The immunomodulatory effects of levamisole were investigated using rainbow trout, Oncor hynchus mykiss. Fish sampled 5 days after intraperitoneal injection of levamisole showed in creased protection against Vibrio anguillarum when a single dose (0.5 mg/kg) was used. Phago cytic activity, chemiluminescence responses and natural killer cell activity in fish treated with levamisole were also increased, compared with those in the saline treated fish. Furthermore, levamisole activated the alternative complement pathway of rainbow trout, although serum bacteri cidal activities against V. anguillarum were not enhanced by the treatment of levamisole. These results indicate that levamisole has immunomodulatory effects in rainbow trout.

Bovine lactofcrrin was evaluated for its ability to enhance resistanec of rainbow trout, Oncorhynchus mykiss (Walbaum), to infection with Vibrio anguillartim. Oral administration of laclotcrrin (UKtmgkg"') to fish dail)' for 3 days prior to an intrapcriloncal challenge with V. anguillarutn resulted in inereascd survival rates, and enhaneed resistance against Streptococcits sp., although to a lesser extent. In laetoferrin-treated fish, an inercase in phagocytic and ehemilumineseent (CL) activities of proncphros cells against V. anguillarum was observed. The phagoeytic and CL aetivities of cells against Streptococcus sp. were also significantly inereascd. However, no in vitro baetcrieidal activities of lactofcrrin against 1'. anguillarum or Streptococcus sp. were observed. This suggests that the Iaetoferrin enhaneed the resistance of the rainbow trout against bacterial infection through the activation of phagocytes.

A butyric acid bacterium, Clostridium butyricum, has been used clinically to prevent disturbances of microbial flora in the human intestine. Oral administration of the bacterial spores has been reported to improve diarrhoea, constipation and abdominal distension in humans. Recently, Young, Kaneda, Mikami, Arai, Igarashi, Saito, Miyoshi & Fuse (1987) showed that C. butyricum had various immunomodulating effects; for example, by stimulating macrophages and natural killer cells, it enhanced protection against Candida albicans infections in mice. In this study, the immunostimulatory effects of C. butyricum when administered orally to rainbow trout, Oncorhynchus mykiss (Walbaum), were examined, and it was estabhshed that administration of the bacterium increased the resistance of fish to Vibrio anguillarum infection.Rainbow trout weighing 50 or 10 g were supplied by the Iwate Experimental Fisheries Station in Iwate Prefecture, Japan. Clostridium butyricum MIYAIRI strain was supplied from Miyarisan Co. Ltd. The bacteria were cultured at 37°C for one day on modified PYG medium (1% peptone, 1% yeast extract and 1% glucose) and collected by centrifugation (5000^, 20min). The bacteria were killed by exposure to air and lyophilization. Vibrio anguillarum (strain PT479), used to challenge the fish, was cultured at 25 °C for 2 days on tryptone agar (TSA) (Nissui) with 1% NaCl.The C butyricum bacterin was suspended in sterile physiological saline (0-85% NaCl) and administered orally by intubation using a catheter. For 3 consecutive days, 300//gkg~* fish~' of the bacterin was administered to the fish. The same concentration of albumin was administered orally to controls, by the same procedure. Fish were infected with V. anguillarum 3 days after treatment and held at a water temperature of 18 °C.The C. butyricum-tTQatcd 50 g fish and albumin-treated controls were challenged by intraperitoneal injection with 01ml V. anguillarum at a concentration of 9-0 X 10^cfum^^ Three fish from each group anaesthetized with MS 222 (3-aminobenzoic acid ester methanesulphonate) were sacrificed at 2, 8, 24 and 48h after the challenge. Blood was collected from the caudal peduncle of the fish with a heparinized syringe and the samples from the three fish were pooled. Kidney and liver samples from the three fish were also pooled. Bacterial levels in the blood, kidney and liver were determined by the plate count method (1% NaCl TSA) (Sakai, Atsuta Sc Kobayashi 1989).Rainbow trout of 10g weight treated with C. butyricum or albumin were anaesthetized with MS-222 and challenged by an intraperitoneal injection of 0-1 ml V. anguillarum, resuspended to either 2 7 x 10' * or lO'^cfumP^ in sterile saline. Each challenge group contained 15 fish; they

The peroxidase-antiperoxidase (PAP) procedure, modified enzyme immunoassay was applied to the detection of Renibacterium salmoninarum. This method did not indicate any cross reactivity with other fish pathogens, and was specific to BKD. The sensitivity of this proceudre was 1 •~ 103 cells/ml which was at least 10 times greater than the indirect fluorescent antibody technique. The PAP procedure was tested in a BKD survey, and gave excellent results when compared with other diagnostic procedures.

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