agar) into duplicate Erlenmeyer flasks containing approximately 150 

 ml of sterile 0.1^ peptone solution in aged sea water. One flask 

 was incubated at 30° C, and the other at 25° C. The culture incubated 

 at 30° C . was deep orange after 24 hours and the one incubated at the 

 lower temperature was yellow. The more intensely pigmented culture 

 was used because it probably contained the greater concentration of 

 bacteria. The ratio of bacterial culture volume to sea water volume 

 varied from a maximum of 33.0 ml of 24-hour culture to 1.0 liter of 

 sea water, which is about the same as used by Bein, to a minimum of 

 one-hundredth of this ratio. Sea water and sterile 0,1^ peptone 

 solution were used as control materials. Each experimental container, 

 2-liter beaker, received 1.0 liter of sea water which was aerated 

 continuously. Two mullet (3 to 4 in. long) were placed in each 

 container. The test fish, which had been maintained in the laboratory 

 for several days, were acclimated in the experimental containers for 

 about 24 hours before beginning the study. The water which became 

 cloudy in all containers by the end of the acclimation period was 

 replaced with fresh sea water shortly before the test materials were 

 added. Samples were collected for bacterial counts about 30 minutes 

 after the experiment was begun. These samples were refrigerated -g- 

 to 2 hours before preparing the pour-plates . 



All fish were alive when the experiment was discontinued 5 days 

 later. A second set of bacterial-count samples were taken at this 

 time . The samples were plated after being refrigerated ^ to_ 3 hoiira. 

 All bacterial plates were prepared with Bein's agar medium and they ' ■ 

 were counted after 6, 12, 15, and 21 days incubation. The number 

 of pigmented subsurface colonies as well as the pigment Intensity 

 of such colonies increased after prolonged incubation. After 21 

 days, however, some colonies began to lose pigment. Bacterial counts 

 of the test materials used in this experiment are listed in table 10. 



Discussion of Results of Studies with Bacteria 



The results of the preliminary experiment with some of the 

 bacteria isolated from unialgal G. brevis cultures suggest that 

 cultures of the two dominant colony tj'pes, both non-pigm.ented, 

 and a sparsely occurring pigmented form are not toxic to fish. 



On the contrary, a "red bacteriimi" isolated from G. brevis - 

 infested water off the west coast of Florida appears to be toxic 

 to fish. Nevertheless, until this bacterium is found m.ore abundantly 

 or its association with fish kills is established, we shall not 

 consider it of importance as a cause of fish mortality occurring 

 off the west coast of Florida. Further toxicity studies with 

 this organism have been discontinued until such time that evidence 

 is obtained to implicate it as a fish-killing agent in nature. 



A bacterium producing red pigment was isolated by Bein 

 (1954) from Indian River on the east coast of Florida at the 



