368 Marine Microbiology 



fluids from several tubes of one kind were pooled and samples 

 were counted in a hemocytometer. From young populations, ten 

 samples were counted and from heavier ones, four samples were 

 counted. When fish tissue culture cells were used in place of KB 

 cells as food source, the cultures had to be washed before inocula- 

 tion because they were grown in Eagle's medium with increased 

 NaCl concentrations. In order to find a comparative basis for the 

 quantitation of food cells of different origin we have started to 

 carry out protein determinations of a control set of feeder cul- 

 tures at the beginning of the growth experiment. This method, 

 however, needs further elaboration. 



Saprozooic Feeding Method 



Two factors have been under investigation in the saprozooic 

 feeding system. The influence of temperature on the growth rate 

 and final yields has been studied with cultures grown at 20 C, 

 29 C and 37 C. Adaptation to media of various salinities was 

 also tested. The array of free amino acids and vitamins present 

 in Eagle's medium did not support growth of our strains. There- 

 fore Melnick's medium, designed for primary monkey kidney 

 cells, was chosen. It consists of Hank's balanced salt solution sup- 

 plemented with 10 per cent lactalbumin and 5 per cent calf 

 serum and an addition of penicillin (200 units/ml) and strepto- 

 mycin (0.10 mg/ml). Before each growth experiment the ciliates 

 were preadapted to the media or temperatures for one week. 

 Fifty thousand animals were introduced into 50 ml aliquots of 

 media. At chosen time intervals, 0.5 ml samples were removed 

 from two sister cultures and counts were made. Depending on the 

 densities of the populations, four to ten counts were carried out. 



Virus Studies 



Since no suitable laboratory strain of a marine fish virus 

 was at hand at this stage of our investigations we had to mak^ 

 use of mammalian viruses for our explorative and methodological 

 studies. The ease of growing and of measuring infectivity titres 

 of poliovirus in KB— which cells sei've as standardized food source 

 —suggested its usefulness for our puipose. Moreover, polio strain 

 Mahoney was found to have a remarkable temperature stability 

 at a range from 20 C to 30 C provided the experimental periods 



