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NO45 - 2

Oncomiracidial Behavior of Heterobothrium okamotoi (Monogenea: Diclidophoridae)

<>Sho Shirakashi, Shuya Nakatsuka, Akihisa Udagawa and Kazuo Ogawa*<>Department of Aquatic Bioscience, Graduate School of Agricultural and Life Sciences,
The University of Tokyo, Tokyo 113-8657, Japan
(Received June 19, 2009)
ABSTRACT―We conducted a series of experiments to understand the behavioral characteristics of oncomiracidium of the monogenean Heterobothrium okamotoi and compared them with those of two other monogenean species, Heteraxine heterocerca and Neobenedenia girellae. Behavior of H. okamotoi oncomiracidium consisted of two alternate phases: a swimming phase with strong ciliary beatings and a stationary phase with ciliary beatings too weak to generate any movements. The duration of one swimming-stationary cycle considerably varied among individuals but tended to increase with larval age. During the 5-day monitoring, length of the swimming phase decreased while that of the stationary phase tended to increase as the larva became older. The locomotion patterns differed significantly among individuals, but as a whole, larvae showed random horizontal movement and no phototactic reaction. The oncomiracidium tended to move downward because it was heavier than seawater and also swam downward at the start of the swimming phase because its anterior part pointed downward when the larva was not swimming. Those behavioral characteristics of larval H. okamotoi are likely to have evolved in order to maximize the chance of encountering its benthic fish host, tiger puffer Takifugu rubripes.

Key words: Heterobothrium okamotoi, oncomiracidium, behavior, Monogenea, Takifugu rubripes

Dynamics of the Bacterial Cold-water Disease Pathogen,Flavobacterium psychrophilum, in Infected Fish Organs and Rearing Water after Warmed Water Treatment

<>Kazuhiro Sugahara1,2*, Erina Fujiwara-Nagata3 and Mitsuru Eguchi3<>1Shiga Prefectural Fisheries Experimental Station, Shiga 522-0057, Japan
2Graduate School of Agriculture, Kinki University, Nara 631-8505, Japan
3Department of Fisheries, Kinki University, Nara 631-8505, Japan
(Received January 30, 2010)

ABSTRACT―Bacterial cold-water disease (BCWD) causes heavy mortality of ayu Plecoglossus altivelis in Japan. To control BCWD of ayu, warmed water treatment at 28ーC has been used. However, the mechanism of this treatment is unknown. Hence, we investigated the growth/survival of Flavobacterium psychrophilum at 15-28ーC under two nutritional conditions: in modified cytophaga (MCY) broth and sterilized underground water. Within 2 days at 28ーC in both the conditions, F. psychrophilum totally lost its colony-forming ability. We also studied distributions of F. psychrophilum in infected fish organs and in rearing water after various treatments. For bacteria detection, the colony-counting method and loop-mediated isothermal amplification (LAMP) method were used. For warmed water treatment, the rearing water was warmed from 18ーC to 23ーC or 28ーC for 3 days. The treatments were started 1 day after immersion infection. In the experimentally infected fish, F. psychrophilum was detected neither in any fish organs nor in the rearing water after the 28ーC treatment for 3 days. These results indicate the effectiveness of the warmed water treatment at 28ーC against BCWD of ayu.

Key words: warmed water treatment, Flavobacterium psychrophilum, bacterial cold-water disease, Plecoglossus altivelis

Antibacterial Effect of Chemical Reagents against Tenacibaculum maritimum

<>Ken-ichi Watanabe1,3* and Toyohiro Nishioka2<>1National Research Institute of Aquaculture, Fisheries Research Agency, Oita 879-2602, Japan
2National Research Institute of Aquaculture, Fisheries Research Agency, Mie 516-0193, Japan
3Present Address: Department of Aquatic Bioscience, Faculty of Bioindustry,
Tokyo University of Agriculture, Hokkaido 099-2493, Japan
(Received July 28, 2009)

ABSTRACT―Minimum inhibitory concentrations (MICs) of 12 chemical reagents were determined for four strains of Tenacibaculum maritimum. Bronopol and stabilized chlorine dioxide showed the lowest MIC (1.0 μg/mL). With 6-h treatment, minimum bactericidal concentrations (MBCs) of bronopol and stabilized chlorine dioxide were 4.0 and ≦8.0 μg/mL, respectively. Both chemicals were non-toxic to five marine fishes in 6-h exposure at concentrations 5 times higher than the MBCs. The two chemicals are the most suitable candidates for antibacterial agents for treatment of T. maritimum infection.

Key words: Tenacibaculum maritimum, tenacibaculosis, bronopol, stabilized chlorine dioxide, antibacterial agent

Involvement of Viral Nervous Necrosis in Larval Mortality of Hatchery-reared Pacific Bluefin Tuna Thunnus olientalis

<>Toyohiro Nishioka1*, Koh-ichiro Mori2, Takuma Sugaya2, Nobuhiro Tezuka3, Takayuki Takebe4, Hitoshi Imaizumi5, Kazunori Kumon6, Shukei Masuma7 and Toshihiro Nakai8<>1National Research Institute of Aquaculture, Fisheries Research Agency, Mie 516-0193, Japan
2National Research Institute of Aquaculture, Fisheries Research Agency, Oita 879-2602, Japan
3Notojima Station, National Center for Stock Enhancement, Fisheries Research Agency,
Ishikawa 926-0216, Japan
4Seikai National Fisheries Research Institute, Ishigaki Tropical Station, Fisheries Research Agency,
Okinawa 907-0451, Japan
5Shibushi Station, National Center for Stock Enhancement, Fisheries Research Agency,
Kagoshima 899-7101, Japan
6Amami Station, National Center for Stock Enhancement, Fisheries Research Agency,
Kagoshima 894-2414, Japan
7Miyazu Station, National Center for Stock Enhancement, Fisheries Research Agency, Kyoto 626-0052, Japan
8Graduate School of Biosphere Science, Hiroshima University, Hiroshima 739-8528, Japan
(Received November 23, 2009)

ABSTRACT―Amami Station of Fisheries Research Agency, Japan has experienced mass mortalities of Pacific bluefin tuna Thunnus olientalis in the process of seed production since around 2000. The mortalities mostly occurred at larval stages. In some cases of the mortalities, but not all cases, the diseased fish was characterized by vacuolation in the central nervous systems and retina, and a betanodavirus (RGNNV genotype) was detected in the affected fish by PCR and FAT methods and culture isolation. The present results suggest that VNN can be a cause of larval mortality of Pacific bluefin tuna.

Key words: betanodavirus, viral nervous necrosis, Thunnus olientalis, Pacific bluefin tuna, mass mortality

Antibody Detection against Red Sea Bream Iridovirus (RSIV) in Yellowtail Seriola quinqueradiata Using ELISA

<>Se Ryun Kwon1, Toyohiko Nishizawa2*, Ikuo Takami3 and Mamoru Yoshimizu1<>1Faculty of Fisheries Sciences, Hokkaido University, Hokkaido 041-8611, Japan
2Department of Aqualife Medicine, Chonnam National University, Yeosu 550-749, Korea
3Nagasaki Prefectural Institute of Fisheries, Nagasaki 851-2213, Japan
(Received December 10, 2009)

ABSTRACT―FBS components in viral culture fluid interfere with the detection of specific antibody in immunized fish. In the present study, we developed a method to eliminate the interference of FBS using a model of red sea bream iridovirus (RSIV) vaccine. In an experiment using yellowtail antiserum against BSA, which is a major component of FBS in cell culture media, antibodies against BSA were completely absorbed via incubation of antiserum in 50% FBS at 25ーC for 1 h. In antibody detection, ELISA with the sera of yellowtail surviving RSIV-challenge, antibodies against RSIV were detected, but those against FBS disappeared via the pre-treatment of the sera. Therefore, antibodies against RSIV are effectively detected using ELISA with the pre-treatment of fish sera.

Key words: antibody detection, ELISA, red sea bream iridovirus, yellowtail

First Isolation of Vibrio tapetis, the Etiological Agent of Brown Ring Disease (BRD), in Manila Clam Ruditapes philippinarum in Japan

<>Tomomasa Matsuyama1*, Takamitsu Sakai1, Ikunari Kiryu1, Kei Yuasa1, Hideki Yasunobu2, Yoshihiro Kawamura2 and Motohiko Sano1<>1National Research Institute of Aquaculture, Fisheries Research Agency, Mie 516-0193, Japan
2Hyogo Prefectural Technology Center for Agriculture, Forestry and Fisheries, Hyogo 674-0093, Japan
(Received January 19, 2010)

ABSTRACT―Mass mortality occurred in Manila clam Ruditapes philippinarum under a hanging cultivation condition from April to July 2008 in Japan. A bacterium was isolated from the affected clams showing brown conchiolin deposits on the inner shell surface and identified as Vibrio tapetis, the causal agent of brown ring disease (BRD) in clams, by bacteriological characterization and genetic analysis. Experimental infection of clams confirmed pathogenicity of the isolated bacterium. These results indicate that the present mortality of cultured clams was caused by BRD. This is the first report on isolation of V. tapetis from diseased R. philippinarum in Japan.

Key words: brown ring disease, BRD, Vibrio tapetis, Ruditapes philippinarum, Manila clam

Detection of Specific Antibodies against Renibacterium salmoninarum by ELISA with p57 Protein as a Capture Antigen

<>Takanori Matsui1, Norihisa Oseko2, Toyohiko Nishizawa3* and Mamoru Yoshimizu1<>1Faculty of Fisheries Sciences, Hokkaido University, Hokkaido 041-8611, Japan
2National Research Institute of Aquaculture, Fisheries Research Agency, Hokkaido 062-0992, Japan
3Department of Aqualife Medicine, Chonnam National University, Yeosu 550-749, Korea
(Received January 21, 2010)

ABSTRACT―We evaluated p57 of Renibacterium salmoninarum (R.s.) as a capture antigen for specific antibody-detection ELISA against R.s.. Using formalin-killed R.s. (FK-R.s.) as ELISA antigens, ELISA values (OD492 nm) of the negative control sera showed high values ranging from 0.12 to 1.59. These values were relatively lower than those of sera from the fish immunized with FK-R.s. (0.16-2.6), but seemed to be still high as negative control sera. Contrarily, when p57 was used as a capture antigen, ELISA values of the negative control sera were all 0.1 or less, while those of the sera from immunized fish ranged from 0.02 to 1.28. It was thus demonstrated that high background of ELISA was enabled to be reduced by p57 as ELISA antigens.

Key words: antibody-detection ELISA, Renibacterium salmoninarum, BKD, capture antigen, O. mykiss, O. masou

Snout Chondroma in a Neon Tetra Paracheirodon innesi

<>Rubina Sirri*, Luciana Mandrioli, Barbara Bacci, Maria Morini and Giuliano Bettini<>Servizio di Prova di Anatomia Patologica, Department of Veterinary Public Health and Animal Pathology, Faculty of Veterinary Medicine, Alma Mater Studiorum University of Bologna, Via Tolara di Sopra,
50, 40064, - Ozzano Emilia (BO), Italy
(Received January 21, 2010)

ABSTRACT―An adult neon tetra Paracheirodon innesi with a history of difficulty in feeding caused by a mass overwhelming the snout is presented. Macroscopically the fish presented a unilateral, single, nodular and well-circumscribed, perlaceous, firm to hard mass in the left snout region. Histologically, tumor tissue was formed mainly by abundant, highly cellular elliptic nests of chondrocytes lying in mature hyaline cartilage matrix. Besides this component there were bubbly appearing large cells (physaliphorous-like cells) and a minority of cells having well defined cytoplasmic boundaries, clear eosinophilic pale cytoplasm and a rounded centrally located nucleus (吐ried egg-like cells・. Alcian-PAS and Masson痴 trichrome stains were applied to demonstrate proteoglycans and collagen of the matrix. Based on topographic localization and histomorphological features, a diagnosis of snout chondroma was made. This case can be added to the other rare spontaneous cartilage tumors of fish described in literature.

Key words: chondroma, Paracheirodon innesi, neon tetra, cartilage tumor, teleost

Pathogenicity of Mycobacterium marinum to Amberjack Seriola dumerili, Red Sea Bream Pagrus major and Mouse

<>Sompoth Weerakhun1, Shinpei Wada1, Kishio Hatai1*, Ayako Sano2, Harunao Nibe3 and Tatsumu Hirae3<>1Laboratory of Fish Diseases, Nippon Veterinary and Life Science University, Tokyo 180-8602, Japan
2Research Center for Pathogenic Fungi and Microbial Toxicoses, Chiba University, Chiba 260-8673, Japan
3Kagoshima Prefecture Fisheries Technology and Development Center, Kagoshima 891-0315, Japan
(Received March 1, 2010)

ABSTRACT―We conducted experimental infection tests of Mycobacterium marinum NJB 0419 isolated from yellowtail Seriola quinqueradiata to amberjack S. dumerili and red sea bream Pagrus major to determine the pathogenicity of the bacterium. As a result, NJB 0419 was pathogenic to both of them. In light of concerns about zoonoses, BALB/c mice were intravenously or subcutaneously injected with NJB 0419. The histopathological findings of examined mouse tissues were normal. Acid-fast bacteria were not found in the tissues and not re-isolated from the tissues. Our data suggest that M. marinum NJB 0419 might not be a strong pathogen of mammals, although more intensive studies are required for confirmation of the pathogenicity.

Key words: Mycobacterium marinum, Seriola dumerili, Pagrus major, mouse

Atypical Aeromonas salmonicida Infection in Sailfin Sandfish

<>Shinpei Wada1*, Kazuki Ozeki1, Osamu Kurata1, Kishio Hatai1, Aya Ishida2 and Kenjiroh Ui2<>1Laboratory of Fish Diseases, Nippon Veterinary and Life Science University, Tokyo 180-8602, Japan
2Oga Aquarium GAO, Akita 010-0673, Japan
(Received April 19, 2010)

ABSTRACT―In 2009, a disease with cumulative mortality of 52.5% was observed in a captive population of sailfin sandfish Arctoscopus japonicus reared at an aquarium in Japan. The affected fish had no apparent lesions on the body surfaces, with the exception of skin abrasions on the lower jaw. The trunk kidney was swollen and bore white miliary nodules. The most prominent histopathological feature of the diseased fish was numerous bacterial colonies in the internal organs. However, no inflammatory cellular response was apparent. Single species of bacteria was isolated from the trunk kidney and spleen, and identified as atypical A. salmonicida based on the biochemical and serological characteristics.

Key words: Arctoscopus japonicus, atypical Aeromonas salmonicida, sailfin sandfish