cally controlled incubator at a preselected temperature, usually 26 °C. 

 One drop of the solution of ^C-labeled protein hydrolysate having a 

 measured activity under a Geiger counter of the order of 0.2 x 10° imp/min 

 is added to each of the test-tubes by means of a Pasteur pippette, and 

 the test-tubes are incubated for 2 hours. Their contents are then fixed 

 with 0.25 ml of formalin and filtered through membrane filters with a 

 pore size 0.2-0.3 mm. After fixation and filtering, 5 ml of physiological 

 solution is filtered to remove the excess portions of the labeled pre- 

 paration. The filters are dried and the radioactivity of bacteria is mea- 

 sured under the Geiger counter. The final dilution in which the radio- 

 activity differs markedly from the control serves as an indicator of the 

 limit of dilution for bacterial reproduction. Using this methodology and 

 various classes of radioactive substances (e.g., phenol) the presence of 

 microorganisms and their number in the water samples may be determined. 



Number of Saprophytic Bacteria 



The number of saprophytic bacteria is the most reliable and sensitive 

 indicator of water pollution by organic substances of household origin. 

 This is a classic method used for about 100 years by sanitary organiza- 

 tions. It was proposed by Koch and has been used for counting of bacteria 

 in water by a number of different workers. The method of determination 

 of the number of saprophitic bacteria is quite simple. In the USSR, 

 standard dry nutrient medium (FPA) is made from fish flesh to which pep- 

 tone, sodium chloride and 15 percent agar-agar are added. 



The medium to be inoculated is prepared in 50-100, or 200 ml flasks 

 depending on the quantity required. The water is taken into sterile 

 glassware by special samplers. The most simple model is the sampler of 

 Meier-Frantsev (Romanenko and Kuznetsov, 1974). Inoculation must be made 

 within one half of an hour after sampling. Samples may be stored in re- 

 frigerator or vacuum flask at low temperature for no longer than a day. 



Inoculation may be of either the surface or depth type. In the former 

 case, the FPA should be melted. In laboratory this is best done in 

 boiling water. The flask with FPA is placed into boiling water until all 

 the medium is molten (not even smallest lumps of solid medium must be 

 left, otherwise the analysis will be spoiled). The medium may also be 

 melted on an open flame of a burner or on electric plate, but the proce- 

 dure must be carefully watched. The medium is then cooled to 40 °C. In 

 practice, microbiologists apply the flask to the cheek, if the medium 

 does not burn, it may be poured into Petri dishes. 



For deep inoculation, the water to be tested is added into sterile 

 Petri dishes by sterile pippettes, then the FPA is poured and all is 

 thoroughly mixed. In this case, bacteria grow throughout the medium. 

 The colonies grown in the depths are physically smaller in size. For sur- 

 face inoculations, the medium is poured into the dishes, and after it 

 solidifies 0.5 to 1 ml of inoculum is added upon the surface and spread 

 over with a help of sterile glass spatula. The dishes are incubated at a 

 room temperature for 10 days. Then the number of colonies are counted, 

 and estimations are made with due consideration to the dilution. 



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