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NO50 - 4

Paradeontacylix buri n. sp. (Trematoda: Aporocotylidae) from Seriola quinqueradiata cultured in Japan with a description of unidentified Paradeontacylix sp. from S. lalandi

Kazuo Ogawa1*, Kousuke Akiyama2 and Daniel Grabner3

2Marine Biological Technology Centre, Nippon Suisan Kaisha, Ltd., Oita 876-1204, Japan
3Department of Aquatic Ecology, University of Duisburg-Essen, D-45141 Essen, Germany

ABSTRACTParadeontacylix buri n. sp. is described based on specimens from the afferent branchial arteries of the Japanese amberjack Seriola quinqueradiata cultured in Mie and Oita Prefectures, Japan. The new species can be differentiated from congeners by a body of up to 4.15 mm with lanceolate tegumental spines of the same size throughout the body, uterus ascending after leaving the oötype and the vitellarium not extending posterior to the ovary. P. buri n. sp. is unique among Paradeontacylix species from Seriola spp. in having the same size tegumental spines throughout the body. A phylogenetic analysis based on the sequences of the internal transcribed spacer 2 and on the 28S rRNA-gene demonstrated that P. buri is grouped with other Paradeontacylix species from Seriola spp., indicating that the enlarged tegumental spines in the posteriormost rows in the other Paradeontacylix spp. from Seriola spp. are not a morphological feature at the generic level. Another blood fluke, Paradeontacylix sp., is described based on a single specimen from the afferent branchial artery of yellowtail amberjack S. lalandi cultured in Oita Prefecture, Japan. This species may be differentiated from the most similar P. buri n. sp. by the cirrus (conical and thick walled in Paradeontacylix sp. vs. spherical and thin-walled in P. buri) and by the uterus (extending posteriorly up to the level of the oötype vs. extending posterior to the oötype).

Key words: Paradeontacylix buri, blood fluke, Seriola quinqueradiata, Seriola lalandi, Japanese amberjack, yellowtail amberjack, new species

Structure of Genetic Loci for Capsular Polysaccharide Biosynthesis in Streptococcus parauberis isolated from Japanese flounder

Chuandeng Tu, Koushirou Suga and Kinya Kanai*

Graduate School of Fisheries and Environmental Sciences, Nagasaki University, Nagasaki 852-8521, Japan

ABSTRACTStreptococcus parauberis is a pathogen of streptococcosis in turbot Scophthalmus maximus and the Japanese flounder Paralichthys olivaceus. S. parauberis isolates from diseased Japanese flounder have been classified into several serological phenotypes. In this study we analyzed the DNA sequence of the genetic locus for capsular polysaccharide (CPS) biosynthesis of S. parauberis KRS02083 (subserotype Ia), NUF1003 (subserotype Ib), NUF1071 (subserotype Ic), NUF1032 (serotype II), 2007-1 (nontypeable/PFGE cluster I) and NUF1095 (nontypeable/PFGE cluster III) to elucidate the genetic basis for serological diversity. As a result, three kinds of cps locus were revealed among serotypes and subserotypes, namely the loci for subserotype Ia, subserotypes Ib/Ic and serotype II. The genetic structure of cps loci suggests that the capsules of S. parauberis are synthesized through the Wzy-dependent pathway. Subserotypes Ib and Ic possessed the same genetic structure, although single-base substitution at several regions or insertion of an IS (insertion sequence) element was found in subserotype Ic. The nontypeable strains, which agglutinated with both serotypes I and II antisera, possessed the same genetic structure as subserotype Ib/Ic or serotype II with single-base substitution at several regions.

Key words: Streptococcus parauberis, streptococcosis, serotype, capsular polysaccharide biosynthesis, cps locus, structure, Paralichthys olivaceus

Pathogenicity and Immunogenicity of Non-agglutinating Lactococcus garvieae with Anti-KG Phenotype Rabbit Serum in Seriola spp.

Yutaka Fukuda1*, Yuya Tue2, Daisaku Oinaka2, Yoshinobu Wada3, Azumi Yamashita4, Shintaro Urasaki5, Sosuke Yoshioka1, Keisuke Kimoto1 and Terutoyo Yoshida2

1Fisheries Research Division, Oita Prefectural Agriculture, Forestry and Fisheries Research Center, Oita 879-2602, Japan
2Faculty of Agriculture, University of Miyazaki, Miyazaki 889-2192, Japan
3Intervet K.K. Central Research Laboratories, Ibaraki 300-0013, Japan
4Fisheries Research Center, Ehime Research Institute of Agriculture, Forestry and Fisheries, Ehime 798-0104, Japan
5Fisheries Division, Ainan Town Office, Ehime 798-4196, Japan

ABSTRACT―Since 2012, Lactococcus garvieae strains which do not agglutinate with anti-KG phenotype rabbit serum have been isolated from cultured yellowtail Seriola quinqueradiata and greater amberjack S. dumerili in Japan. In this study, pathogenicity of non-agglutinating L. garvieae was confirmed by intraperitoneal injection into these fish species. Cross-protective responses of yellowtail to the KG phenotype and non-agglutinating strains were also examined. Yellowtail were immunized with formalin-killed cells (FKC) prepared from both types. Three weeks after immunization, the fish were challenged with the KG phenotype and non-agglutinating strains. Each type of FKC provided effective protection against infection with the homologous strain. The protection level of the KG phenotype FKC was relatively lower against the non-agglutinating strain than against the homologous KG phenotype and the non-agglutinating type FKC was ineffective against the KG phenotype. In conclusion, we propose two serotypes (I and II) of L. garvieae isolated from marine fish species in Japan. Serotype I which agglutinates with anti-KG phenotype serum is typical L. garvieae and divided into Ia and Ib, which are KG (capsulated) and KG+ (non-capsulated) phenotypes, respectively. Serotype II which does not agglutinate with the anti-KG- serum is a new type in fish pathogenic L. garvieae.

Key words: Lactococcus garvieae, non-agglutinating, serotype, pathogenicity, immunogenicity, Seriola quinqueradiata, Seriola dumerili

Fimbriae Expression by Edwardsiella tarda in High-salt Culture Conditions

Indah Istiqomah1, Yasuhiko Kawato2, Mahmoud Mostafa Mahmoud3, Jun Okuda4, Motoshige Yasuike5, Yoji Nakamura5, Atushi Fujiwara5, Koichi Sahiro6, Masahiro Sadakane6 and Toshihiro Nakai1*

1Graduate School of Biosphere Science, Hiroshima University, Hiroshima 739-8528, Japan 2Diagnosis and Training Center for Fish Diseases, National Research Institute of Aquaculture, Fisheries Research Agency, Mie 516-0193, Japan 3Faculty of Veterinary Medicine, Assiut University, Assiut 71526, Egypt 4Department of Microbiology, Kagawa Prefectural University of Health Sciences, Kagawa 761-0123, Japan 5Research Center for Aquatic Genomics, National Research Institute of Fisheries Science, Fisheries Research Agency, Kanagawa 236-8648, Japan 6Graduate School of Engineering, Hiroshima University, Hiroshima 739-8527, Japan

ABSTRACTEdwardsiella tarda causes a serious disease known as edwardsiellosis in farmed freshwater and marine fishes. We previously reported that the hemagglutination and cell adherence activities of E. tarda increase in high-salt culture conditions (3% NaCl). This paper describes the induction of fimbriae expression in E. tarda in such high-salt (3% NaCl) culture conditions, using two different phenotypic strains of E. tarda, FK1051 (motile) and MEE0309 (non-motile), isolated from diseased fish. Both strains exhibited faster growth in liquid medium supplemented with 0% to 2% NaCl and slower growth in the 3% NaCl conditions. Hemagglutination activity against guinea pig erythrocytes was detected only in the 2% NaCl and/or 3% NaCl cultures. Electron microscopy revealed two types of fimbriae. The first type was thick (ca. 9 nm) and appeared only in the 0% NaCl culture of FK1051, and the second type was thin (ca. 4 nm) and appeared in the 3% cultures of both strains. The expressions of the major fimbrial subunit gene (etfA) in both strains were significantly higher in the 3% NaCl cultures than in the 0% NaCl cultures. The present results suggest that the thin type of fimbriae is involved in the cell adherence of E. tarda.

Key words: Edwardsiella tarda, fimbriae, major fimbrial subunit gene, etfA, adherence, hemagglutination, electron microscopy

A Multiplex PCR Assay for Differentiation of Streptococcus parauberis Serotypes

Chuandeng Tu, Koushirou Suga and Kinya Kanai*

ABSTRACT―A multiplex PCR assay for differentiation of Streptococcus parauberis serotypes was developed. The three primer pairs for subserotypes Ia and Ib/Ic and serotype II were designed from the serotype-specific sequences of the wzy gene in the loci for capsular polysaccharide biosynthesis. All of 188 S. parauberis isolates from Japanese flounder showed positive reaction with the expected size of PCR product for each serotype, which was consistent with the results of agglutination test using rabbit antisera. The nontypeable isolates, which are not differentiated by agglutination test, could be identified as subserotype Ib/Ic or serotype II. Other streptococci including S. parauberis derived from cow and major bacterial fish pathogens showed negative reaction.

Key words: Streptococcus parauberis, multiplex PCR, serotyping, streptococcosis