pathogenicity factors. The most important biological property of Shigella, which determines their pathogenicity, is the ability to invade epithelial cells, multiply in them and cause their death. This effect can be detected using a keratoconjunctival test (introduction of one loop of a Shigella culture (2–3 billion bacteria) under the lower eyelid of a guinea pig causes the development of serous-purulent keratoconjunctivitis), as well as by infection of cell cultures (cytotoxic effect) or chicken embryos (their death) , or intranasally in white mice (development of pneumonia). The main pathogenicity factors of Shigella can be divided into three groups:

1) factors determining interaction with the epithelium of the mucous membrane;

2) factors that ensure resistance to humoral and cellular defense mechanisms of the macroorganism and the ability of Shigella to reproduce in its cells;

3) the ability to produce toxins and toxic products that determine the development of the pathological process itself.

The first group includes adhesion and colonization factors: their role is played by pili, outer membrane proteins and LPS. Adhesion and colonization are promoted by enzymes that destroy mucus - neuraminidase, hyaluronidase, mucinase. The second group includes invasion factors that promote the penetration of Shigella into enterocytes and their reproduction in them and in macrophages with the simultaneous manifestation of a cytotoxic and (or) enterotoxic effect. These properties are controlled by the genes of a plasmid with a molecular weight of 140 MD (it encodes the synthesis of outer membrane proteins that cause invasion) and the chromosomal genes of Shigella: kcp A (causes keratoconjunctivitis), cyt (responsible for cell destruction), as well as other genes not yet identified. Protection of Shigella from phagocytosis is provided by the surface K-antigen, antigens 3, 4 and lipopolysaccharide. In addition, lipid A of Shigella endotoxin has an immunosuppressive effect: it suppresses the activity of immune memory cells.

The third group of pathogenicity factors includes endotoxin and two types of exotoxins found in Shigella - Shiga and Shiga-like exotoxins (SLT-I and SLT-II), the cytotoxic properties of which are most pronounced in S. dysenteriae 1. Shiga and Shiga-like toxins have also been found in other serotypes S. dysenteriae, they are also formed S. flexneri, S. sonnei, S. boydii, EHEC and some salmonella. The synthesis of these toxins is controlled by the tox genes of converting phages. Type LT enterotoxins are found in Shigella Flexner, Sonne and Boyd. Their LT synthesis is controlled by plasmid genes. Enterotoxin stimulates the activity of adenylate cyclase and is responsible for the development of diarrhea. Shiga toxin, or neurotoxin, does not react with the adenylate cyclase system, but has a direct cytotoxic effect. Shiga and Shiga-like toxins (SLT-I and SLT-II) have a molecular weight of 70 kDa and consist of subunits A and B (the latter of 5 identical small subunits). The receptor for toxins is a glycolipid of the cell membrane.

The virulence of Shigella Sonne also depends on a plasmid with a molecular weight of 120 MD. It controls the synthesis of about 40 outer membrane polypeptides, seven of them are associated with virulence. Shigella Sonne, having this plasmid, form phase I colonies and are virulent. Cultures that have lost the plasmid form phase II colonies and lack virulence. Plasmids with a molecular weight of 120–140 MD were found in Shigella Flexner and Boyd. Shigella lipopolysaccharide is a strong endotoxin.

Features of epidemiology. The source of infection is only humans. No animals in nature suffer from dysentery. Under experimental conditions, dysentery can only be reproduced in monkeys. The method of infection is fecal-oral. Routes of transmission: water (predominant for Shigella Flexner), food, especially milk and dairy products (predominant route of infection for Shigella Sonne), and household contact, especially for the species S. dysenteriae.

A feature of the epidemiology of dysentery is a change in the species composition of pathogens, as well as Sonne biotypes and Flexner serotypes in certain regions. For example, until the end of the 30s. XX century to a share S. dysenteriae 1 accounted for up to 30–40% of all cases of dysentery, and then this serotype began to occur less and less often and almost disappeared. However, in the 1960s - 1980s. S. dysenteriae reappeared on the historical arena and caused a series of epidemics that led to the formation of three hyperendemic foci of it - in Central America, Central Africa and South Asia (India, Pakistan, Bangladesh and other countries). The reasons for the change in the species composition of dysentery pathogens are probably associated with changes in collective immunity and changes in the properties of dysentery bacteria. In particular, the return S. dysenteriae 1 and its widespread distribution, which caused the formation of hyperendemic foci of dysentery, is associated with its acquisition of plasmids that caused multidrug resistance and increased virulence.

Features of pathogenesis and clinic. The incubation period for dysentery is 2–5 days, sometimes less than a day. The formation of an infectious focus in the mucous membrane of the descending part of the large intestine (sigmoid and rectum), where the dysentery pathogen penetrates, is cyclical in nature: adhesion, colonization, introduction of Shigella into the cytoplasm of enterocytes, their intracellular reproduction, destruction and rejection of epithelial cells, release of pathogens into the lumen intestines; after this, the next cycle begins - adhesion, colonization, etc. The intensity of the cycles depends on the concentration of pathogens in the parietal layer of the mucous membrane. As a result of repeated cycles, the inflammatory focus grows, the resulting ulcers, connecting, increase the exposure of the intestinal wall, as a result of which blood, mucopurulent lumps, and polymorphonuclear leukocytes appear in the feces. Cytotoxins (SLT-I and SLT-II) cause cell destruction, enterotoxin – diarrhea, endotoxins – general intoxication. The clinical picture of dysentery is largely determined by what type of exotoxin is produced to a greater extent by the pathogen, the degree of its allergenic effect and the immune status of the body. However, many questions of the pathogenesis of dysentery remain unclear, in particular: the features of the course of dysentery in children of the first two years of life, the reasons for the transition of acute dysentery to chronic, the significance of sensitization, the mechanism of local immunity of the intestinal mucosa, etc. The most typical clinical manifestations of dysentery are diarrhea, frequent urge: in severe cases, up to 50 or more times a day, tenesmus (painful spasms of the rectum) and general intoxication. The nature of the stool is determined by the degree of damage to the large intestine. The most severe dysentery is caused by S. dysenteriae 1, most easily - Sonne dysentery.

Post-infectious immunity. As observations of monkeys have shown, after suffering from dysentery, strong and fairly long-lasting immunity remains. It is caused by antimicrobial antibodies, antitoxins, increased activity of macrophages and T-lymphocytes. Local immunity of the intestinal mucosa, mediated by IgAs, plays a significant role. However, immunity is type-specific; strong cross-immunity does not occur.

Laboratory diagnostics. The main method is bacteriological. The material for research is feces. Pathogen isolation scheme: inoculation on differential diagnostic media Endo and Ploskirev (in parallel on enrichment medium followed by inoculation on Endo and Ploskirev media) to isolate isolated colonies, obtaining a pure culture, studying its biochemical properties and, taking into account the latter, identification using polyvalent and monovalent diagnostic agglutinating sera. The following commercial serums are produced.

1. To Shigella, which does not ferment mannitol:

To S. dysenteriae 1 And 2

To S. dysenteriae 3 – 7(polyvalent and monovalent),

To S. dysenteriae 8 – 12(polyvalent and monovalent).

2. To Shigella fermenting mannitol:

to typical antigens S. flexneri I, II, III, IV, V, VI,

to group antigens S. flexneri 3, 4, 6, 7, 8– polyvalent,

to antigens S. boydii 1 – 18(polyvalent and monovalent), to antigens S. sonnei I phase, II phase,

to antigens S. flexneri I–VI+ S. sonnei– polyvalent.

To quickly identify Shigella, the following method is recommended: a suspicious colony (lactose-negative on Endo medium) is subcultured on TSI medium (English. triple sugar iron) – three-sugar agar (glucose, lactose, sucrose) with iron to determine H2S production; or to a medium containing glucose, lactose, sucrose, iron and urea. Any organism that breaks down urea after 4 to 6 hours of incubation is most likely a member of the genus Proteus and may be excluded. A microorganism that produces H2S or has a urease or acid-forming joint (fermentes lactose or sucrose) can be excluded, although H2S-producing strains should be investigated as possible members of the genus Salmonella. In all other cases, the culture grown on these media should be examined and, if fermenting glucose (colour change), isolated in pure form. At the same time, it can be studied in a glass agglutination reaction with appropriate antisera to the genus Shigella. If necessary, other biochemical tests are performed to check genus membership. Shigella, and also study mobility.

To detect antigens in the blood (including as part of the CEC), urine and feces, the following methods can be used: RPGA, RSK, coagglutination reaction (in urine and feces), IFM, RAGA (in blood serum). These methods are highly effective, specific and suitable for early diagnosis.

For serological diagnosis, the following can be used: RPHA with the corresponding erythrocyte diagnostics, immunofluorescence method (indirect modification), Coombs method (determining the titer of incomplete antibodies). An allergy test with dysenterine (a solution of protein fractions of Shigella Flexner and Sonne) is also of diagnostic value. The reaction is taken into account after 24 hours. It is considered positive in the presence of hyperemia and infiltrate with a diameter of 10–20 mm.

Treatment. The main attention is paid to restoring normal water-salt metabolism, rational nutrition, detoxification, rational antibiotic therapy (taking into account the sensitivity of the pathogen to antibiotics). A good effect is achieved by early use of a polyvalent dysentery bacteriophage, especially tablets with a pectin coating, which protects the phage from the action of gastric juice HCl; In the small intestine, pectin dissolves, phages are released and exert their effect. For preventive purposes, the phage should be given at least once every three days (the period of its survival in the intestine).

The problem of specific prevention. To create artificial immunity against dysentery, various vaccines were used: from killed bacteria, chemical, alcohol, but all of them turned out to be ineffective and were discontinued. Vaccines against Flexner's dysentery have been created from live (mutant, streptomycin-dependent) Shigella Flexner; ribosomal vaccines, but they also have not found widespread use. Therefore, the problem of specific prevention of dysentery remains unresolved. The main way to combat dysentery is to improve the water supply and sewerage system, ensure strict sanitary and hygienic regimes in food enterprises, especially the dairy industry, in child care institutions, public places and in maintaining personal hygiene.