Supprasion of Bacterial Growth by Sea Water 573 



water, rather than bacterial growth. In the present study, care- 

 ful control of all growth parameters, with the source of sea waters 

 as a variable, demonstrates the sensitivity of the test organisms to 

 differences in the water. 



MATERIALS AND METHODS 



Samples of pelagic sea water were collected at various depths 

 with plastic samplers having a capacity of tln'ee to four liters 

 (20). Bottom water samples were obtained by pouring off the 

 water above the sediment collected by a Phleger Bottom Sampler 

 ( 12 ) , care being taken not to disturb the sediment or to allow 

 any sediment particles to pass into the sample. Surface sea water 

 was obtained off the end of the Scripps Institution Pier (more 

 than 1000 feet beyond high-tide level) with a clean plastic 

 bucket. 



The samples were dispensed immediately into sterile, Pyrex 

 reagent bottles, and were examined in the laboratory within four 

 hrs after collection. They were filtered through Whatman No. 2 

 paper in a Buchner funnel to remove large organisms and partic- 

 ulate matter, and were then filter-sterilized by passage through 

 sterile Morton (UF) fritted-glass filters. Autoclaving was found 

 to diminish the inhibitory properties of sea water. 



The test bacteria. Staphylococcus aureus (ATCC 6538), 

 Bacillus subtilis, and Micrococcus lijsodeikticus (ATCC 4698), 

 have been maintained in the laboratory of Dr. C. E. ZoBell for 

 a period of ten years or longer. Escherichia coli was isolated by 

 Mrs. L. R. Berger in 1957 while working in this laboratory, and 

 its identity was confirmed on Levine's EMB agar medium. Ser- 

 ratia marinorubra was isolated and maintained in this laboratory 

 (24). 



The sea water was tested in three ways with artificial sea 

 water (9) used as the control: 



(a) A sea-water concentration of 18 per cent (two ml 

 added to nine ml of stock nutrient Medium I) was tested with 

 the five test bacteria (0.15 ml inocula of twelve hour nutrient 

 broth cultures grown at 30 C). The final concentiations per liter 

 of nutrients were approximately: yeast extract (Difco), 0.82 g; 

 proteose peptone (Difco), 1.64 g; glucose, 8.2 g; K,HP04, 0.82 



