Biochemical and serological typing of pathogens of food toxic infections. Variegated series microbiology Biochemical properties of salmonella
Magnesium medium Rappaport-Vassiliadis, 500 g/pack, Cat. No. 107700
- SalmonellaenrichmentbrothRAPPAPORT- Rappaport's magnesium medium, 500 g/pack, Cat. No. 110236
- Tetrathionate enrichment broth Muller-Kauffman- Tetrathionate broth, 500 g/pack, Cat. No. 110863
- Muller-
KauffmannTetrathionate-
NovobiocinBroth (MKTTn)
- Tetrathionate broth with novobiocin, 500 g/pack, Cat. No. 105878
- BismuthsulfiteagarWILSON-
BLAIR- Bismuth-sulfite agar, 500 g/pack, Cat. No. 105418
- Hektoenentericagar- Hectoene agar, 500 g/pack, Cat. No. 111681
- SS-
agar- Salmonella-Shigella agar, 500 g/pack, Cat. No. 107667
- BPLS (Brilliant-
greenPhenol-
redLactoseSucroseAgar)
- Lactose sucrose agar with brilliant green and phenol red, 500 g/pack, Cat. No. 107232
- XLD-agar- Xylose-lysine-deoxycholate agar for the isolation and differentiation of pathogenic enterobacteria, 500 g/pack, Cat. No. 105287
- XLT4 agar- Xylose-lysine with tergitol 4 (completely inhibits the growth of Proteus), 500 g/pack, Cat. No. 113919
- XLT4 Supplement- Selective additive to XLT4 agar medium, 100 ml/pack, Cat. No. 108981
- Rambach agar- Chromogenic Rambach agar for salmonella, 250 definitions, 4 bottles x 250 ml, Cat. No. 107500
- MSRV (Modified semi-solid Rappaport-Vassiliadis)- Semi-liquid medium for the accelerated isolation of salmonella from products and raw materials, 500 g/pack, Cat. No. 109878
- MSRV Selective Supplement- Selective additive (novobiocin, 10 mg) to the MSRV medium, 10 bottles/pack, Cat. No. 109874
- Salmosyst broth base- Non-selective enrichment, 500 g/pack, Cat. No. 110153
- Salmosyst selective tablets- Selective enrichment of Salmonella - tetrathionate medium in tablets, 250 tablets/pack, Cat. No. 110141
- KLIGLER agar- Kligler agar, 500 g/l, Cat. No. 103913
- Triple sugar iron agar- Three-sugar agar with iron, 500 g/pack, Cat. No. 103915
- Lysine iron agar- Lysine agar with iron for differentiation and identification of enterobacteria; determination of lysine decarboxylase and hydrogen sulfide, 500 g/pack, Cat. No. 111640
- SIMMONS citrate agar- Simmons Citrate Agar for identification, 500 g/l, Cat. No. 102501
- Singlepath Salmonella- Express test (20 min) for salmonella, 25 tests/pack, Cat. No. 104140
Salmonellaspp.)
The genus Salmonella belongs to the family Enterobacteriaceae. The bacteria are rod-shaped, gram-negative, asporogenic, predominantly motile. Globally, Salmonella is one of the most common foodborne pathogens, infecting most types of raw foods (eg, meat, eggs, plant products). The bacteria's resistance to desiccation, coupled with their high heat resistance, poses a problem in protecting most dry and semi-dry foods from the pathogen. According to the food legislation of most countries, salmonella is not allowed in 25 g of food product. To obtain a simple answer (yes/no) about the presence of salmonella in food and feed using traditional microbiological methods, a total of up to 5 days must be spent. For products that need to be sold quickly, this means a significant delay. To meet the requirements of food manufacturers for the rapid sale of finished products and reduce storage costs, it is necessary to use innovative, accelerated methods for detecting salmonella. Thus, accelerated methods for the analysis of Salmonella are of increasing interest. Rapid tests offered for these purposes must be specific, sensitive, convenient and cost-effective. Depending on the number of samples studied and the requirements for the specificity of the method, the choice can be made either in favor of molecular analysis of bacterial DNA by PCR (usually for a high-throughput laboratory) or in favor of immunodiagnostic tests (when analyzing a smaller number of samples).
Salmonellosis
Most people infected with Salmonella develop diarrhea, fever, and abdominal pain 12 to 72 hours after infection. The illness usually lasts 4-7 days, and most patients recover without treatment. However, in some cases the illness may be so severe that hospitalization is required. In such patients, Salmonella infection can spread from the intestines to the blood and other parts of the body. If the patient is not treated with antibiotics in a timely manner, this can lead to death. Older adults, infants, and those with compromised immune systems will be affected first by the most severe form of the disease.
Pathogenicity
Salmonella is one of the most dangerous pathogens of intestinal infections in humans and farm animals. According to WHO, up to 1.3 billion cases of salmonellosis are registered every year in the world, while the dynamics of the disease in the population tends to increase. With absence effective treatment fatal cases in humans range from 1-3 to 10-15%. Every year in the United States, the disease is registered in 1.4 million people, and the material costs associated with the consequences and prevention of the disease are estimated at $1-2.3 billion. In Russia, this disease ranks second in the structure of acute intestinal infections.
Taxonomy
Genus Salmonella, numbering over 2,500 serovars (serotypes), is part of the Enterobacteriaceae (enterobacteria) family. According to the modern classification based on DNA analysis, the genus Salmonella includes the pathogenic species S. enterica for humans and warm-blooded animals. This species is divided into 6 subspecies. Subspecies S. enterica subsp. Enteritidis is a causative agent of foodborne diseases. Subspecies S. enterica subsp. Typhi, S. enterica subsp. Paratyphi A, B are the causative agents of typhoid fever, paratyphoid A and B in humans.
Morphology
Salmonella cells are motile (thanks to flagella), asporogenic, gram-negative straight rods (0.5-1x1-3 µm) with rounded ends. There are also non-motile individuals and strains. They do not form a capsule; they are facultative anaerobic chemoorganotrophs with oxidative and fermentative metabolism. They grow well on simple nutrient and bile-containing media. On solid media they can form R-form (rough) and S-form (smooth) colonies; on liquid media they give diffuse turbidity. Colonies in S-shape are medium in size, shiny, translucent, with a bluish tint. When inoculating blood, the best liquid enrichment medium is bile broth; when inoculating biomaterials (feces, bile, urine) containing additional flora, selenite broth. On lactose-containing differential media, bacteria form colorless colonies, on bismuth-sulfite agar - black colonies.
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Biochemical characteristics
Salmonella have pronounced biochemical activity characteristic of the genus. To identify them, it is important to take into account the following biochemical properties:
1) fermentation of glucose and other carbohydrates (mannitol, maltose) to acid and gas (subspecies S.t.yphi releases only acid)
2) absence of fermentation of lactose, sucrose, salicin and urea,
3) react with methyl rot, produce hydrogen sulfide, do not form indole (as a rule) - oxidase is negative, catalase is positive, the Voges-Proskauer reaction is negative,
4) temperature optimum for growth is 35-37 o C, growth completely stops at 5 o C; optimum pH=7.2-7.4.
According to the serological classification, the vast majority of Salmonella serovars (serotypes) pathogenic for humans belong to groups A, B, C, D and E. Salmonella are typed using the Kauffman-White agglutination reaction. For its production, hyperimmune sera or monoclonal antibodies to Salmonella are used. The diagnosis of salmonellosis and epidemiological analysis of pathogens are based on serotyping.
Sources and factors of transmission of infection
Salmonellosis (typhoid fever, paratyphoid fever, gastroenteritis, septicemia, etc.) are widespread foodborne diseases of animals and humans with a fecal-oral transmission mechanism. In humans, foodborne toxic infections are accompanied by damage gastrointestinal tract and dehydration of the body. The infecting dose is from 1000 to 10 thousand cells. The permanent habitat is the intestines of humans and warm-blooded animals, which are a reservoir of infection. Contaminated food products and raw materials, as well as water, are the main sources and factors of transmission of the pathogen. The pathogen enters food products from contaminated raw materials. Soil is involved in the contact route of infection transmission. The presence of salmonella in soil or water always indicates contamination of these environments with the feces of infected people and/or animals - birds, cattle, pigs, cats, dogs, pigeons. Their infection with salmonella ranges from 6-7 to 80%.
Main sources of infection
In the United States, salmonellosis accounts for about 9% of foodborne infections, and a significant part of the population of this country is its asymptomatic bacteria carriers. Contaminated poultry products, meat and meat products, milk, cheese, butter, vegetables and fruits, semi-finished products and seasonings (mayonnaise, egg powder, creams, etc.) are the main sources of salmonellosis. Food products are also contaminated by bacteria during cooking, contact with carriers, production equipment, and animal carriers (flies, mouse-like rodents, pets). The duration of viability of Salmonella depends on the type of product and environmental conditions. Thus, bacteria survive on the surface of vegetables and fruits for 5-10 days, in milk - up to 20 days, in beer and kefir - up to 2 months, in sausages, meat (including salted), butter- from 2 to 6 months, in cheeses - up to 1 year, in frozen meat - up to 2-3 years.
Pathogenesis
The different nature of the disease is explained by the multiplicity of pathogenicity factors (endo-, exotoxins, etc.), which have not yet been sufficiently studied. All virulent Salmonella produce endotoxin, which induces the development of fever in infected people (with an increase in temperature to 39-40 o). After oral infection, once in the small intestine, salmonella invade the intestinal mucosa and, multiplying in macrophages, form the primary focus of infection. At the end of the incubation period (10-14 days after infection), Salmonella enters the bloodstream and causes bacteremia. The causative agents of typhoid and paratyphoid fever spread throughout the body through the bloodstream, settling in the cells of the liver, spleen, lungs, bone marrow, as well as the gallbladder. By the end of the 2nd week from the onset of the disease, the pathogen is released from the patient’s body in urine, feces, breast milk, and saliva. Immunity to the disease is not formed.
Detection methods
An important role is played by specific prevention of salmonellosis, which consists in carrying out veterinary, sanitary, sanitary and hygienic and anti-epidemic measures. Prevention is accompanied by permanent control of the pathogen in food, fodder, raw materials and water.
Classic method
Media for isolation of Salmonella
- Buffered peptone water
- Rappaport-Vassiliadis environment
- Selenite environment
- Tetrathionate broth
- Xylose-lysine-deoxycholate agar
- Diamond Green Agar
- Bismuth sulfite agar
Classical method for studying salmonella using nutrient media. However, due to the length of the analysis procedure, one classical method is not enough to determine the pathogen.
Immunochromatographic expresstests
To speed up the detection of salmonella, significantly reduce labor costs and save resources, in recent decades, accelerated methods for identifying the pathogen have been developed, tested and widely used abroad. Accelerated methods can significantly (by 24-48 hours) reduce the duration of research. Possessing high sensitivity, they provide reliable detection of Salmonella in the analyzed material.
Normative documents
- MP 24 FC 976 Methods for identifying pathogenic microorganisms using immunochromatographic rapid tests produced by Merck (Germany)
- GOST R 50455-92. Meat and meat products. Salmonella detection (arbitration method)
- GOST R 52814-2007 (ISO 6579:2002). Food products. Method for identifying bacteria of the genus Salmonella.
- GOST R 53665-2009. Poultry meat, offal and semi-finished poultry meat products. Salmonella detection method;
- SP 3.1.7.2616-10. Sanitary and epidemiological rules. Prevention of salmonellosis. M.: Rospotrebnadzor. 2010.18 p.
The textbook consists of seven parts. Part one – “General Microbiology” – contains information about the morphology and physiology of bacteria. Part two is devoted to the genetics of bacteria. Part three – “Microflora of the biosphere” – examines the microflora environment, its role in the cycle of substances in nature, as well as human microflora and its significance. Part four – “The Study of Infection” – is devoted to the pathogenic properties of microorganisms, their role in the infectious process, and also contains information about antibiotics and their mechanisms of action. Part five – “The Doctrine of Immunity” – contains modern ideas about immunity. The sixth part – “Viruses and the diseases they cause” – provides information about the basic biological properties of viruses and the diseases they cause. Part seven - “Private medical microbiology” - contains information about the morphology, physiology, pathogenic properties of pathogens of many infectious diseases, as well as modern methods their diagnosis, specific prevention and therapy.
The textbook is intended for students, graduate students and teachers of higher medical educational institutions, universities, microbiologists of all specialties and practicing doctors.
5th edition, revised and expanded
Specific prevention not used, although various vaccines from killed and live (mutant) strains have been proposed S. typhimurium.
This genus of the Enterobacteriaceae family includes more than 2,000 different bacteria that cause diseases in humans and animals. These diseases are called salmonellosis. Salmonella are similar in morphological, cultural and enzymatic properties, but differ in antigenic structure.
Salmonella is divided into monopathogenic and polypathogenic. The first include the pathogens of typhoid fever, paratyphoid A and paratyphoid B. Only humans suffer from these diseases. The second group includes pathogens that infect humans and animals.
S. typhi was first discovered by Ebert (1880) in the organs of a person who died of typhoid fever. Ashar and Bansod (1886), in diseases similar to typhoid fever, isolated bacteria from the pus and urine of patients that differed in biochemical and serological properties from the causative agents of typhoid fever. They were named S. paratyphi A and S. paratyphi B. Almost simultaneously, the American scientist D. Salmon (1885) first described the causative agents of swine cholera (S.choleraesuis). Subsequently, many similar bacteria were described, united in the genus Salmonella, named after the scientist who described them.
Morphology. All Salmonella are small, 1.0-3.0 × 0.6-0.8 µm rods with rounded ends. Gram negative. Motile, peritrichous. They do not form spores or capsules.
Cultivation. Salmonella are facultative anaerobes. They are not demanding on nutrient media. They grow well on MPA and MPB at 37°C (from 20 to 40°C) and pH 7.2-7.4 (from 5.0 to 8.0). On MPA they form delicate, translucent, slightly convex, shiny colonies; on MPB they form uniform turbidity.
During the initial culture of material from patients (feces, urine, vomit, blood, bile), slow growth of Salmonella is often noted. To accumulate them, they are inoculated on enrichment media: selenite broth, Müller’s medium, Kaufman’s medium. Elective (selective) media are also used: bile (10-20%) and Rappoport medium.
On differential diagnostic media Endo, EMS, Ploskirev, salmonella grow in the form of colorless colonies, since they do not break down lactose included in the medium. On bismuth-sulfite agar after 48 hours they form black colonies that leave a mark after they are removed with a loop (except for Salmonella paratyphoid A).
In freshly isolated cultures of S. paratyphi B, after incubation in a thermostat for 18-20 hours and keeping at room temperature for 1-2 days, a mucous shaft is formed at the periphery of the colony.
Enzymatic properties. Salmonella break down glucose, mannitol, and maltose to form acid and gas. An exception is the causative agent of typhoid fever (S. typhi), which breaks down these sugars only into acid. Salmonella does not ferment lactose and sucrose. Proteolytic properties: most salmonella break down protein media with the formation of hydrogen sulfide (the causative agents of paratyphoid A are distinguished by the absence of this property). Indole is not formed. Gelatin is not liquefied.
Toxigenicity. Salmonella contains endotoxin - a lipopolysaccharide-protein complex.
Antigenic structure and classification. At the beginning of the 20th century, scientists noticed the different nature of Salmonella antigens. Kaufman (1934), based on the results of an agglutination reaction of different Salmonella with a set of sera, divided all Salmonella into groups and types and proposed a diagnostic scheme for their antigenic structure. In accordance with this scheme, Salmonella are currently identified.
Salmonella contains two antigenic complexes: O and H, O-antigen - lipopolysaccharide-protein complex; it is heat stable and inactivated by formaldehyde. The H-antigen is associated with flagella and is of a protein nature; it is thermolabile, inactivated by alcohol and phenol, but resistant to formaldehyde.
All Salmonella are divided into O-groups, each of which is characterized by the presence of certain O-antigens: the main one, indicated by an Arabic numeral (2, 4, 7, 8, 9, etc.), and additional ones, common to several O-groups ( 1, 12). Currently, more than 60 O-groups are known, designated by capital letters of the Latin alphabet (A, B, C, D, E, etc.).
S. typhi also contains a Vi antigen, which is located more superficially in the microbial cell than the O antigen and prevents agglutination of the culture with O serum. This antigen is heat labile. Its presence was associated with the virulence of the pathogen. The Vi antigen is also contained in S. paratyphi C cells.
Salmonella H-antigens have two phases. Salmonella of different serovars of the same O-group have a different first phase of the H-antigen, which is designated by lowercase letters of the Latin alphabet: a, b, c, d, eh ... u, z, etc. The second phase of the H-antigen is usually designated by Arabic letters numbers: 1, 2, 5, 6, 7 and lowercase Latin letters. The combination of various O- and H-antigens determines the antigenic structure of cultures and their name.
In practical work, to determine the antigenic structure of Salmonella, adsorbed monoreceptor agglutinating sera are used, which contain antibodies to one antigen. An agglutination reaction is performed on glass and, based on the presence of agglutination with certain sera, the antigenic structure of the isolated culture is characterized. For example, a culture is agglutinated by O-sera “9” and “12” and H-serum “d”; find a serovar with the same antigenic composition (S. typhi) in the scheme, perform an additional reaction with Vi-serum
There are sets of specific Salmonella phages that lyse only Salmonella of the corresponding phage. To determine the phage product of S. typhi cultures containing the Vi antigen, 45 phages are produced in our country; for S. paratyphi B-11 phages; S. paratyphi A - 6, etc. These studies are carried out to determine the source and routes of transmission of infection.
Resistance to environmental factors. Salmonella is quite resistant. At a temperature of 100° C they die instantly, at 60-70° C - in 10-15 minutes. They tolerate low temperatures well and can be stored in clean water and ice for several months; in smoked and salted meat - up to 2 months. Resistant to drying, long-lasting in dust.
Under the influence of disinfectants they die within a few minutes (2-5% phenol solution, 1:1000 mercuric solution, 3-10% chloramine solution).
Animal susceptibility. Most Salmonella cause diseases in humans and many species of animals and birds (polypathogenic).
Typhoid fever, paratyphoid A and B
Source of infection. A sick person and a bacteria carrier.
Transmission routes. Infectious agents are transmitted through objects contaminated with human secretions, through hands, water, and food. Pathogens are often carried by flies. Depending on the route of transmission, outbreaks of typhoid fever and paratyphoid fever are distinguished between household, waterborne and foodborne outbreaks.
Pathogenesis. Infection occurs through the mouth. From the oral cavity, microorganisms enter the stomach, where they are partially destroyed under the influence of gastric juice and enzymes. The remaining salmonella enter the small intestine, penetrate the lymphoid tissue of the small intestine (group lymphatic and solitary follicles), in which they multiply during the incubation period (10-14 days). By the end of this period, pathogens enter the lymph and blood (bacteremia) and spread throughout the body. During this period, they are localized in the lymphoid tissue of internal organs, the macrophage system, liver, spleen, and bone marrow. Salmonella accumulates in the gallbladder, where it finds favorable conditions for reproduction, since bile is a good breeding ground for these bacteria. At the same time, they enter the small intestine for the second time and, affecting already sensitized lymphoid tissue (group lymphatic and solitary follicles), cause the formation of specific typhoid ulcers (Fig. 42).
During the period of bacteremia, some microorganisms are destroyed, this releases endotoxin and intoxication phenomena occur: the temperature rises, general malaise, weakness, headache, etc. appear. From the end of the 2nd and beginning of the 3rd week, salmonella begin to be excreted from the body in feces , urine, saliva, etc.
The period of convalescence (recovery) is characterized by cleansing the body of the pathogen, increased phagocytic activity of cells, and accumulation of antibodies in the blood.
However, with typhoid fever and paratyphoid fever, bacterial excretion often does not end with the patient’s recovery - bacterial carriage is formed. Chronic inflammatory phenomena in the gallbladder contribute to the survival of salmonella in the bile and their long-term excretion from the body (sometimes up to several years).
Immunity. Post-infectious immunity is quite intense and long-lasting. Recurrent diseases are rare. During the course of the disease, antibodies are produced: at the end of the 1st week, agglutinins, precipitins and other types of antibodies appear. Their number increases, reaching a maximum on the 14-15th day of illness. Antibodies remain in the blood serum of a person who has been ill for a long time.
The activity of phagocytes and other cellular defense factors are also important in the formation of the immune state of the body.
Prevention. Maintaining personal hygiene and carrying out all sanitary and hygienic measures: supervision of water supplies, control of food products and enterprises Catering.
Specific prevention. A chemical vaccine containing full antigens of the causative agents of typhoid fever, paratyphoid A and B and tetanus toxoid (TAB"te). There is also a typhoid alcohol vaccine enriched with Vi-antigen, the administration of which for prophylactic purposes gives a good effect. In the outbreak of the disease, persons in contact with patients are given typhoid bacteriophage.
Treatment. Antibiotics: chloramphenicol, tetracycline, etc.
Foodborne illnesses
When consuming products contaminated with Salmonella of various serovars (except S. typhi, S. paratyphi A and B), foodborne illnesses occur.
Sources of infection. Animals and birds sick with salmonellosis, or healthy ones, in whose body, without causing harm, there are salmonella.
Transmission routes. Infection occurs through consumption of meat, meat products, eggs, milk, and dairy products infected with salmonella. The most dangerous is the consumption of food in which Salmonella multiply and die and endotoxin accumulates.
Pathogenesis. Once entering the body through the mouth, salmonella penetrates the digestive tract. In this case, a significant part of the bacteria dies and endotoxin is released, which can penetrate into the blood. Symptoms of damage to the gastrointestinal tract and general toxicosis appear. The disease lasts no more than 4-5 days; sometimes those who have recovered become carriers of salmonella.
Immunity short-lived. Various antibodies accumulate in the blood of patients and convalescents: agglutinins, precipitins, etc. There are a lot of Salmonella serovars, and immunity is specific, that is, it is directed only against one pathogen, so a person can get salmonellosis again.
Prevention. Constant strict veterinary and sanitary control over livestock, slaughter and cutting of carcasses, storage and processing of meat and meat products. It is necessary to strictly observe the sanitary regime and personal hygiene in public catering establishments.
Specific prevention. People in areas of foodborne illness should be given Salmonella polyvalent bacteriophage.
Treatment. The main therapeutic agent is detoxification of the body - administration of large amounts of fluid, gastric lavage. Antibiotics are also used.
Hospital-acquired salmonella infection
The causative agent of nosocomial Salmonella infection is most often S. typhimurim. There are also “hospital” outbreaks caused by S. heidelberg, S. derby, etc. Although the morphological and cultural properties of these pathogens do not differ from the properties of other Salmonella, there are some biological features, characteristic of them. For example, pathogens of nosocomial infections belong to certain biovars; they are more pathogenic for white mice, etc.
Sources of infection. More often a bacteria carrier, less often a patient.
Transmission routes. Indirect contact predominates (toys, underwear, patient care items). Less common are airborne dust and food transmission routes.
Pathogenesis. The disease develops against a background of weakening of the body and a decrease in its immune activity. The pathogen enters the body orally or through the respiratory tract, which determines the development of the pathological process: dysfunction of the gastrointestinal tract with dehydration or damage to the respiratory system, bacteremia, septic complications. Young children are the first to get sick.
Immunity. Produced only in relation to one Salmonella serovar.
Prevention. Strict compliance with the sanitary and hygienic regime in medical institutions.
Specific prevention. If a nosocomial Salmonella infection occurs, children who have been in contact with the patient should be given Salmonella polyvalent bacteriophage.
Treatment. Symptomatic.
Control questions
1. What are the morphological, cultural and enzymatic properties of Salmonella?
2. What is the classification of Salmonella based on?
3. What diseases are caused by salmonella?
Microbiological examination
Purpose of the study: isolation of pathogens and determination of Salmonella serovar.
Material for research
2. Bowel movements.
4. Duodenal contents.
Depending on the stage of the disease, different material is examined.
The contents of roseola, bone marrow, sputum and material obtained at autopsy - pieces of organs - can also be examined.
For toxic infections, the material for research can be rinsing waters stomach, vomit, food residues.
Regardless of the nature of the material taken for research, from the moment the pure culture is isolated, the research is carried out according to the general scheme.
Basic research methods
1. Bacteriological (Fig. 43).
2. Serological.
Progress of the study
Second day of the study
Remove the dishes from the thermostat (incubation for 18-24 hours) and examine the grown colonies with the naked eye and using a magnifying glass. Several (5-6) suspicious colonies are isolated on Olkenitsky or Russell medium. Inoculation is carried out as follows: carefully, without touching the edges of the tube, the removed colony is introduced into the condensation liquid, then the entire beveled surface of the medium is inoculated with streaks and an injection is made into the depth of the column to detect gas formation. The injection should be made into the center of the agar column.
Test tubes with cultures are placed in a thermostat. If the material under study was sown on an enrichment medium, then after 18-24 hours it is sown from the enrichment medium onto plates with differential media. Further research is carried out according to the general scheme.
1 (A black mark remains in place of the removed colonies (the color of the medium changes).)
Third day of the study
Remove the test tubes with the cultures from the thermostat and examine the growth pattern.
The composition of the combined media includes lactose, glucose, sometimes urea and an indicator. The breakdown of glucose occurs only under conditions of anaerobiosis. Therefore, the beveled surface of the medium does not change during the breakdown of glucose, and the column is painted in a color corresponding to the indicator. Bacteria that break down lactose and urea change the color of the entire medium.
If the isolated cultures ferment lactose or break down urea, changing the color of the entire medium, then they are not salmonella and a negative answer can be given.
The culture that breaks down only glucose is subjected to further study: smears are made, stained with Gram and microscopically examined. If gram-negative rods are present in smears, their mobility and enzymatic properties are studied.
Motility can be determined in a hanging drop or a crushed drop, and can also be determined by the growth pattern in semi-solid Hiss medium or in 0.2% agar. If there is motility during sowing by injection, growth on the medium is diffuse, the medium becomes cloudy.
To detect enzymatic activity, inoculation is carried out on Hiss media, MPB, and peptone water. Indicator papers are lowered (under the stopper) into test tubes with the latter media to determine indole and hydrogen sulfide. They also do a culture test for litmus milk.
Fourth day of research
Biochemical activity is taken into account based on the result of fermentation of carbohydrate and other media (see Table 33).
Note. k - acid formation; kg - formation of acid and gas; u - alkali; + presence of property; - lack of property.
Having determined the morphological, cultural and enzymatic properties of the isolated culture, it is necessary to analyze the antigenic structure (Table 34).
Serological identification of Salmonella begins with an agglutination reaction on glass with polyvalent O-serum A, B, C, D, E. In the absence of agglutination, the isolated culture is tested with polyvalent O-serum for rare groups of Salmonella. If there is a positive reaction with one of the sera, the culture is tested with each O-serum included in the polyvalent serum to determine the O-serogroup. Having established that a culture belongs to the O-group, its H-antigens are determined with sera of the first and then the second phase (Table 35).
The culture of Salmonella typhus is also tested with Vi-serum. Typhoid fever pathogens containing Vi antigen are tested with Vi phages (there are 86 of them). Determining the phagotype is of great epidemiological importance (see Fig. 43).
Phagotyping technique. 1st method. 20-25 ml of agar is poured into Petri dishes and dried with open lids in a thermostat. The bottom of the cup is divided into sectors. The name of the phage is written on each sector. A 4-6 hour broth culture is studied because it contains more Vi antigen. Apply 8-10 drops of broth culture to the surface of the agar and rub it over the surface of the agar with a glass spatula. The cups with the crops are dried with the lids open in a thermostat. A drop of the corresponding type phage is applied to each sector. After the drops have dried, the cups are placed in a thermostat for 18-24 hours. The result is measured with the naked eye or using a magnifying glass through the bottom of the cup.
The presence of lysis of a culture by one or more typical phages makes it possible to determine whether the isolated strain belongs to a specific phage type.
2nd method. The culture is applied dropwise onto the nutrient medium. After the culture has dried in a thermostat, a drop of a typical phage is applied to each drop. Place it in the thermostat.
The degree of lysis is expressed using the four-cross system.
Control questions
1. What material is examined for typhoid fever, paratyphoid fever and toxic infections?
2. In what period of the disease is the blood culture method used?
3. During what period of the disease with typhoid and paratyphoid fever are feces and urine examined?
4. What differential diagnostic media are used to inoculate the test material?
5. What media are used for the accumulation of salmonella?
6. What is determined by monoreceptor O-sera and what by monoreceptor H-sera?
1. Study from the table. 32 growth pattern of salmonella on differential media. Look at the teacher for plates inoculated with Salmonella typhus on Endo, Ploskirev media, and bismuth-sulfite agar. Draw the colonies with colored pencils and show them to your teacher.
2. Take salmonella culture, O- and H-monoreceptor sera from the teacher. Perform an agglutination reaction on glass. Take into account the reaction and show it to the teacher.
The isolated culture gave a positive agglutination reaction with O-serum 4. With which H-sera should an agglutination reaction be performed if you think that this is a culture of Salmonella paratyphoid B?
Serological diagnosis of typhoid and paratyphoid fever
Vidal reaction. From the second week of the disease, antibodies against the infectious agent accumulate in the blood of patients. To identify them, the patient’s blood serum is examined in an agglutination reaction. Killed Salmonella cultures - diagnosticums - are used as antigens.
To perform the Widal reaction, the patient’s serum, a set of diagnostic kits, and an isotonic sodium chloride solution are used.
Blood (2-3 ml) from the pulp of the finger or the cubital vein is collected in a sterile tube and delivered to the laboratory. In the laboratory, the test tube is placed in a thermostat for 20-30 minutes to form a clot, then a Pasteur pipette is used to circle the clot to separate it from the wall of the test tube, and it is placed in the cold for 30-40 minutes. The separated serum is sucked off and used to perform an agglutination reaction with diagnostics from Salmonella typhus and paratyphoid fever. Blood can be centrifuged to obtain serum.
When an infectious process occurs - typhoid fever or paratyphoid fever - the body produces O- and H-antibodies to the same antigens of the pathogen.
O-antibodies appear first and disappear quite quickly. H-antibodies last a long time. The same thing happens during vaccination, therefore a positive Widal reaction with O- and H-antigens indicates the presence of the disease, and a reaction only with H-antigens can occur in both those who have recovered from the disease (anamnestic reaction) and those vaccinated (vaccine reaction). Based on this, the Widal reaction is performed separately with O- and H-antigens (diagnosticums).
Since typhoid fever and paratyphoid A and B are clinically similar, to identify the nature of the disease, the patient’s serum is tested simultaneously with diagnostics from Salmonella typhus and paratyphoid A and B.
The Widal reaction is widely used because it is simple and does not require special conditions.
The reaction can be carried out in two ways: dropwise and volumetric (see Chapter 12). In practice, the volumetric method is more often used. When performing a linear agglutination reaction, the number of rows must correspond to the number of antigens (diagnosticums). The causative agent of the disease is considered to be a microorganism from which the diagnosticum was agglutinated by the patient’s serum. Sometimes group agglutination is noted, since the causative agents of typhoid and paratyphoid fever have common group antigens. In this case, the result of the reaction in the series in which agglutination is noted in a larger dilution of serum is considered positive (Table 36).
Note. In practice, the Widal reaction is performed with four diagnosticums: typhoid fever “O” and “H”, and paratyphoid fever A and B with diagnosticums “ON”.
If agglutination occurs only in small dilutions of serum - 1:100, 1:200, then to distinguish the reaction during the disease from the vaccination or anamnestic reaction, they resort to repeating the agglutination reaction after 5-7 days. In a patient, the antibody titer increases, but in a vaccinated or recovered person it does not change. Thus, an increase in antibody titer in the blood serum serves as an indicator of the disease.
In response to the introduction of typhoid pathogens possessing the Vi antigen into the body, Vi agglutinins appear in the patient’s blood. They are detected from the 2nd week of illness, but their titer usually does not exceed 1:10. The detection of Vi-antibodies is associated with the presence of typhoid pathogens in the body, therefore the determination of these antibodies is of great epidemiological importance, as it allows the identification of bacteria carriers.
Vi-hemagglutination reaction. This is the most sensitive reaction for detecting antibodies.
The principle of the reaction is that human (group I) or sheep erythrocytes, after special treatment, can adsorb Vi-antigen on their surface and thereby acquire the ability to agglutinate with the corresponding Vi-antibodies.
Red blood cells with antigens adsorbed on the surface are called erythrocyte diagnosticums.
To perform the Vi-hemagglutination reaction, take:
1) the patient’s blood serum (1-2 ml); 2) erythrocyte Salmonella Vi-diagnosticum; H) Vi-serum; 4) O-serum; 5) isotonic sodium chloride solution.
The reaction is carried out in agglutination tubes or in plastic plates with wells.
Blood is taken from the patient in the same way as for the Vidal reaction. Serum is obtained. Two-fold serial dilutions are prepared from the serum, starting from 1:10 to 1:160.
0.5 ml of each dilution is added to the well and 0.25 ml of erythrocyte diagnosticum is added. The reaction is performed in a volume of 0.75 ml.
The controls are: 1) standard agglutinating monoreceptor serum + diagnosticum - the reaction must be positive up to the serum titer; 2) diagnosticum in isotonic sodium chloride solution (control) - the reaction should be negative.
The contents of the wells are thoroughly mixed, placed in a thermostat for 2 hours and left at room temperature until the next day (18-24 hours).
Accounting begins with control. The reaction is assessed depending on the degree of agglutination of the diagnosticum.
The results are taken into account according to the four-cross system:
Red blood cells are completely agglutinated - sediment at the bottom of the well in the form of an “umbrella”;
+++ “umbrella” is smaller, not all red blood cells are agglutinated;
++ the “umbrella” is small, at the bottom of the hole there is a sediment of non-agglutinated erythrocytes;
The reaction is negative; the red blood cells did not agglutinate and settled to the bottom of the well in the form of a button.
Control questions
1. At what period of the disease is the Vidal reaction performed?
2. What ingredients are needed to perform the Widal reaction?
3. What diagnostics are used to perform the Vidal reaction?
4. Which serological reaction is the most sensitive in diagnosing typhoid paratyphoid infections?
5. What diagnosticum is used when performing the Vi-hemagglutination reaction?
6. What serum is used to determine the presence of Vi-antigen in the culture under study?
7. What is the significance of determining the Vi-phagotype?
Take O- and H-diagnostics from Salmonella typhus, paratyphoid A and paratyphoid B and the patient’s serum from the teacher. Give Vidal's reaction.
Culture media
EMS, Ploskireva, and bismuth-sulfite agar media are produced by the medical industry in the form of dry powder. They are prepared according to the directions on the label: weigh out a certain amount of powder, pour in the appropriate amount of water, boil and pour into sterile Petri dishes.
Russell Wednesday. Add 40 g of dry nutrient medium to 950 ml of distilled water and add 5 g of nutrient agar. Heat until boiling and the powders dissolve. 1 g of x is dissolved in 50 ml of distilled water. hours of glucose and add to the prepared mixture. The medium is poured into sterile test tubes of 5-7 ml, sterilized with flowing steam (2 days for 2 minutes) and beveled so that a column remains. Russell's medium with mannitol and sucrose is prepared in the same way.
Olkenitsky's medium from dry agar. 2.5 g of dry nutrient agar is melted in 100 ml of distilled water. All the ingredients specified in the recipe (label) are added to the agar cooled to 50°C. The medium poured into test tubes is sterilized with flowing steam (3 days, 20 minutes each) and then mowed down. The prepared medium should be pale pink in color.
13.1.3 Salmonella
In 1880, the German researcher K. Ebert first described the bacterium that causes typhoid fever. In 1884, this microorganism was isolated and carefully studied by G. Gaffki.
A similar pathogen that causes disease in pigs was discovered in 1885 by D. Salmon. Subsequently, the entire genus to which these bacteria belong was named Salmonella, and the pathogen was named S. choleraesuis .
Further, Salmonella, the causative agents of animal diseases and food toxic infections in humans, was identified - S.enteritidis(A. Gartner, 1888) and S.typhimurium(K. Kensch and E. Nobel, 1898).
Later, in 1900, G. Schottmuller studied in detail Salmonella - the causative agents of human paratyphoid infections - S. paratyphi B or S . schottmuelleri. In turn, the causative agent of paratyphoid A was isolated and studied by A. Brion and G. Kaiser.
Classification
According to modern taxonomy, the genus Salmonella includes only 2 types - S. enterica And S. bongori. Pathogenic representatives belong only to the species S. enterica.
View S. enterica includes subspecies enterica, salamae, arizonae, diarizonae, houtenae And indica. More than 99% of human diseases are caused by Salmonella subspecies enterica.
Salmonella are extremely variable antigenically. More than 2500 serovars are known. For a long time, bacterial serovars were considered different types, which are designated separately.
Only serovars of the subspecies have proper names enterica. At the same time, the names of most of their variants have become commonly used in medical practice.
Serovars of other subspecies are designated by numbers.
In humans, Salmonella is caused by anthroponotic ( typhoid fever, paratyphoid fever) and zooanthroponotic infections ( salmonella).
The causative agent of typhoid fever is S. enterica serovar Typhi. Its short name, taking into account the name of the serovar, is S. Typhi (indicated in a non-italic font with a capital letter).
The causative agents of paratyphoid diseases are S. Paratyphi A, S. Paratyphi B, S. Paratyphi C.
The main serovars that cause salmonellosis are S. Enteritidis and S. Typhimurium. Many other variants can also cause these diseases (S. Choleraesuis, S. Heidelberg, S. Derby, etc.)
Morphology
All Salmonella are gram-negative motile rods, have multiple pili and flagella (peritrichs), do not form spores, and may have a polysaccharide capsule.
Cultural properties
Facultative anaerobes, chemoorganotrophs.
Capable of growing at temperatures from 8 to 45 0 C.
They reproduce well on simple nutrient media. On MPA they form translucent, colorless colonies.
Bile media are selective (bile broth, liquid Rapoport medium with glucose, bile salts and Andrade indicator). Capable of growing in selenite broth.
In liquid media, S-forms cause uniform turbidity.
On differential diagnostic media, Endo, Levin, and McConkey form colorless colonies, because Salmonella does not break down lactose.
The selective medium for salmonella is bismuth-sulfite agar, where they grow in the form of black shiny colonies.
Biochemical properties
Salmonella ferment carbohydrates (glucose, maltose, mannitol, arabinose, mannose) to produce acid and gas. Does not ferment lactose or sucrose.
Unlike other serovars, S. Typhi does not produce gas during the fermentation of carbohydrates.
When proteins are broken down, they form hydrogen sulfide, with the exception of S. Paratyphi A. They do not form indole.
Oxidase negative, catalase positive
Antigenic structure and Kaufman-White classification
Salmonella have 3 main antigens: O-AG, N-AG, and some - capsular Vi-AG.
O-antigen heat-stable, can withstand boiling for 2.5 hours. It is a cell wall LPS that has endotoxin properties.
H-antigen– flagellated, thermolabile, destroyed at a temperature of 75-100 o C. It is a flagellin protein.
Unlike other enterobacteriaceae, it has 2 phases: first – specific and the second - nonspecific. The phases are separate antigens that are determined by different genes. Most Salmonella are biphasic. There are monophasic salmonellae that express only one variant of H-AG.
F. Kaufman and P. White classified Salmonella according to their antigenic structure.
According to O-AG, all salmonella are divided into 67 groups (A, B, C, D, E, etc.). One group includes salmonella that have a common O-antigen determinant, indicated by a number.
According to H-AG, Salmonella are divided into serovars within groups. Specific phase 1 of the H-antigen is indicated in Latin lowercase letters, phase 2 - in Arabic numerals (or together with Latin letters). Based on the 1st phase of the H-antigen, the serovar is directly determined.
Vi-AG belongs to the group of superficial or capsular AGs. In most cases, it is found only in S. Typhi, rarely in S. Paratyphi C and S. Dublin.
It is thermolabile, completely destroyed when boiled for 10 minutes, and partially inactivated at a temperature of 60 o C for 1 hour.
Salmonella that have the Vi antigen are lysed by typhoid Vi bacteriophages. Phage typing is carried out to determine the source of infection, which is of epidemiological significance. About 100 phagotypes are known. The Vi-AG polysaccharide provides specific interaction with Vi phages.
Pathogenicity factors
Salmonella have at least 10 genetic islands of pathogenicity, which can be found in many pathogens. In addition, S.Typhi has main island of pathogenicity, distinguishing it from other representatives.
Two main islands of pathogenicity play a leading role in the pathogenesis of infections SPI-1 And SPI-2 , localized in the nucleoid. Some of the genes on these islands were obtained as a result of transduction of temperate bacteriophages.
Both islands are responsible for the formation of structures IIItype of secretion(injectis And effector proteins invasion), but these structures are different .
Island effector moleculesSPI-1 are responsible for the penetration of the pathogen into epithelial cells and the development of enterocolitis.
Some of them form injectisome(or acupuncture complex). The rest, after contact of bacteria with the epithelium, enter the cells using injectisomes.
They rearrange the actin of the cellular cytoskeleton, which leads to the formation of folds on the surface of M-cells of Peyer's patches and intestinal epithelium. Thus, epithelial cells acquire the ability to capture bacteria that penetrate inside by macropinocytosis.
In addition, SPI-1 island virulence proteins activate membrane channels in the epithelium, which increases chloride secretion and leads to diarrhea.
When these molecules enter macrophages, they activate caspase-1. On the one hand, this stimulates the production of pro-inflammatory cytokines (IL 1, neutrophil chemokine IL 8, etc.) On the other hand, the death of macrophages through apoptosis is activated. Thus, these proteins cause an immunoinflammatory process in the intestinal wall with the penetration of neutrophils there.
Effector molecules another island of pathogenicity SPI-2 responsible for the survival of bacteria inside phagocytes and cells of affected organs. Thus, they determine the development not of the local, but systemic salmonellosis infections.
SPI-2 island proteins also form injectisome. After the pathogen enters the phagocyte, it is located inside the vacuole, where it is able to multiply. Effector molecules inhibit respiratory burst enzymes, which ensures long-term bacterial survival. In addition, these proteins maintain the structure of the wall of vacuoles containing Salmonella.
Another island of pathogenicity SPI-3 encodes enzymes that provide salmonella with magnesium cations. It is also necessary for the survival of bacteria inside phagocytes.
When destroyed, Salmonella is released endotoxin, which through TLR-4 receptors on cells stimulates the release of pro-inflammatory cytokines. It has a pyrogenic effect and damages the vascular endothelium.
Some pathogens are capable of producing enterotoxins which cause secretory diarrhea.
Main pathogenicity island S.Typhi determines the invasiveness of the pathogen, as well as the ability to produce capsular Vi-AG.
Two S.Typhi plasmids contain antibiotic resistance genes. In addition, some Salmonella have a set of genes for multiple resistance to antibiotics, which are located in the nucleoid.
Resistance
In the external environment, Salmonella retain their viability for a long time: in open water they live for up to 120 days, in sea water for up to a month, in soil for up to 9 months, in room dust for up to 1.5 years, in sausages for 2-4 months, in frozen meat and eggs up to 1 year. Salmonella not only persists in products, but also multiplies (milk, sour cream, cottage cheese, minced meat). Flies can play a role in food contamination.
Bacteria tolerate low temperatures well, but are sensitive to high temperatures - when heated to 60 0 C they die after 30 minutes, at 100 0 C - almost instantly. Disinfectants (chloramine, hypochlorite, Lysol) in normal concentrations kill pathogens within a few minutes.
Characteristics of diseases
Salmonella causes 3 groups of lesions: typhoid and paratyphoid fever, Salmonella gastroenteritis And septicemia. Their development depends on the virulence of the pathogen, its infectious dose and the state of immunity of the macroorganism. For typhoid fever to occur, 10 3 -10 5 microbial cells are required. For the development of salmonellosis, the infecting dose is significantly higher - 10 6 -10 9 bacteria, but with high virulence of the pathogen or with a person’s immunodeficiency state, the number of bacteria can be many times less.
Typhoid fever and paratyphoid diseases
Typhoid fever And paratyphoid– these are spicy infectious diseases, which are characterized by inflammatory damage to the small intestine with destruction of lymphoid tissue and ulceration, bacteremia, fever, general intoxication, enlargement of the spleen and liver.
Typhoid fever is the most severe.
This disease poses a serious health problem, especially in developing countries. Every year, from 15 to 30 million cases of typhoid fever occur in the world, with 250 to 500 thousand deaths recorded. In developing countries, it mainly affects children and young people. In developed countries, the disease occurs in sporadic cases.
Typhoid fever and paratyphoid A – anthroponotic infections, the reservoir of which is man. The causative agents of paratyphoid B and C have also been isolated from some animals and birds.
Sources of infection are patients or bacteria carriers who excrete the pathogen in their feces, urine, or saliva. Main mechanism of infection– fecal-oral (water, food and contact-household routes).
Incubation period can last up to 2-3 weeks.
When ingested orally, the bacteria penetrate the protective barriers of the stomach and enter the small intestine ( infection phase). The most important role in pathogenesis is played by invasive system proteinsIIItype of secretion(see above). Some of the invasion proteins exhibit translocase activity - form injectisome and ensure the penetration of Salmonella into epithelial M cells and enterocytes. The rest block the metabolism of infected cells, leading to disruption of their function. There is an increase in the production of chemokines (for example, IL-8) and other pro-inflammatory cytokines by enterocytes and intestinal macrophages.
Salmonella remain viable in the vacuoles of affected cells and cause apoptosis of macrophages by activating caspase-1.
As a result of disruption of the hemolymphatic barrier, Salmonella enters the blood ( bacteremia phase). The causative agents of typhoid fever survive and multiply in phagocytes, and after the death of the latter in large quantities enter the blood. Wherein Vi-AG inhibits the action of serum and phagocytic bactericidal factors.
At this time, clinical symptoms of the disease appear ( first week of illness). The temperature rises to 39-40 o. Under the influence of the bactericidal properties of the blood and due to phagocytosis, salmonella are destroyed and released endotoxin, which affects microcirculation vessels and has a pronounced neurotropic effect. In severe cases, as a result of damage to the central nervous system, status typhosus(severe headache, insomnia, severe weakness, apathy, impaired consciousness, even coma). Intestinal damage is accompanied by swelling and desquamation of the epithelium. A disorder of the autonomic nervous system is accompanied by flatulence and abdominal pain. Diarrhea develops.
At 2 weeks of illness (the height of the disease) Salmonella spreads through the blood to internal organs, affecting the liver, gallbladder, spleen, kidneys, a rash appears on the skin. From the 2nd week, salmonella with bile again enter the small intestine, the lymphoid formations of which are already sensitized with salmonella antigens. As a result, there is autoimmune inflammatory response, sometimes necrosis forms in places where lymphoid cells accumulate. Necrosis of the mucous membrane can result in bleeding and intestinal perforation.
After the height of the disease there is a gradual extinction of clinical manifestations diseases. The release of pathogens from the body occurs in feces, urine, sweat, saliva, and breast milk (in nursing women). The immune response ensures the gradual elimination of Salmonella.
Patients treated with antibiotics are discharged from the hospital no earlier than the 21st day of normal temperature. Before discharge, a three-time bacteriological examination of feces and urine and a single examination of bile are carried out.
Usually the disease ends recovery. Mortality does not exceed 0.5-1%. However, in the absence of adequate medical care, isolated outbreaks of typhoid fever in tropical countries have had mortality rates exceeding 30%.
Paratyphoid fevers A and B proceed more favorably. Their clinical symptoms are similar. In general, these diseases are characterized by a milder course compared to typhoid fever.
Immunity
After an infection, immunity is generally stable, but there may be relapses and repeated diseases.
Recovery does not always end with complete freedom from the pathogen. More than 2% of patients have bacterial carriage. Since the bacteria are resistant to bile, they are concentrated in the gallbladder, isolated from the action of immune factors. Such pathogens produce increased amounts of Vi-AG. Capable of persisting inside macrophages.
Bacteria carriers are dangerous as sources of infection. They can retain pathogens for many months. In chronic carriers, a deficiency of IgM antibodies against O-AG was detected.
Carriage of paratyphoid pathogens occurs more often than with typhoid fever, but it lasts less – within a few weeks.
Laboratory diagnosis of typhoid fever
Use bacteriological And serological methods which are carried out taking into account the period of the infectious process.
Material for highlighting are blood ( blood culture), excreta ( coproculture), urine ( urine culture), duodenal contents, bile ( biculture), scraping roseola, bone marrow.
IN bacteriological research early method is the isolation of the pathogen from the blood (blood culture) during the period of bacteremia (the first week of the disease).
Blood is inoculated into bile broth or Rapoport medium in a ratio of 1:10 (to reduce the bactericidal properties of blood proteins). On the 2nd day, subculture is carried out on Endo or Levin medium, or bismuth sulfite agar. Suspicious (transparent or black depending on the media) colonies are subcultured onto agar slants or one of the combined media (Olkenitsky, Ressel, Kligler). On these media, for primary identification, glucose fermentation, the ability to form gases, the release of hydrogen sulfide, and the absence of urease are determined.
At the same time, morphology and tinctorial properties are studied.
Biochemical properties are determined. Bacteria of the typhoid-paratyphoid group do not decompose sucrose, lactose, and do not form indole.
When isolating cultures that have enzymatic properties characteristic of Salmonella, their antigenic structure is studied in a glass agglutination reaction with O- and H-diagnostic antisera, sensitivity to antibiotics is determined, and phage typing is performed.
For serological For the diagnosis of typhoid fever and paratyphoid fever from the 5th to 7th day of the disease, RPGA with O- and N-erythrocyte diagnosticums is mainly used. A reaction with a titer of 1:160 or higher is considered positive. When examined in the RPGA, the antibody titer increases over the course of the disease.
It is possible to use the Widal agglutination reaction with O- and H-monodiagnosticums to specific pathogens (positive reaction titer - 1:200 and above). Serological diagnosis is retrospective.
To identify bacteria carriers use RPGA with erythrocyte Vi-diagnosticum (reaction titer – 1:40). They study biliary and coproculture. Phagotyping is carried out with Vi-1 antigen.
During epidemic outbreaks of typhoid fever, RIF and ELISA are used for express diagnostics to detect hypertension in the blood, bone marrow and other material.
Treatment of typhoid fever
Etiotropic therapy is carried out immediately after the clinical diagnosis is established. Fluoroquinolones are used for treatment. In case of resistance to them, third generation cephalosporins and azithromycin are used.
Levomycetin and co-trimoxazole are currently used less frequently due to the spread of multidrug-resistant strains. Pathogenetic treatment includes infusion-detoxification therapy.
Prevention
Sanitary, hygienic and anti-epidemic measures are being carried out aimed at neutralizing sources of infection, suppressing transmission routes, and increasing the body's immunity.
For specific immunoprophylaxis of typhoid fever, 3 types of vaccines have been developed. Inactivated vaccines are used (50-70% effective); a live attenuated vaccine from the Tu21a strain has been developed (has a greater protective effect, is at the stage of clinical trials). A polysaccharide vaccine made from the Vi-antigen of S. typhi is effective (For example, Vianvac pr-va Russian Federation), used according to epidemiological indications, the protective effect lasts up to 2 years.
Salmonella
Salmonella– a group of polyetiological acute infectious diseases of humans, animals and birds, characterized by predominant damage to the gastrointestinal tract, diarrhea and bacteremia.
The most common clinical form of Salmonella infection is salmonella gastroenteritis. The main causative agents of gastroenteritis are: S. Enteritidis, S. Choleraesuis, S. Anatum, S. Derby, although diseases can be caused by many other variants of bacteria.
A much more severe form is generalized salmonella infection - septicemia. Its leading pathogen is S. Typhimurium.
Most pathogens are isolated from various animals (the main reservoir) and humans.
Source of infection humans are most often poultry (50%), especially chickens and ducks, as well as their eggs (salmonella can penetrate the shell inside). Salmonella carriage has been detected in livestock, dogs, cats, rodents, and in many wild animals and birds. Infected animals excrete bacteria in their urine and feces, milk, and saliva, polluting the environment.
Basic transmission route salmonella - food. Diseases occur in humans due to the consumption of meat products (beef, pork - up to 20% of cases, poultry), eggs, and less often - fish, vegetables, fruits, shellfish, crayfish, crabs.
Meat can become infected endogenously during the life of the animal during its illness, as well as exogenously during transportation, processing, and storage. Sometimes food becomes infected due to improper cooking or cooking.
Failure to comply with sanitary and hygienic standards may result in contact-household path transmission, which is typical for nosocomial outbreaks salmonellosis. Such outbreaks have been noted in maternity institutions, surgical, children's and other hospitals. In hospital-acquired salmonellosis, S. typhimurium is more often isolated and S. Haifa. In the Republic of Belarus, salmonella infections account for more than 50% of all cases of hospital infections
The causative agents of hospital-acquired salmonellosis are highly resistant to chemotherapeutic drugs and antibiotics.
Children under 1 year of age and persons with various immunodeficiencies are most susceptible to salmonellosis.
The incubation period of the disease is from 2-6 hours to 2-3 days (on average 7-24 hours).
The pathogenesis of salmonellosis is determined by the virulence factors of the pathogens. Among them, the most important role is played by invasive proteins of type III secretion.
Some of the invasion proteins ensure the penetration of Salmonella into intestinal epithelial cells and their survival inside vacuoles. In addition, they stimulate the release of proinflammatory cytokines and chemokines from affected cells and apoptosis of macrophages.
Inside macrophages, bacteria not only multiply, but also partially die with the release of endotoxin, which affects the neurovascular system of the intestine and increases the permeability of cell membranes.
Within 1 hour from the penetration of salmonella into the cells, pronounced neutrophilic infiltration of the intestinal wall develops. Intestinal inflammation is accompanied by the release of protein from affected enterocytes, increased secretion of chlorides with the development of profuse diarrhea.
Some Salmonella can produce enterotoxin, which, through an increase in cAMP content in enterocytes, stimulates the excretion of chlorides, which aggravates diarrhea.
In most cases, at this stage the infectious process can be completed ( gastrointestinal form).
In severe cases, bacteremia and generalization of infection occur, which leads to septicemia.
This form of salmonellosis is most typical for S. Typhimurium and S. Enteritidis. Its development is determined by virulence proteins, which are encoded by the pathogenicity island SPI-2 . These proteins suppress phagocytosis, which ensures the survival and reproduction of bacteria inside phagocytes, their penetration into the blood and parenchymal organs.
As a result, Salmonella can cause dystrophic changes in the affected organs (spleen, liver) with the formation of secondary purulent foci.
Usually the disease ends in recovery, but septic forms of infection can lead to death.
Immunity
Post-infectious immunity is short-lived, unstable, and type-specific. Agglutinins, precipitins, bacteriolysins and other antibodies are found in the serum of patients and convalescents. A disease caused by one serovar does not create immunity to others, and a previous infection does not exclude reinfection.
Laboratory diagnosis of salmonellosis
The basis of diagnosis is bacteriological method. For research they take various materials: feces, vomit, gastric lavage, urine, food debris, as well as the starting products used for its preparation; washouts from various equipment and objects.
To diagnose septicemia, blood is examined.
Selenite broth, selenite agar, and 20% bile broth are used as enrichment media. Among the differential diagnostic media for primary cultures and cultures from enrichment media, selective media (bismuth sulfite agar or brilliant green agar) and differential diagnostic media (Endo and Levina) are distinguished. Suspicious colonies are subcultured into tubes with one of the combined media (Olkenitsky, Kligler, Ressel) and onto an MPA slant.
The morphological, tinctorial, and biochemical properties of pathogens are studied.
With cultures grown on MPA, they carry out serological typing according to the Kaufman-White scheme. An agglutination reaction is performed on glass with O- and H-agglutinating antisera. Based on the results of the reaction, a final bacteriological diagnosis is made.
Serological diagnostics are rarely used (RA, RPGA).
ELISA methods have been developed for Salmonella antigen detection in blood and urine.
Treatment
Pathogenetic therapy for salmonellosis is aimed at detoxification, restoration of water-electrolyte balance and hemodynamics. Antibacterial therapy for mild forms of gastroenteritis is not indicated. In case of generalized infection, fluoroquinolones are prescribed; in case of resistance to them, third generation cephalosporins (ceftriaxone) are prescribed.
In the complex treatment of salmonellosis, it is possible to use a polyvalent salmonella bacteriophage.
Prevention
Includes veterinary-sanitary, sanitary-hygienic and anti-epidemic measures. In the event of an intrahospital outbreak of salmonellosis, a special operating regime for the treatment and prevention facility is established.
Vaccine prevention has not been developed.
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