論文 - 田中 龍聖
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Ryusei Tanaka, Natsumi Kanzaki, Robin M. Giblin-Davis
Nematology 27 ( 1 ) 37 - 48 2024年11月
担当区分：筆頭著者 記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Nematology
Schistonchus pumilae was first reported from the syconi of a creeper fig, Ficus pumila, from Miyazaki, Japan, and the species was described based on propagative (plant-parasitic) females and males. Thus, the premature female (entomophilic form) of the species was not described. In the present study, male syconi of F. pumila were collected from the type locality of the nematode (Miyazaki, Japan), and the premature females of S. pumilae were collected from inside the syconi and also from the emerging pollinator fig wasps (Wiebesia pumilae). Although only 26 female wasps were recovered for dissections, seven of them harboured 1-7 nematodes each. Nematodes associated with the wasp were all premature females, and their typological and morphometric characters were similar with those isolated from inside the syconi. In addition, the premature females were slightly smaller than propagative females and had a less developed gonad, suggesting that the nematode is a phoretic associate, not a parasite. Regardless of its phylogenetic status, the carrier insect usage of S. pumilae is somewhat similar to two Ficophagus species, and clearly different from S. caprifici, which propagates in the host wasp haemocoel as a parasite. Additionally, the molecular profiles of S. pumilae (SSU and D1-D3 LSU of ribosomal RNA genes) were re-sequenced using a cloning technique, and the information was updated for phylogenetic analysis in the future.
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Kawahara K, Inada T, Tanaka R, Dayi M, Makino T, Maruyama S, Kikuchi T, Sugimoto A, Kawata M
Genome biology and evolution 15 ( 4 ) 2023年4月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Genome biology and evolution
Why the recently discovered nematode Caenorhabditis inopinata differs so greatly from its sibling species Caenorhabditis elegans remains unknown. A previous study showed that C. inopinata has more transposable elements (TEs), sequences that replicate and move autonomously throughout the genome, potentially altering the expression of neighboring genes. In this study, we focused on how the body size of this species has evolved and whether TEs could affect the expression of genes related to species-specific traits such as body size. First, we compared gene expression levels between C. inopinata and C. elegans in the L4 larval and young adult stages-when growth rates differ most prominently between these species-to identify candidate genes contributing to their differences. The results showed that the expression levels of collagen genes were consistently higher in C. inopinata than in C. elegans and that some genes related to cell size were differentially expressed between the species. Then, we examined whether genes with TE insertions are differentially expressed between species. Indeed, the genes featuring C. inopinata-specific TE insertions had higher expression levels in C. inopinata than in C. elegans. These upregulated genes included those related to body size, suggesting that these genes could be candidates for artificial TE insertion to examine the role of TEs in the body size evolution of C. inopinata.
DOI： 10.1093/gbe/evad063
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Ko P.P., Haraguchi M., Hara T., Hieu D.D., Ito A., Tanaka R., Tanaka M., Suzumura T., Ueda M., Yoshida A., Maruyama H., Nagayasu E.
Parasitology International 92 102663 2023年2月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Parasitology International
Strongyloides is a genus of parasitic nematodes of vertebrates comprising approximately 50 documented species, each with various host ranges. Among these, three species (S. stercoralis, S. fuelleborni, and S. cebus) are known to infect primate hosts. S. fuelleborni typically infects non-human primates in the Old World. To complement the existing information on the global genetic structure of this species, we conducted a genotyping study of S. fuelleborni samples collected from rhesus macaques in Myanmar, Japanese macaques in Japan, and some zoo-kept primates. This study identified a novel haplotype group in isolates from the Myanmar rhesus macaques. Subsequently, we obtained the complete or nearly complete mitochondrial genome sequences of S. fuelleborni, S. cebus (Strongyloides of New World monkeys), and S. vituli (Strongyloides of cattle). Phylogenetic analysis based on concatenated mitochondrial protein sequences of various Strongyloides species indicated a close relationship between S. fuelleborni, S. vituli and S. papillosus (Strongyloides in sheep and cattle). S. cebus is quite distantly related to both S. fuelleborni and S. stercoralis, which led to the hypothesis that the three primate Strongyloides species evolved independently as parasites of primates.
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Kanzaki N., Tanaka R., Giblin-Davis R.M.
Nematology 25 ( 5 ) 509 - 529 2023年
掲載種別：研究論文（学術雑誌） 出版者・発行元：Nematology
A Schistonchus species was isolated from the syconia of the creeping fig, Ficus pumila, collected from Miyazaki, Japan. The nematode was considered an undescribed species based on its typological characters, molecular profiles (near full length of small subunit and D3 expansion segment of large subunit of RNA) and phylogenetic status inferred from these two loci. The new species is characterised by the presence of flagellate sperm, spicule possessing small condylus, triangular rostrum with bluntly pointed tip and clear dorsal and ventral limbs, structure and arrangement of male caudal papillae, i.e., presence of papilliform P3 and P3a and small and gland opening-like P4 (glandpapillae), relatively short post-uterine sac and female tail forming elongate conoid. It is phylogenetically close to S. hirtus. However, the new species is distinguished from other nominal Schistonchus species by its female tail shape forming elongate conoid, the structure and arrangement of male caudal papillae and several other typological characters. In addition, the host fig of the nematode, F. pumila, is a creeper fig species belonging to subgenus Synoecia (section Rhizocladus; subsection Plagiostigma), from which no fig-Associated nematode has been reported so far, i.e., the present study is the first report of Schistonchus (and other syconia-inhabiting nematodes) from the Ficus subgenus Synoecia. Keywords
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Sun S., Kanzaki N., Dayi M., Maeda Y., Yoshida A., Tanaka R., Kikuchi T.
BMC Genomics 23 ( 1 ) 765 2022年12月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：BMC Genomics
Background: The first metazoan genome sequenced, that of Caenorhabditis elegans, has motivated animal genome evolution studies. To date > 50 species from the genus Caenorhabditis have been sequenced, allowing research on genome variation. Results: In the present study, we describe a new gonochoristic species, Caenorhabditis niphades n. sp., previously referred as C. sp. 36, isolated from adult weevils (Niphades variegatus), with whom they appear to be tightly associated during its life cycle. Along with a species description, we sequenced the genome of C. niphades n. sp. and produced a chromosome-level assembly. A genome comparison highlighted that C. niphades n. sp. has the smallest genome (59 Mbp) so far sequenced in the Elegans supergroup, despite being closely related to a species with an exceptionally large genome, C. japonica. Conclusions: The compact genome of C. niphades n. sp. can serve as a key resource for comparative evolutionary studies of genome and gene number expansions in Caenorhabditis species.
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ノキシノブ葉面から分離された菌類
柴田 紗帆, 岸本 圭子, 田中 龍聖, 竹本 周平, 廣岡 裕吏
日本森林学会大会発表データベース 133 ( 0 ) 639 2022年5月
記述言語：日本語 掲載種別：研究論文（研究会，シンポジウム資料等） 出版者・発行元：日本森林学会
葉の表面を指す葉面（phylloplane）は，陸上微生物最大の生息場所の1つと考えられている．この葉面上には，植物を基質とする菌類だけでなく，葉面上の他の生物，花粉や塵といった堆積物などを基質とする多様な菌類種が存在すると考えられている．しかしながら，葉面に存在する菌類（葉面菌）は胞子や酵母，菌糸などの単純な形態で存在することや，その単純な形態ゆえに種同定が極めて難しいことから，生態学的役割について未解明な点が多い．そこで本研究では，葉面菌の生態学的役割解明に向けたそれら菌の同定基盤の構築を目指し，林内で容易に観察されるノキシノブ（常緑シダ植物の一種）の葉面より分離された菌類の多様性解明を行なった．異なる地域や時期に採取したノキシノブ71標本の葉面から420菌株を分離し，rDNA LSU領域の塩基配列を用いてそれら菌株の簡易同定を行なった．そして，そのうち分離頻度の高い17種について，詳細な種同定を行った結果，約半数にあたる8種は未記載種と考えられた．今後，葉面菌の生態学的役割を明らかにするためには，これら多様な菌類のさらなる分類学的研究が必要である．
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Dayi M., Kanzaki N., Sun S., Ide T., Tanaka R., Masuya H., Okabe K., Kajimura H., Kikuchi T.
Scientific Reports 11 ( 1 ) 6720 2021年12月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Scientific Reports
Caenorhabditis auriculariae, which was morphologically described in 1999, was re-isolated from a Platydema mushroom-associated beetle. Based on the re-isolated materials, some morphological characteristics were re-examined and ascribed to the species. In addition, to clarify phylogenetic relationships with other Caenorhabditis species and biological features of the nematode, the whole genome was sequenced and assembled into 109.5 Mb with 16,279 predicted protein-coding genes. Molecular phylogenetic analyses based on ribosomal RNA and 269 single-copy genes revealed the species is closely related to C. sonorae and C. monodelphis placing them at the most basal clade of the genus. C. auriculariae has morphological characteristics clearly differed from those two species and harbours a number of species-specific gene families, indicating its usefulness as a new outgroup species for Caenorhabditis evolutionary studies. A comparison of carbohydrate-active enzyme (CAZy) repertoires in genomes, which we found useful to speculate about the lifestyle of Caenorhabditis nematodes, suggested that C. auriculariae likely has a life-cycle with tight-association with insects.
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Nearly complete genome assembly of the pinewood nematode bursaphelenchus xylophilus strain Ka4C1 査読あり 国際共著
Dayi M., Sun S., Maeda Y., Tanaka R., Yoshida A., Tsai I.J., Kikuchi T.
Microbiology Resource Announcements 9 ( 42 ) 2020年10月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Microbiology Resource Announcements
Bursaphelenchus xylophilus has been destroying pine forests in East Asia and western Europe. Here, we report its nearly complete genomic sequence containing five ;12-Mb scaffolds and one ∼15-Mb scaffold representing six chromosomes. Large repeat regions that were previously unidentified are now reasonably integrated, particularly in the ∼15-Mb scaffold.
DOI： 10.1128/MRA.01002-20
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井龍 友美, 田中 龍聖, 吉賀 豊司
日本線虫学会誌 50 ( 1 ) 1 - 7 2020年9月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：日本線虫学会
シヘンチュウは様々な無脊椎動物の寄生虫であるが、その宿主と関係する生活史についての情報は限られている。我々は一化性で定着性のベニツチカメムシから分離されたシヘンチュウについて、生活環と寄生が宿主に及ぼす影響について調査した。シヘンチュウは6カ所中5カ所の採集場所で検出され、寄生率は0％から16％だった。多くの場合は単寄生だったが重寄生もみられた。シヘンチュウは終齢若虫から離脱し、宿主は離脱後にすぐに死亡した。本調査では、成虫からの離脱は観察されなかった。宿主とシヘンチュウの体重の間には相関はみられなかった。寄生の有無によって宿主の生体重や形態的・行動的変化はなく、離脱までは宿主は影響を受けないことが示唆された。系統解析によって、佐賀エリアのみ、および全ての異なるベニツチカメムシ個体群から検出される、2種類のシヘンチュウが存在することが明らかとなった。
DOI： 10.3725/jjn.50.1
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Mondal S.I., Akter A., Koga R., Hosokawa T., Dayi M., Murase K., Tanaka R., Shigenobu S., Fukatsu T., Kikuchi T.
Frontiers in Microbiology 11 840 2020年5月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Frontiers in Microbiology
Diverse animals, including insects, harbor microbial symbionts within their gut, body cavity, or cells. The subsocial parastrachiid stinkbug Parastrachia japonensis is well-known for its peculiar ecological and behavioral traits, including its prolonged non-feeding diapause period and maternal care of eggs/nymphs in an underground nest. P. japonensis harbors a specific bacterial symbiont within the gut cavity extracellularly, which is vertically inherited through maternal excretion of symbiont-containing white mucus. Thus far, biological roles of the symbiont in the host lifecycle has been little understood. Here we sequenced the genome of the uncultivable gut symbiont “Candidatus Benitsuchiphilus tojoi.” The symbiont has an 804 kb circular chromosome encoding 606 proteins and a 14.5 kb plasmid encoding 13 proteins. Phylogenetic analysis indicated that the bacterium is closely related to other obligate insect symbionts belonging to the Gammaproteobacteria, including Buchnera of aphids and Blochmannia of ants, and the most closely related to Ishikawaella, an extracellular gut symbiont of plataspid stinkbugs. These data suggested that the symbiont genome has evolved like highly reduced gamma-proteobacterial symbiont genomes reported from a variety of insects. The presence of genes involved in biosynthesis pathways for amino acids, vitamins, and cofactors in the genome implicated the symbiont as a nutritional mutualist, supplementing essential nutrients to the host. Interestingly, the symbiont’s plasmid encoded genes for thiamine and carotenoid synthesis pathways, suggesting the possibility of additional functions of the symbiont for protecting the host against oxidative stress and DNA damage. Finally, possible involvement of the symbiont in uric acid metabolism during diapause is discussed.
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Tanaka S., Dayi M., Maeda Y., Tsai I., Tanaka R., Bligh M., Takeuchi-Kaneko Y., Fukuda K., Kanzaki N., Kikuchi T.
Scientific Reports 9 ( 1 ) 6080 2019年12月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Scientific Reports
© 2019, The Author(s). The pine wood nematode Bursaphelenchus xylophilus is the causal agent of pine wilt disease, one of the most devastating forest diseases in East Asian and West European countries. The lifecycle of B. xylophilus includes four propagative larval stages and gonochoristic adults which are involved in the pathogenicity, and two stages of dispersal larvae involved in the spread of the disease. To elucidate the ecological roles of each developmental stage in the pathogenic life cycle, we performed a comprehensive transcriptome analysis using RNA-seq generated from all developmental stages of B. xylophilus and compared transcriptomes between stages. We found more than 9000 genes are differentially expressed in at least one stage of the life cycle including genes involved in general nematode biology such as reproduction and moulting but also effector genes likely to be involved in parasitism. The dispersal-stage transcriptome revealed its analogy to C. elegans dauer and the distinct roles of the two larval stages from each other regarding survival and transmission. This study provides important insights and resources to understand B. xylophilus parasitic biology.
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Biology and genome of a newly discovered sibling species of Caenorhabditis elegans 査読あり 国際共著
Kanzaki N., Tsai I., Tanaka R., Hunt V., Liu D., Tsuyama K., Maeda Y., Namai S., Kumagai R., Tracey A., Holroyd N., Doyle S., Woodruff G., Murase K., Kitazume H., Chai C., Akagi A., Panda O., Ke H., Schroeder F., Wang J., Berriman M., Sternberg P., Sugimoto A., Kikuchi T.
Nature Communications 9 ( 1 ) 3216 2018年12月
担当区分：筆頭著者 記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Nature Communications
© 2018, The Author(s). A ‘sibling’ species of the model organism Caenorhabditis elegans has long been sought for use in comparative analyses that would enable deep evolutionary interpretations of biological phenomena. Here, we describe the first sibling species of C. elegans, C. inopinata n. sp., isolated from fig syconia in Okinawa, Japan. We investigate the morphology, developmental processes and behaviour of C. inopinata, which differ significantly from those of C. elegans. The 123-Mb C. inopinata genome was sequenced and assembled into six nuclear chromosomes, allowing delineation of Caenorhabditis genome evolution and revealing unique characteristics, such as highly expanded transposable elements that might have contributed to the genome evolution of C. inopinata. In addition, C. inopinata exhibits massive gene losses in chemoreceptor gene families, which could be correlated with its limited habitat area. We have developed genetic and molecular techniques for C. inopinata; thus C. inopinata provides an exciting new platform for comparative evolutionary studies.
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Assessment of the behaviour and survival of nematodes under low oxygen concentrations 査読あり 国際共著
Kitazume H., Dayi M., Tanaka R., Kikuchi T.
PLoS ONE 13 ( 5 ) e0197122 2018年5月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：PLoS ONE
© 2018 Kitazume et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Oxygen is required for the completion of almost all known metazoan lifecycles, but many metazoans harbour abilities to withstand varying degrees and periods of hypoxia. Caenorhabditis elegans, one of the most popular model organism is extensively used as a model for the study of hypoxia and anoxia biology and it has been found that this nematode is capable of tolerance to varying degrees of hypoxia. Considering the extremely high diversity of nematodes, the effects of low oxygen concentration and mechanisms of adaptation to oxygen depletion differ among species. In this study, we used a simple assay to examine anoxia tolerance in four nematode species, including three free-living and one plant parasitic nematode. We found that the plant parasitic nematode Bursaphelenchus xylophilus can survive more than 14 days under anoxic conditions. Comparisons of behaviour during anoxia induction and the repertoire of oxygen sensation genes among the tested species suggested the existence of different oxygen sensation systems between B. xylophilus and C. elegans, which quickly introduce suspended animation in response to oxygen depletion to survive long-term anoxia.
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Kanzaki N., Kiontke K., Tanaka R., Hirooka Y., Schwarz A., Müller-Reichert T., Chaudhuri J., Pires-Dasilva A.
Scientific Reports 7 ( 1 ) 11135 2017年12月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Scientific Reports
© 2017 The Author(s). The co-existence of males, females and hermaphrodites, a rare mating system known as trioecy, has been considered as an evolutionarily transient state. In nematodes, androdioecy (males/hermaphrodites) as found in Caenorhabditis elegans, is thought to have evolved from dioecy (males/females) through a trioecious intermediate. Thus, trioecious species are good models to understand the steps and requirements for the evolution of new mating systems. Here we describe two new species of nematodes with trioecy, Auanema rhodensis and A. freiburgensis. Along with molecular barcodes, we provide a detailed analysis of the morphology of these species, and document it with drawings and light and SEM micrographs. Based on morphological data, these free-living nematodes were assigned to a new genus, Auanema, together with three other species described previously. Auanema species display convergent evolution in some features with parasitic nematodes with complex life cycles, such as the production of few males after outcrossing and the obligatory development of dauers into self-propagating adults.
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Kikuchi T., Hino A., Tanaka T., Aung M., Afrin T., Nagayasu E., Tanaka R., Higashiarakawa M., Win K., Hirata T., Htike W., Fujita J., Maruyama H.
PLoS Neglected Tropical Diseases 10 ( 12 ) e0005253 2016年12月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：PLoS Neglected Tropical Diseases
© 2016 Kikuchi et al. The helminth Strongyloides stercoralis, which is transmitted through soil, infects 30–100 million people worldwide. S. stercoralis reproduces sexually outside the host as well as asexually within the host, which causes a life-long infection. To understand the population structure and transmission patterns of this parasite, we re-sequenced the genomes of 33 individual S. stercoralis nematodes collected in Myanmar (prevalent region) and Japan (non-prevalent region). We utilised a method combining whole genome amplification and next-generation sequencing techniques to detect 298,202 variant positions (0.6% of the genome) compared with the reference genome. Phylogenetic analyses of SNP data revealed an unambiguous geographical separation and sub-populations that correlated with the host geographical origin, particularly for the Myanmar samples. The relatively higher heterozygosity in the genomes of the Japanese samples can possibly be explained by the independent evolution of two haplotypes of diploid genomes through asexual reproduction during the auto-infection cycle, suggesting that analysing heterozygosity is useful and necessary to infer infection history and geographical prevalence.
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Genome-Wide Analyses of Individual Strongyloides stercoralis (Nematoda: Rhabditoidea) Provide Insights into Population Structure and Reproductive Life Cycles 査読あり
Taisei Kikuchi , Akina Hino , Teruhisa Tanaka , Myo Pa Pa Thet Hnin Htwe Aung,Tanzila Afrin, Eiji Nagayasu, Ryusei Tanaka, Miwa Higashiarakawa, Kyu Kyu Win, Tetsuo Hirata,Wah Win Htike, Jiro Fujita,Haruhiko Maruyama
PLOS Neglected Tropical Diseases 2016年12月
記述言語：英語 掲載種別：研究論文（学術雑誌）
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Venison, another source of Paragonimus westermani infection 査読あり
Yoshida A., Matsuo K., Moribe J., Tanaka R., Kikuchi T., Nagayasu E., Misawa N., Maruyama H.
Parasitology International 65 ( 6 ) 607 - 612 2016年12月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Parasitology International
© 2016 Elsevier Ireland Ltd Paragonimiasis is a typical food-borne parasitic disease, endemic in most parts of Asia, with sporadic case reports from American and African countries. The major source of infection is undercooked freshwater crab or crayfish, though consumption of wild boar meat is also responsible for the infection in Japan, because wild boar is a paratenic host for Paragonimus westermani. Recently, living juveniles of P. westermani were isolated from muscle of a sika deer, Cervus nippon, in Japan, raising the possibility that venison has been another source of infection. In order to clarify the potential contribution of venison consumption to the occurrence of paragonimiasis, we analysed dietary histories of those paragonimiasis patients in whose diagnoses we were involved between 2001 and 2015. Among 380 patients, freshwater crab had been consumed by 208 patients, wild boar meat by 190, and wild deer meat by 76 patients before the onset of the disease. Overall contribution of wild deer meat was estimated to be 6.8% to 20.0%, although in Oita and Gifu Prefectures, where a substantial proportion of patients had consumed raw venison, the contribution of venison consumption was much higher (27.5 to 62.1% and 42.1 to 78.9% in Oita and Gifu Prefectures, respectively). We demonstrated P. westermani-specific antibodies in the sera of 4 out of 160 sika deer from Gifu Prefecture, strongly suggesting that these deer were infected with P. westermani.
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Venison, another source of Paragonimus westermani infection 査読あり
Yoshida A, Matsuo K, Moribe J, Tanaka R, Kikuchi T, Nagayasu E, Misawa N, Maruyama H.
Parasitology International 2016年12月
記述言語：英語 掲載種別：研究論文（学術雑誌）
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Tsai I., Tanaka R., Kanzaki N., Akiba M., Yokoi T., Espada M., Jones J., Kikuchi T.
Molecular Plant Pathology 17 ( 1 ) 77 - 83 2016年1月
担当区分：筆頭著者 記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Molecular Plant Pathology
© 2016 BSPP and John Wiley & Sons Ltd. Summary: Drastic physiological and morphological changes in parasites are crucial for the establishment of a successful infection. The nematode Bursaphelenchus xylophilus is the pathogenic agent of pine wilt disease, and little is known about the physiology and morphology in this nematode at the initial stage of infection. In this study, we devised an infection system using pine stem cuttings that allowed us to observe transcriptional and morphological changes in the host-infecting phytophagous phase. We found that 60 genes enriched in xenobiotic detoxification were up-regulated in two independent post-inoculation events, whereas down-regulation was observed in multiple members of collagen gene families. After 48h of inoculation, the tails in some of the adult females exposed to the host changed in morphology. These results suggest that B.xylophilus may change its physiology and morphology to protect itself and to adapt to the host pine wood environment.
DOI： 10.1111/mpp.12261
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田中 克, 田中 龍聖, 竹内 祐子, 神崎 菜摘, 菊地 泰生, 福田 健二
日本森林学会大会発表データベース 127 ( 0 ) 2016年
記述言語：日本語 掲載種別：研究論文（研究会，シンポジウム資料等） 出版者・発行元：日本森林学会
マツノザイセンチュウは東アジアや西ヨーロッパで甚大な被害を及ぼすマツ材線虫病（以下マツ枯れ）の病原体である。本線虫は増殖型と分散型の二型性の生活環を持ち、増殖型はマツの衰弱・枯死に関与する一方、分散型はマツ枯れ伝播に関与する。特に分散型IV期幼虫（以下D<sub>IV</sub>）は、媒介昆虫であるマツノマダラカミキリからの誘導を受けて出現し、マツからマツへと運ばれるため、疫学上重要な役割を担う。本研究ではD<sub>IV</sub>の生理的機能を遺伝子発現の面から解明するため、RNA-seq法を用いた線虫全ステージの比較解析を行い、D<sub>IV</sub>で特徴的な発現を示す遺伝子の同定を試みた。その結果、線虫全ステージ合計で15,251遺伝子、D<sub>IV</sub>では12,730遺伝子の発現が確認された。D<sub>IV</sub>は、増殖型各ステージと比較して774~3,448遺伝子（約5~21%）の発現が有意に異なった。また、D<sub>IV</sub>の前段階である分散型III期幼虫との比較においても、2,347遺伝子（約14%）の発現が有意に異なった。これらのことから、D<sub>IV</sub>が形態や生態だけでなく遺伝子発現においても特徴的であることが明らかとなった。講演では上記の遺伝子の機能も含めて、D<sub>IV</sub>の遺伝子発現様式を総合的に論じる。
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Ogura N., Takeuchi T., Kikuchi T., Tanaka R., Yamaguchi M., Dayi M.
Nematology 17 ( 5 ) 515 - 522 2015年1月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Nematology
© 2015 Koninklijke Brill NV, Leiden. Subanguina moxae, belonging to the subfamily Anguininae, is an obligate parasite of Artemisia plants, which are widely used as cooking herbs and in traditional medicine in East Asia. Because the nematode is distributed throughout East Russia and East Asia, there is concern about the potential for significant damage to commercial farming; however, details about its biology remain unclear. To investigate the genetic diversity of S. moxae, we developed 2243 simple sequence repeat (SSR) markers using Illumina short reads of the genomic DNA.We validated 100 randomly selected markers indicating their robustness and examined polymorphisms among nematode populations sampled from four different locations in Japan. These SSR markers will be a useful tool for understanding the population structure and transmission patterns of this parasitic nematode.
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マツノザイセンチュウ各ステージの比較トランスクリプトーム解析
田中 克, 田中 龍聖, 菊地 泰生, 竹内 祐子
日本森林学会大会発表データベース 126 ( 0 ) 2015年
記述言語：日本語 掲載種別：研究論文（研究会，シンポジウム資料等） 出版者・発行元：日本森林学会
マツノザイセンチュウ（以下線虫）は東アジアや西ヨーロッパを中心に甚大な被害を及ぼすマツ材線虫病の病原体である。本研究では、線虫の各生育ステージの機能的役割を遺伝子発現の面から解明することを目的として、RNA-seq法による遺伝子発現様式の網羅的な比較解析を行った。卵を含む増殖型各ステージからRNAを抽出し、次世代シーケンサーによる塩基配列の解読を経て、得られたcDNA配列データを統計解析ソフトにより解析した。今回の実験では増殖型全ステージで計14,825遺伝子の発現が確認された。これらの遺伝子について、ステージ間で発現レベルが有意に異なる遺伝子数及びステージ間の相関係数の算出、GO解析を用いた遺伝子の機能解析を行った。その結果、増殖型3期幼虫と増殖型4期幼虫は遺伝子発現において高い類似度を示した。また、増殖型4期幼虫では植物細胞壁分解酵素の活性や宿主植物の免疫機構打破に関わる遺伝子群の発現が上昇しており、本病発病過程において重要な役割を果たしていることが示唆された。雌雄成虫はそれぞれ卵及び精子形成に関わる遺伝子の発現が上昇しており、生殖活動に重点を置いていることが明らかとなった。
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Assessment of helminth biodiversity in wild rats using 18S rDNA based metagenomics 査読あり 国際共著
Tanaka R., Hino A., Tsai I., Palomares-Rius J., Yoshida A., Ogura Y., Hayashi T., Maruyama H., Kikuchi T.
PLoS ONE 9 ( 10 ) e110769 2014年10月
担当区分：筆頭著者 記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：PLoS ONE
© 2014 Tanaka et al. Parasite diversity has important implications in several research fields including ecology, evolutionary biology and epidemiology. Wide-ranging analysis has been restricted because of the difficult, highly specialised and time-consuming processes involved in parasite identification. In this study, we assessed parasite diversity in wild rats using 18S rDNA-based metagenomics. 18S rDNA PCR products were sequenced using an Illumina MiSeq sequencer and the analysis of the sequences using the QIIME software successfully classified them into several parasite groups. The comparison of the results with those obtained using standard methods including microscopic observation of helminth parasites in the rat intestines and PCR amplification/sequencing of 18S rDNA from isolated single worms suggests that this new technique is reliable and useful to investigate parasite diversity.
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Kanzaki N., Tanaka R., Giblin-Davis R., Davies K.
PLoS ONE 9 ( 6 ) e99241 2014年6月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：PLoS ONE
A new nematode species, Bursaphelenchus sycophilus n. sp. is described. The species was found in syconia of a fig species, Ficus variegata during a field survey of fig-associated nematodes in Japan. Because it has a well-developed stylet and pharyngeal glands, the species is considered an obligate plant parasite, and is easily distinguished from all other fungalfeeding species in the genus based upon these characters. Although B. sycophilus n. sp. shares an important typological character, male spicule possessing a strongly recurved condylus, with the "B. eremus group" and the "B. leoni group" of the genus, it was inferred to be monophyletic with the "B. fungivorus group". The uniquely shaped stylet and well-developed pharyngeal glands is reminiscent of the fig-floret parasitic but paraphyletic assemblage of "Schistonchus". Thus, these morphological characters appear to be an extreme example of convergent evolution in the nematode family, Aphelenchoididae, inside figs. Other characters shared by the new species and its close relatives, i.e., lack of ventral P1 male genital papilla, female vulval flap, and papilla-shaped P4 genital papillae in males, corroborate the molecular phylogenetic inference. The unique biological character of obligate plant parasitism and highly derived appearance of the ingestive organs of Bursaphelenchus sycophilus n. sp. expands our knowledge of the potential morphological, physiological and developmental plasticity of the genus Bursaphelenchus. © 2014 Kanzaki et al.
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Tanaka S., Tanaka R., Akiba M., Aikawa T., Maehara N., Takeuchi Y., Kanzaki N.
Nematology 16 ( 3 ) 259 - 281 2014年3月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Nematology
A Bursaphelenchus species was isolated from a Japanese native wood-boring weevil, Niphades variegatus, and dead Pinaceae trees. The nematode is associated with the weevils as dauer (dispersal third stage) juveniles and the dauers enter the weevil tracheal system forming an abnormal expansion on the weevil trachea (atrium). Thus, the nematode is hypothesised to be an amensal/phoretic associate of the weevil because the abnormal expansion appeared to inhibit weevil respiration. The propagative stages of the nematode are associated with dead trees (wood and bark materials) and are thought to feed there on naturally propagated fungi.Morphologically, the new species is considered an undescribed species close to B. antoniae, B. chengi and B. hylobianum. Within these four species, the new species, which is described herein as B. niphades n. sp., is closest to B. chengi, i.e., the typological character of these two species are almost identical to each other and is distinguished by some minor characters (structure of the male P4 genital papillae and spicule length). The molecular phylogenetic analysis supported the morphological observations. Bursaphelenchus niphades n. sp. formed a well supported subclade with the four species and is intermediate between B. hylobianum and B. chengi; however, it is distinguished by the molecular sequences of some ribosomal RNA genes. Because three of these four species are associated with weevil species, the subclade is considered a 'weevil-associated' species group. © Koninklijke Brill NV, Leiden, 2014.
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Specialist versus generalist life histories and nucleotide diversity in Caenorhabditis nematodes 査読あり 国際共著
Li S., Jovelin R., Yoshiga T., Tanaka R., Cutter A.
Proceedings of the Royal Society B: Biological Sciences 281 ( 1777 ) 20132858 2014年1月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Proceedings of the Royal Society B: Biological Sciences
Species with broad ecological amplitudes with respect to a key focal resource, niche generalists, should maintain larger and more connected populations than niche specialists, leading to the prediction that nucleotide diversity will be lower and more subdivided in specialists relative to their generalist relatives. This logic describes the specialist-generalist variation hypothesis (SGVH). Some outbreeding species of Caenorhabditis nematodes use a variety of invertebrate dispersal vectors and have high molecular diversity. By contrast, Caenorhabditis japonica lives in a strict association and synchronized life cycle with its dispersal host, the shield bug Parastrachia japonensis, itself a diet specialist. Here, we characterize sequence variation for 20 nuclear loci to investigate how C. japonica's life history shapes nucleotide diversity. We find that C. japonica has more than threefold lower polymorphism than other outbreeding Caenorhabditis species, but that local populations are not genetically disconnected. Coupled with its restricted range, we propose that its specialist host association contributes to a smaller effective population size and lower genetic variation than host generalist Caenorhabditis species with outbreeding reproductive modes. A literature survey of diverse organisms provides broader support for the SGVH. These findings encourage further testing of ecological and evolutionary hypotheses with comparative population genetics in Caenorhabditis and other taxa. © 2014 The Author(s) Published by the Royal Society. All rights reserved.
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Woodruff G., Tanaka R., Kanzaki N.
Nematology 16 ( 10 ) 1153 - 1166 2014年1月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Nematology
© 2014 Koninklijke Brill NV, Leiden. Teratodiplogaster variegatae n. Sp. is described and illustrated from Ficus variegata on Ishigaki Island, Okinawa, Japan. Teratodiplogaster variegatae n. Sp. is characterised by its scoop-like lip, the presence of large sac-like receptaculum seminis in the female gonads, and eight pairs of male genital papillae with an arrangement of (P1, P2, P3, vs, C, (P4, P5d), (P6, P7), P8d, Ph). It is distinguished from its close relatives T. fignewmani and T. martini by its stomatal morphology, spicule and gubernaculum morphology, structure of female reproductive organs, and the characteristic star-shaped appendage on the tail tip of males and females. Molecular phylogenetic analyses based upon near-full-length SSU and D2-D3 expansion segments of LSU corroborate a monophyletic origin of the genus Teratodiplogaster.
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Comparative genomics of taphrina fungi causing varying degrees of tumorous deformity in plants 査読あり
Tsai I., Tanaka E., Masuya H., Tanaka R., Hirooka Y., Endoh R., Sahashi N., Kikuchi T.
Genome Biology and Evolution 6 ( 4 ) 861 - 872 2014年1月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Genome Biology and Evolution
Taphrina fungi are biotrophic plant pathogens that cause plant deformity diseases. We sequenced the genomes of four Taphrina species-Taphrinawiesneri, T. deformans, T. flavorubra, andT. populina-which parasitize Prunus, Cerasus, and Populus hosts with varying severity of disease symptoms. High levels of gene synteny within Taphrina species were observed, and our comparative analysis further revealed that these fungi may utilize multiple strategies in coping with the host environment that are also found in some specialized dimorphic species. These include species-specific aneuploidy and clusters of highly diverged secreted proteins located at subtelomeres. We also identified species differences in plant hormone biosynthesis pathways, which may contribute to varying degree of disease symptoms. The genomes provide a rich resource for investigation into Taphrina biology and evolutionary studies across the basal ascomycetes clade. © 2014 The Author(s).
DOI： 10.1093/gbe/evu067
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Kanzaki N., Tanaka R.
Nematology 15 ( 8 ) 975 - 990 2013年12月
担当区分：最終著者 記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Nematology
A Sheraphelenchus species was isolated from a sample of sap exuding from a scar on the bark of Quercus serrata. Besides its generic characteristics, i.e., posteriorly located vulva in the female, male spicule with conspicuous dorsal limb and male tail with spike-like projection, the new species is characterised by a short stylet with a small basal swelling. The near-full-length of 18S and D2/D3 expansion segments of ribosomal RNA genes (near-full SSU and D2/D3LSU) were determined as its molecular barcode sequences and the phylogenetic status of the species (= genus Sheraphelenchus) was estimated using the near-full SSU. The molecular phylogenetic analysis revealed that the new species is included in the genus Bursaphelenchus, and is close to B. anatolius and B. kevini. Although Sheraphelenchus is phylogenetically inferred to occur within Bursaphelenchus, the new species is described herein as S. sucus n. sp., thereby retaining the genus Sheraphelenchus until additional data about morphological similarities between these two taxa are obtained. © Koninklijke Brill NV, Leiden, 2013.
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Kanzaki N., Tanaka R., Ikeda H., Taki H., Sugiura S., Matsumoto K.
Journal of Parasitology 99 ( 4 ) 639 - 649 2013年8月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Journal of Parasitology
The nematode family Aphelenchoididae is a highly divergent group that contains plant parasites, predators, fungal feeders, and insect parasites. It is taxonomically separated into 7 subfamilies. Although molecular phylogenetic relationships among 6 of the subfamilies have been clarified, the phylogenetic position of the subfamily Entaphelenchinae, which is composed of endoparasites of insects, remains unclear. Here, a new entaphelenchid species, Peraphelenchus orientalis n. sp., was isolated from the body cavities of burying beetles, Nicrophorus quadripunctatus, with a 14% prevalence (5 out of 36). The phylogenetic position of the subfamily within the family Aphelenchoididae was determined using morphological and molecular data for the new species. The rRNA sequences suggested that the new species belongs to Clade 3 of Aphelenchoididae, which mostly consists of predators and insect parasites. Although molecular sequences from other entaphelenchid species were not available, the subfamily appears to be monophyletic. The new species is characterized by the absence of a functional rectum and anus and by the presence of 3 pairs of male genital papillae, a loosely coiled male body, and a W-shaped male spicule. Compared with the original description of Peraphelenchus necrophori, P. orientalis n. sp. has significant typological differences, possibly because of misinterpretations during the original description of P. necrophori. Excluding these questionable characters, i.e., presence-absence of functional rectum and anus and number of male genital papillae, the new species is distinguished from P. necrophori by minor morphological characters and morphometric values. © American Society of Parasitologists 2013.
DOI： 10.1645/12-118.1
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Negative gravitactic behavior of Caenorhabditis japonica dauer larvae 査読あり
Okumura E., Tanaka R., Yoshiga T.
Journal of Experimental Biology 216 ( 8 ) 1470 - 1474 2013年4月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Journal of Experimental Biology
Gravity on Earth is a constant stimulus and many organisms are able to perceive and respond to it. However, there is no clear evidence that nematodes respond to gravity. In this study, we demonstrated negative gravitaxis in a nematode using dauer larvae (DL) of Caenorhabditis japonica, which form an association with their carrier insect Parastrachia japonensis. Caenorhabditis japonica DL demonstrating nictation, a typical host-finding behavior, had a negative gravitactic behavior, whereas non-nictating C. japonica and C. elegans DL did not. The negative gravitactic index of nictating DL collected from younger nematode cultures was higher than that from older cultures. After a 24?h incubation in M9 buffer, nictating DL did not alter their negative gravitactic behavior, but a longer incubation resulted in less pronounced negative gravitaxis. These results are indicative of negative gravitaxis in nictating C. japonica DL, which is maintained once initiated, seems to be affected by the age of DL and does not appear to be a simple passive mechanism. © 2013. Published by The Company of Biologists Ltd.
DOI： 10.1242/jeb.075739
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Sugiura S., Tanaka R., Taki H., Kanzaki N.
Biological Conservation 159 206 - 213 2013年3月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Biological Conservation
Forest loss and degradation can impact on ecosystem functions. Decomposition of vertebrate carcasses is an important ecosystem function in forests, and burying beetles and scavenging vertebrates is known to contribute to the provision of this function in forests. However, the relative importance of burying beetles and scavenging vertebrates in the removal of vertebrate carcasses has rarely been explored in terms of the impact of forest loss and degradation. Here, we experimentally placed mouse carcasses on forest floors to investigate burial and removal rates by forest animals during a 7-day period at 31 sites (natural broad-leaved forests and conifer plantations in an urban/agricultural matrix) in central Japan. Ninety-six percent of dead mice disappeared with 7. days of being placed on the forest floor. Rates of carcass disappearance increased with forest area surrounding the study sites. Field observations and camera trapping showed that two species of Nicrophorus burying beetles and six species of scavenging vertebrates contributed to the carcass burial and removal. Burial rates by beetles increased with forest area, while removal rates by vertebrates decreased with forest area. Model comparisons showed that the area of both natural forests and plantations surrounding the study sites influenced burial and removal rates. Therefore, forest loss influenced the disappearance rates of vertebrate carcasses. Different responses to forest loss between burying beetles and scavenging vertebrates could maintain the carcass burying/removing function at a landscape level. © 2012 Elsevier Ltd.
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Propagation of Caenorhabditis japonica in the nest of its carrier insect, Parastrachia japonensis 査読あり
Okumura E., Ishikawa Y., Tanaka R., Yoshiga T.
Zoological Science 30 ( 3 ) 174 - 177 2013年3月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Zoological Science
We demonstrated the disembarkation of the bacterial-feeding nematode Caenorhabditis japonica dauer larvae (DL) from adult Parastrachia japonensis female insects and observed the propagation of nematodes in artificial insect nests. Our results clarify the process of propagation in this nematode species and provide insights into the nematode-insect relationship. Quiescent C. japonica DL resumed their mobility only at > 99.9% relative humidity (RH) at 25°C in the presence or absence of the carrier insect. In artificial nests with > 99.9% RH, DL resumed their mobility and the number of DL on female insects decreased gradually after oviposition, although numerous DL remained on the insects. Very few DL were detected on mother insects after hatching. Nematode propagation was observed on the egg mass after hatching and on nymphal carcasses; the total number of nematodes in the nest increased dramatically after this point. These results indicate that humidity is an important factor for disembarkation of C. japonica DL and that C. japonica propagates in the nest of P. japonensis where it feeds on the remains of eggs and nymph carcasses, indicating that C. japonica and P. japonensis have a unique phoretic and necromenic association. © 2013 Zoological Society of Japan.
DOI： 10.2108/zsj.30.174
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Nematodes associated with dryocoetes uniseriatus (Coleoptera: Scolytidae) 査読あり
Shimizu A., Tanaka R., Akiba M., Masuya H., Iwata R., Fukuda K., Kanzaki N.
Environmental Entomology 42 ( 1 ) 79 - 88 2013年2月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Environmental Entomology
We examined the nematode fauna associated with a species of bark beetle, Dryocoetes uniseriatus Eggers, as part of a biodiversity survey of forest beetle-associated nematodes. Collections were made in a pine stand at an experimental forest station in Ibaraki, Japan, from April to July of 2011; we examined the nematode association in 273 insects collected during this time. In total, 68% of the insects were associated with at least one species of nematode. Six species of nematodes, including two phoretic microbe feeders (Bursaphelenchus rainulfi Braasch & Burgermeister and Micoletzkya sp.), one insect parasite and nematode predator (Devibursaphelenchus cf. eproctatus), one insect parasite (Contortyhnchus sp.), one insect parasite and potential microbe feeder (unidentified rhabditid parasite), and one potential insect parasite and fungal feeder (B. sinensis) were recovered from the beetles. D. cf. eproctatus was enclosed in nematangia on the backsides of the elytra, B. rainulfi was isolated from the backsides of the elytra or enclosed in nematangia, Micoletzkya sp. was isolated from under the elytra, Contortylenchus sp. and a rhabditid parasite parasitized the body cavity, and B. sinensis was found in the digestive tract of the insect. The association patterns of the nematode species varied seasonally, although definitive interactions among species (e.g., segregation, competition) were not observed. © 2013 Entomological Society of America.
DOI： 10.1603/EN12165
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Okumura E., Tanaka R., Yoshiga T.
Journal of Experimental Biology 216 ( 4 ) 568 - 572 2013年2月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Journal of Experimental Biology
Host recognition is crucial during the phoretic stage of nematodes because it facilitates their association with hosts. However, limited information is available on the direct cues used for host recognition and host specificity in nematodes. Caenorhabditis japonica forms an intimate association with the burrower bug Parastrachia japonensis. Caenorhabditis japonica dauer larvae (DL), the phoretic stage of the nematode, are mainly found on adult P. japonensis females but no other species. To understand the mechanisms of species-specific and female carrier-biased ectophoresy in C. japonica, we investigated whether C. japonica DL could recognize their hosts using nematode loading and chemoattraction experiments. During the loading experiments, up to 300 C. japonica DL embarked on male and female P. japonensis, whereas none or very few utilized the other shield bugs Erthesina fullo and Macroscytus japonensis or the terrestrial isopod Armadillidium vulgare. In the chemoattraction experiments, hexane extracts containing the body surface components of nymphs and both adult P. japonensis sexes attracted C. japonica DL, whereas those of other shield bugs did not. Parastrachia japonensis extracts also arrested the dispersal of C. japonica DL released at a site where hexane extracts were spotted on an agar plate; i.e. >50% of DL remained at the site even 60min after nematode inoculation whereas M. japonensis extracts or hexane alone did not have the same effect. These results suggest that C. japonica DL recognize their host species using direct chemical attractants from their specific host to maintain their association. © 2013. Published by The Company of Biologists Ltd.
DOI： 10.1242/jeb.073593
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Species-specific and female host-biased ectophoresy in the roundworm Caenorhabditis japonica 査読あり
Yoshiga T., Ishikawa Y., Tanaka R., Hironaka M., Okumura E.
Naturwissenschaften 100 ( 2 ) 205 - 208 2013年2月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Naturwissenschaften
Caenorhabditis japonica is a bacteriophagous nematode species that was discovered on the semi-social burrower bug, Parastrachia japonensis, which demonstrates egg-guarding and provisioning behaviors. To understand the life history of C. japonica in relation to P. japonensis, we demonstrated the specificity of this association and fluctuations in nematode number on the insect throughout the year. C. japonica dauer larvae (DL), larvae in a nonfeeding diapause stage, were predominantly found as clumps on the adult female insects but rarely found on the male insects in all populations examined. This female-biased association was consistent throughout the year, but after the nymphs hatched, nematodes were not detected on the mother insects showing provisioning behavior. DL appeared on the nymphs, and the number of DL on the newly emerged female insects gradually increased thereafter. C. japonica has never been detected on other invertebrates collected from the P. japonensis habitat thus far. Our data suggest that the life cycles of C. japonica and P. japonensis are synchronized. © 2013 Springer-Verlag Berlin Heidelberg.
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Kanzaki N., Ragsdale E., Herrmann M., Mayer W., Tanaka R., Sommer R.
Nematology 14 ( 8 ) 933 - 947 2012年12月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Nematology
A new species of diplogastrid nematode, isolated in a previous survey of nematodes associated with stag beetles in Japan, is described as Parapristionchus giblindavisi n. gen., n. sp. Parapristionchus n. gen. differs from other diplogastrid genera chiefly by its stomatal morphology. Distinguishing the genus are the presence of a claw-like dorsal tooth in both the eurystomatous and stenostomatous forms and the division of the cheilostom into 12 plates lacking apical flaps. According to phylogenetic analysis of nine ribosomal protein gene sequences, Parapristionchus n. gen. shows deep divergence from other known genera. Molecular evidence strongly supports P. giblindavisi n. gen., n. sp. + Pristionchus spp. as monophyletic with respect to all other diplogastrids examined. Congruent with a clade of P. giblindavisi n. gen., n. sp. + Pristionchus spp. is the shared presence of a bifurcate P7 genital papilla. Discovery and description of a close sister group to Pristionchus, a model biological system, enables character polarisation in macroevolutionary studies of Pristionchus nematodes. © Koninklijke Brill NV, Leiden, 2012.
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Kanzaki N., Tanaka R., Sahashi N.
Environmental Entomology 41 ( 4 ) 828 - 832 2012年8月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Environmental Entomology
The virulence of two species (three isolates) of pine wood nematodes in stressed adult Japanese red and black pines was tested using an inoculation experiment. Three thousand nematodes, Bursaphelenchus xylophilus isolate 'Ka4' (virulent), isolate 'C14-5' (less virulent) or B. mucronatus isolate 'M' (less virulent), were inoculated into 15-yr-old naturally shaded and stressed pine trees planted in two separated experimental stands, in which shelterwood cutting had not been carried out after planting. The inoculation was conducted at the beginning of August 2010, and the trees were then visually examined every month for disease symptoms until February 2011. Trees that died during the experimental period were cut and brought back to the laboratory for reisolation attempts of the pathogenic nematodes. The Ka4 isolate killed all inoculated trees within one to 2-mo, whereas C14-5 and M each killed about half. The inoculated nematodes were reisolated from all the dead trees and some of the surviving ones. These results corroborated those of previous research by using small saplings (i.e., that less virulent nematodes can kill shaded (stressed) trees). Further, a feeding preference experiment using their vector beetles, Monochamus alternatus Hope, showed that both healthy and stressed trees are at equal risk of being used by beetles and hence of getting infected by the nematodes. Therefore, pine wilt disease in shading-stressed trees is assumed to occur in susceptible pine trees in natural pine stands. The dead tree is used by insects as an oviposition resource. © 2012 Entomological Society of America.
DOI： 10.1603/EN12031
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Simple and quick methods for nematode DNA preparation 査読あり
Tanaka R., Kikuchi T., Aikawa T., Kanzaki N.
Applied Entomology and Zoology 47 ( 3 ) 291 - 294 2012年8月
担当区分：筆頭著者 記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Applied Entomology and Zoology
A DNA extraction kit, ISOHAIR® (Nippon Gene), which was originally developed for preparing DNA from hair and nail samples, was used to prepare nematode DNA for PCR and sequencing analyses. The methods provided here, which involved digesting (resolving) a single nematode individual in a tube containing the mixed enzyme solution, enabled the DNA to be prepared within 20 min. The prepared DNA was suitable for PCR amplification of near-full-length small subunit ribosomal RNA (ca 1. 7 kb), of the D2/D3 expansion segments of large subunit RNA (ca. 0. 7 kb), and of partial mitochondrial COI (ca. 0. 7 kb) genes, followed by sequencing analysis. Furthermore, the prepared material could be preserved in a freezer (-30 °C) for at least 20 months, and more than 300 PCR reactions could be performed from a single individual nematode. © 2012 The Japanese Society of Applied Entomology and Zoology.
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Tanaka R., Okumura E., Kanzaki N., Yoshiga T.
Experimental Gerontology 47 ( 5 ) 388 - 393 2012年5月
担当区分：筆頭著者, 責任著者 記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Experimental Gerontology
The nematode dauer larva (DL) is a non-aging diapause stage. The DL of the model nematode . Caenorhabditis elegans has been studied as a model system for aging and longevity. However, information on DL in other nematode species is limited. In this study, the survivorship, storage, energy consumption, and oxidative stress tolerance of . Caenorhabditis japonica DL were examined. . C. japonica is a close relative of . C. elegans, but has species-specific phoretic associations with the shield bug . Parastrachia japonensis. Also, its DL has a much longer lifespan than . C. elegans in a biological setting. However, when . C. japonica DLs were detached from their phoretic host, they did not survive more than 10. days while more than 80% of . C. elegans survived under the same conditions. Also, . C. japonica DL showed more active movement (swimming) and lower tolerance to oxidative stress than . C. elegans DL. Because the concentration of triacylglycerol (TAG), the energy source of nematodes, did not decrease significantly during the experiment, exhaustion of the energy reservoir did not cause the low survivorship of . C. japonica. Instead, low tolerance to oxidizing stress and increased production of reactive oxygen species in . C. japonica were the main causes of the reduced survivorship. The fact that . C. japonica DL cannot survive away from its insect host indicates that its longevity is increased by unknown factors derived from the host. Despite these significant differences between . C. japonica and . C. elegans, these two species are phylogenetically closely related (they are derived from a common ancestor). Therefore, . C. japonica could be a good comparative system for . C. elegans, and further physiological and molecular analyses of . C. japonica DL may provide important information about the internal and external factors affecting the longevity of nematodes in general. © 2012 Elsevier Inc.
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Diversity of stag beetle-associated nematodes in Japan 査読あり
Kanzaki N., Taki H., Masuya H., Okabe K., Tanaka R., Abe F.
Environmental Entomology 40 ( 2 ) 281 - 288 2011年4月
記述言語：英語 掲載種別：研究論文（学術雑誌） 出版者・発行元：Environmental Entomology
A survey of stag beetle (Lucanidae)-associated nematodes was conducted in Japan to construct a preliminary species inventory and to obtain their molecular profiles as a first step for elucidating their host fidelity and ecological roles. Eight species of stag beetles (94 individuals in total), Dorcus rubrofemoratus, D. montivagus, D. striatipennis, D. rectus, D. titanus, Prismognathus angularis, Prosopocoilus inclinatus, and Lucanus maculifemoratus were collected from 12 localities in Japan, dissected and examined for nematode associates. Isolated nematodes were used to attempt the establishment of voucher cultures. Successfully cultured nematodes were observed under a light microscope for morphological identification at genus or species level, and were sequenced for their near full length SSU and D2/D3 LSU. All investigated lucanid individuals harbored at least one species of nematode, and eight species (Bursaphelenchus sp., Koerneria luziae, K. lucani, Pristionchus cf. pacificus, Pristionchus sp., Pseudodiplogasteroides composites, Pseudodiplogasteroides sp., and Rhabditidoides sp.: 40 populations of nematodes in total), were successfully cultured from the insects. Within these eight species, four species were considered to predominate and had low carrier specificity and wide distribution, i.e., K. luziae, P. composites, Pristionchus sp. and Rhabditidoides sp. were isolated from various species of beetles and from several different localities, and K. luziae, K. lucani, and P. composites have been previously described from native lucanids in Germany. Considering the recent world-wide trade of living insects, especially lucanids as pets, we suggest the use of caution to prevent the invasion of their associated nematodes and other associated organisms. © 2011 Entomological Society of America.
DOI： 10.1603/EN10182