Creatine kinase (CK) isoenzymes were examined as diagnostic indicators for pathophysiological changes in yellowtail infected with Lactococcus garvieae. Three isoenzymes of CK similar to those found in human were detected by agarose electrophoresis. CK isoenzymes in the brain, heart and intestine consisted of CK-MM, CK-MB and CK-BB. CK-BB was not detected in other tissues or plasma. Normal yellowtail plasma contained CK-MM and CK-MB, while CK-BB and CK-SP appeared in the plasma of infected fish. This phenomenon was found in both naturally and artificially infected fish. These results suggest that appearance of CK-BB and CK-SP in the plasma may reflect the pathophysiological changes due to the infection. Therefore, we consider that the examination of plasma CK isoenzymes is a pathophysiological diagnostic tool for the streptococcal infection in yellowtail.
A 37 kDa major outer membrane protein (MOMP) was purified from the outer membrane fraction ofEdwardsiella tardaEF-1 strain. The protein had a glycoprotein profile and showed immunogenicity to Japanese eelAnguilla japonica. Immunoblot analysis by using an eel antiserum raised against this MOMP revealed no antigenic cross-reaction with other MOMPs of this bacterium. Eel antisera against 37 kDa MOMP extracted from EF-1 and V1 strains showed cross agglutination to formalin-killed cells and heated cells between the two strains. The results indicate that the heat-resistant antigen, 37 kDa, MOMP is located on the cell surface and has a common antigenic determinant. Immunization with the purified 37 kDa MOMP of EF-1 and V1 strains increased resistance in eel against challenge by intraperitoneal injection with live EF-1 strain. The results show that this 37 kDa MOMP is one of the protective antigens ofE. tarda. As the MOMP was found in two strains of different serotypes, it is probably contributed as a promising vaccine antigen against edwardsiellosis caused byE. tarda.
Genetic variability among strains of Flavobacterium columnare (formerly Flexibacter columnaris) was characterized using restriction fragment length polymorphism (RFLP) analysis of 16S rDNA, 16S rDNA sequencing, and DNA-DNA hybridization. A total of 23 Fl. columnare strains isolated from diseased fishes in various areas were divided into three distinct genomic groups. The main group, genomovar 1 consisted of 20 strains, including the type strain. Two strains were assigned to genomovar 2, and one to genomovar 3. However, as we found their phenotypic characteristics to be identical, there is no evidence to classify strains of these genomovars as belonging to new species or sub-species.
This study was undertaken to determine whether marine birnavirus (MABV) can induce apoptosis in vitro in cell lines established from four different fish species. MABV Y-6 strain was inoculated onto chinook salmon embryo (CHSE-214), red sea bream kidney (RSBK-2), fathead minnow caudal peduncle (FHM) and epithelioma papulosum cyprini (EPC) cell lines at a multiplicity of infection (m.o.i.) of 0.1. At 0, 12, 24, 36, 48, 72, and 96 h post-infection, the cells were harvested and used for DNA fragmentation analysis, apoptotic cell counts and determination of virus titer by plaque assay. MABV infection appeared to induce the typical features of apoptosis such as nuclear and cytoplasmic condensation, DNA fragmentation and formation of apoptotic bodies in CHSE-214 and RSBK-2 cells, but not in FHM and EPC cells. The concentration of free virus increased immediately after infection, whereas the apoptotic cell ratio and cell-associated virus titer scarecely increased until 24 h. Apoptotsis increased after 36 h when monitored by counting of apoptotic cells and analysis of DNA. The results suggest that MABV replicating to high concentrations during an early stage of infection induces apoptosis in later stages.