488 



FISHERY BULLETIN OF THE FISH AND WILDLIFE SERVICE 



significant role in the development of G. brevis 

 blooms should not be overlooked. Such organ- 

 isms maj' contribute appreciably to the nutrition 

 of G. brevis. For example, some of the bacteria 

 isolated from unialgal G. brevis cultures produce 

 vitamin Bij-active substances (Starr et al., 

 1957). Vitamin Bjj apparently stimulates the 

 growth of G. brevis in sea-water-base medium 

 (Wilson and Collier, 1955). 



Once fish kills are initiated by blooms of G. 

 brevis, excessive bacterial growth resulting from 

 the increased availability of organic matter may 

 possibly cause the blooms to decline in isolated 

 situations. Bacteria could unfavorably affect G. 



brevis blooms by producing substances toxic to 

 this organism, by competing for nutritive sub- 

 stances, and by adversely altering the pH. We 

 have frequently observed the failure of unialgal 

 G. brevis to grow in tubes in which the medium 

 became cloudy with bacterial growth. 



Another role for which bacteria must be con- 

 sidered is that of a detoxicating agent. Shilo and 

 Aschner (1953) found that bacteria decreased the 

 toxicity of cultures of Prymnesium parvum, a 

 marine and brackish water chrysomonad that is 

 toxic to fish. .Similarly, bacterial activity may 

 influence the toxicity of G. brevis in the laboratory 

 and in nature. 



TOXICITY STUDIES WITH BACTERIA 



Although the available evidence indicated that 

 Gymnodinium brevis causes the toxicity of unialgal 

 cultures, direct proof was lacking prior to the 

 development of bacteria-free cultures. In addi- 

 tion to the bacterial studies previously considered, 

 tl^e possibility that bacteria may cause all or 

 some of the toxic efi'ects was investigated by test- 

 ing pure cultures of some of the bacteria from 

 unialgal G. brevis cultures. Furthermore, toxicity 

 tests were conducted with pure cultures of an 

 unidentified red-pigment-producing bacterium iso- 

 lated from Florida water and Flavobacterium 

 piscicida Bein. Bein (1954) suggested that F. 

 piscicida, a chromogenic bacterium, possibly was 

 a cause of mass fish mortality associated with 

 discolored water off the west coast of Florida. 



BACTERIA ISOLATED FROM UNIALGAL 

 GYMNODINIUM BREVIS CULTURES 



Because of the preliminary nature of these 

 studies and the crudeness of the quantitative 

 bacterial estimates, onty a summary will be pre- 

 sented. Tlie bacleria used have not been identi- 

 fied; presently they are being characterized 

 morphologically and physiologically by Dr. T. J. 

 Starr. The test fish were Gulf killifish (Fundulus 

 similis'). 



Two test fish per container (1 -liter beaker) were 

 subjected to about 500 ml. of nonaerated test 

 materials. Bacterial suspensions were prepared 

 by adding 16.5 ml. of a 24-hour culture to 500 ml. 

 of filtered Galveston Bay water. Control ma- 

 terials consisted of the same ratio of sterile culture 

 medium and bay water as well as bay water alone. 

 Crude estimates of bacterial concentrations were 



made by preparing a pour-plate of 0.02 to 0.03 

 ml. of a sample collected shortly after the fish 

 were added. A second sample was taken either 

 after both fish died or after 23 hours if at least one 

 fish survived this period. The colonies were too 

 numerous to count in most of the plates prepared 

 from the second set of samples. Therefore, not 

 even rough estimates could be made for the 

 bacterial concentrations. 



The most-abundant colony type isolated from 

 unialgal G. brevis cultures on Spencer's peptone 

 sea-water agar and Bein's peptone agar is gener- 

 ally a convex, white, opaque colony produced by 

 Gram-negative rods. This colony type may 

 represent several different species and physiologi- 

 cal types of bacteria. Two separate isolates of 

 the white, opaque colony did not give evidence of 

 being toxic to F. siinilis. The initial bacterial 

 concentration was in the order of 0.5 to 1 million 

 per ml. 



A flat, white, transluscent colony with an irides- 

 cent sheen, also produced by Gram-negative rods, 

 is usually the second most-abundant colony type 

 isolated from unialgal G. brevis cultures. An 

 initial concentration of approximately 1 million 

 bacteria per ml. of this colony type gave no 

 evidence of toxicity to F. si7nilis. 



Chromogenic bacteria constitute only a small 

 portion of the bacterial flora of unialgal cultm-es 

 of G. brevis; however, yellow-pigment-producing 

 bacteria become abundant in cultures treated with 

 dihydrostreptomycin sulfate. They dominate in 

 G. brevis cultures treated with 500 to 1 ,000 Mg. of 

 this antibiotic per ml., and often occur in nearly 

 pure cultm-e. This antibiotic may cnliance the 



