BGKP. Bacteria of the group of Escherichia coli (coliforms) include genera Escherichia(typical representative E. coli), Citrobacter(typical representative C. colicitrovorum), Enterobacter(a typical representative of E. aerogenes), which are united in one family Enterobacteriaceae due to common properties.

General characteristics of BGKP: - sticks gram-negative, short; - not spore-forming; - on End's medium they give red colonies with a metallic sheen - E.coli, red - enterobacteria, pink - citrobacteria, b / color - lactose - negative. biochemical properties. Most bacteria of the Escherichia coli group (ECG) do not liquefy gelatin, coagulate milk, break down peptones with the formation of amines, ammonia, hydrogen sulfide, and have high enzymatic activity against lactose, glucose and other sugars, as well as alcohols. They do not have oxidase activity. Sustainability. The bacteria of the Escherichia coli group are neutralized by conventional pasteurization methods (65-75°C). At 60°C, Escherichia coli dies within 15 minutes. A 1% solution of phenol causes the death of the microbe in 5-15 minutes. Sanitary and indicative value. Bacteria of the genus Escherichia- constant. intestinal inhabitants of humans and animals, and their detection in water and PP is evidence of fresh faecal contamination. Bacteria of the genera Citrobacter and Enterobacter r can be found everywhere: in the soil, on plants, less often in the intestines. It is believed that they are the result of changes in ischerichia after their exposure to the external environment and therefore are indicators of older faecal contamination. BGKP value:

In raw milk indicates - on the epidemiological danger

A few hours later at 8-10 o C - a violation of the conditions of storage and sale, protractors.

Appeared BGKP after pasteurization is regarded as the 2nd contamination

The presence of BGKP in the finished product indicates - poor washing and disinfection of equipment.

GenusSalmonella . Salmonellosis are among the most common toxicoinfections. Finding Salmonella is always indicative of faecal contamination. Salmonella are resistant to high concentrations of sodium chloride (especially in media containing protein) and desiccation. Retain their viability in room dust, in various soils (97 months), in the water of open reservoirs (up to 45 days). Being in PP, especially in meat, Salmonella is very resistant to heat treatment. Salting and smoking meat have little effect on Salmonella. During the reproduction of Salmonella in milk, its appearance and taste do not change; pasteurization of milk for 30 minutes at 85ºС under production conditions contributes to the complete destruction of these bacteria. A person becomes infected with salmonella as a result of the consumption of meat and meat products. Milk and dairy products are much less likely to cause food poisoning. Infection of milk mainly occurs through contaminated dishes, milking machines, hands of milkers, etc. Salmonellosis pathogens can get into food products made from vegetable raw materials (salads and table sauces) not only during the production process, but also with food ingredients, in particular with dry vegetable seasonings and spices.

BGKP identification:

● Seeding on the enrichment medium - Kessler, simultaneous identification by gaseous formations: there is gas formation - BKGP is possible;

● Identification of CGB on Endo medium: Take 1 ml from gas (+) tubes and inoculate on Endo solid medium, identify CGB colonies by color, differentiate by genera depending on the color of the colonies: If there are red, pink and pale pink cultures - it means there are BGKP, if there are no colonies - there is no BGKP. If there are colonies, but colorless - suspicion of pathogens. Further, the genera of BGKP are identified by color: 1) red - with metallic. shade. - Escherichia 2) pink - Enterobacter 3) pale pink - with mucus - Klebsiela 4) pale pink - citrobacter, cerrations 5) colorless (lactose (-)) - Proteus 6) transparent small - pathogenic

● Identification on the Coser medium: growing on the medium with glucose/citric acid, T=43°C, 24h. M / o citrate (+) change the color of the dye from green to cornflower blue. M / o citrate (-) do not change color.

Determined by the number of positive samples in 3 test tubes.

Salmonella- pathogenic, analyzed in 25 g of the product, they should not be there. Serves as an indicator of pathogens.

Salmonella detection is carried out in 4 stages

1) primary (direct) sowing - Sowing on the environment of End and Ploskirav for a day and T = 37 0 C. On cf. Enda - transparent colonies,

2) enrichment (inoculation on liquid selective media, temperature control)

3) sowing from the enrichment medium after enrichment on dense diagnostic media, temperature control - on cf. Ploskirava - transparent, but smaller than on Endo medium

4) confirmation by establishing the enzymatic and serological properties of Salmonella

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1. Literature review

.1 Taxonomy of Escherichia coli

scientific classification

Domain: Bacteria

Type: Proteobacteria

Class: Gamma Proteobacteria

Order: Enterobacteriales

Family: Enterobacteriaceae

Genus: Escherichia

Species: Coli (E. coli)

International scientific name

Escherichia coli (Migula 1895)

1.2 The structure and chemical composition of a bacterial cell

The internal organization of a bacterial cell is complex. Each systematic group of microorganisms has its own specific structural features.

The bacterial cell is covered with a dense membrane. This surface layer, located outside the cytoplasmic membrane, is called the cell wall. The wall performs protective and supporting functions, and also gives the cell a permanent, characteristic shape (for example, the shape of a rod or coccus) and is the outer skeleton of the cell. This dense shell makes bacteria related to plant cells, which distinguishes them from animal cells that have soft shells. Inside the bacterial cell, the osmotic pressure is several times, and sometimes tens of times higher than in the external environment. Therefore, the cell would quickly rupture if it were not protected by such a dense, rigid structure as the cell wall.

The thickness of the cell wall is 0.01-0.04 µm. It is from 10 to 50% of the dry mass of bacteria. The amount of material from which the cell wall is built changes during bacterial growth and usually increases with age.

Murein (glycopeptide, mucopeptide) is the main structural component of the walls, the basis of their rigid structure in almost all bacteria studied so far. This is an organic compound of a complex structure, which includes sugars that carry nitrogen - amino sugars and 4-5 amino acids. Moreover, the amino acids of cell walls have an unusual shape (D-stereoisomers), which is rarely found in nature.

Using the method of staining, first proposed in 1884 by Christian Gram, bacteria can be divided into two groups: gram-positive, gram-negative .

Gram-positive organisms are able to bind certain aniline dyes, such as crystal violet, and after treatment with iodine and then with alcohol (or acetone), retain the iodine-dye complex. The same bacteria in which this complex is destroyed under the influence of ethyl alcohol (cells become discolored) are gram-negative.

The chemical composition of the cell walls of Gram-positive and Gram-negative bacteria is different. In gram-positive bacteria, the cell walls include, in addition to mucopeptides, polysaccharides (complex, high-molecular sugars), teichoic acids (complex in composition and structure, compounds consisting of sugars, alcohols, amino acids and phosphoric acid). Polysaccharides and teichoic acids are associated with the framework of the walls - murein. We do not yet know what structure these constituent parts of the cell wall of gram-positive bacteria form. With the help of electronic photographs, thin sections (layering) were not found in the walls of gram-positive bacteria. Probably, all these substances are very closely related to each other.

The walls of gram-negative cells contain a significant amount of lipids (fats) associated with proteins and sugars in complex complexes - lipoproteins and lipopolysaccharides. In general, there is less murein in the cell walls of gram-negative bacteria than in gram-positive bacteria. The wall structure of Gram-negative bacteria is also more complex. Using an electron microscope, it was found that the walls of these bacteria are multilayered.

The inner layer is murein. Above it is a wider layer of loosely packed protein molecules. This layer is in turn covered by a layer of lipopolysaccharide. The top layer is made up of lipoproteins.

The cell wall is permeable: through it, nutrients freely pass into the cell, and metabolic products are released into the environment. Large molecules with high molecular weight do not pass through the shell.

The cell wall of many bacteria is surrounded on top by a layer of mucous material - a capsule. The thickness of the capsule can be many times greater than the diameter of the cell itself, and sometimes it is so thin that it can only be seen through an electron microscope - a microcapsule.

The capsule is not an obligatory part of the cell, it is formed depending on the conditions in which the bacteria enter. It serves as a protective cover of the cell and participates in water exchange, protecting the cell from drying out.

By chemical composition, capsules are most often polysaccharides. Sometimes they consist of glycoproteins (complex complexes of sugars and proteins) and polypeptides (genus Bacillus), in rare cases - of fiber (genus Acetobacter).

Mucous substances secreted into the substrate by some bacteria determine, for example, the mucous-viscous consistency of spoiled milk and beer.

The entire contents of a cell, with the exception of the nucleus and cell wall, is called the cytoplasm. The liquid, structureless phase of the cytoplasm (matrix) contains ribosomes, membrane systems, mitochondria, plastids and other structures, as well as reserve nutrients. The cytoplasm has an extremely complex, fine structure (layered, granular). With the help of an electron microscope, many interesting details of the structure of the cell have been revealed.

The outer lipoprotein layer of the bacterial protoplast, which has special physical and chemical properties, is called the cytoplasmic membrane.

Inside the cytoplasm are all vital structures and organelles.

The cytoplasmic membrane plays a very important role - it regulates the flow of substances into the cell and the release of metabolic products to the outside.

Through the membrane, nutrients can enter the cell as a result of an active biochemical process involving enzymes. In addition, the synthesis of some components of the cell occurs in the membrane, mainly the components of the cell wall and the capsule. Finally, the most important enzymes (biological catalysts) are located in the cytoplasmic membrane. The orderly arrangement of enzymes on membranes makes it possible to regulate their activity and prevent the destruction of some enzymes by others. Ribosomes are attached to the membrane - structural particles on which protein is synthesized. The membrane is made up of lipoproteins. It is strong enough and can provide the temporary existence of a cell without a shell. The cytoplasmic membrane makes up to 20% of the dry mass of the cell.

In electron photographs of thin sections of bacteria, the cytoplasmic membrane appears as a continuous strand about 75 Å thick, consisting of a light layer (lipids) enclosed between two darker ones (proteins). Each layer has a width of 20-30A. Such a membrane is called elementary.

Between the plasma membrane and the cell wall there is a connection in the form of desmoses - bridges. The cytoplasmic membrane often gives invaginations - invaginations into the cell. These invaginations form special membrane structures in the cytoplasm called mesosomes. Some types of mesosomes are bodies separated from the cytoplasm by their own membrane. Numerous vesicles and tubules are packed inside such membranous sacs. These structures perform a variety of functions in bacteria. Some of these structures are analogues of mitochondria. Others perform the functions of the endoplasmic reticulum or the Golgi apparatus. By invagination of the cytoplasmic membrane, the photosynthetic apparatus of bacteria is also formed. After invagination of the cytoplasm, the membrane continues to grow and forms stacks, which, by analogy with plant chloroplast granules, are called thylakoid stacks. These membranes, which often fill most of the cytoplasm of a bacterial cell, contain pigments (bacteriochlorophyll, carotenoids) and enzymes (cytochromes) that carry out the process of photosynthesis.

The cytoplasm of bacteria contains ribosomes - protein-synthesizing particles with a diameter of 200A. There are more than a thousand of them in a cage. Ribosomes are made up of RNA and protein. In bacteria, many ribosomes are located freely in the cytoplasm, some of them can be associated with membranes.

The cytoplasm of bacterial cells often contains granules of various shapes and sizes. However, their presence cannot be considered as some kind of permanent feature of the microorganism, usually it is largely associated with the physical and chemical conditions of the environment. Many cytoplasmic inclusions are composed of compounds that serve as a source of energy and carbon. These reserve substances are formed when the body is supplied with a sufficient amount of nutrients, and, conversely, are used when the body enters conditions that are less favorable in terms of nutrition.

In many bacteria, the granules are composed of starch or other polysaccharides - glycogen and granulosa. Some bacteria, when grown on a sugar-rich medium, have droplets of fat inside the cell. Another widespread type of granular inclusions is volutin (metachromatin granules). These granules are composed of polymetaphosphate (reserve substance, including phosphoric acid residues). Polymetaphosphate serves as a source of phosphate groups and energy for the body. Bacteria accumulate volutin more often under unusual nutritional conditions, such as on a medium that does not contain sulfur. Sulfur droplets are found in the cytoplasm of some sulfur bacteria.

In addition to various structural components, the cytoplasm consists of a liquid part - a soluble fraction. It contains proteins, various enzymes, t-RNA, some pigments and low molecular weight compounds - sugars, amino acids.

As a result of the presence of low molecular weight compounds in the cytoplasm, a difference arises in the osmotic pressure of the cellular contents and the external environment, and this pressure may be different for different microorganisms. The highest osmotic pressure was noted in gram-positive bacteria - 30 atm, in gram-negative bacteria it is much lower than 4-8 atm.

In the central part of the cell, the nuclear substance, deoxyribonucleic acid (DNA), is localized.

Bacteria do not have such a nucleus as in higher organisms (eukaryotes), but there is its analogue - the "nuclear equivalent" - the nucleoid , which is an evolutionarily more primitive form of organization of nuclear matter. Microorganisms that do not have a real nucleus, but have its analogue, belong to prokaryotes. All bacteria are prokaryotes. In the cells of most bacteria, most of the DNA is concentrated in one or more places. In bacteria, DNA is less densely packed than in true nuclei; A nucleoid does not have a membrane, a nucleolus, or a set of chromosomes. Bacterial DNA is not associated with the main proteins - histones - and is located in the nucleoid in the form of a bundle of fibrils.

Some bacteria have adnexal structures on their surface; the most widespread of them are flagella - the organs of movement of bacteria.

The flagellum is anchored under the cytoplasmic membrane by two pairs of discs. Bacteria can have one, two, or many flagella. Their location is different: at one end of the cell, at two, over the entire surface. Bacterial flagella have a diameter of 0.01-0.03 microns, their length can be many times greater than the length of the cell. Bacterial flagella are made up of a protein, flagellin, and are twisted helical filaments.

1.3 Morphology of Escherichia coli and its representatives

coli microflora

E. coli is a polymorphic facultative anaerobic short (length 1-3 microns, width 0.5-0.8 microns) gram-negative bacillus with a rounded end. Strains in smears are arranged randomly, without forming spores and peritrichs. Some strains are microencapsulated and pili, found widely in the lower gut of warm-blooded organisms. Most strains of E. coli are harmless, but the O157:H7 serotype can cause severe food poisoning in humans.

Bacteria of the Escherichia coli group grow well on simple nutrient media: meat-peptone broth (MPB), meat-peptone agar (MPA). On Endo's medium, flat red colonies of medium size form. Red colonies can be with a dark metallic luster (E. coli) or without luster (E. aerogenes).

They have a high enzymatic activity against lactose, glucose and other sugars, as well as alcohols. They do not have oxidase activity. According to the ability to break down lactose at a temperature of 37 ° C, bacteria are divided into lactose-negative and lactose-positive Escherichia coli (LCE), or coliforms, which are formed according to international standards. Fecal Escherichia coli (FEC) stand out from the LEC group, capable of fermenting lactose at a temperature of 44.5 ° C. fecal pollution.

Common coliform bacteria (CBC) are gram-negative, non-spore-forming rods capable of growing on differential lactose media, fermenting lactose to acid, aldehyde and gas at a temperature of 37 +/- 1°C for 24 - 48 hours.

Coliform bacteria (coliforms) - a group of gram-negative rods, mainly living and multiplying in the lower digestive tract of humans and most warm-blooded animals (for example, livestock and waterfowl). They usually enter water with faecal effluents and are able to survive in it for several weeks, although they (in the vast majority) do not reproduce.

Thermotolerant coliform bacteria play an important role in evaluating the effectiveness of water purification from faecal bacteria. It is E. coli (E. coli) that serves as a more accurate indicator, since not only fecal water can serve as a source of some other thermotolerant coliforms. At the same time, the total concentration of thermotolerant coliforms in most cases is directly proportional to the concentration of E. coli, and their secondary growth in the distribution network is unlikely (unless there are sufficient nutrients in the water, at temperatures above 13 °C.

Thermotolerant coliform bacteria (TCB) - are among the common coliform bacteria, have all their characteristics and, in addition, are able to ferment lactose to acid, aldehyde and gas at a temperature of 44 +/- 0.5 ° C for 24 hours.

They include the genus Escherichia and, to a lesser extent, individual strains of Citrobacter, Enterobacter, and Klebsiella. Of these organisms, only E. coli is specifically of fecal origin, and it is always present in large quantities in human and animal feces and is rarely found in water and soil that have not been subjected to fecal contamination. It is believed that the detection and identification of E. coli provides sufficient information to establish the faecal nature of the contamination.

Coliforms are found in large quantities in domestic wastewater, as well as in surface runoff from livestock farms. In water sources used for centralized drinking and household water supply, the number of total coliforms is allowed no more than 1000 units (CFU / 100 ml, CFU - colony-forming units), and thermotolerant coliforms - no more than 100 units. In drinking water, coliforms should not be detected in a 100 ml sample. Coliforms may be accidentally introduced into the distribution system, but not more than 5% of samples taken during any 12-month period, provided that E. coli is absent.

The presence of coliform organisms in water indicates insufficient purification, secondary pollution, or the presence of excess nutrients in the water.

2. Materials and research methods

When examining relatively pure microbially water for the presence of pathogenic microorganisms, it is necessary to concentrate the desired microflora, which is contained in a negligible amount in water. Detection of causative agents of intestinal infections in the water of open reservoirs and wastewater against the background of the prevailing mass of saprophytic microflora is most effective when the desired bacteria are concentrated in accumulation media that inhibit the growth of accompanying microflora. Therefore, when analyzing water that has a different degree of general microbial contamination, certain methods are used to isolate pathogenic microflora.

Open waters are usually characterized by a significant content of suspended solids, i.e. turbidity, often color, low salt content, relatively low hardness, the presence of a large amount of organic substances, relatively high oxidizability and a significant content of bacteria . Seasonal fluctuations in the quality of river water are often very sharp. During the flood period, turbidity and bacterial contamination of water greatly increase, but its hardness (alkalinity and salinity) usually decreases. Seasonal changes in water quality largely affect the nature of the operation of water treatment facilities in certain periods of the year.

The number of microbes in 1 ml of water depends on the presence of nutrients in it. The more polluted the water with organic residues, the more microbes it contains. Especially open reservoirs and rivers are rich in microbes. The largest number of microbes in them is in the surface layers (in a layer of 10 cm from the water surface) of coastal zones. With distance from the coast and increasing depth, the number of microbes decreases.

River silt is richer in microbes than river water. There are so many bacteria in the very surface layer of silt that a kind of film is formed from them. This film contains many filamentous sulfur bacteria, iron bacteria, they oxidize hydrogen sulfide to sulfuric acid and thus prevent the inhibitory effect of hydrogen sulfide (fish death is prevented).

Rivers in urban areas are often natural recipients of household and fecal sewage, so the number of microbes sharply increases within the boundaries of settlements. But as the river moves away from the city, the number of microbes gradually decreases, and after 3-4 tens of kilometers it again approaches its original value. This self-purification of water depends on a number of factors: mechanical sedimentation of microbial bodies; reduction in water of nutrients assimilated by microbes; the action of the direct rays of the sun; consumption of bacteria by protozoa, etc.

Pathogens can enter rivers and reservoirs with sewage. Brucellosis bacillus, tularemia bacillus, poliomyelitis virus, foot-and-mouth disease virus, as well as causative agents of intestinal infections - typhoid bacillus, paratyphoid bacillus, dysentery bacillus, vibrio cholerae - can remain in water for a long time, and water can become a source of infectious diseases. Especially dangerous is the ingress of pathogenic microbes into the water supply network, which happens when it malfunctions. Therefore, sanitary biological control has been established for the state of reservoirs and the tap water supplied from them.

2.1 Hydrometric float method for measuring and determining the speed of water flow

To measure and determine the speed of water flow, there is a float method, which is based on tracking the movement of an object lowered into the stream (float) using instruments or with the naked eye. Floats are dropped into the water on small rivers from the shore or from a boat. The stopwatch determines the time and passage of the float between two adjacent sections, the distance between which is known. The surface current velocity is equal to the velocity of the float. By dividing the distance traveled by the float by the time of observation, the flow velocity is obtained.

2.2 Water sampling, storage and transport of samples

Water samples for bacteriological analysis are taken in compliance with the rules of sterility: in sterile bottles or sterile devices - bottles in the amount of 1 liter.

For the selection of water from open reservoirs, wastewater, water from pools, wells, the so-called bottle bottle is convenient.

Guidelines for the detection of pathogens of intestinal infections of a bacterial nature in water.

When sampling water from open reservoirs, the following points should be provided: at the place of stagnation and at the place of the fastest flow (from the surface and at a depth of 50 - 100 cm).

Bottle bottle. Bathometers are devices of various designs for taking water samples from different depths. In the classical form, these are cylinders that can be lowered to a certain depth, closed and removed there. It is not easy to make a classic bottle on your own. But instead, you can use a simple glass or plastic bottle with a narrow neck, weighted with some kind of load and plugged with a cork, ideally - cork. Ropes are tied to the neck of the bottle and to the cork. Having lowered the bottle to the desired depth (the main thing is that it sinks, this is what the load is for), you need to pull out the cork - therefore, you should not plug it tightly. After giving the bottle time to fill at the desired depth (1-2 minutes), it is pulled to the surface. This should be done as vigorously as possible - with a high lifting speed and a narrow neck, water from the overlying layers will practically not get inside.
Samples brought to the surface with a bathometer should also be “thickened” using a plankton net, and then the volume of filtered water should be calculated. Since this volume should be as large as possible, the bottle should be made as large as possible, for example using a 2-liter glass or plastic bottle or some other large vessel with a narrow neck. On the rope to which the bottle is tied, marks should also be made every meter - to determine the depth of sampling.

The first control point at the dam (the beginning of the beach) is the fence point (TK1).

The second control point at the boat station (the end of the beach) is the fence point (TK2).

T31 - the first control point at the dam (the beginning of the beach) T32 - the second control point at the boat station (the end of the beach)

2.3 Storage and transport of samples

Samples should be analyzed in the laboratory as soon as possible after collection.

The analysis should be carried out within 2 hours after sampling.

If the sample delivery time and storage temperature cannot be met, the sample should not be analyzed.

2.4 Preparing glassware for analysis

Laboratory glassware should be thoroughly washed, rinsed with distilled water until detergents and other impurities are completely removed, and dried.

Test tubes, flasks, bottles, vials must be closed with silicone or cotton-gauze stoppers and packed in such a way as to prevent contamination after sterilization during operation and storage. Caps can be metal, silicone, foil or thick paper.

New rubber stoppers are boiled in 2% sodium bicarbonate solution for 30 minutes and washed 5 times with tap water (boiling and washing are repeated twice). Then the corks are boiled for 30 minutes in distilled water, dried, wrapped in paper or foil and sterilized in a steam sterilizer. Previously used rubber stoppers are disinfected, boiled for 30 minutes in tap water with a neutral detergent, washed in tap water, dried, mounted and sterilized.

Pipettes with inserted cotton swabs should be placed in metal cases or wrapped in paper.

Petri dishes in the closed state should be placed in metal cases or wrapped in paper.

Prepared dishes are sterilized in a dry oven at 160-170°C for 1 hour, counting from the moment the specified temperature is reached. Sterilized dishes can only be removed from the drying cabinet after it has cooled below 60 °C.

After performing the analysis, all used cups and test tubes are decontaminated in an autoclave at (126±2)°C for 60 minutes. Pipettes are disinfected by boiling in a 2% NaHC03 solution.

After cooling, the remains of the media are removed, then the cups and test tubes are soaked, boiled in tap water and washed, followed by rinsing with distilled water.

Pre-prepared ENDO nutrient agar is poured into Petri dishes and set to solidify.

2.5 Membrane filter method

Method for determining the number of E.coli cells per unit volume of liquid (coli-index); the essence of the method consists in filtering the analyzed liquid through membrane filters that trap bacteria, after which these filters are placed on a solid nutrient medium and the bacteria colonies grown on it are counted.

Membrane filter preparation

Membrane filters should be prepared for analysis in accordance with the manufacturer's instructions.

Preparation of the filter apparatus

The filter apparatus is wiped with a cotton swab moistened with alcohol and flambéed. After cooling, a sterile membrane filter is placed on the lower part of the filter apparatus (table) with flambéed tweezers, pressed with the upper part of the device (glass, funnel) and fixed with a device provided for by the design of the device.

In the membrane filter method, a certain amount of water is passed through a special membrane with a pore size of about 0.45 µm.

As a result, all bacteria present in the water remain on the membrane surface. After that, the membrane with bacteria is placed on a special nutrient medium (ENDO). After that, the Petri dishes were turned over and placed in a thermostat for a certain time and temperature. Common coliform bacteria (CBC) were incubated at a temperature of 37 +/- 1°C for 24-48 hours. hours.

The medium is photosensitive. Therefore, all inoculated cups are protected from light.

During this period, called the incubation period, the bacteria have the opportunity to multiply and form well-defined colonies that are already easy to count.

At the end of the incubation period, the crops are viewed:

a) the absence of microbial growth on the filters or the detection of colonies on them that are not characteristic of bacteria of the intestinal group (spongy, membranous with an uneven surface and edge), allows at this stage of the analysis to complete the study (18-24 hours) with a negative result for the presence of intestinal rods in the analyzed volume of water;

b) if colonies characteristic of Escherichia coli (dark red with or without a metallic sheen, pink and transparent) are found on the filter, the study is continued and microscoped.

If the growth of round colonies of crimson color with a metallic sheen with a diameter of 2.0-3.0 mm - Escherichia coli 3912/41 (055: K59);

If the growth of round colonies of crimson color with a diameter of 1.5-2.5 mm with a fuzzy metallic sheen - Escherichia coli 168/59 (O111:K58)

2.6 Accounting for results

After an incubation period of 48 hours for common coliform bacteria and 24 hours for thermotolerant bacteria, colonies grown on plates are counted.

Colonies that grew on the surface as well as in the depth of the agar were counted using a loupe with a fivefold magnification or a special device with a magnifying glass. To do this, the dish is placed upside down on a black background and each colony is marked from the side of the bottom with ink or glass ink.

To confirm the presence of OKB, examine:

all colonies if less than 5 colonies grew on the filters;

at least 3 - 4 colonies of each type.

To confirm the presence of TKB, all typical colonies are examined, but no more than 10.

Count the number of colonies of each type.

Calculation and presentation of results.

The result of the analysis is expressed as the number of colony forming units (CFU) of common coliform bacteria in 100 ml of water. To calculate the result, sum the number of colonies confirmed as total coliforms grown on all filters and divide by 3.

Since this method of water analysis involves only determining the total number of colony-forming bacteria of different types, its results cannot unambiguously judge the presence of pathogenic microbes in the water. However, a high microbial count indicates a general bacteriological contamination of water and a high probability of the presence of pathogenic organisms.

Each selected isolated colony is examined for Gram affiliation.

Gram stain

Gram stain is of great importance in the taxonomy of bacteria, as well as for the microbiological diagnosis of infectious diseases. A feature of the Gram stain is the unequal ratio of various microorganisms to the dyes of the triphenylmethane group: gentian, methyl or crystal violet. Microorganisms belonging to the group of gram-positive Gram (+), such as staphylococci, streptococci, give a strong connection with the indicated dyes and iodine. Stained microorganisms do not discolour when exposed to alcohol, as a result of which, with additional Gram (+) fuchsin staining, microorganisms do not change their originally adopted purple color. Gram-negative Gram (-) microorganisms (bacteroids, fusobacteria, etc.) form a compound that is easily destroyed under the action of alcohol with gentian crystal or methylene violet and iodine, as a result of which they become discolored and then stained with fuchsin, becoming red.

Reagents: carbolic solution of gentian violet or crystal violet, aqueous solution of Lugol, 96% ethyl alcohol, water-alcohol solution of fuchsin.

Coloring technique. A piece of filter paper is placed on a fixed smear and a carbolic solution of gentian violet is poured on it from 1/2 to 1 minute. The dye is drained and, without washing off, Lugol's solution is poured for 1 minute. Drain Lugol's solution and rinse the drug in 96% alcohol for 1/2 to 1 minute until the dye stops leaving. Washed with water. Additionally stain with diluted fuchsin from 1/2 to 1 minute. Drain the dye, wash and dry the drug.

3. Research results

.1 Microbiological analysis of water in Lake Pechersk (for example,E. coli) in the spring period (May) of the 2009-2013 study.

As a result of three-time water intake at two sampling points (PZ1 - at the beginning of the beach, near the dam, PZ2 - the end of the beach, boat station), we calculated the average indicators of OKB and TKB, the results of which are presented in Table 3.1.

Table 3.1. Average indicators of OKB and TKB in the water of Lake Pechersk for May 2013

The index of E.coli bacteria content according to OKB at the beginning and at the end of May in TZ1 (near the dam) does not differ, amounting to 195 CFU / cm 3, which is 3.3 times less compared to the water sample taken in TK2 (near the boat station ) at the beginning of May and 4.3 times more at the end of May.

The study of the dynamics of the content of Escherichia coli in the water of Lake Pechersk for May 2013, according to the SES data, confirmed the correctness of our own research and showed that the TCA indicator in TK2 is 3.4 times higher than in TK1 (according to our own results, 3.3 times more).

The study of changes in OKB and TKB indicators for the month of May from 2009 to 2013. showed a wide variation in indicators, which is clearly shown in Figures 3.1 - 3.2

Analysis of data from the health care institution "Mogilev Zonal Center for Hygiene and Epidemiology" for the beginning of May 2008-2013.

At the end of the data analysis for the beginning of May 2008-2013, we found that in 2008-2012 there were more OKBs in TK1 than in TK2.

Analysis of data from the health care institution "Mogilev Zonal Center for Hygiene and Epidemiology" for the end of May 2008-2013.

Common coliform bacteria according to SanPiN must be absent in 100 ml of drinking water

According to SanPiN, thermotolerant fecal coliforms should be absent in 100 ml of the studied drinking water.

For open reservoirs, according to the Design Bureau, no more than 500 CFU per 100 ml of water, according to the TKB, not more than 100 CFU per 100 ml of water.

The presence of Escherichia coli in the water confirms the fecal nature of the contamination.

According to the results of measurements in the summer low water, coliform bacteria are present in small quantities, usually from one hundred to several hundred units, and only during periods of floods briefly increase to 1000 or more units.

Low values ​​in summer may be due to several factors:

) intense solar radiation, which is detrimental to bacteria;

) increased pH values ​​in summer (usually pH > 8 in summer, in winter< 8) за счет развития фитопланктона;

) the release of phytoplankton metabolites into the water, which inhibit the bacterial flora.

With the beginning of the autumn-winter season, these factors are significantly weakened, and the number of bacteria rises to the level of several thousand units. The greatest extremes occur during periods of snowmelt, especially during floods, when the meltwater washes away bacteria from the catchment surface.

The total number of colony-forming bacteria in the middle of summer is lower than in the spring-autumn period, which is associated with intense solar radiation, which is detrimental to bacteria.

Rivers in urban areas are often natural recipients of household and fecal sewage, so the number of microbes sharply increases within the boundaries of settlements. But as the river moves away from the city, the number of microbes gradually decreases, and after 3-4 tens of kilometers it again approaches its original value.

The largest number of microbes in open water bodies is found in the surface layers (in a layer of 10 cm from the water surface) of coastal zones. With distance from the coast and increasing depth, the number of microbes decreases.

River silt is richer in microbes than river water. There are so many bacteria in the very surface layer of silt that a kind of film is formed from them. This film contains many filamentous sulfur bacteria, iron bacteria, they oxidize hydrogen sulfide to sulfuric acid and thus prevent the inhibitory effect of hydrogen sulfide (fish death is prevented).

Conclusion

coli bacterium pathogen

To find and identify E. coli, a microbiological analysis of samples was carried out for the beginning of May 2013. A statistical analysis of the data of the health care institution "Mogilev Zonal Center for Hygiene and Epidemiology" for the beginning of May 2008-2012 was also carried out.

At the end of the analysis, it was found that the number of bacteria of the Escherichia coli group calculated by us does not exceed the permissible norm.

At the end of the statistical analysis of the data of the health care institution "Mogilev Zonal Center for Hygiene and Epidemiology" for 2008-2012, it was found that coliform bacteria are present in small quantities in the summer low water period. The total number of colony-forming bacteria in the middle of summer is lower than in the spring-autumn period, since intense solar radiation, which is detrimental to bacteria, and with the onset of the autumn-winter season, the number of bacteria rises to the level of several thousand units. The greatest extremes occur during periods of snowmelt, especially during floods, when the meltwater washes away bacteria from the catchment surface.

Bibliography

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Dolgonosov B.M., Dyatlov D.V., Suraeva N.O., Bogdanovich O.V., Gromov D.V., Korchagin K.A. Aqua CAD information modeling system - a tool for managing technological regimes at a waterworks // Water Supply and Sanitary Engineering. 2003. No. 6. pp. 26-31.

Dolgonosov B.M., Khramenkov S.V., Vlasov D.Yu., Dyatlov D.V., Suraeva N.O., Grigorieva S.V., Korchagin K.A. Forecast of indicators of water quality at the inlet of a waterworks // Water Supply and Sanitary Engineering 2004. No. 11. pp. 15-20.

Kochemasova Z.N., Efremova S.A., Rybakova A.M. Sanitary microbiology and virology. M.: Medicine, 1987.

SanPiN 2.1.5.980-00. Water disposal of populated areas, sanitary protection of water bodies. Hygienic requirements for the protection of surface waters.

SanPiN 2.1.4.1074-01. Drinking water. Hygienic requirements for water quality of centralized drinking water supply systems. Quality control.

MUK 4.2.1018-01. Control methods. Biological and microbiological factors. Sanitary and microbiological analysis of drinking water.

Table of contents of the subject "Sanitary and Microbiological Study of the Soil. Microflora of Reservoirs.":

Between groups of sanitary-indicative microorganisms there are no clearly defined boundaries. Some microorganisms are indicators of both faecal and oral contamination. Some are indicators of self-purification processes. In this regard, all SMPs are regarded as indicators of biological pollution.

Group A of sanitary-indicative microorganisms. Includes inhabitants of the intestines of humans and animals. Microorganisms are regarded as indicators of faecal contamination. It includes BGKP - Escherichia, Enterococcus, Proteus, Salmonella. Also included in group A are sulfite-reducing clostridia (Clostridium petfringens and others), thermophiles, bacteriophages, bacteroids, Pseudomonas aeruginosa, candida, akinetobacter and aeromonads.

Group B of sanitary-indicative microorganisms. Includes inhabitants of the upper respiratory tract and nasopharynx. Microorganisms are regarded as indicators of oral contamination. It includes green, a- and (3-streptococci, staphylococci (plasma-coagulating, licitinase-positive, hemolytic and antibiotic-resistant; in some cases, the type of Staphylococcus aureus is also determined).

Group C of sanitary-indicative microorganisms. Includes saprophytic microorganisms living in the external environment. Microorganisms are regarded as indicators of self-purification processes. It includes proteolytic bacteria, ammonifying and nitrifying bacteria, some spore-forming bacteria, fungi, actinomycetes, cellulose bacteria, bdellovibrios, and blue-green algae.

The main groups of sanitary-indicative microorganisms

To the main sanitary-indicative microorganisms include BGKP, enterococci, proteas, salmonella, Clostridium perfringens, thermophilic bacteria and bacteriophages of enterobacteria (coliphages).

Bacteria of the Escherichia coli group

coli marked the beginning of the entire SPM group. The BGKP includes various representatives of the Enterobacteriaceae family. Depending on the purpose and object of the study, various requirements are imposed on the sanitary-indicative BGKP. They are conditionally divided into three subgroups and, under various circumstances, the fact of their presence is used for bacteriological characteristics of an object or substrate.

Subgroup I Escherichia coli includes BGKP, which are trying to identify, but which should not be in the study of objects and substrates that are "clean" in nature or become pure as a result of their processing (for example, thermal). The group of objects with such properties includes the following. Drinking (artesian, tap chlorinated, well) and distilled water (taken from a distiller or pipeline). Thermally processed food products (cutlets, sausages, fish, etc.). Analyze samples taken from the thickness of the product.

Milk(taken from the pasteurizer before entering the milk pipelines), soups, sauces, compotes, main courses (selected from boilers). Washouts selected during the control of the effectiveness of disinfection treatment in due time (not earlier than 45 minutes and not later than 1 hour after treatment).

Bacteria of this subgroup of Escherichia coli ferment lactose and glucose or only glucose to gas at 37 ° C and do not show oxidase activity. This subgroup includes Escherichia ha//, Klebsiella, Citrobacter, Enterobacter and other members of the Enterobacteriaceae family. Their presence is allowed in objects that do not belong to the category of "clean".

Subgroup II Escherichia coli includes CGBs indicating temporally undetermined faecal contamination. Microorganisms ferment lactose and glucose to acid and gas at 43-44.5 °C. This subgroup includes bacteria (E. coli, Klebsiella, citrobacter, enterobacter, etc.) that have retained the ability to form gas at elevated temperatures. Similar requirements are imposed on BGKP if it is impossible to protect the substrate from contamination. At the same time, one should limit oneself to determining only indicators of epidemiological distress. Such objects include: water from open reservoirs, wastewater, soil and all food products for which there is a high risk of contamination after heat treatment. In such cases, solid food products (surface layer), liquid food products, second and third dishes for distribution, washings from equipment and utensils are examined. Crops are cultivated at 43-44.5 °C. E. coli is differentiated from other bacteria by its ability to ferment lactose and glucose or only glucose.

Subgroup III Escherichia coli includes CGBs indicating fresh faecal contamination. A distinctive feature of this group of bacteria is the ability to break down lactose to gas at 43-44.5 "C.

Coliform bacteria are always present in the digestive tract of animals and humans, as well as in their waste products. They can also be found on plants, soil and water, where contamination is a major problem due to the possibility of infection by diseases caused by various pathogens.

Harm to the body

Are coliform bacteria harmful? Most of them do not cause disease, however, some rare strains of E. coli can cause serious illness. In addition to humans, sheep and cattle may also be infected. It is worrying that contaminated water, in its external characteristics, is no different from ordinary drinking water in taste, smell and appearance. Coliform bacteria are found even in which is considered to be flawless in every sense. Testing is the only reliable way to find out about the presence of pathogenic bacteria.

What happens when discovered?

What to do if coliform bacteria or any other bacteria are found in drinking water? In this case, repair or modification of the water supply system will be required. When used for disinfection, mandatory boiling is provided, as well as retesting, which can confirm that the contamination was not eliminated if it was thermotolerant coliform bacteria.

indicator organisms

Common coliforms are often referred to as indicator organisms because they indicate the potential presence of pathogenic bacteria in water, such as E. coli. While most strains are harmless and live in the intestines of healthy humans and animals, some can produce toxins, cause serious illness, and even death. If pathogenic bacteria are present in the body, the most common symptoms are gastrointestinal upset, fever, abdominal pain, and diarrhea. Symptoms are more pronounced in children or older family members.

safe water

If there are no common coliform bacteria in the water, then it can be assumed with almost certainty that it is microbiologically safe to drink.
If they were found, then it would be justified to conduct additional tests.

Bacteria love warmth and moisture.

Temperature and weather conditions also play an important role. For example, E. coli prefers to live on the surface of the earth and loves warmth, thus coliform bacteria in drinking water appear as a result of movement in underground streams during warm and humid weather conditions, while the smallest number of bacteria will be found in the winter season.

Impact chlorination

To effectively destroy bacteria, chlorine is used, which oxidizes all impurities. Its amount will be affected by water characteristics such as pH and temperature. On average, the weight per liter is approximately 0.3-0.5 milligrams. It takes approximately 30 minutes to kill common coliform bacteria in drinking water. Contact time can be reduced by increasing the dose of chlorine, but this may require additional filters to remove specific tastes and odors.

Harmful ultraviolet light

Ultraviolet rays are considered a popular disinfection option. This method does not involve the use of any chemical compounds. However, this agent is not used where the total coliform bacteria exceed one thousand colonies per 100 ml of water. The device itself consists of a UV lamp surrounded by a sleeve of quartz glass through which a liquid flows, irradiated with ultraviolet light. The raw water inside the apparatus must be completely clean and free from any visible contaminants, blockages or turbidity to allow exposure of all harmful organisms.

Other cleaning options

There are many other treatment methods used to disinfect water. However, they are not recommended as long term for various reasons.

• Boiling. At 100 degrees Celsius for one minute, bacteria are effectively killed. This method is often used to disinfect water during emergencies or when needed. This takes time and is an energy intensive process and is generally only applied in small amounts of water. This is not a long-term or permanent option for water disinfection.
• Ozonation. In recent years, this method has been used as a way to improve water quality, eliminate various problems, including bacterial contamination. Like chlorine, ozone is a strong oxidizing agent that kills bacteria. But at the same time, this gas is unstable, and it can only be obtained with the help of electricity. Ozone units are generally not recommended for disinfection because they are much more expensive than chlorination or UV systems.
• Iodization. The once popular disinfection method has recently been recommended only for short-term or emergency water disinfection.

thermotolerant coliform bacteria

This is a special group of living organisms that are able to ferment lactose at 44-45 degrees Celsius. These include the genus Escherichia and some species of Klebsiella, Enterobacter and Citrobacter. If foreign organisms are present in the water, this indicates that it has not been sufficiently cleaned, re-contaminated, or contains nutrients in excess. When they are detected, it is necessary to check for the presence of coliform bacteria that are resistant to elevated temperatures.

Microbiological analysis

If coliforms were found, then this may indicate that they got into the water. Thus, various diseases begin to spread. In contaminated drinking water, strains of Salmonella, Shigella, Escherichia coli and many other pathogens can be found, ranging from mild digestive tract disorders to severe forms of dysentery, cholera, typhoid fever and many others.

Household sources of infection

The quality of drinking water is monitored, it is regularly checked by specialized sanitary services. And what can an ordinary person do to protect himself and protect himself from unwanted infection? What are the sources of water pollution in the home?

1. Water from the cooler. The more people touch this device, the more likely it is that harmful bacteria will enter. Studies show that the water in every third cooler is simply teeming with living organisms.
2. Rainwater. Surprisingly, the moisture collected after the rain is a favorable environment for the development of coliform bacteria. Advanced gardeners do not use such water even for watering plants.
3. Lakes and reservoirs are also at risk, since all living organisms multiply faster in stagnant water, and not just bacteria. An exception is the oceans, where the development and spread of harmful forms is minimal.
4. Pipeline condition. If the sewers have not been changed and cleaned for a long time, this can also lead to trouble.

Who are BGKP and where do they live

GOST for coliform bacteria

An interstate standard has been developed for methods for detecting and determining the number of coliform microbes. This GOST ensures food safety. Any product included in the GOST list must undergo laboratory tests. After laboratory tests proving the acceptable values ​​of BGKP, the products are sold. Mandatory research is subject to:

• Water.
• Canned food.
• Meat products.
• Pet food.
• Crockery and equipment.

It is important to know that GOST does not apply to milk and dairy products. All milk and other dairy products purchased in bulk or in bulk must be pasteurized to kill coliforms. Pasteurization - heating up to + 80⁰С for 30 minutes.

GOST obliges to monitor the sanitary and bacteriological state of water. Water intake to determine the presence of BGKP is made from:

• City water supply system.
• Open water reservoirs (rivers, seas, reservoirs).
• Sources of drinking water (wells, springs).
• Swimming pools.
• Wastewater (before and after treatment).

Wash your hands!

All types of bacteria of the Escherichia coli group die when boiled or pasteurized. Escherichia and salmonella toxins will not remain in milk, meat and water at temperatures above + 60⁰С. Door handles or table surfaces should be wiped with a disinfectant solution. Coliform bacteria are instantly killed by alcohol or another antibacterial agent. But the most reliable way to prevent intestinal diseases according to GOST and life experience is hand washing with soap. The alkaline environment of soap destroys the walls of microbes. If it is not possible to wash your hands, for example, on the road, use disinfectant wet wipes or hand gel.