A visceral mycosis in cultured masu salmon (Oncorhynchus masou) caused by a species of Ochroconis

Ochronconis sp. infection was found in masu salmon (Oncorhynchus masou) with visceral mycosis in Japan. The external and internal clinical signs were reddening of the anal area, swelling of the abdomen due to accumulation of ascitic fluid in the abdominal cavity and extensive swelling of the posterior kidney. Many pale brown, septate hyphae were found in the kidney by direct microscopical examination; these were usually not found in the other organs of infected fish. Histopathological examination of the kidney revealed large granulomas with the fungal hyphae and giant cells. The isolated fungus was identified as a species of the genus Ochroconis and was compared with O. tshawytschae, a known fish pathogen. Based on morphological and growth characteristics, we believe that these cases resulted from infection with a different species.     

Atkinsiella parasitica sp. nov. isolated from a rotifer,Brachionus plicatilis

A new species ofAtkinsiella (Lagenidiales, Oomycetes),Atkinsiella parasitica is described and illustrated. It was isolated from the eggs and bodies of a rotifer,Brachionus plicatilis, with fungal infection in 1992. The rotifer was reared in a hatchery as the first food supply for seed production of crustaceans and fishes. The fungus is characterized by producing monoplanetic, lateral biflagellate zoospores, and infrequently branched discharge tubes. Optimum temperature for the fungus was 25°C. The fungus was considered an obligate marine fungus, because its growth was observed only on PYGS medium including seawater.     

Lagenidium myophilum infection in the coonstripe shrimp,Pandalus hypsinotus

A fungal infection occurred in juvenile coonstripe shrimps,Pandalus hypsinotus, cultured at Hokkaido Institute of Mariculture, Hokkaido, Japan. The fungus was identified asLagenidium myophilum, the same fungus that had previously been isolated from the abdominal muscle of adult northern shrimps,Pandalus borealis, and larvae of the coonstripe shrimp. Histopathologically, numerous nonseptate hyphae were observed in the lesions, and melanized hemocytes were present within the blackened areas. The optimum temperature for growth of the present strain was 25–30°C, and the optimum NaCl concentration for growth was 0.5–1.0%. Its biological characteristics were compared with those ofLagenidium myophilum isolated from diseased larval coonstripe shrimp and adult northern shrimp. The fungus was pathogenic toward shrimps of the genusPandalus, which live in deep sea areas. The fungus could infect shrimps at various stages, from larva to adult.     

Genotypic characteristics of a Mycobacterium sp. isolated from yellowtail Seriola quinqueradiata and striped jack Pseudocaranx dentex in Japan

In Japan, a Mycobacterium marinum‐like mycobacterium was isolated from the yellowtail, Seriola quinqueradiata. The species was identified as M. marinum by a commercial mycobacterial DNA‐DNA hybridization kit. Nevertheless, PCR restriction analysis of the DNA of its RNA polymerase β‐subunit gene definitively showed that this Mycobacterium sp. was M. ulcerans. PCR analysis revealed the genotypic characteristics of M. ulcerans in the Mycobacterium sp., only the mup053 gene sequence being absent, as has been found previously in other piscine mycobacteria such as M. marinum strains DL240490 and DL045 and M. pseudoshottsii. With one exception, this Mycobacterium sp. and M. pseudoshottsii had identical 16S rRNA gene sequences, which is also probably true of M. marinum strains DL240490 and DL045. Similarly, according to comparisons of the 16S rRNA gene, ITS region, and hsp65 gene sequences, this Mycobacterium sp. is more closely related to M. pseudoshottsii than to M. ulcerans or M. marinum. A PCR product of approximately 2000 bp was amplified from region of difference 9 in the Mycobacterium sp. The nucleotide sequence revealed insertion of IS2404, the sequence of which is 1366 bp long. The novel single nucleotide polymorphisms identified in this region distinguished this Mycobacterium sp. from M. marinum strain DL240490 and M. pseudoshottsii. The present findings raise the possibility that these species have a common ancestor. Further studies are required to improve our understanding of the relationship between their geographical origin and genetic diversity.     

Saprolegniosis in salmonids and their eggs in Japan

An epizootic of the fungal infection saprolegniosis that occurred in freshwater-cultured salmons and their eggs at some hatcheries in Hokkaido (Japan) was investigated. In almost all cases, the initial clinical sign was characterized by the growth of cotton-like mycelia on the fishs' body surface, especially the head, adipose fin, and caudal fin, but the mycelia were not visible to the naked eye in the internal organs. Thirty-three strains isolated from lesions were classified in the genus Saprolegnia according to their morphological and biological characteristics on hemp seed cultures at various temperatures. Fifteen of the strains were identified as Saprolegnia parasitica, 16 were identified as S. salmonis, and two were identified as S. australis.     

ATP-induced calcium oscillations and change of P2Y subtypes with culture conditions in HeLa cells

ATP, UTP, ADP and UDP induced intracellular Ca(2+) responses and oscillations in HeLa cells that sometimes lasted over 1 h. The response is due to the activation of P2Ys, G-protein coupled ATP receptors, because the oscillations persisted for several minutes even in Ca(2+)-free solution, and suramin and PPADS, antagonists of ATP receptors, partially inhibited the response. The potency of these nucleotides varied with the culture or cell conditions, i.e. UTP was generally most potent but in some cases UDP was more potent; responses to UDP were variable while those to ATP were constant. In addition, Ca(2+) responses to ATP and UDP were additive. These findings suggested the existence of two or more subtypes of P2Ys in HeLa cells. RT-PCR experiments revealed the existence of P2Y(2), P2Y(4) and P2Y(6). Recovery from starvation (culture in FBS-free medium overnight and re-addition of FBS) increased the responses to UTP and UDP but not to ATP, suggesting that the number or activity of P2Y(6) and/or P2Y(4) receptors may increase with cell proliferation in HeLa cells.     

Hydrogen-Deuterium Exchange Effects on β-Endorphin Release from AtT20 Murine Pituitary Tumor Cells

Abundant evidences demonstrate that deuterium oxide (D₂O) modulates various secretory activities, but specific mechanisms remain unclear. Using AtT20 cells, we examined effects of D₂O on physiological processes underlying β-endorphin release. Immunofluorescent confocal microscopy demonstrated that 90% D₂O buffer increased the amount of actin filament in cell somas and decreased it in cell processes, whereas β-tubulin was not affected. Ca²⁺ imaging demonstrated that high-K⁺-induced Ca²⁺ influx was not affected during D₂O treatment, but was completely inhibited upon D₂O washout. The H₂O/D₂O replacement in internal solutions of patch electrodes reduced Ca²⁺ currents evoked by depolarizing voltage steps, whereas additional extracellular H₂O/D₂O replacement recovered the currents, suggesting that D₂O gradient across plasma membrane is critical for Ca²⁺ channel kinetics. Radioimmunoassay of high-K⁺-induced β-endorphin release demonstrated an increase during D₂O treatment and a decrease upon D₂O washout. These results demonstrate that the H₂O-to-D₂O-induced increase in β-endorphin release corresponded with the redistribution of actin, and the D₂O-to-H₂O-induced decrease in β-endorphin release corresponded with the inhibition of voltage-sensitive Ca²⁺ channels. The computer modeling suggests that the differences in the zero-point vibrational energy between protonated and deuterated amino acids produce an asymmetric distribution of these amino acids upon D₂O washout and this causes the dysfunction of Ca²⁺ channels.