Bacterial Counts 



Coliform counts were made by a widely used modification [1] of the 

 Standard Total Coliform Membrane Filter Procedure [2]. The counts were 

 made with field monitoring equipment developed and supplied by the 

 Millipore Corporation, Bedford, MA. The equipment included a syringe with 

 two one-way valves. With the syringe, water samples were drawn through 

 membrane filters having pores sufficiently small (0.45 millimicron) to 

 retain most microorganisms. The membrane filters were mounted in 

 ''Millipore Field Monitors'' which are sterile, disposable, plastic 

 devices, serving both as filter holders and culture chambers. The 

 membranes are supported on absorbent pads for retaining culture medium. 



The water samples to be counted were serially diluted (Figure 5) 

 in test tubes, each containing 9 ml of an autoclaved 3:1 mixture of 

 distilled water and seawater. The diluted fractions of the samples were 

 then drawn through the membrane filters contained in the Millipore Field 

 Monitors (Figure 6). In this step, all of the bacteria in the diluted 

 fractions were deposited on the surfaces of the membrane filters. The 

 absorbent pads on which the membrane filters were mounted were each 

 moistened with 0.8 ml of sterile ' 'MF-Endo Broth,'' a proprietary medium 

 prepared specifically for the isolation and identification of coliform 

 bacteria. The monitors were then closed and incubated at 35 C for 24 

 hours. 



Each coliform bacterium on the surface of the membrane filters 

 multiplies many times in the course of 24 hours and ultimately forms 

 a macroscopically visible aggregate or colony. Most other bacteria species 

 do not multiply on the selective MF-Endo Broth. The number of colonies 

 which develop is assumed to be the same as the number of coliform bacteria 

 in the serially diluted fraction of the water sample that was filtered. 

 Because most samples were counted in duplicate and several dilutions 

 were made of each, several estimates were obtained of the number of 

 microorganisms in a unit volume of each sample. When the results were 

 averaged, greatest reliance was placed on plate counts in the range of 

 20 through 80. 



EXPERIMENTAL RESULTS 



Effect of Sea Salts on the Survival of E. coli 



In preparation for experiments which were to be conducted at sea, 



several preliminary tests or exercises were conducted in the laboratory. 



In the first exercise, one BACT-CHEK disc was placed in a test tube 



containing a 3:1 mixture of distilled water and seawater. The test tube 



o 

 was incubated for 2 hours at 35 C. At the end of that time the disc was 



sufficiently soft to disintegrate when the test tube was shaken. The 



bacteria were then uniformly distributed throughout the tube. The 



contents of the tube was added to a 1 -liter flask filled with filtered 



autoclaved seawater. Bacterial counts were made at various intervals and 



the following counts were obtained: 



