Ploskirev's medium (also called bactoagar F) is a breeding ground for the cultivation of certain microorganisms, mainly shigella and salmonella. Infected materials are used as a source for it: urine, bile, stool.
History
Nikolai Ivanovich Ploskirev - Soviet microbiologist and Honored Doctor of the RSFSR. He was born in 1869 and died in 1948, having worked most of his life in his hometown of Tomsk.
In 1888, he, a future scientist, did not graduate from the sixth grade of the school, enlisted in the military. He could finish his studies only in 1892, after which he passed the exams for admission to the Imperial University of Tomsk, studied at the Faculty of Medicine. In 1898 he received the title of doctor.
Ploskirev participated in the Russian-Japanese war, and after it worked in the Tomsk hospital. In 1910, he was appointed to the post of head of the Tomsk Dermatovenerologic Hospital and for thirty years was its permanent head. Also, the scientist was one of the founders of the Dermatovenerologic Dispensary in Tomsk.
Nikolai Ivanovich Ploskirev wrote a series of works devoted to the fight against sexually transmitted diseases.
Composition
Ploskirev's medium is a material for the cultivation of intestinal bacteria, which means it must contain several types of nutrients. It is produced in dry form.
A rather large share in its total mass is pancreatic sprat hydrolyzate (10.4 g / l). Sodium disodium hydrogen citrate (8.5 g / l) accounts for slightly less. It also contains dry nutritious broth and milk sugar (8.62 and 7.3 g / l).
The second name of Ploskirev's medium is Bactoagar J. Its composition contains agar in the amount of 6.94 g / l. The content of anhydrous serovatistisodium sodium is 5.1 g / l. There is dehydrated disodium phosphate - 2.1 g / l, sodium bile salts of about 3.46 g / l, and soda ash - 2.4 g.
Less than a gram contains indicators - neutral red (0.05) and brilliant green (0.0002). The iodine content is 0.13.
Application
The study of the biochemical processes of bacteria plays a very important role in the diagnosis of diseases that cause anaerobic organisms. There are hundreds of species in this family. They are almost identical in morphology and cultural properties, and the most reliable way to distinguish them from each other is to study biochemistry.
The study begins with the inoculation of pathological material into the nutrient medium. It is differential pathological for the intestinal group of bacteria, and in addition to meat and peptone agar, it also includes indicator and lactose. The ability to process lactose is an important sign of the differentiation of enterobacteria. If this is detected, then the pH shifts to the side of acidity and an indicator lights up, which stains the colony.
Other nutrient media may be used in other countries, but their mechanism of action corresponds to the three most common in Russia - the environments of Endo, Levin and Ploskirev's environment.
Method Description
Abroad, an analogue of Ploskirev’s environment is the so-called MacConkey Agar. In the finished form, the solution is transparent, has a light pinkish-yellow hue. Colonies capable of processing lactose in the Ploskirev environment give a red (lingonberry) color. If the bacterium cannot process it, the colony is either colorless or has a weak color.
Due to the fact that the Ploskirev medium contains inhibitor substances (brilliant green, bile salts, iodine), it actually completely inhibits the growth of gram-positive flora, and on the first day it significantly inhibits the growth of eshechiria, as well as another usually associated microflora.
Second phase
Next, the colony of interest is selected, and it is seeded on the environment of primary differentiation and accumulation. Sowing media should contain several substrates. The bacterial culture must detect enzymatic activity in relation to them, in addition, the media are located in test tubes so that there are two sections:
- the one on which the agar is mowed;
- with a column.
The colony of interest to the researcher is populated on the beveled part with a dense stroke, and in the column with an injection. At this stage, the environments of Ressel, Kligler or Olkenitsky are used. As the differential diagnostic is used and the Ploskirev environment.
Microbiology and pathogens
Agar Ploskirev is an important factor in the detection of infections caused by enterobacteria. For example, colonies of microorganisms that are the causative agents of bacterial dysentery are grown and differentiated on it. These are anaerobic organisms belonging to the genus Schigella.
Like everyone who is part of the family of intestinal bacteria, shigella are in the form of sticks, two micrometers in size. They do not form capsules and spores, do not have flagella - this allows them to be distinguished from salmonella, which, in turn, are mobile. They grow well on the simplest environments, at temperatures slightly above room temperature (35-37 ° C) and 7.4 pH. By the nature of growth, the colonies do not differ from salmonella.
As mentioned above, microorganisms may have practically no morphological differences, but vary significantly in the biochemical processes of life. Thus, excluding Shigella Newcastle, fermentation of carbohydrates results in the formation of acid without gas. With the exception of Shigella Sonne, they cannot ferment lactose, but break down glucose. Also, their key features include the fact that they can reduce nirates to nitrites. Their colonies do not break down urea, and gelatin is not liquefied in the nutrient medium.
Seed collection and preparation
To detect the causative agents of dysentery, microbiological studies of infected media, that is, the feces of the patient, are necessary. After receiving the material, it is necessary to do the sowing as soon as possible. If this is not possible, the source must be placed in a preservative substance - phosphate buffer mixture or glycerin. At a temperature of 4 ° C they can be stored for no more than a day. The collection of material must be done using a rectal glass tube, which is inserted into the rectum.
For the study, purulent-mucous pieces are selected from the material, which should be washed in two to three test tubes of isotonic sodium chloride solution.
The application of pathogenic bacteria to the Ploskirev medium is carried out with a glass spatula. It is necessary to rub it into the agar in a small area, then tear off the spatula from the medium, and rub the residual material into the unseeded surface. If sowing is done in several cups, then in each of them you need to sow a new portion.
If there are no mucopurulent pieces in the secretions taken for analysis, this does not mean that pathogenic microorganisms are not present there. In this case, it is necessary to emulsify the feces in 10 ml of sodium chloride solution (concentration 0.85%), then sow one or two drops on Ploskirev's medium. Non-emulsified stool can be seeded in selenite broth. It is also used if instead of feces there are vomit or flushing water.
Microbiological diagnosis
At the first stage of microbiological research , the pathogenic bacteria are inoculated in two cups in order to then observe how shigella grow. On Ploskirev’s environment, one sowing is performed, the second on Levin or Endo.
Due to the emergence of Shigella strains resistant to antibiotics, chloramphenicol is added to the media. Then, during the day, incubation takes place in a thermostat at a temperature of 37 ° C.
On the second day, you need to study the grown colonies. Those that did not grow colorless on the differential diagnostic medium must be weeded out on Ressel's medium or a short "motley row". Further research is carried out according to the algorithm of primary crops on them. Shigella colonies on Ploskirev's medium grow in the form of transparent, colorless and small columns. They are of two types:
- flattened with serrated edges;
- roundish, convex, with a characteristic moist shine.
Three to four colonies must be microscopic and examined for motility. If the latter is not found, they are transferred to the Olkenitsky environment to highlight a pure culture. In the absence of characteristic Shigella colonies or if no growth is observed at all, it is necessary to sow from the selenite broth on Endo or Ploskirev agar. If typical colonies are present in sufficient quantity, an indicative agglutination reaction is set up. A mixture of Sonne and Flexner serums is used, the reaction takes place on glass.
Further research
On the third day, changes are noted that have occurred in the colonies on the environment of Ressel. If there is a culture that does not decompose lactose, and ferments glucose to form acid, it is separated and examined, microscopy is performed, and also the sowing on the "motley row" is carried out, but this time deployed. The agglutination reaction is also put in order to serological identification.
On the last day, a conclusion can be made on the basis of changes in the "motley row" (whether carbohydrate fermentation occurs), as well as the results of the agglutination reaction.
Storage and preparation
The preparation of Ploskirev agar is as follows:
- 55 g of dry matter is stirred in a liter of distilled water.
- Boil for two to three minutes, until the agar is completely melted.
- Poured into Petri dishes (sterility optional) with a layer of 5-6 mm.
- The cups are left open for an hour and a half at room temperature (18-25 degrees). After this time, the medium will congeal sufficiently and dry.
Ploskirev's medium is photosensitive and hygroscopic. The powder must be stored in sealed packaging, the relative humidity in the room should not exceed 60%. The optimum storage temperature is from 2 ° C to 25 ° C. It is necessary to observe the storage rules given, otherwise inaccurate research results are possible.