Annual catch landings of the Manila clam Ruditapes philippinarum have decreased in Japan since the early 1980s because of the decreased size of many Manila clam populations. It is hypothesized that Perkinsus olseni infection contributed to these decreases. However, the effect of Perkinsus infection on the survival and physiology of Manila clam is poorly understood. We examined the influence of Perkinsus infection on clearance rate, tolerance against high water temperature, and burrowing activity in a captive adult clam population. Infection intensity did not affect clearance rates, burrowing activity, or tolerance against high water temperature. In addition, we monitored infection intensity in wild, adult clams from a tidal flat lacated in western Seto Inland Sea over a one-year period. Infection levels were relatively stable in the wild population between June 2004 and May 2005, but we noted a significant decrease in the infection intensity at the end of the spawning period in November 2004. However, in the present study, clear evidence showing the negative impact of Perkinsus on clam physiology and survival was not obtained.
Bacterial cold-water disease (BCWD) caused by Flavobacterium psychrophilum leads to heavy mortality of ayu Plecoglossus altivelis in Japan. Previously, a 28°C warmed water treatment was shown to be an effective treatment for BCWD, since F. psychrophilum can not grow and maintain colonies at this temperature. However, it was unclear whether the bacteria might resuscitate after the treatment was over. Therefore, we investigated colony formation, membrane potential and pathogenicity of F. psychrophilum at various temperatures (15°C, 23°C, 28°C and 33°C) in sterilized underground water. Within 2 days at 28°C, F. psychrophilum completely lost their colony-forming abilities but still maintained their membrane potentials. It seemed that these cells entered into viable but non-culturable (VBNC) state. However, experimental ayu infection revealed that VBNC F. psychrophilum cells were unable to cause BCWD, suggesting that the cells were progressing towards death at 28°C.
We evaluated the efficacy of povidone-iodine treatments in preventing vertical transmission of Flavobacterium psychrophilum in the eggs of masu salmon Oncorhynchus masou, Japanese char Salvelinus pluvius, coho salmon O. kisutch and rainbow trout O. mykiss. When newly spawned eggs were immersion-challenged with F. psychrophilum (> 109 CFU/mL) and subsequently disinfected with povidone-iodine (50 ppm in PBS, 15 min) prior to fertilization, the pathogen was not detected in the contents of the eggs. Without this pre-fertilization disinfection with povidone-iodine, however, procedures such as water-hardening of immersion-challenged eggs in povidone-iodine (50-100 ppm in water) and rinsing of the F. psychrophilum-contaminated eggs with PBS were insufficient to control the intra-ovum infection. In a separate experiment using coho salmon eggs challenged with F. psychrophilum, pre-fertilization disinfection was effective to prevent bacterial cold-water disease in the produced fry. Surface disinfection of unfertilized eggs immediately after spawning is thus essential to prevent vertical transmission of F. psychrophilum.
The optimum conditions for the expression of WSSV (= PRDV) rVP26 and rVP28 in E. coli cells were investigated, especially the IPTG concentration (0.001-1.0 mM) and inducement temperatures (25°C and 37°C). For both rVP26 and rVP28, the strongest intensity of expression was observed in medium containing 0.01 mM IPTG at 37°C. The growth of the transformed bacteria expressing rVP26 or rVP28 in media containing 0.01 mM IPTG was the almost same as that in medium without IPTG, but clearly delayed than that in the medium containing 1.0 mM IPTG. The productivity was increased 9-fold in rVP26 and 6-fold in rVP28 when the IPTG concentration was reduced from 1.0 mM to 0.01 mM. Next, we investigated the minimum doses of rVPs required to induce phylaxis of kuruma shrimp Penaeus japonicus against WSD (= PAV) by an experimental challenge. The results show that at least 10 μg/g shrimp/day of rVPs should be orally administered to kuruma shrimp for 15 days in order to induce effective phylaxis.
Heterobothrium ecuadori is a gill monogenean infecting the gills of the bullseye puffer fish Sphoeroides annulatus and feeding fish blood. This experimental study was performed to describe the juvenile stages of H. ecuadori and to determine the time required to reach the adult stage at 20-23°C. Uninfected bullseye puffer fish were exposed to oncomiracidia and sampled at intervals to obtain worms of different developmental stages. Between 5 and 10 days after the exposure, the first and second pairs of clamps were observed in worms along with the emergence of the third clamp. Four pairs of clamps were observed on day 15, and primordial male copulatory organ, testes and ovary were found on day 20. Adult parasites started to appear between 25 and 30 days post-infection. The time required for egg laying was estimated to be 33 days, based on the first collection of eggs in the rearing water.
Hafnia alvei is a Gram-negative bacillus that belongs to the family Enterobacteriaceae and has been isolated from different fish species. H. alvei causes a wide variety of diseases, including hemorrhagic septicemia, ikidney ilesions and mortality in different fish species, as rainbow trout Oncorhynchus mykiss, cherry salmon O. masou, and brown trout Salmo trutta. The aim of this study is to investigate the interaction of different H. alvei strains with RTG-2 by gentamicin protection and immunofluorescence assays. Our results demonstrated a considerable potential of H. alvei to attach to RTG-2 cells, and that the internalized strains remained viable in the cells for 48 h.