Recently, a microsporidian infection was found in the trunk muscle of hatchery-bred juvenile spotted halibut Verasper variegatus. The disease occurred from mid-July to the end of September 2007 and the cumulative mortality reached approximately 20%. Infected fish showed the external sign of a concave body surface on the eyed side, and the microsporidian parasite formed numerous ‘cysts’ in the muscle. These characteristics resembled ‘beko’ disease caused by Microsporidium seriolae from cultured yellowtail Seriola quinqueradiata, Microsporidium sp. from red sea bream Pagrus major (RSB) and Microsporidium sp. from gilthead sea bream Sparus aurata (GSB), but several differences were observed in associated pathological findings such as internal hemorrhage around the ‘cysts’. Average dimensions of spores from spotted halibut (SH) were 3.07 × 2.13 μm, which were relatively smaller than those of M. seriolae, Microsporidium sp. RSB and Microsporidium sp. GSB, but the ranges of measurements overlapped among them. Molecular analysis of rDNA sequences suggested that the present parasite, provisionally named as Microsporidium sp. SH, was distinct from the other known species.
Japanese eel Anguilla japonica and European eel A. anguilla, 65 individuals each, were exposed in a tank to 8,600 oncomiracidia of Pseudodactylogyrus bini and P. anguillae, monogeneans originally recorded on Japanese eel from Asia. Infection was monitored weekly for 6 weeks to compare susceptibility to these monogenean infections between the two eel species. Approximately 40% of the oncomiracidia infected the eels. In the six samplings, both P. bini and P. anguillae were always found in greater numbers on the gills of European eel than on Japanese eel. The first generation of parasites matured within 2 weeks, laying eggs in the tank, and the second generation appeared on the gills of the eels on the third week. European eel was more susceptible not only to the first but also to the second and later generations of parasites than Japanese eel. Besides, both parasites grew faster on European eel than on Japanese eel. This is the first experimental evidence that there are substantial differences in the susceptibility to P. spp. infection between European eel, an unnatural host in Japan, and Japanese eel, the natural host in this country.
From middle August to early October in 2007, mortalities in wild ayu Plecoglossus altivelis occurred in the rivers of Tokyo Metropolis, Yamaguchi Prefecture and Hiroshima Prefecture, Japan. Hemorrhagic ascites was observed in almost all the examined fish. Some fish showed reddening of body surface, anus or bases of fins and exophthalmos. A single species of bacteria was isolated from the kidney, spleen and ascites of all the examined fish and identified as Edwardsiella ictaluri. Pathogenicity against ayu of the bacterial isolates was demonstrated by experimental infections, with the LD50 being approximately 104 CFU/fish. The dead fish exhibited similar disease signs to those of naturally affected fish. From these results, it was concluded that the present mortality of wild ayu was caused by E. ictaluri infection. This is the first report on E. ictaluri infection of fish in Japan.
A Gram-negative bacterium was isolated from diseased wild ayu Plecoglossus altivelis, which were caught in rivers in Japan from August to October in 2007. All four isolates examined showed the same morphological, physiological and biochemical characteristics and were classified into the genus Edwardsiella. The isolates were differentiated from E. tarda in respect of its negative production of indole and no growth at 37°C, and different from a reference strain of E. ictaluri (JCM1680) in its positive production of hydrogen sulfide. All isolates were completely identical in the partial nucleotide sequences of 16S rDNA, a type 1 fimbrial gene (etfA) and a heat shock protein gene (dnaJ), and these sequences showed high similarity (100%, 99.7% and 100%, respectively) with E. ictaluri but low similarity (99.7%, 92.5% and 87.3%, respectively) with E. tarda. Based on these phenotypic and genetic characteristics, the present isolates from ayu were identified as E. ictaluri.
Edwardsiellosis was induced in a population of Japanese flounder Paralichthys olivaseus by cohabitation with the flounder intraperitoneally inoculated with Edwardsiella tarda. When coefficient of variation of hematological and immunological parameters of the blood increased and no death caused by edwardsiellosis was observed, 1/3 individuals in the population were transferred to a tank under an improved condition where fish density was lowered to 1/3 and exchange rate of rearing sea water was increased three times than before (Group A). Another 1/3 individuals were transferred to another tank under the same improved condition when 10% individuals died of edwardsiellosis (Group B). The remaining 1/3 individuals were reared under the non-improved conditions (positive control). Cumulative mortality in Group A was 57.8%, significantly lower than that in Group B (74.4%; p<0.05) or positive control (80.0%; p<0.01). This result suggests that improvements of rearing conditions at an early stage of edwardsiellosis reduce the mortality caused by the disease.
Overwintering ayu Plecoglossus altivelis (n = 23) were captured in Sho-river, Toyama Prefecture in April, 2007, and kept under a crowded condition (120 g fish/L). When the fish became moribund or dead, bacteria were isolated from the kidney of the fish using a modified Cytophaga agar plate, and the isolates were examined by the PCR-RFLP method using primers targeting gyrA, gyrB and PPIC genes of Flavobacterium psychrophilum. The isolates obtained from nine individuals of the fish were identified as F. psychrophilum and classified into two genotypes of A/R/QR and A/S/QR. An isolate from each genotype was virulent to ayu. These results showed that overwintering ayu carry virulent F. psychrophilum, suggesting a source of infection for newcomers of ayu in this spring.
A one-step, single tube, real-time loop-mediated isothermal amplification (real-time LAMP) assay was developed to detect a non-structural protein encoding gene of infectious hypodermal and haematopoietic necrosis virus (IHHNV). The real-time LAMP method for IHHNV is simple and rapid with specific amplification within 60 min at 63°C employing four primers. This method requires less time than the PCR method and is specific for IHHNV when analysed with PRDV (= WSSV), YHV and TSV DNA/cDNA, and healthy shrimp DNA. Sensitivity analysis revealed this method is capable of detecting as few as 102-103 copies/μL, suggesting it can be used as an alternative quantitative detection method for IHHNV in diagnostic laboratories.