NOTES 



ISOLATION AND DESCRIPTION OF TWO 



VIBRIOS PATHOGENIC TO PACIFIC 



SALMON IN PUGET SOUND, WASHINGTON 



Vibrio anguillarum (Bergman 1909) is recognized 

 worldwide as a saltwater pathogen in fish (An- 

 derson and Conroy 1970). Most epizootics caused 

 by marine bacteria have been attributed to this 

 organism (Rucker 1959; Sindermann 1966). This 

 note describes recent mortalities resulting from 

 vibriosis of Pacific salmon in the marine waters of 

 Puget Sound, Wash., and heterogeneity observed 

 in vibrios isolated from diseased fish. 



The National Marine Fisheries Service 

 (NMFS) is engaged in the experimental culture of 

 Pacific salmon in salt water at the NMFS Aqua- 

 culture Experiment Station near Manchester, 

 Wash. Epizootics caused by marine vibrios have 

 occurred regularly in cultured salmon during the 

 spring and summer months; the organisms were 

 also isolated from diseased fish on a minor scale 

 in every month during fall and winter (Novotny 

 1975). Vibrios originally isolated from diseased 

 fish at Manchester were typical of Vibrio anguil- 

 larum (Evelyn 1971); strain 775 was represen- 

 tative. 



In November 1973, a commercial salmon farm 

 in the Manchester area suffered a high mortality 

 of pen-reared, 0-age, 250-g coho salmon, On- 

 corhynchus kisutch. Past experience with vib- 

 riosis in the area indicated that the first serious 

 outbreaks usually began in April when water 

 temperatures exceeded 9°C and continued until 

 water temperatures dropped below 12°C in early 

 October (Novotny 1975). Water temperatures in 

 November 1973 were 10° to 11°C; therefore, prob- 

 lems from vibriosis were not anticipated. 



Mortalities also began to occur at about the 

 same time, although not on an epizootic scale, in 

 coho salmon held at the NMFS facility at Man- 

 chester. These fish had been vaccinated in late 

 spring by injecting a heat-killed bacterin pre- 

 pared from V anguillarum lib. Oral antibiotics 

 were administered, but the period required to 

 bring the disease under control appeared to be 

 almost twice that usually required for V. an- 

 guillarum. 



Diseased fish sampled from the NMFS pens and 



the commercial farm exhibited the common signs 

 of vibriosis, most notably a hemorrhagic sep- 

 ticemia. Bacteria characterized as vibrios were 

 consistently isolated from dead or dying fish, but 

 the growth rate of the isolated bacteria was mar- 

 kedly different from that of the typical V. anguil- 

 larum. Also, this bacterium was not agglutinated 

 by rabbit anti-V anguillarum lib serum in rapid 

 slide agglutination tests. 



The new isolates were confirmed as pathogens 

 by injecting pure cultures of them into salmon. 

 All the injected fish died and the organism was 

 routinely re-isolated from kidneys. We desig- 

 nated this bacterium as Vibrio sp. 1669. 



In June 1974, NMFS conducted cooperative 

 vaccination tests with a second commercial salm- 

 on farm in the Manchester area. Approximately 

 280,000 coho salmon smolts were injected with a 

 heat-killed bacterin of V. anguillarum 115 at 

 least 2 wk prior to their transfer to saltwater 

 pens. Mortalities were exceptionally low until 

 late August (less than 6% from all causes and less 

 than 2% from vibriosis). At that time the rate of 

 mortality began to increase and Vibrio sp. 1669 

 was isolated. 



Further tests were made in early August 1974, 

 when 450 0-age sockeye salmon, O. nerka, smolts 

 were transferred to NMFS saltwater pens. One 

 pen contained 150 unvaccinated control fish, and 

 two pens contained 150 fish each that had been 

 vaccinated in fresh water with a heat-killed V. an- 

 guillarum lib bacterin. After 50 days in the 

 saltwater pens, 95% of the unvaccinated fish had 

 died. During the same period the mortalities in 

 the vaccinated lots were 9% and 27% (Figure 1). 

 Vibrios isolated from the vaccinated fish were 

 only of the 1669 type, based on results of slide 

 agglutination tests. 



Materials and Methods 



Samples of kidney, eye, or spleen from freshly 

 dead or moribund fish were streaked on trypti- 

 case soy agar (TSA) (Difco)i with 1% salt added, 

 or on 50% seawater cytophaga agar (Pacha and 



'Reference to trade names does not imply endorsement by 

 the National Marine Fisheries Service, NOAA. 



447 



