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NO40 - 1 (2005)

Diseases of Cultured Kuruma Shrimp in Japan: a Review

<>Kazuo Momoyama1* and Kiyokuni Muroga2<>1Inland Sea Division, Yamaguchi Prefectural Fisheries Research Center, Aio-Futashima,
Yamaguchi 754-0893, Japan
2Graduate School of Agricultural Science, Tohoku University,
Sendai 981-8555, Japan
(Received January 24, 2005)
ABSTRACT輸fter the establishment of a large scale production method for penaeid shrimp larvae in the middle of 1960s, the culture production of kuruma shrimp Penaeus japonicus rapidly increased and attained the peak of 3,020 metric tons in 1988 in Japan. Development of formulated diet and introduction of a double-harvest system were the main factors for the expansion. In addition to aquaculture, 300 million juveniles of P. japonicus have been produced at public sea farming centers and stocked in coastal waters to enhance the natural resources every year since the late 1970s. The number of shrimp pathogens reported in Japan is smaller than that reported from overseas, mainly because almost one shrimp species, P. japonicus has been farmed in Japan. Baculoviral mid-gut gland necrosis (BMN), penaeid acute viremia (PAV) (= white spot disease: WSD) and vibriosis (Vibrio penaeicida infection) have been known as major infectious diseases of cultured shrimp in Japan. BMN was a serious problem in larval production at hatcheries from the early 1970s to the mid 1980s, but the disease has not occurred since 1993, due to the dissemination of the practical countermeasure, egg washing. Vibriosis was prevalent especially from the late 1980s to the early 1990s when the culture system became intensive, and the losses by this disease were estimated to be 20-30% of annual shrimp production. PAV, which was introduced into Japan with live stock of young P. japonicus from China in 1993, has been not only giving serious economic losses to the shrimp aquaculture industry, but also causing various troubles in sea farming operation for shrimp. In this paper eight infectious and three non-infectious diseases of P. japonicus reported in Japan are reviewed.

Key words: review, Penaeus japonicus, kuruma shrimp, BMN, vibriosis, PAV, WSD

The Efficacy of Inactivated Virus Vaccine against Viral Nervous Necrosis (VNN)

<>Hirofumi Yamashita1, Yoshiyuki Fujita1, Hidemasa Kawakami2 and Toshihiro Nakai3*<>1Ehime Prefectural Fisheries Experimental Station, Uwajima 798-0104, Japan
2Ehime Prefectural Fish Disease Control Center, Uwajima 798-0087, Japan
3Graduate School of Biosphere Science, Hiroshima University,
Higashihiroshima 739-8528, Japan
(Received November 18, 2004)
ABSTRACT裕he efficacy of inactivated betanodavirus as a vaccine against viral nervous necrosis (VNN) was evaluated using juvenile sevenband grouper Epinephelus septemfasciatus. Fish were intraperitoneally injected once with formalin-inactivated redspotted grouper nervous necrosis virus (RGNNV). Virus-neutralizing antibodies were detected in the vaccinated fish from Day 10 to the end of the experimental period (Day 160), showing 1: 2,000 or higher mean antibody titers from Day 21 to Day 77. The vaccinated and unvaccinated control fish were challenged by intramuscular injection with the homologous virus at 14, 35 and 74 days post-vaccination. The vaccinated fish showed significantly lower mortalities at any challenges than the control fish, with the RPS (relative percent survival) values 67 or higher. A field trial, in which fish were exposed to natural infection in net pens, also resulted in higher survival rates in the vaccinated fish (RPS = 85) during the experimental period of 9 weeks. This high induction of neutralizing antibodies and protection indicates the potential of the inactivated virus vaccine against VNN.

Key words: betanodavirus, viral nervous necrosis, viral encephalopathy and retinopathy, vaccination, Epinephelus septemfasciatus, inactivated vaccine, VNN, VER

Development of a PCR-based Method for the Detection of Enteric Myxozoans Causing the Emaciation Disease of Cultured Tiger Puffer

<>Tetsuya Yanagida1, Mark A. Freeman1, Yoshinori Nomura2, Ikuo Takami3, Yukitaka Sugihara3, Hiroshi Yokoyama1* and Kazuo Ogawa1<>1Department of Aquatic Bioscience, Graduate School of Agricultural and Life Sciences,
The University of Tokyo, Tokyo 113-8657, Japan
2Kumamoto Prefectural Fisheries Research Center, Amakusa,
Kumamoto 869-3603, Japan
3Nagasaki Prefectural Institute of Fisheries, Taira,
Nagasaki 851-2213, Japan
(Received December 14, 2004)
ABSTRACT輸 polymerase chain reaction (PCR)-based method was developed to detect enteric myxozoans which cause the emaciation disease of cultured tiger puffer Takifugu rubripes. Three primer sets were designed based on the sequences of small subunit ribosomal DNA of Enteromyxum leei, Enteromyxum fugu and Leptotheca fugu. All of the primer sets specifically
amplified the target DNA of each species. The PCR successfully detected parasite DNA not only from the intestinal mucosa of killed fish but also from gut contents collected from live fish using a swab inserted into the anus. These PCR analyses were more sensitive to detection of parasites than microscopic observation on the intestinal imprints. The present PCR method is useful for rapid diagnosis of the myxosporean emaciation disease.

Key words: Enteromyxum, Leptotheca, PCR, SSU rDNA, Takifugu rubripes, myxosporean emaciation disease, diagnosis

Virus Surveillance of Wild Marine Fish Collected in Coastal Areas of Hokkaido, Japan

<>Kenichiro Kobayashi, Jacob A. Wani, Hisae Kasai, Toyohiko Nishizawa and Mamoru Yoshimizu*<>Graduate School of Fisheries Sciences, Hokkaido
University, Hakodate, 041-8611 Japan

(Received August 16, 2004)

ABSTRACT輸 total 645 wild marine fish of 40 species were collected at the north (Wakkanai and Haboro), south (Hakodate) and east (Akkeshi) coasts of Hokkaido from 2000 to 2004 to conduct surveillance of fish viruses. Four fish cell lines (RTG-2, EPC, CHSE-214, FHM) were used for virus isolation from the brain, kidney and spleen of fish. Aquabirnavirus, which was identified as yellowtail ascites
virus by a neutralization test, was isolated only from ten fish of four species caught at Akkeshi coastal area in 2002.
These species were saffron cod Eleginus gracilis, snowy sculpin Myoxocephalus blandti, Japanese dace Tribolodon hakonensis and rainbow smelt Osmerus eperlanus mordax. Other viruses including viral hemorrhagic septicemia virus were not isolated.

Key words: fish virus, surveillance, aquabirnavirus, YTAV, wild fish

Larval Attachment and Development of the Monogenean Neoheterobothrium hirame under Low Water Temperature

<>Sho Shirakashi1, Tomoyoshi Yoshinaga1, Masakazu Oka2 and Kazuo Ogawa1*<>1Department of Aquatic Bioscience, Graduate School of Agricultural and Life Sciences, The University of
Tokyo, Tokyo 113-8657, Japan
2Kamiura Station, National Center for Stock Enhancement, Fisheries Research Agency, Oita 879-2602, Japan

(Received October 10, 2004)

ABSTRACT勇ffects of low water temperature (below 10゚C) on the oncomiracidial attachment and its subsequent development of Neoheterobothrium hirame were investigated. The cumulative attached larvae to the gill pieces from olive flounder Paralichthys olivaceus was reduced by 30% at 5゚C when compared with 20゚C. At 8゚C the parasite development on flounder was significantly retarded compared at 20゚C, and considerable number of worms disappeared from the host before reaching maturation. These results suggest low water temperature is a factor limiting the population growth of N. hirame in cold temperature regions.

Key words: Neoheterobothrium hirame, Paralichthys olivaceus, water temperature, survival, development

Improvement of a PCR Method with the Sph I-5 Primer Set for the Detection of Koi Herpesvirus (KHV)

<>Kei Yuasa1*, Motohiko Sano1, Jun Kurita1, Takafumi Ito1 and Takaji Iida2<>1Inland Station, National Research Institute of Aquaculture, Fisheries Research Agency,
Tamaki, Mie 519-0423, Japan
2National Research Institute of Aquaculture, Fisheries Research Agency,
Nansei, Mie 516-0193, Japan
(Received February 2, 2005)
ABSTRACT悠mprovement of a PCR method using the Sph I-5 primer set, which has been used for detection of KHV in Japan, was carried out to reduce negative factors, such as the appearance of non-specific reactions and the comparatively long reaction time. Moreover, there was a wrong
sequence in the original reverse primer. The modified PCR protocol with the corrected primer set is as follows: initial denaturation at 94゚C for 30 s, 40 cycles of amplification (denaturation at 94゚C for 30 s, annealing at 63゚C for 30 s and elongation at 72゚C for 30 s) and final elongation at 72゚C for 7 min. The improved PCR reduced non-specific reactions and the total reaction time to almost half reguired by the original one.

Key words: polymerase chain reaction, PCR, koi herpesvirus, KHV, diagnosis