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Ayu Plecoglossus altivelis altivelis is a valuable osmeroid species for inland fishery in Japan. It is classified into two ecological forms of amphidromous migrating between rivers and sea and landlocked migrating between rivers and lakes or dam reservoirs. The number of dams and their reservoirs has remarkably increased in the twenty-first century under climate change, because of their respective roles in hydropower generation with negligible carbon emissions and in flood control. Dam reservoirs therefore become increasingly important as inland nursery grounds of ayu. In this study, we investigated the reproduction status of landlocked ayu migrating in the Haidzuka Dam reservoir and the Tabusa River in western Japan by molecular phylogenetic analysis based on population structure and demographic history for year cohort dynamics. A total of 849 individuals were collected monthly from October 2018 to September 2021 according to an annual life cycle of ayu. Nucleotide sequences of the partial mitochondrial DNA control region yielded 31 haplotypes, consisting of 4 shared haplotypes among the 2019, 2020 and 2021 cohorts and 27 unique haplotypes. The overall haplotype diversity and nucleotide diversity were calculated to be relatively low at 0.3503 ± 0.0206 and 0.0077 ± 0.0045, respectively, suggesting a founder event by dominant haplotypes. Star-shaped radiational haplotypes from dominant shared haplotypes on the median-joining network likely support a founder event. Although pairwise Ф ST values were determined to be very low among the year cohorts, only the 2019 cohort was found to have a significant difference from the 2020 and 2021 cohorts, for both of which Tajima's D values were also statistically significant. For the overall population, multimodal mismatch distribution and negative Tajima's D and Fu's Fs values in the neutrality test suggested population expansion or population subdivision. The native riverine population in the Tabusa River suffered habitat fragmentation and population bottleneck from dam construction, and therefore severe founder effect remained behind the artificially landlocked population with a low level of genetic diversity in the Haidzuka Dam reservoir., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Sapovirus (SaV) infections are a public health problem because they cause acute gastroenteritis in humans of all ages, both sporadically and as outbreaks. However, only a limited amount of SaV sequence information, especially whole-genome sequences for all the SaV genotypes, is publicly available. Therefore, in this study, we determined the full/near-full-length genomic sequences of 138 SaVs from the 2001 to 2015 seasons in 13 prefectures across Japan. The genogroup GI was predominant (67%, n = 92), followed by genogroups GII (18%, n = 25), GIV (9%, n = 12), and GV (6%, n = 9). Within the GI genogroup, four different genotypes were identified: GI.1 (n = 44), GI.2 (n = 40), GI.3 (n = 7), and GI.5 (n = 1). We then compared these Japanese SaV sequences with 3,119 publicly available human SaV sequences collected from 49 countries over the last 46 years. The results indicated that GI.1, and GI.2 have been the predominant genotypes in Japan, as well as in other countries, over at least four decades. The 138 newly determined Japanese SaV sequences together with the currently available SaV sequences, could facilitate a better understanding of the evolutionary patterns of SaV genotypes.

The cheilostome bryozoan Cauloramphus magnus is common in the rocky intertidal habitat from southeastern Alaska to northern Japan. We examined its phylogeography by analyzing 576 bp of the mitochondrial COI ( cox1 ) gene sequenced for 298 colonies from 16 localities in northern Japan. A maximum-likelihood phylogeny detected three main clades (A, B, C). Clades A and B occurred throughout the study area but differed in frequency, haplotype diversity, and haplotype distribution; each resolved into three divergent subclades (AI-III, BI-III). Clade A shared none among 15 haplotypes between the Pacific and Sea of Japan sides of Hokkaido. In contrast, Clade B (29 haplotypes) was thrice as common as Clade A among samples, with haplotype B28 common on both sides. Divergent Clade C (nine haplotypes) was detected only at Rumoi. K2P divergences of 12.3-28.3% among Clades A-C suggest these are distinct biological species, a conclusion supported by different inferred evolutionary histories. A bPTP species delimitation analysis indicated nine phylogenetic species among the sequences included in our phylogeny (AI-III, BI-III, C, and one specimen each from Alaska and the Commander Islands), with K2P divergences of 3.9-6.5% among subclades in A or B. Statistical and principal components analyses suggested weak morphological differentiation between Clades A + B and C, although overlapping ranges of measurements preclude identification to clade; these three clades are morphologically cryptic. For taxonomy, we suggest retaining the name C. magnus for lineages within this species complex across its range, followed by a clade designation, if known.

Bovine respiratory syncytial virus (BRSV) strains that were detected in Kagoshima prefecture and isolated in Hokkaido between 2017 and 2019, together with a BRSV vaccine strain, were subjected to full-genome sequencing. The BRSV strains identified in Japan were found to be genetically close to each other but distant from the vaccine strains. The deduced amino acids at positions 206 and 208 of the glycoprotein (G protein), which form one of the major epitopes of the recent Japanese BRSV strains, were different from those of the vaccine strains. Therefore, the recent Japanese BRSV strains might be antigenically different from the BRSV vaccine strains., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

The Bioinformation and DNA Data Bank of Japan (DDBJ) Center (https://www.ddbj.nig.ac.jp) maintains database archives that cover a wide range of fields in life sciences. As a founding member of the International Nucleotide Sequence Database Collaboration (INSDC), our primary mission is to collect and distribute nucleotide sequence data, as well as their study and sample information, in collaboration with the National Center for Biotechnology Information in the United States and the European Bioinformatics Institute. In addition to INSDC resources, the Center operates databases for functional genomics (GEA: Genomic Expression Archive), metabolomics (MetaboBank), and human genetic and phenotypic data (JGA: Japanese Genotype-Phenotype Archive). These databases are built on the supercomputer of the National Institute of Genetics, whose remaining computational capacity is actively utilized by domestic researchers for large-scale biological data analyses. Here, we report our recent updates and the activities of our services., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)

We evaluated the utility of single nucleotide polymorphism (SNP) markers for parentage testing in Breton (BR) and Percheron (PR) horses in Japan using the proposed International Society for Animal Genetics (P-ISAG) 147 SNP panel and 414 autosomal SNPs. Genomic DNA was extracted from 98 horses of two breeds, BR (n = 47) and PR (n = 51), and sequenced using next-generation sequencing. The average minor allele frequencies for the P-ISAG panel for BR and PR were 0.306 and 0.301, respectively. The combined probabilities of exclusion (PEs) given two parents and one offspring: exclude a relationship (PE01) and given one parent and one offspring: exclude their relationship (PE02) were over 0.9999 for both breeds. Using the P-ISAG panel, no exclusion or doubtful cases were identified in 35 valid parent-offspring pairs, suggesting that the P-ISAG panel is helpful for parentage verification in both breeds. In contrast, as 0.18% of falsely accepted parentages were observed in the parentage discovery cases, additional markers such as the combination of the P-ISAG panel and 414 autosomal SNPs (561-SNP set) presented here should be used to identify valid parent-offspring pairs of horses with unknown parentage relationships., (© 2023 Japanese Society of Animal Science.)

HLA-B*15:01:74 differs from B*15:01:01:01 by one nucleotide in exon 2 at position 470., (© 2022 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-B*13:01:01:09 differs from HLA-B*13:01:01:01 by one nucleotide in intron 4 at positions 1913., (© 2022 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

A*24:02:01:111 differs from A*24:02:01:01 by two nucleotides in intron 7, positions 2784 and 2785., (© 2022 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The Ryukyu Islands, an island chain in southwestern Japan, originated from land masses that separated from the Eurasian continent due to the formation of sea barriers about 1.55 million years ago. In this study, we investigated the phylogenetic relationships of the operculate land snail genus Cyclophorus (Caenogastropoda: Cyclophoridae) in the Ryukyu Archipelago and surrounding regions based on DNA sequence data. According to our results, all studied Cyclophorus specimens from Japan form a monophyletic group containing eight subclades. Six of these subclades were found only on the Ryukyu Islands. On most islands of the Ryukyu Archipelago, no more than one Cyclophorus subclade was recorded, which may be due to limited ecological niche space and competition. No subclade was found to occur on both sides of the Watase Line, a regional zoogeographical boundary. Divergence times were estimated based on a time-calibrated phylogeny. We found that multiple splits among the Japanese Cyclophorus subclades predate the emergence of major sea barriers in the Ryukyu Archipelago. Vicariance due to sea barrier formation, as assumed for many other taxa from the region, was thus likely not the main driver for subclade divergence in these snails. Instead, certain geographical features might have shaped the diversification of subclades prior to sea barrier formation. Given that Cyclophorus populations were also present on islands that have never been connected to other land masses, the snails must have colonized them via oversea dispersal. As not all nominal taxa corresponded to monophyletic groups, our molecular phylogenetic approach revealed that a taxonomic revision of the Japanese Cyclophorus fauna is necessary. The eight subclades may be regarded as potential species-level groups based on COI p-distances. A canonical discriminant analysis using shell morphological data revealed slight differences among the subclades., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Pulsating broodsacs of Leucochloridium sp. (Trematoda: Leucochloridiidae) were found from amber snails (Succinea lauta) in Iwate, the northern part of Honshu, Japan. A pattern with red-brown vertical stripes was characteristic of the broodsac. Very similar broodsacs were already detected from Okinawa Islands, the southern archipelago of Japan, and tentatively identified as Leucochloridium cf. passeri. A phylogenetic analysis based on DNA sequences of mitochondrial cytochrome c oxidase subunit 1 (cox1) showed that Leucochloridium sp. is different at species level from L. cf. passeri and that both species are related to Leucochloridium vogtianum from Europe. In this study the definitive identification of larval Leucochloridium sp. was impossible, but the resulting phylogeny confirmed that at least 4 species of Leucochloridium are distributed in Japan, depending on locality and climate. The DNA barcode generated in this study will be useful in detecting the adult stage of Leucochloridium sp. from birds., (Copyright © 2021 Elsevier B.V. All rights reserved.)

HIV-1 subtype/circulating recombinant form (CRF) distribution of HIV-1-positive specimens for evaluating HIV in vitro diagnostics (IVDs) was examined and compared with the HIV-1 epidemic in Japan. The nucleotide sequences of the gag-pol region of 173 plasma specimens (84, provided in 2007, and 89 in 2013-2015) were determined. HIV-1 subtype/CRF classification was performed based on the phylogenetic analyses of the sequences. The subtype/CRF distribution resulting in this study was similar to that of a previous epidemiological report. Three CRF02_AG and one unique recombinant form, including subtype G and A regions, were observed in the 2013 and 2014 specimens, except in the 2007 specimens. The reference panel consisting of these specimens was practical for the evaluation of HIV IVDs in Japan.

Background: In this study, we identified a novel glycophorin variant (GP.MOT) in a Mi a -positive Japanese blood donor. The proband with this glycophorin variant was discovered by antigen screening of samples from 475,493 Japanese blood donors using monoclonal anti-Mi a ., Study Design and Methods: Standard serological techniques and flow cytometry were performed. GP.MOT RBCs were examined by immunoblotting using anti-GPA, anti-MUT or anti-Mur. Genome DNA was extracted from whole blood, and the GYPA/GYPB was analyzed by polymerase chain reactions and Sanger sequencing., Results: The MNS blood group of the proband was M + N +  w S-s + with the presence of other low-frequency antigens including Mi a , Mur, MUT, and KIPP. A 43-kDa molecule, which is almost equivalent in size to glycophorin A (GPA), was identified by immunoblotting using monoclonal anti-MUT and anti-Mur. Sanger sequencing clearly indicated that the proband had two different GYPA*M alleles at SNP rs62334651 (GYPA*M232 + 55A and GYPA*M232 + 55G), as well as a GYP(B-A) hybrid allele (GYP*MOT) with breakpoints located on pseudoexon 3 of GYPB from c.210 to c.219., Discussion: We identified a hybrid glycophorin GP.MOT with the deduced unique amino acid sequence GPB (20-45)-GPΨB (46-70)-GPA (71-149), which has not been previously reported., (© 2021 AABB.)

To further classify the oomycete viruses that have been discovered in recent years, we investigated virus infection in the plant-parasitic oomycete Globisporangium ultimum in Japan. Double-stranded RNA detection, high-throughput sequencing, and RT-PCR revealed that the G. ultimum isolate UOP226 contained two viruses related to fusarivirus and totivirus, named Pythium ultimum RNA virus 1 (PuRV1) and Pythium ultimum RNA virus 2 (PuRV2), respectively. Phylogenetic analysis of the deduced amino acid sequence of the RNA-dependent RNA polymerase (RdRp) showed that fusari-like PuRV1 belonged to a different phylogenetic group than Plasmopara viticola lesion-associated fusari virus (PvlaFV) 1-3 from oomycete Plasmopara viticola . Codon usage bias of the PuRV1 RdRp gene was more similar to those of fungi than Globisporangium and Phytophthora , suggesting that the PuRV1 ancestor horizontally transmitted to G. ultimum ancestor from fungi. Phylogenetic analysis of the deduced amino acid sequence of the RdRp of toti-like PuRV2 showed a monophyletic group with the other toti-like oomycete viruses from Globisporangium , Phytophthora, and Pl. viticola . However, the nucleotide sequences of toti-like oomycete viruses were not so homologous, suggesting the possibility of convergent evolution of toti-like oomycete viruses.

Intermediate-sized insertions are one of the structural variants contributing to genome diversity. However, due to technical difficulties in identifying them, their importance in disease pathogenicity and gene expression regulation remains unclear. We used whole-genome sequencing data of 174 Japanese samples to characterize intermediate-sized insertions using a highly-accurate insertion calling method (IMSindel software and joint-call recovery) and obtained a catalogue of 4254 insertions. We constructed an imputation panel comprising of insertions and SNVs from all samples, and conducted imputation of intermediate-sized insertions for 82 publicly-available Japanese samples. Positive Predictive Value of imputation, evaluated using Nanopore long-read sequencing data, was 97%. Subsequent eQTL analysis predicted 128 (~ 3.0%) insertions as causative for gene expression level changes. Enrichment analysis of causal insertions for genome regulatory elements showed significant associations with CTCF-binding sites, super-enhancers, and promoters. Among 17 causal insertions found in the same causal set with GWAS hits, there were insertions associated with changes in expression of cancer-related genes such as BRCA1, ZNF222, and ABCB10. Analysis of insertions sequences revealed that 461 insertions were short tandem duplications frequently found in early-replicating regions of genome. Furthermore, comparison of functional importance of intermediate-sized insertions with that of intermediate-sized deletions detected in the same sample set in our previous study showed that insertions were more frequent in genic regions, and proportion of functional candidates was smaller in insertions. Here, we characterize a high-confidence set of intermediate-sized insertions and indicate their importance in gene expression regulation. Our results emphasize the importance of considering intermediate-sized insertions in trait association studies.

This study assessed whether invasive group B Streptococcus (GBS) isolates were similar to non-invasive isolates from adult patients. Invasive and non-invasive GBS isolates were collected from three hospitals and two laboratory centers between January 2015 and October 2019. The isolates were identified by 16S rRNA amplicon sequencing and amplification of the GBS-specific dltS gene. The virulence gene profiles, capsular genotypes, sequence types (STs)/clonal complexes (CCs), and antimicrobial resistance (AMR) phenotypes/genotypes were determined for the 72 invasive and 50 non-invasive isolates that were comparatively analyzed. We observed a significantly decreased rate of rib detection in the invasive isolates compared to that in the non-invasive isolates (77.8% vs. 92.0%, P < 0.05). Additionally, we found significant differences in the prevalence of CC1 (23.6% vs. 46.0%, P < 0.05) and CC26 (12.5% vs. 2.0%, P < 0.05) between invasive and non-invasive populations. However, there were no significant differences in the comparative data of the virulence gene profiles, capsular genotypes, other STs/CCs, and AMR phenotypes/genotypes between the two populations. These findings suggest that both invasive and non-invasive isolates share similar features in terms of virulence gene profile, capsular genotype, ST/CC, and AMR genotype/phenotype (except for the rates of rib detection and CC1/CC26 prevalence).

Identifying different species of the genus Atractylodes which are commonly used in Chinese and Japanese traditional medicine, using chromatographic approaches can be difficult. 1 H NMR metabolic profiling of DNA-authenticated, archived rhizomes of the genus Atractylodes was performed for genetic and chemical evaluation. The ITS region of the nuclear rDNA was sequenced for five species, A. japonica, A. macrocephala, A. lancea, A. chinensis, and A. koreana. Our samples had nucleotide sequences as previously reported, except that part of the A. lancea cultivated in Japan had a type 5, hybrid DNA sequence. Principal component analysis (PCA) using 1 H NMR spectra of extracts with two solvent systems (CD 3 OD, CDCl 3 ) was performed. When CDCl 3 extracts were utilized, the chemometric analysis enabled the identification and classification of Atractylodes species according to their composition of major sesquiterpene compounds. The 1 H NMR spectra using CD 3 OD contained confounding sugar peaks. PCA removal of these peaks gave the same result as that obtained using CDCl 3 and allowed species distinction. Such chemometric methods with multivariate analysis of NMR spectra will be useful for the discrimination of plant species, without specifying the index components and quantitative analysis on multi-components.

NTRK fusion-positive tumors are known to be highly sensitive to TRK inhibitors, such as larotrectinib and entrectinib. Therefore, identification of patients who can potentially benefit from these inhibitors is important; however, the frequency of NTRK fusions in Japanese patients with colorectal cancer (CRC) is unknown. We performed pan-TRK staining using TMA-based immunohistochemistry (IHC) on samples from 971 consecutive Japanese CRC cases from a single institution. Positive cases were further analyzed using NanoString and subsequent targeted RNA sequencing. We found three positive cases using TRK-IHC. Furthermore, the Nanostring assay supported the presence of NTRK fusion in these cases. Subsequent targeted RNA-sequencing and RT-PCR revealed two cases with TPM3-NTRK1 and one with TPR-NTRK1. The TNM stages of these cases were stage I, stage IIA, and stage IIIB, and two showed microsatellite instability-high status. Next-generation sequencing analysis using Cancer hotspot panel revealed TP53 and SMAD4 mutations in separate cases. IHC of β-catenin did not show nuclear accumulation. We found three cases (0.31%) of CRC with NTRK1 fusion among 971 consecutive Japanese CRC cases. No potential driver alterations other than NTRK fusion were identified in these three patients.

Pleioblastus mosaic virus (PleMV) is a tentative member of the genus Potyvirus in the family Potyviridae and was discovered in bamboo with mosaic symptoms in Tokyo, Japan. Since no information on the genome sequence of PleMV has been reported, its taxonomic position has long been uncertain. Here, we report the first complete genome sequences of two distinct PleMV isolates. Excluding the 3'-terminal poly(A) tail, their genomic RNA sequences consist of 9,634 and 9,643 nucleotides (nt); both contain a large open reading frame (ORF) encoding a polyprotein and a small ORF termed PIPO. The large ORFs of the two isolates share 79.2% and 87.6% sequence identity at the nucleotide (nt) and amino acid (aa) level, respectively, and were found to have the highest nt and aa sequence identity (69.0% and 69.9%) to the potyvirus johnsongrass mosaic virus (JGMV). Phylogenetic analysis showed that PleMV is most closely related to JGMV but forms its own clade. These results suggest that PleMV is a distinct member of the genus Potyvirus.

Background: Spinal muscular atrophy (SMA) is a neuromuscular disease caused by homozygous deletion of SMN1 exons 7 and 8. However, exon 8 is retained in some cases, where SMN2 exon 7 recombines with SMN1 exon 8, forming a hybrid SMN gene. It remains unknown how the hybrid SMN gene contribute to the SMA phenotype., Method: We analyzed 515 patients with clinical suspicion for SMA. SMN1 exons 7 and 8 deletion was detected by PCR followed by enzyme digestion. Hybrid SMN genes were further analyzed by nucleotide sequencing. SMN2 copy number was determined by real-time PCR., Results: SMN1 exon 7 was deleted in 228 out of 515 patients, and SMN1 exon 8 was also deleted in 204 out of the 228 patients. The remaining 24 patients were judged to carry a hybrid SMN gene. In the patients with SMN1 exon 7 deletion, the frequency of the severe phenotype was significantly lower in the patients with hybrid SMN gene than in the patients without hybrid SMN gene. However, as for the distribution of SMN2 exon 7 copy number among the clinical phenotypes, there was no significant difference between both groups of SMA patients with or without hybrid SMN gene., Conclusion: Hybrid SMN genes are not rare in Japanese SMA patients, and it appears to be associated with a less severe phenotype. The phenotype of patients with hybrid SMN gene was determined by the copy number of SMN2 exon 7, as similarly for the patients without hybrid SMN gene., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.)

The Bioinformation and DDBJ Center (DDBJ Center, https://www.ddbj.nig.ac.jp) provides databases that capture, preserve and disseminate diverse biological data to support research in the life sciences. This center collects nucleotide sequences with annotations, raw sequencing data, and alignment information from high-throughput sequencing platforms, and study and sample information, in collaboration with the National Center for Biotechnology Information (NCBI) and the European Bioinformatics Institute (EBI). This collaborative framework is known as the International Nucleotide Sequence Database Collaboration (INSDC). In collaboration with the National Bioscience Database Center (NBDC), the DDBJ Center also provides a controlled-access database, the Japanese Genotype-phenotype Archive (JGA), which archives and distributes human genotype and phenotype data, requiring authorized access. The NBDC formulates guidelines and policies for sharing human data and reviews data submission and use applications. To streamline all of the processes at NBDC and JGA, we have integrated the two systems by introducing a unified login platform with a group structure in September 2020. In addition to the public databases, the DDBJ Center provides a computer resource, the NIG supercomputer, for domestic researchers to analyze large-scale genomic data. This report describes updates to the services of the DDBJ Center, focusing on the NBDC and JGA system enhancements., (© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The International Nucleotide Sequence Database Collaboration (INSDC; http://www.insdc.org/) has been the core infrastructure for collecting and providing nucleotide sequence data and metadata for >30 years. Three partner organizations, the DNA Data Bank of Japan (DDBJ) at the National Institute of Genetics in Mishima, Japan; the European Nucleotide Archive (ENA) at the European Molecular Biology Laboratory's European Bioinformatics Institute (EMBL-EBI) in Hinxton, UK; and GenBank at National Center for Biotechnology Information (NCBI), National Library of Medicine, National Institutes of Health in Bethesda, Maryland, USA have been collaboratively maintaining the INSDC for the benefit of not only science but all types of community worldwide., (© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Severe fever with thrombocytopenia syndrome (SFTS) is caused by SFTS virus (SFTSV). Although SFTS originated in China, it is an emerging infectious disease with prevalence confirmed in Japan, Korea, and Vietnam. The full-length genomes of 51 Korean SFTSV isolates from 2013 to 2016 were sequenced, and the sequences were deposited into a public database (GenBank) and analyzed to elucidate the phylogeny and evolution of the virus. Although most of the Korean SFTSV isolates were closely related to previously reported Japanese isolates, some were closely related to previously reported Chinese isolates. We identified one Korean strain that appears to have resulted from multiple inter-lineage reassortments. Several nucleotide and amino acid variations specific to the Korean isolates were identified. Future studies should focus on how these variations affect virus pathogenicity and evolution.

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) which emerged in the city of Wuhan, Hubei Province, China, has spread worldwide and is threatening human life. The detection of SARS-CoV-2 is critical for preventing new outbreaks, curbing disease spread, and managing patients. Currently, a reverse-transcription polymerase chain reaction (RT-PCR) assay is used to detect the virus in clinical laboratories. However, although this assay is considered to have high specificity, its sensitivity is reportedly as low as 60-70 %. Improved sensitivity is, therefore, urgently required., Methods: We used the primers and single-quencher probes recommended by the CDC (N1, N2 and N3) in the USA and the NIID (N1 and N2) in Japan. In addition, we designed double-quencher probes according to the virus sequence provided by the NIID to develop a further assay (termed the YCH assay [N1 and N2]). Using these assays, we conducted RT-PCR with serially diluted DNA positive controls to assess and compare the detection sensitivity of the three assays. Furthermore, 66 nasopharyngeal swabs were tested to determine the diagnostic performances., Results: The threshold cycle (Ct) value of the RT-PCR was relatively low for the CDC and YCH assays compared with the NIID assay. Serial dilution assays showed that both the CDC and YCH assays could detect low copy numbers of the DNA positive control. The background fluorescence signal at the baseline was lower for the YCH assay compared with the NIID assay. We assessed the diagnostic performance between single- (NIID) and double-quencher (YCH) probes using 66 nasopharyngeal swabs. When the results of YCH-N2 assay were used as a reference, each assay detected SARS-CoV-2 with positive percent agreements of 56 % for NIID-N1, 61 % for YCH-N1, and 94 % for NIID-N2, and 100 % negative percent agreements for NIID-N1, YCH-N1 and NIID-N2., Conclusion: Double-quencher probes decreased the background fluorescence and improved the detection sensitivity of RT-PCR for SARS-CoV-2., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The first study reporting the morphological characterization of Sarcocystis sybillensis was performed in 1983; however, without any molecular analysis. Sarcocystis nipponi has been recently described as a species synonymic to S. sybillensis. We reconfirmed the presence of S. sybillensis in Japanese sika deer (Cervus nippon) captured in its native territory; and performed its molecular and phylogenetic characterization. The morphological characteristics of the sarcocysts were consistent with those of S. nipponi and S. sybillensis described in the first report. However, the nucleotide sequence of 18S rRNA gene of S. sybillensis showed only 91.9% identity to that of S. nipponi, suggesting low homology among the concerned Sarcocystis spp. Accordingly, S. sybillensis was found to occupy a clade distinct from that of S. nipponi in a phylogenetic tree of Sarcocystis. Therefore, the present study provides essential information on 18S rRNA-based molecular characterization of S. sybillensis and disproves the existing notion of morphology-based species synonymity of S. sibillensis and S. nipponi. These results also suggest that S. sybillensis belongs to type 2 Sarcocystis.

The diversity in our genome is crucial to understanding the demographic history of worldwide populations. However, we have yet to know whether subtle genetic differences within a population can be disentangled, or whether they have an impact on complex traits. Here we apply dimensionality reduction methods (PCA, t-SNE, PCA-t-SNE, UMAP, and PCA-UMAP) to biobank-derived genomic data of a Japanese population (n = 169,719). Dimensionality reduction reveals fine-scale population structure, conspicuously differentiating adjacent insular subpopulations. We further enluciate the demographic landscape of these Japanese subpopulations using population genetics analyses. Finally, we perform phenome-wide polygenic risk score (PRS) analyses on 67 complex traits. Differences in PRS between the deconvoluted subpopulations are not always concordant with those in the observed phenotypes, suggesting that the PRS differences might reflect biases from the uncorrected structure, in a trait-dependent manner. This study suggests that such an uncorrected structure can be a potential pitfall in the clinical application of PRS.

Tick-borne viruses have emerged recently in many parts of the world, and the discoveries of novel tick-borne viruses have been accelerated by the development of high-throughput sequencing technology. In this study, a cost-efficient small benchtop next-generation sequencer, the Illumina MiniSeq, was used for the RNA virome analysis of questing ticks collected from Hokuriku District, Japan, and assessed for their potential utility in a tick-borne virus surveillance system. We detected two phleboviruses [Kabuto Mountain virus (KAMV) and Okutama tick virus (OKTV)], a coltivirus [Tarumizu tick virus (TarTV)], and a novel iflavirus [Hamaphysalis flava iflavirus (HfIFV)] from tick homogenates and/or cell culture supernatants after virus isolation processes. The number of sequence reads from KAMV and TarTV markedly increased when cell culture supernatants were used, indicating a successful isolation of these viruses. In contrast, OKTV and HfIFV were detected only in tick homogenates but not from cell culture supernatants, suggesting a failure to isolate these viruses. Furthermore, we performed genomic and phylogenetic analyzes of these detected viruses. OKTV and some phleboviruses discovered recently by NGS-based methods were probably deficient in the M genome segment, which are herein proposed as M segment-deficient phlebovirus (MdPV). A phylogenetic analysis of phleboviruses, including MdPV, suggested that Uukuniemi and Kaisodi group viruses and kabutoviruses evolved from an ancestral MdPV, which provides insights into the evolutionary dynamics of phleboviruses as emerging pathogens., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

Morphological and genetic analyses were performed on seven species of the family Capillariidae (Nematoda: Trichocephalida: Trichinelloidea), viz. Eucoleus perforans, Eucoleus contortus, Aonchotheca bursata, Baruscapillaria obsignata, Capillaria anatis, Capillaria phasianina, and Capillaria spinulosa, detected in poultry from Japan and Indonesia. Two Eucoleus spp., perforating the esophageal mucosa of the Japanese green pheasant farmed in Japan (E. perforans) and domestic goose in Indonesia (E. contortus), were morphologically characterized. Furthermore, we observed substantial nucleotide differences between their 18S ribosomal RNA gene (rDNA), revealing maximum identity (97.27%) over the 1797-bp length. Similarly, B. obsignata in the small intestine of Japanese green pheasants in Japan, a chicken, geese, domestic pigeons, and a turkey in Indonesia, and C. anatis in the ceca of chickens in Indonesia were morphologically and molecular-genetically compared with previously reported isolates of these species in Japan. Aonchotheca bursata collected from the small intestine of the Japanese green pheasant was also molecular-genetically characterized for the first time; however, sequencing of the 18S rDNA of C. phasianina from the cecum of the same bird was unsuccessful. Capillaria worms in the ceca of a domestic duck and geese in Indonesia were identified as C. spinulosa, which had only previously been recorded in wild birds of the Anseriformes in the Palaearctic region. Morphologically, this species was closest to Capillaria pudendotecta recorded from the ceca of wild and captive swans, except for the lack of vaginal appendages in female worms and shorter esophagi relative to the body length in both male and female worms. Phylogenetically, these two species were closely related, although substantial nucleotide changes were noted. The 18S rDNA nucleotide sequences of the species isolated here were consistent with the recent taxonomic system established for Capillariidae based primarily on the morphology of male caudal ends.

The Izumi plain in Kagoshima Prefecture, Japan, is an overwintering site for migratory ducks and endangered cranes. We have surveyed avian influenza viruses (AIVs) in this area since 2012 and isolated low-pathogenic AIVs (LPAIVs) of various subtypes every winter season. H3N8 LPAIVs were isolated during the 2012/13 and 2016/17 seasons, and H4N6 LPAIVs were isolated during the 2012/13 and 2013/14 seasons. In the 2017/18 season, one H3N8 and two H4N6 LPAIV strains were isolated from environmental water samples. Genetic and phylogenetic analysis for each gene segment from these H3N8 and H4N6 LPAIVs suggested that our isolates were genetic reassortants generated by intermixing between AIVs circulating not only in Eurasia but also in Africa and/or North America. Comparison of the genetic constellations of our three isolates with their counterparts isolated during previous seasons from the Izumi plain revealed a drastic transition in the genetic constellations of both subtypes. These findings emphasize the importance of continuous surveillance of AIVs on the Izumi plain.

The A(H1N1)pdm09 virus emerged in 2009 and continues to circulate in human populations. Recent A(H1N1)pdm09 viruses, that is, A(H1N1)pdm09 viruses circulating in the post-pandemic era, can cause more or less severe infections than those caused by the initial pandemic viruses. To evaluate the changes in pathogenicity of the A(H1N1)pdm09 viruses during their continued circulation in humans, we compared the nucleotide and amino acid sequences of ten A(H1N1)pdm09 viruses isolated in Japan between 2009 and 2015, and experimentally infected mice with each virus. The severity of infection caused by these Japanese isolates ranged from milder to more severe than that caused by the prototypic pandemic strain A/California/04/2009 (CA04/09); however, specific mutations responsible for their pathogenicity have not yet been identified., Competing Interests: Y.K. has received speaker’s honoraria from Toyama Chemical and Astellas; grant support from Chugai Pharmaceuticals, Daiichi Sankyo Pharmaceutical, Toyama Chemical, Tauns Laboratories, Otsuka Pharmaceutical, and Kyoritsu Seiyaku; and is a founder of FluGen. All of the other authors declare that they have no conflicts of interest.

The genus Chloromyxum (Myxozoa: Myxosporea: Bivalvulida) is defined as having ridged or smooth bivalvular myxospores containing four polar capsules, with/without caudal filaments. Currently containing more than 140 nominal species, this genus is reasonably speciose with myxospores of unique but heterogeneous morphology. Recent phylogenetic studies have demonstrated its polyphyletic nature. During our myxosporean survey of freshwater fish, a new coelozoic myxosporean species, Chloromyxum trilineatum n. sp., was detected in the gall bladder of the pale chub, Zacco platypus (Cypriniformes: Cyprinidae), which originated from central Japan. Spores were subspherical, measuring 8.5-9.1 (8.8) μm in length, 7.6-8.2 (8.0) μm in width, and 6.8-7.8 (7.4) μm in thickness (n = 20). The valvular surface was smooth and three or four distinct ridges ran parallel to the suture line. Four almost equal polar capsules, 2.9-3.8 (3.3) μm in length and 1.6-2.4 (2.0) μm in width, assembled at the apical part of the spores. The partial nucleotide sequence of the 18S ribosomal RNA gene, 2014 bp in length, was closest to that of morphologically distinct Chloromyxum ellipticum, infecting the gall bladder of grass carp (Ctenopharyngodon idella) in China with 96.99% (1673/1725) identity and three insertion/deletion (indel) sites, followed by Chloromyxum legeri, infecting the gall bladder of common carp (Cyprinus carpio) in the Czech Republic with 89.97% (1803/2004) identity and 14 indel sites. Other myxosporean species, including Chloromyxum spp. from the gall bladder or urinary system of freshwater and marine fish, were phylogenetically distant from the present species.

Albinism, which is commonly inherited as an autosomal recessive trait, is characterized by a reduction or absence of melanin in the eyes, skin, and hair. To date, more than 20 causal genes for albinism have been identified; thus, the accurate diagnosis of albinism requires next-generation sequencing (NGS). In this study, we analyzed 46 patients who tested negative for oculocutaneous albinism (OCA)1-4 and Hermansky-Pudlak syndrome (HPS)1 based on conventional analysis, in addition to 28 new Japanese patients, using NGS-based targeted resequencing. We identified a genetic background for albinism in 18 of the 46 patients (39%), who were previously tested negative according to the conventional analysis. In addition, we unveiled a genetic predisposition toward albinism in 23 of the 28 new patients (82%). We identified six patients with rare subtypes of albinism, including HPS3, HPS4, and HPS6, and found 12 novel pathological mutations in albinism-related genes. Furthermore, most patients who were not diagnosed with albinism by the NGS analysis showed mild manifestations of albinism without apparent eye symptoms and harbored only one heterozygous mutation, occasionally in combination with skin-color associated gene variants., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Species of the fungal genus Colletotrichum are among the most devastating pathogens of agricultural crops in the world. Based on DNA sequence data (ITS, GAPDH, CHS-1, ACT, TUB2) and morphology, we revealed Colletotrichum isolates infecting the oil crop Perilla frutescens, commonly known as shiso, to represent a previously unknown species of the C. destructivum species complex and described it as C. shisoi. We found that C. shisoi appears to be able to adopt a hemibiotrophic lifestyle, characterised by the formation of biotrophic hyphae followed by severe necrotic lesions on P. frutescens, but is less virulent on Arabidopsis, compared to its close relative C. higginsianum which also belongs to the C. destructivum species complex. The genome of C. shisoi was sequenced, annotated and its predicted proteome compared with four other Colletotrichum species. The predicted proteomes of C. shisoi and C. higginsianum, share many candidate effectors, which are small, secreted proteins that may contribute to infection. Interestingly, C. destructivum species complex-specific secreted proteins showed evidence of increased diversifying selection which may be related to their host specificities.

To gain insight into the origin, evolution, dissemination and viral factors affecting HTLV-1-associated diseases, knowing the complete viral genome sequences is important. So far, no full-length HTLV-1 genome sequence has been reported from Iran. Here we report the complete nucleotide sequence of HTLV-1 viruses isolated from adult T cell leukemia/lymphoma (ATLL) patients from this region. The genome size of HTLV-1-MhD (Mashhad) was found to be 9036 bp and sequence analysis of the LTR region showed that it belongs to cosmopolitan subtype A. Comparing the sequences with isolates from another endemic area (HTLV-1ATK) revealed variations in the U3 region (~3.4%), while there was 99.1% and 97.0% similarity in R and U5 regions, respectively. The nucleotide sequences of HTLV-1 gag, pro and pol genes had a difference of 1.1% compared with HTLV-1 ATK with 16 nucleotides replaced in the gag and 27 in the pol regions. There was no variability in the amino acid sequences in the p24 gag , however three residues were different in the p19 gag and one in the p15 gag . The nucleotide sequence of env showed a divergence of 1.5% compared to ATK with 22-nucleotide variation. The HTLV-1-MhD Tax, p13, p30, and p12 had 99.1, 100, 98.8, and 98%, respectively similarity with the prototype strain. Four amino acid changes were detected in ORF1 and ORF2 products p12 and p30, respectively, while the p13 region showed 100% conservation. The nucleotide identity between the isolates of Mashhad and those isolated from France, Germany, China, Canada and Brazil was 99.1%, 99.2%, 97.9%, 99% and 99.3%, respectively. Four amino acid changes compared with HTLV-1ATK from Japan were detected in ORF1 and ORF2 products p12 and p30, respectively, while the p13 region showed 100% conservation. This data could provide information regarding the evolutionary history, phylogeny, origin of the virus and vaccine design., (Copyright © 2019. Published by Elsevier B.V.)

Myomaker is a membrane protein that plays a crucial role in the fusion of myoblasts during muscle growth. DNA methylation, a significant factor, regulates gene expression. The aim of this study was to examine the methylation and mRNA expression patterns of the myomaker gene during 8 different postnatal developmental stages in the Japanese flounder (L: 7 days post hatch (dph); M1: 21 dph; M2: 28 dph; M3: 35 dph; J1: 90 dph; J2: 180 dph; A1: 24 months; A2: 36 months). Muscle tissue samples were taken from Japanese flounder at different postnatal development stages to measure the extent of DNA methylation and gene expression. Methylation level in the promoter and exon 1 of myomaker was measured using bisulfite sequencing, and the relative expression of myomaker during each developmental stage was measured by quantitative PCR. The relative expression levels of myomaker were up-regulated from stages L to M2, M3 to J2, and methylation of myomaker was negatively correlated with mRNA expression. Furthermore, the CpG site located at -26 bp in the promoter was the lowest methylated region in all developmental stages. These results offer a basis for understanding the mechanism by which myomaker regulates muscle formation during postnatal development., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Learedius learedi Price, 1934 , is a blood fluke found in sea turtles, and the adult fluke parasitizes the cardiovascular system of the host. In this study we surveyed 46 green sea turtles, Chelonia mydas , on the Ogasawara Islands, Japan, and blood flukes were detected in the heart and blood vessels of 26 turtles. The flukes were identified as L. learedi based on a detailed morphological description. In addition, molecular identification and characterization of the parasite were performed. The nucleotide sequences of nuclear internal transcribed spacer 2 ( ITS2 ) regions were almost identical to those of L. learedi reported previously, but not to those of Hapalotrema spp., which is the closest related genus. The nucleotide sequences of the 28S ribosomal DNA region formed a single clade with those of the reference L. learedi in the phylogenetic tree, but not with those of Hapalotrema spp. Therefore, the nucleotide sequences of ITS2 and 28S are robust markers for distinguishing L. learedi from other species. The nucleotide sequences of the mitochondrial cytochrome c oxidase subunit 1 ( COI ) region were analyzed to evaluate the genetic variations in L. learedi . The COI haplotypes revealed the extremely high genetic diversity of the species as well as the host turtles on the Ogasawara Islands. The haplotype frequency in the mitochondrial DNA of the green sea turtles on the Ogasawara Islands is known to be significantly different from those in other Pacific rookeries. Although the number of analyzed flukes is small in this study, no haplotype was close to that in other areas; on the basis of the data, we hypothesized that L. learedi differentiated along with the host turtles on the Ogasawara Islands.

Philophthalmid eyeflukes are cosmopolitan parasites of birds and occasionally of mammals, including humans. A gravid adult of Philophthalmus sp. was found from the bulbar conjunctiva of a 64-yr-old woman in Japan, who was diagnosed with acute conjunctivitis. The parasite was morphologically most similar to Philophthalmus hegeneri , but distinctive in lacking an esophagus and in having clearly lobed testes. The DNA sequence analysis of genes for nuclear 28S ribosomal RNA and mitochondrial cytochrome c oxidase subunit 1 supported the identification at generic level. The morphological and molecular analyses strongly suggest that the eyefluke from a human in Japan should be treated as an undescribed species of Philophthalmus . The occurrence of human philophthalmosis is very rare. As far as we know, a total of 11 human cases have been reported worldwide to date.

Background: Flathead fishes of the genus Platycephalus are economically important demersal fishes that widely inhabit the continental shelves of tropical and temperate sea waters. This genus has a long history of taxonomic revision, and recently four Platycephalus species (Platycephalus sp. 1, Platycephalus sp. 2, P. indicus, and P. cultellatus) in the northwestern Pacific Ocean (NWP) have been recognized and redescribed. However, many aspects of their systematics and evolutionary history are unclear., Results: A total of 411 individuals were sampled from 22 different sites across their distributions in the NWP. Three mitochondrial loci were sequenced to clarify the phylogeny and phylogeographic history of the fishes. The results showed significant differentiation of four Platycephalus species in the NWP with well-supported clades in which Platycephalus sp. 1 and Platycephalus sp. 2 were the closest, clustered with P. cultellatus, while their genetic relationship with P. indicus was the furthest. There were significant genealogical branches corresponding to P. indicus but not to other Platycephalus lineages. We further examined the phylogeographic patterns of 16 Platycephalus sp. 1 populations along the coastlines of China and Japan. A total of 69 haplotypes were obtained, with 23 shared among populations. One dominant haplotypic group, with a modest lineage structure and low levels of haplotype diversity and nucleotide diversity, was observed among Platycephalus sp. 1 populations. The demographic history reconstruction suggested a Platycephalus sp. 1 population expansion event dating back to the late Pleistocene., Conclusions: Distributional rang variations may be the crucial factors shaping the genetic relationships of the genus Platycephalus. Reproductive schooling and potential egg/larval dispersal ability, coupled with the effects of ocean currents, are responsible for the present phylogeographic pattern of Platycephalus sp. 1.

A flexuous virus was detected in a Cnidium officinale plant in Japan showing mosaic symptoms. The virus was assigned to the genus Potexvirus based on analysis of its complete nucleotide sequence. The genomic RNA of the virus was 5,964 nucleotides in length, excluding the 3'-terminal poly(A) tail. It contained five open reading frames (ORFs), consistent with other members of Potexvirus. The ORF sequences differ from those of previously reported potexviruses. Phylogenetic analysis indicated that the polymerase of the virus is closely related to that of strawberry mild yellow edge virus; and the CP, to those of both yam virus X and vanilla virus X. We propose that this virus be designated as "cnidium virus X" (CnVX).

In 2018, a patient was diagnosed with Shimokoshi type scrub typhus in Yamagata Prefecture, Japan. The causative pathogen was likely a variant type because 43 (8.3%) of 521 deduced amino acid sequences of the 56-kDa type-specific antigen (TSA) were different from those of the Shimokoshi prototype strain. The patient's paired sera showed low antibody titers against the Shimokoshi prototype strain. Two cases of scrub typhus reported in the Tohoku region during 2011-2012 also involved the same 56-kDa TSA gene sequence. These findings suggest the presence of diversity in Shimokoshi type Orientia tsutsugamushi, which may impede the laboratory diagnosis of scrub typhus., (© 2019 The Societies and John Wiley & Sons Australia, Ltd.)

Mycoplasma bovis isolates belonging to the sequence type 5 (ST5) group, the dominant group in Japan since 1999, were low susceptible to 16-membered macrolides and tetracyclines and were confirmed to have a guanine-to-adenine transition mutation at position 748 in the 23S rRNA gene ( rrl ) and adenine-to-thymine transversion mutations at positions 965 and 967 in the 16S rRNA gene ( rrs ) ( Escherichia coli numbering). Moreover, isolates of ST93 and ST155, members of the ST5 group, were low susceptible to lincosamides and azithromycin and showed an adenine-to-guanine transition mutation at position 2059 of rrl Isolates of ST93 were additionally low susceptible to spectinomycin and showed a cytosine-to-adenine transversion mutation at position 1192 of rrs Strains of the ST5 group seem to spread to Japan and Europe from North America with imported cows, while strains of ST93 and ST155 originated in Japan. Melting curve analysis using hybridization probes revealed the existence of point mutations involved in decreased susceptibility to macrolides, lincosamides, and spectinomycin, as demonstrated by changes in the melting curve shape and/or decreases in the melting peak temperature, so the susceptibility to these antimicrobials can be assessed on the same day. For decreased susceptibility to fluoroquinolones to exist, nonsynonymous mutations in the DNA gyrase gene ( gyrA ) and topoisomerase IV gene ( parC ) had to coexist. The combination of amino acid substitutions of serine at position 83 in gyrA and serine at position 80 in parC resulted in particularly low susceptibility to fluoroquinolones. IMPORTANCE Mycoplasma bovis is the main causal species of bovine mycoplasmal disease and leads to significant economic losses because of its severe symptoms, strong infectivity, and refractoriness. As for mastitis, culling cows with intramammary infections is a general countermeasure to prevent spreading. The conventional antimicrobial susceptibility test for mycoplasma is time-consuming and troublesome, but no quick and easy method for grasping the antimicrobial susceptibility of the causal strain exists at present. Treatment without antimicrobial susceptibility information may be one reason why M. bovis strains in Japan.M. bovis strains in Japan., (Copyright © 2019 American Society for Microbiology.)

In order to develop new domestic production of Rhei Rhizoma (RR) from Rheum specimens cultivated in the Sugadaira Medicinal Plant Cultivation Test Field (SMPCF), the ITS sequences of 12 SMPCF specimens and Chinese Rheum specimens of four species, as well as RR samples produced in North Korea, China and Japan, were determined by subcloning and their sequences were compared. As the ITS sequences of 10 SMPCF specimens showed significant intra-individual polymorphism, identification of pseudogenes was conducted by detecting the three motifs of the 5.8S sequence and the stability of the 5.8S secondary structure. Approximately 46% of sequences obtained from the SMPCF specimens were putative pseudogenes. The maximum likelihood tree based on ITS sequences showed three main groups-the outer group and inner clusters I and II; clones from 10 SMPCF specimens including putative pseudogenes belonged to the outer group. Cluster I was composed of two clades, one including clones from R. officinale specimens and R. palmatum-derived samples with matK genotype Rp9, and another including clones from R. coreanum-derived samples. Cluster II consisted of three clades, one including clones from R. palmatum specimens with genotype Rp5, another including clones mainly from R. tanguticum specimens with genotype Rt4, and the third including clones from R. palmatum or R. tanguticum specimens with various matK genotypes. Clones from SMPCF specimen RC5 showed a close relationship with those from R. tanguticum specimens with matK genotype Rt4, whereas those from specimen RC9 related to R. coreanum-derived samples. As a result, specimens RC5 and RC9 were considered as candidates for the development of domestic RR.

Several surveys of Sarcocystis infection in sika deer in Japan have shown a high prevalence, but the identification has been unclear because molecular data have been lacking or have been limited to 18S ribosomal RNA gene sequences. Thus, in our previous study based on such sequences, some Sarcocystis isolates from sika deer were not clearly separated from other species in the phylogenetic analysis. In the present study, we therefore characterized sarcocyst isolates from sika deer (Cervus nippon centralis) at the mitochondrial cytochrome c oxidase subunit I gene (cox1). Moreover, we developed a multiplex PCR based on cox1 sequences of all species found, so that we could rapidly identify sarcocysts of these species. Twenty-one sarcocysts from nine sika deer were examined. Five distinct cox1 sequence types, each with a high sequence identity (> 99%), were found, and the sarcocysts could thus be classified into five species. Based on the sequence comparisons and the phylogeny, Sarcocystis spp. of types 1, 3, and 5 are considered to represent three new species, which were most closely related to Sarcocystis silva/Sarcocystis truncata, Sarcocystis entzerothi, and Sarcocystis iberica/Sarcocystis venatoria, respectively. There was a slight uncertainly whether Sarcocystis sp. with type 2 sequences represented a new species or was identical to Sarcocystis tarandi. Type 4 sequences showed 99% identity with those of Sarcocystis pilosa from sika deer in Lithuania and have therefore been assigned to this species. In the multiplex PCR, type-specific fragments were successfully amplified for all five Sarcocystis spp., indicating that this assay may be useful for a rapid identification of sarcocysts of these species.

Two species of Endoclita C. . R. Felder, 1874 from Taiwan are evaluated for their taxonomic status: Endoclita meifenga Buchsbaum Grehan sp. n. is described as new for a unique male from the central mountains of Taiwan, and E. kosemponis (Strand, 1916), stat. rev. is elevated to species status. We show that the forewing markings and genitalic characteristics of E. meifenga sp. n. distinguish this species from all other Endoclita. The external appearance of E. kosemponis is similar to E. sinensis (Moore, 1877) but the two species show distinct differences in the male and female genitalia. Preliminary comparison of E. sinensis from Taiwan and Japan suggests they represent more than one species. A sample of the COI gene was sequenced for E. meifenga sp. n. and the primary types of E. meifenga sp. n. and E. kosemponis sp. n. are illustrated here for the first time. The taxonomic status of the non endemic Taiwanese E. davidi (Poujade, 1886) and E. sinensis needs future evaluation.

Fanconi anemia (FA) is a genetically and clinically heterogeneous disorder that predisposes patients to bone marrow failure (BMF), myelodysplastic syndromes (MDS), and acute myeloid leukemia (AML). To study which genetic and phenotypic factors predict clinical outcomes for Japanese FA patients, we examined the FA genes, bone marrow karyotype, and aldehyde dehydrogenase-2 (ALDH2) genotype; variants of which are associated with accelerated progression of BMF in FA. In 88 patients, we found morphologic MDS/AML in 33 patients, including refractory cytopenia in 16, refractory anemia with excess blasts (RAEB) in 7, and AML in 10. The major mutated FA genes observed in this study were FANCA (n = 52) and FANCG (n = 23). The distribution of the ALDH2 variant alleles did not differ significantly between patients with mutations in FANCA and FANCG. However, patients with FANCG mutations had inferior BMF-free survival and received hematopoietic stem cell transplantation (HSCT) at a younger age than those with FANCA mutations. In FANCA, patients with the c.2546delC mutation (n = 24) related to poorer MDS/AML-free survival and a younger age at HSCT than those without this mutation. All patients with RAEB/AML had an abnormal karyotype and poorer prognosis after HSCT; specifically, the presence of a structurally complex karyotype with a monosomy (n = 6) was associated with dismal prognosis. In conclusion, the best practice for a clinician may be to integrate the morphological, cytogenetic, and genetic data to optimize HSCT timing in Japanese FA patients.

A new species, Fibularia coffea sp. nov., occurs from shallow waters in Japan. This new species is distinguished from the other species of Fibularia by the following characters: test height is low, oral surface is slightly depressed toward the peristome, number of pores of petal III continues to increase with the test growth, reaching over 30 at TL &gt; 7.5 mm, and black pigments form symmetric pentaradial on aboral surface in living animals. Two further Japanese species, Fibularia japonica and F. ovulum, are redescribed based on the type specimens (F. japonica) and additional specimens (F. ovulum), respectively. A tabular key to the extant species of Fibularia is also provided. A partial fragment of the mitochondrial gene cytochrome oxidase subunit I (COI) of the type specimens of F. coffea sp. nov. and the additional specimen of F. japonica was sequenced for barcoding in future works.

To date, 26 Kudoa spp. (Myxozoa: Myxosporea: Multivalvulida) have been recorded in edible marine fishes in Japan. In the future, it is likely that even more marine fish multivalvulid myxosporeans will be characterized morphologically and genetically, which will aid the precise understanding of their biodiversity and biology. We examined 60 individuals of six fish species collected from the Philippine Sea off Kochi or from the border between the Philippine Sea and East China Sea around Miyako Island, Okinawa, i.e., the southern part of Japan. Newly collected parasite species included Kudoa yasunagai from the brain of Japanese meagre (Argyrosomus japonicus) and Japanese parrotfish (Calotomus japonicus), Kudoa miyakoensis n. sp. and Kudoa thalassomi from the brain and trunk muscle, respectively, of bluespine unicornfish (Naso unicornis), and Kudoa igami from the trunk muscle of Carolines parrotfish (Calotomus carolinus), African coris (Coris gaimard), and Pastel ringwrasse (Hologymnosus doliatus). With the exception of Japanese parrotfish for K. yasunagai, all these fish are new host records for each kudoid species. Notable variation in the number of shell valves (SV) and polar capsules (PC) was observed for all four kudoid species. In particular, spores with seven or eight SV/PC were prominent in K. igami isolates, despite the original Japanese parrotfish-derived description characterizing it as having spores with six, or less commonly five, SV/PC. However, molecular genetic characterization based on the ribosomal RNA gene (rDNA) and mitochondrial DNA (cytochrome c oxidase subunit 1 and ribosomal RNA small and large subunits) found no significant differences in the nucleotide sequences of isolates with different phenotypical features as far as examined in the present study. A newly erected species, K. miyakoensis n. sp., was determined to be phylogenetically closest to brain-parasitizing species, such as K. chaetodoni, K. lemniscati, and K. yasunagai based on rDNA nucleotide sequences, but differed from them morphologically.

A novel lytic bacteriophage, Escherichia phage EcS1, was isolated from sewage samples collected in Higashi-Hiroshima, Japan. The complete genome sequence of EcS1 was determined using the Illumina Miseq System. The whole genome of EcS1 was found to be 175,437 bp in length with a mean G+C content of 37.8%. A total of 295 genes were identified as structural, functional, and hypothetical genes. BLAST analysis of the EcS1 genomic sequence revealed the highest identity (79%; query cover of 73-74%) to three T4-related phages that infect Serratia sp. ATCC 39006. Host range experiments revealed that EcS1 has lytic effects on three pathogenic strains of Shigella spp. and a pathogenic strain of Salmonella enterica as well as on E. coli strains. However, two strains of Serratia marcescens showed resistance to this phage. Phylogenetic trees for phage tail fiber protein sequences revealed that EcS1 is closely related to Enterobacteriaceae-infecting phages. Thus, EcS1 is a novel phage that infects several pathogenic strains of the family Enterobacteriaceae.

Background: Although a number of recent studies have uncovered remarkable diversity of microbes associated with plants, understanding and managing dynamics of plant microbiomes remain major scientific challenges. In this respect, network analytical methods have provided a basis for exploring "hub" microbial species, which potentially organize community-scale processes of plant-microbe interactions., Methods: By compiling Illumina sequencing data of root-associated fungi in eight forest ecosystems across the Japanese Archipelago, we explored hubs within "metacommunity-scale" networks of plant-fungus associations. In total, the metadata included 8080 fungal operational taxonomic units (OTUs) detected from 227 local populations of 150 plant species/taxa., Results: Few fungal OTUs were common across all the eight forests. However, in each of the metacommunity-scale networks representing northern four localities or southern four localities, diverse mycorrhizal, endophytic, and pathogenic fungi were classified as "metacommunity hubs," which were detected from diverse host plant taxa throughout a climatic region. Specifically, Mortierella (Mortierellales), Cladophialophora (Chaetothyriales), Ilyonectria (Hypocreales), Pezicula (Helotiales), and Cadophora (incertae sedis) had broad geographic and host ranges across the northern (cool-temperate) region, while Saitozyma/Cryptococcus (Tremellales/Trichosporonales) and Mortierella as well as some arbuscular mycorrhizal fungi were placed at the central positions of the metacommunity-scale network representing warm-temperate and subtropical forests in southern Japan., Conclusions: The network theoretical framework presented in this study will help us explore prospective fungi and bacteria, which have high potentials for agricultural application to diverse plant species within each climatic region. As some of those fungal taxa with broad geographic and host ranges have been known to promote the survival and growth of host plants, further studies elucidating their functional roles are awaited.