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Motokawa, Shogo,Narasaki, Yukie,Song, Jun-Young,Yokoyama, Yoshihiro,Hirose, Euichi,Murakami, Shoko,Jung, Sung-Ju,Oh, Myung-Joo,Nakayama, Kei,Kitamura, Shin-Ichi Elsevier 2018 Parasitology international Vol.67 No.2
<P><B>Abstract</B></P> <P>The ciliate <I>Miamiensis avidus</I> causes scuticociliatosis in Japanese flounder <I>Paralichthys olivaceus</I>. We previously reported three serotypes of this ciliate distinguishable by serotype-specific antigenic polypeptides (serotype I, 30kDa; serotype II, 38kDa; serotype III, 34kDa). In this study, we determined the localization site of the serotype-specific polypeptides in the ciliate and determined the genes encoding the polypeptides, using the isolates IyoI (serotype I), Nakajima (serotype II), and Mie0301 (serotype III). SDS-PAGE and immunoblot analysis of cilia, membrane proteins, and cytoskeletal elements of the ciliates revealed that the polypeptides were abundant in the former two. Scanning electron microscopy of ciliates immobilized by homologous antiserum showed morphological changes in the cilia. These evidences suggested that the polypeptides were ciliary membrane immobilization antigens. The ciliary genes identified showed low identity scores—<51.5% between serotypes. To differentiate the serotypes, we designed serotype-specific PCR primer sets based on the DNA sequences. The PCR-based serotyping results were completely consistent with conventional serotyping methods (immobilization assay and immunoblot analysis). Twenty of 21 isolates were classified as either serotype I or II, and one isolate was undistinguishable. The combination of species-specific PCR previously reported and three serotype-specific PCR could be useful for identifying, serotyping, and surveillance for occurrences of new serotypes of <I>M. avidus</I>.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Three serotypes of <I>M. avidus</I> were reported in Japan and Korea. </LI> <LI> Serotype-specific polypeptides were identified as ciliary membrane proteins. </LI> <LI> The nucleotide sequences of ORFs were determined. </LI> <LI> Serotype-specific PCR revealed that the pandemic serotypes were serotype I and II. </LI> </UL> </P>
Kimura, Satoshi,Nakayama, Kei,Wada, Masahisa,Kim, Ung-Jin,Azumi, Kaoru,Ojima, Takao,Nozawa, Akino,Kitamura, Shin-Ichi,Hirose, Euichi Inter-Research 2015 Diseases of aquatic organisms Vol.116 No.2
<P>Soft tunic syndrome is a fatal disease in the edible ascidian Halocynthia roretzi, causing serious damage to ascidian aquaculture in Korea and Japan. In diseased individuals, the tunic, an integumentary extracellular matrix of ascidians, softens and eventually tears. This is an infectious disease caused by the kinetoplastid flagellate Azumiobodo hoyamushi. However, the mechanism of tunic softening remains unknown. Because cellulose fibrils are the main component of the tunic, we compared the contents and structures of cellulose in healthy and diseased tunics by means of biochemical quantification and X-ray diffractometry. Unexpectedly, the cellulose contents and structures of cellulose microfibrils were almost the same regardless of the presence or absence of the disease. Therefore, it is unlikely that thinning of the microfibrils occurred in the softened tunic, because digestion should have resulted in decreases in crystallinity index and crystallite size. Moreover, cellulase was not detected in pure cultures of A. hoyamushi in biochemical and expressed sequence tag analyses. These results indicate that cellulose degradation does not occur in the softened tunic.</P>