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NO45 - 3

Long-term Dynamics of Infection of Fish Eggs and Larvae with the Endoparasite Ichthyodinium sp. (Dinoflagellata) in Nha Trang Bay, Vietnam

<>A. M. Shadrin1*, D. S. Pavlov2 and M. V. Kholodova2<>1M. V. Lomonosov Moscow State University, Vorob弾vy Gory, Moscow 119992, Russia
2A. N. Severtsov Institute of Ecology and Evolution, Leninsky prospect,
33 Moscow 117071, Russia
(Received August 31, 2009)
ABSTRACT―Results of long-term investigation of infection prevalence of fish eggs and larvae with the endoparasite Ichthyodinium sp. (Dinoflagellata) in Nha Trang Bay (South China Sea, Vietnam) are presented. The parasite was identified on the base of morphological characteristics and phylogenetic analysis with 18S rRNA sequences. After the first record in 1993 the infection rate steadily increased till 2004, remainded high in 2004-2007, decreased in 2008-2009 and again boosted in 2010. Seasonal and annual dynamics of the infection rate was described. The parasite showed different degrees of prevalence in different taxonomic groups of fishes. The infection rates were different among closely related species. This is the first report on long-term dynamics of Ichthyodinium infection in Southeast Asia.

Key words: Ichthyodinium, Dinoflagellate, endoparasite, phylogenetic analysis, prevalence, Vietnam

In vitro and In vivo Efficacy of Antifungal Agents against Acremonium sp.

<>Pham Minh Duc1,2, Shinpei Wada1, Osamu Kurata1 and Kishio Hatai1*<>1Laboratory of Fish Diseases, Nippon Veterinary and Life Science University,
Tokyo 180-8602, Japan
2College of Aquaculture and Fisheries, Cantho University, Can Tho City, Viet Nam
(Received April 8, 2010)

ABSTRACT―Acremonium sp. NJM 0672, isolated from diseased mantis shrimp Oratosquilla oratoria, was susceptible in vitro to three kinds of antifungal agents: voriconazole, amphotericin B and terbinafine hydrochloride. Voriconazole was selected to treat kuruma prawn Penaeus japonicus, which had been intramuscularly injected with 0.1 mL of 5.0 × 104 conidia/mL of Acremonium sp. Voriconazole was administered orally at doses of 6 and 2 mg/kg body weight per day for 7 consecutive days, or intramuscularly injected at doses of 4 and 2 mg/kg body weight per day for 3 consecutive days. Both treatments were begun at 6 h after injection of the conidial suspension. The gross features, mortality and histopathological findings demonstrated that voriconazole was an efficient antifungal agent against Acremonium sp.

Key words: Acremonium sp., antifungal agent, voriconazole, Penaeus japonicus, kuruma prawn, Oratosquilla oratoria

Monitoring of Outbreaks of Bacterial Cold Water Disease among Ayu in a River Where Asymptomatic Carriers of Flavobacterium psychrophilum Were Released

<>Akira Kumagai1*, Akatsuki Nawata2 and Yuichi Taniai2<>1Miyagi Prefecture Fisheries Technology Institute, Miyagi 986-2135, Japan
2Miyagi Prefecture Fisheries Technology Institute, Freshwater
Fisheries Experimental Station, Miyagi 981-3625, Japan
(Received May 7, 2010)

ABSTRACT―Occurrences of bacterial cold-water disease (BCWD) caused by Flavobacterium psychrophilum were monitored among released and wild ayu Plecoglossus altivelis in Hirose River in 2006 and 2007 in order to clarify the infection source. When hatchery-reared juvenile ayu were examined immediately prior to release, F. psychrophilum was not isolated. However, when fish from the same lots were reared until maturation, the bacterium was isolated from ripe fish, indicating that the released juvenile ayu were carriers. In epizootiological investigations in Hirose River, outbreaks of BCWD occurred from June to August in both years, but the source of BCWD seemed to differ: released ayu in 2006 and wild ayu in 2007. Additionally, the disease occurred in released ayu by the pathogen carried by themselves in 2007. The disease subsided in August when water temperature exceeded 20ーC. Thereafter, the prevalence of F. psychrophilum in ripe fish in the river increased to above 90% between October and November in both years.

Key words: bacterial cold-water disease, Flavobacterium psychrophilum, Plecoglossus altivelis, ayu, infection source, BCWD, epizootiology

Activity of Granulocytes and Chemokines in the Leukocyte-encapsulation Response of Japanese Flounder Paralichthys olivaceus

<>Osamu Kurata1*, Nilubol Kitancharoen2, Atushi Fujiwara3, Chihaya Nakayasu3,Shinpei Wada1 and Kishio Hatai1<>1Laboratory of Fish Diseases, Department of Veterinary Science, Nippon Veterinary and Life Science University, Tokyo 180-8602, Japan
2Department of Fisheries, Faculty of Agriculture, Khon Kaen University, Tambon Naimuang,
Amphur Muang, Khon Kaen 40002, Thailand
3National Research Institute of Aquaculture, Fisheries Research Agency,
Mie 519-0423, Japan
(Received July 1, 2010)

ABSTRACT―The role of leukocytes from the kidney of Japanese flounder Paralichthys olivaceus in the encapsulation response was investigated using an in vitro model. The aggregation response of leukocytes to the target fish pathogen Ichthyophonus hoferi was observed over 12 h. The cellular aggregates mainly comprised granulocytes, which were characterized by intracellular peroxidase expression. Moreover, studies of isolated granulocytes demonstrated their adhesion to I. hoferi and formation of an initial layer in the encapsulation response. We observed that mRNA expression levels of three CC-chemokines (CCL3, CCL4 and CC-CLM) and one CXC-chemokine (IL-8) increased during the encapsulation process. Among these chemokines, CC-CLM and IL-8 were actively produced by granulocytes upon stimulation by I. hoferi. Flounder granulocytes therefore appeared to play an important role in the process by releasing specific chemokines upon pathogen recognition, thereby inducing subsequent cellular recruitment leading to encapsulation.

Key words: cytokine, Paralichthys olivaceus, granulomatous inflammation, leukocyte aggregation, neutrophil, peroxidase

Detection and Identification of Miamiensis avidus Causing Scuticociliatosis by PCR

<>Nahoko Tange1*, Jun-Young Song2,3 and Shin-Ichi Kitamura2,3<>1Department of Coastal Fisheries, Tottori Prefectural Fisheries Experimental Station,
Tottori 689-0602, Japan
2Graduate School of Science and Engineering, Ehime University, Ehime 790-8577, Japan
3Centre for Marine Environmental Studies (CMES), Ehime University, Ehime 790-8577, Japan
(Received December 4, 2009)

ABSTRACT―We designed a PCR primer set for specific detection of Miamiensis avidus causing scuticociliatosis in Japanese flounder Paralichthys olivaceus. In the PCR targeting the small subunit ribosomal RNA gene of M. avidus, the expected PCR product with 1,433 bp was amplified from four isolates of M. avidus including three different serotypes, but not from other scuticociliates including Pseudocohnilembus persalinus, P. hargisi and Uronema marinum. Detection limit of the present PCR was 125 fg of genomic DNA. When a group of Japanese flounder suffering the disease was examined by the PCR, 90% of symptomatic fish and 50% of asymptomatic fish were found positive. Thus it is considered that the PCR is useful for detection and identification of M. avidus and consequently for diagnosis of scuticociliatosis in Japanese flounder.

Key words: scuticociliatosis, Miamiensis avidus, PCR, Paralichthys olivaceus, detection

Pathogenicity of Plectosporium oratosquillae and Acremonium sp. Isolated from Mantis Shrimp Oratosquilla oratoria against Kuruma Prawn Penaeus japonicus

<>Pham Minh Duc, Shinpei Wada, Osamu Kurata and Kishio Hatai*<>Laboratory of Fish Diseases, Nippon Veterinary and Life Science University, Tokyo 180-8602, Japan
(Received March 1, 2010)

ABSTRACT―This study was carried out to assess the pathogenicity of Plectosporium oratosquillae NJM 0662 and Acremonium sp. NJM 0672, both of which were isolated from mantis shrimp Oratosquilla oratoris, to kuruma prawn Penaeus japonicus by intramuscular injection of conidial suspensions. These fungi caused mortality in the injected kuruma prawn. The diseased kuruma prawn showed numerous black spots in the gills. Histopathologically, hyphae in the gill filaments and the injected sites were encapsulated by hemocytes. The results indicate that these two fungi are pathogenic against kuruma prawn.

Key words: Plectosporium oratosquillae, Acremonium sp., Penaeus japonicus, pathogenicity, kuruma prawn

Nephroblastoma in Two Siamese Fighting Fish Betta splendens

<>Eric D. Lombardini1*, Mac Law2 and Bridget S. Lewis1<>1Armed Forces Institute of Pathology (AFIP), Department of Veterinary Pathology, Washington, DC 20306, USA
2North Carolina State University, Laboratory for Toxicological and Molecular Pathology, Raleigh, NC 27606, USA
(Received March 4, 2010)

ABSTRACT―In 2005 and 2008, renal nephroblastomas were diagnosed in two adult male Siamese fighting fish Betta splendens. In both cases the masses effaced approximately 90 percent of renal tissue and histopathologically, were composed of a triptych of diagnostic features, to include: nests of cells suggestive of blastema, a primitive mesenchyme as well as an epithelial component forming glomeruloid structures. To our knowledge, these cases represent the first reports of nephroblastoma in Siamese fighting fish.

Key words: Nephroblastoma, Wilms・tumor, Betta splendens

Improvement of PCR Targeting 16S Ribosomal DNA of Piscirickettsia salmonis

<>Takamitsu Sakai1*, Akira Kumagai2, Hiroto Ohta2, Norihisa Oseko1, Motohiko Sano1 and Takaji Iida1<>1National Research Institute of Aquaculture, Fisheries Research Agency, Mie 516-0193, Japan
2Miyagi Prefecture Fisheries Technology Institute, Miyagi 986-2135, Japan
(Received April 26, 2010)

ABSTRACT―Detection of Piscirickettsia salmonis by the PCR targeting 16S rDNA has been applied to the diagnosis for piscirickettsiosis. However, some nucleotide substitutions were observed in the primer regions of the published PCR among several strains of P. salmonis. Thus, we designed a new PCR primer set targeting the 16S rDNA regions and a modified PCR program. The predicted amplicon was specifically generated from P. salmonis but not from 19 other bacterial species. Sensitivity of the present PCR system was also improved. It was confirmed that P. salmonis was detectable in the kidney and liver of the experimentally infected coho salmon Oncorhynchus kisutch by the present system.

Key words: Piscirickettsia salmonis, PCR, 16S rDNA

Multiple Drug Resistant Isolates of Vibrio Carrying the Transferable R Plasmid from Shrimp Farms in Thailand

<>Nutcharnart Tipmongkolsilp, Hidehiro Kondo, Ikuo Hirono and Takashi Aoki*<>Laboratory of Genome Science, Graduate School of Marine Science and Technology, Tokyo University of Marine Science and Technology, Tokyo 108-8477, Japan
(Received June 16, 2010)

ABSTRACT―One thousand forty-nine bacterial isolates, which were collected from shrimp farms in Thailand, were resistant to antimicrobial agents: ampicillin (ABPC), chloramphenicol (CP), kanamycin (KM), oxolinic acid (OA), streptomycin (SM) and tetracycline (TC). One hundred forty-two isolates were identified as Vibrio. These isolates were conjugated with Escherichia coli for detection of transferable R plasmid. Six patterns of transferable R plasmids, which encoded resistance to the combinations of one to five drugs (ABPC, CP, SM, sulfamonomethoxine, TC and trimethoprim), were found from 16 drug-resistant isolates of Vibrio. This is the first report of transferable R plasmids from Vibrio spp. in shrimp farms in Thailand.

Key words: Vibrio, antimicrobial resistance, transferable R plasmid, shrimp