Publications by authors named "Hiroto Shinomiya"

36 Publications

The incubation period of the SARS-CoV-2 B.1.1.7 variant is shorter than that of other strains.

J Infect 2021 Jun 16. Epub 2021 Jun 16.

Ehime Prefectural Institute of Public Health and Environmental Science, 8-234 Sanbancho, Matsuyama city, Ehime, 790-0003, Japan.

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http://dx.doi.org/10.1016/j.jinf.2021.06.011DOI Listing
June 2021

Subtype screening of bla genes using bipartite primers for DNA sequencing.

Jpn J Infect Dis 2021 Mar 31. Epub 2021 Mar 31.

Department of Microbiology, Ehime Prefectural Institute of Public Health and Environmental Science, Japan.

Genes conferring carbapenem resistance have spread worldwide among gram-negative bacteria. Subtyping of these genes has epidemiological value due to the global cross-border movement of people. Subtyping of bla genes that frequently detected in Japan appears to be important in public health settings; however, there are few useful tools for this purpose. We developed a subtyping screening tool based on PCR direct sequencing, which targets the internal sequences of almost all bla genes. The tool used bipartite multiplex primers with M13 universal sequences at the 5'-end. According to in silico analysis, among the 78 known IMP-type genes, except for bla, 77 detected genes were estimated to be differentiated. In vitro evaluation indicated that sequences of amplicons of IMP-1, IMP-6, IMP-7, and IMP-20 templates were identical to their respective subtypes. Even if the amplicons were small or undetectable through the first PCR, sufficient amplicons for DNA sequencing were obtained through a second PCR using the M13 universal primers. In conclusion, our tool can be possibly used for subtype screening of bla, which is useful for the surveillance of bacteria with bla in clinical and public health settings or environmental fields.
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http://dx.doi.org/10.7883/yoken.JJID.2020.926DOI Listing
March 2021

Isolation and plasmid characterisation of Salmonella enterica serovar Albany harbouring mcr-5 from retail chicken meat in Japan.

FEMS Microbiol Lett 2020 08;367(15)

Bacteriology Section, Division of Microbiology, Osaka Institute of Public Health, 1-3-69 Nakamichi, Higashinari-ku, Osaka, Japan.

The emergence of plasmid-mediated colistin resistance genes (mcr), which is occurring in numerous countries, is a worldwide concern, primarily because colistin is a last-resort antibiotic. Compared to E. coli, prevalence of mcr genes in Salmonella is unclear in Japan. Here we screened for mcr-1-5 genes in our collection of Salmonella strains isolated from retail meat products collected in Japan from 2012 through 2016. We found that Salmonella Albany strain 27A-368 encodes mcr-5 and that mcr genes were undetectable among the remaining 202 isolates. The resistance plasmid p27A-368 was transferred by conjugation to S. Infantis and was stably retained as a transconjugant. Whole-genome sequencing revealed that mcr-5 resided on a 115 kb plasmid (p27A-368). The plasmid backbone of p27A-368 is more similar to that of pCOV27, an ESBL-encoding plasmid recovered from avian pathogenic E. coli, rather than pSE13-SA01718 of S. Paratyphi B that encodes mcr-5. Further, mcr-5 is located on a transposon, and its sequence is similar to that of pSE13-SA01718. A phylogenetic tree based on single nucleotide variants implies a relationship between 27A-368 and S. Albany isolated in Southeast Asian countries.
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http://dx.doi.org/10.1093/femsle/fnaa127DOI Listing
August 2020

Single-Tube Multiplex Polymerase Chain Reaction for the Detection of Genes Encoding Enterobacteriaceae Carbapenemase.

Jpn J Infect Dis 2020 Mar 29;73(2):166-172. Epub 2019 Nov 29.

Department of Microbiology, Ehime Prefectural Institute of Public Health and Environmental Science.

A multiplex PCR assay in a single tube was developed for the detection of the carbapenemase genes of Enterobacteriaceae. Primers were designed to amplify the following six carbapenemase genes: bla, bla, bla, bla, bla, and bla. Of 70 bla variants, 67 subtypes were simulated to be PCR-positive based on in silico simulation and the primer-design strategy. After determining the optimal PCR conditions and performing in vitro assays, the performance of the PCR assay was evaluated using 51 and 91 clinical isolates with and without carbapenemase genes, respectively. In conclusion, the combination of multiplex PCR primers and QIAGEN Multiplex PCR Plus Kit was used to determine the best performance for the rapid and efficient screening of carbapenemase genes in Enterobacteriaceae. The assay had an overall sensitivity and specificity of 100%. This PCR assay compensates for the limitations of phenotypic testing, such as antimicrobial susceptibility testing and the modified carbapenem inactivation method, in clinical and public health settings.
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http://dx.doi.org/10.7883/yoken.JJID.2019.041DOI Listing
March 2020

Characterization of the Ca-coordination structures of L- and T-plastins in combination with their synthetic peptide analogs by FTIR spectroscopy.

Sci Rep 2019 03 12;9(1):4217. Epub 2019 Mar 12.

Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, University of Tokyo, Tokyo, 113-8657, Japan.

FTIR spectroscopy was employed to characterize the coordination structures of divalent cations (M = Ca or Mg) bound by L- and T-plastins, which contain two EF-hand motifs. We focused on the N-terminal headpieces in the L- and T-plastins to analyze the regions of COO stretching and amide-I in solution. The spectral profiles indicated that these headpieces have EF-hand calcium-binding sites because bands at 1551 cm and 1555 cm were observed for the bidentate coordination mode of Glu at the 12th position of the Ca-binding site of Ca-loaded L-plastin and T-plastin, respectively. The amide-I profile of the Mg-loaded L-plastin headpiece was identical with that of the apo L-plastin headpiece, meaning that L-plastin has a lower affinity for Mg. The amide-I profiles for apo, Mg-loaded and Ca-loaded T-plastin suggested that aggregation was generated in protein solution at a concentration of 1 mM. The implications of the FTIR spectral data for these plastin headpieces are discussed on the basis of data obtained for synthetic peptide analogs corresponding to the Ca-binding site.
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http://dx.doi.org/10.1038/s41598-019-40889-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6414500PMC
March 2019

Predicting Directions of Changes in Genotype Proportions Between Norovirus Seasons in Japan.

Front Microbiol 2019 5;10:116. Epub 2019 Feb 5.

Kitasato Institute for Life Sciences, Kitasato University, Minato, Japan.

The norovirus forecasting system (NOROCAST) has been developed for predicting directions of changes in genotype proportions between human norovirus (HuNoV) seasons in Japan through modeling herd immunity to structural protein 1 (VP1). Here 404 nearly complete genomic sequences of HuNoV were analyzed to examine whether the performance of NOROCAST could be improved by modeling herd immunity to VP2 and non-structural proteins (NS) in addition to VP1. It was found that the applicability of NOROCAST may be extended by compensating for unavailable sequence data and observed genotype proportions of 0 in each season. Incorporation of herd immunity to VP2 and NS did not appear to improve the performance of NOROCAST, suggesting that VP1 may be a suitable target of vaccines.
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http://dx.doi.org/10.3389/fmicb.2019.00116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6370659PMC
February 2019

Seroprevalence of severe fever with thrombocytopenia syndrome (SFTS) virus antibodies in humans and animals in Ehime prefecture, Japan, an endemic region of SFTS.

J Infect Chemother 2018 Oct 13;24(10):802-806. Epub 2018 Jul 13.

Department of Microbiology, Ehime Prefectural Institute of Public Health and Environmental Science, 8-234 Sanbancho, Matsuyama-city, Ehime, 790-0003, Japan. Electronic address:

Severe fever with thrombocytopenia syndrome (SFTS) was first identified as an emerging tick-borne infectious disease caused by the SFTS virus (SFTSV) in China and has also been found to be endemic to Japan and South Korea, indicating that SFTS is of great concern in East Asia. The aim of the present study was to determine the seroprevalence of SFTSV antibodies in humans and animals in SFTS-endemic regions of Japan. One of 694 (0.14%) healthy persons over 50 years of age and 20 of 107 (18.7%) wild and domestic animals in Ehime prefecture of western Japan were determined to be seropositive for SFTSV antibodies by virus neutralization test and ELISA, respectively. The seropositive person, a healthy 74-year-old woman, was a resident of the southwest part of Ehime prefecture engaged in citriculture and field work. This woman's sample exhibited neutralizing activity against SFTSV although she had neither a clear experience with tick bites nor SFTS-like clinical illness. These findings indicate that most people living in the endemic regions are not infected with SFTSV and suggest that most of the SFTS patients reported so far do not reflect the tip of an iceberg of people infected with SFTSV, but at the same time, that SFTSV infection does not always induce severe SFTS-associated symptoms. These findings also suggested that SFTSV has been maintained in nature within animal species and ticks.
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http://dx.doi.org/10.1016/j.jiac.2018.06.007DOI Listing
October 2018

Characterization of a novel thogotovirus isolated from Amblyomma testudinarium ticks in Ehime, Japan: A significant phylogenetic relationship to Bourbon virus.

Virus Res 2018 04 13;249:57-65. Epub 2018 Mar 13.

Department of Medical Entomology, National Institute of Infectious Diseases, 1-23-1 Toyama, Shinjuku-ku, Tokyo 162-8640, Japan; Department of Agricultural and Environmental Biology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Yayoi 1-1-1, Bunkyo-ku, Tokyo 113-8657, Japan. Electronic address:

The genus Thogotovirus, as represented by Thogoto virus and Dhori virus, comprises a group of arthropod-borne viruses, most members of which are transmitted by ticks. Here we report the genetic and biological characterization of a new thogotovirus, designated Oz virus (OZV), isolated from the hard tick Amblyomma testudinarium in Ehime, Japan. OZV efficiently replicated and induced a cytopathic effect in Vero cells, from which enveloped pleomorphic virus particles were formed by budding. OZV could also replicate in BHK-21 and DH82 cells and caused high mortality in suckling mice after intracerebral inoculation. Phylogenetic analyses of six viral proteins indicated that OZV is clustered with Dhori and related viruses, and is most closely related in glycoprotein (GP) and matrix protein (M) sequences to Bourbon virus, a human-pathogenic thogotovirus discovered recently in the United States. Our findings emphasize the need for understanding the geographic distribution and ecology of OZV and related viruses and for reevaluation of the medical and public health importance of thogotoviruses.
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http://dx.doi.org/10.1016/j.virusres.2018.03.004DOI Listing
April 2018

Evaluation of a surface plasmon resonance imaging-based multiplex O-antigen serogrouping for Escherichia coli using eleven major serotypes of Shiga -toxin-producing E. coli.

J Infect Chemother 2018 Jun 28;24(6):443-448. Epub 2018 Feb 28.

Department of Clinical Laboratory Medicine, Kyoto University Graduate School of Medicine, 54 Syogoin Kawahara-cho, Sakyo-ku, Kyoto 606-8507, Japan.

The early detection of Shiga toxin-producing Escherichia coli (STEC) is important for early diagnosis and preventing the spread of STEC. Although the confirmatory test for STEC should be based on the detection of Shiga toxin using molecular analysis, isolation permits additional characterization of STEC using a variety of methods, including O:H serotyping. The conventional slide agglutination O-antigen serogrouping used in many clinical laboratories is laborious and time-consuming. Surface plasmon resonance (SPR)-based immunosensors are commonly used to investigate a large variety of bio-interactions such as antibody/antigen, peptide/antibody, DNA/DNA, and antibody/bacteria interactions. SPR imaging (SPRi) is characterized by multiplexing capabilities for rapidly screening (approximately 100 to several hundred sensorgrams in parallel) molecules. SPRi-based O-antigen serogrouping method for STEC was recently developed by detecting the interactions between O-antigen-specific antibodies and bacterial cells themselves. The aim of this study was to evaluate its performance for E. coli serogrouping using clinical STEC isolates by comparing the results of slide agglutination tests. We tested a total of 188 isolates, including O26, O45, O91, O103, O111, O115, O121, O128, O145, O157, and O159. The overall sensitivity of SPRi-based O-antigen serogrouping was 98.9%. Only two O157 isolates were misidentified as nontypeable and O121. The detection limits of all serotypes were distributed between 1.1 × 10 and 17.6 × 10 CFU/ml. Pulsed-field gel electrophoresis (PFGE) revealed the heterogeneity of the examined isolates. In conclusion, SPRi is a useful method for the O-antigen serogrouping of STEC isolates, but the further evaluation of non-O157 minor serogroups is needed.
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http://dx.doi.org/10.1016/j.jiac.2018.01.008DOI Listing
June 2018

Genetic Analysis of Human Norovirus Strains in Japan in 2016-2017.

Front Microbiol 2018 18;9. Epub 2018 Jan 18.

Infectious Disease Surveillance Center, National Institute of Infectious Diseases, Musashimurayama, Japan.

In the 2016/2017 winter season in Japan, HuNoV GII.P16-GII.2 strains (2016 strains) emerged and caused large outbreaks of acute gastroenteritis. To better understand the outbreaks, we examined the molecular evolution of the gene and region in 2016 strains from patients by studying their time-scale evolutionary phylogeny, positive/negative selection, conformational epitopes, and phylodynamics. The time-scale phylogeny suggested that the common ancestors of the 2016 strains gene and region diverged in 2006 and 1999, respectively, and that the 2016 strain was the progeny of a pre-2016 GII.2. The evolutionary rates of the gene and region were around 10 substitutions/site/year. Amino acid substitutions (position 341) in an epitope in the P2 domain of 2016 strains were not found in pre-2016 GII.2 strains. Bayesian skyline plot analyses showed that the effective population size of the gene in GII.2 strains was almost constant for those 50 years, although the number of patients with NoV GII.2 increased in 2016. The 2016 strain may be involved in future outbreaks in Japan and elsewhere.
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http://dx.doi.org/10.3389/fmicb.2018.00001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5778136PMC
January 2018

Phylogeny and Immunoreactivity of Norovirus GII.P16-GII.2, Japan, Winter 2016-17.

Emerg Infect Dis 2018 01;24(1):144-148

During the 2016-17 winter season in Japan, human norovirus GII.P16-GII.2 strains (2016 strains) caused large outbreaks of acute gastroenteritis. Phylogenetic analyses suggested that the 2016 strains derived from the GII.2 strains detected during 2010-12. Immunochromatography between 2016 strains and the pre-2016 GII.2 strains showed similar reactivity.
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http://dx.doi.org/10.3201/eid2401.170284DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5749477PMC
January 2018

A patient with severe fever with thrombocytopenia syndrome and hemophagocytic lymphohistiocytosis-associated involvement of the central nervous system.

J Infect Chemother 2018 Apr 11;24(4):292-297. Epub 2017 Nov 11.

Department of Virology 1, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan. Electronic address:

Severe fever with thrombocytopenia syndrome (SFTS), a severe infectious disease caused by novel bunyavirus, SFTS virus (SFTSV), is endemic to China, Korea, and Japan. Most SFTS patients show abnormalities in consciousness. Pathological findings in the central nervous system (CNS) of SFTS patients are not reported. A 53-year-old Japanese man was admitted to Uwajima City Hospital with an 8-day history of fever and diarrhea. Laboratory tests revealed leukopenia, thrombocytopenia, and liver enzyme elevation. He was diagnosed as having severe fever with thrombocytopenia syndrome (SFTS) following detection of the SFTSV genome in his blood. Bone marrow aspiration revealed hemophagocytic lymphohistiocytosis. He suffered progressive CNS disturbance and died on day 13 from onset of first symptoms. The SFTSV genome load in blood and levels of certain cytokines increased over the disease course. Necrotizing lymphadenitis with systemic lymphoid tissues positive for nucleocapsid protein (NP) of SFTSV was revealed by immunohistochemical (IHC) analysis. SFTSV-NP-positive immunoblasts were detected in all organs examined, including the CNS, and in the vascular lumina of each organ. Parenchymal cells of all organs examined were negative for SFTSV-NP on IHC analysis. Microscopic examination of the pons showed focal neuronal cell degeneration with hemosiderin-laden macrophages around extended microvessels with perivascular inflammatory cell infiltration and intravascular fibrin deposition. Autopsy confirmed this patient with SFTS was positive for systemic hemophagocytic lymphohistiocytosis including in the CNS. This patient's neurological abnormalities may have been caused by both functional and organic abnormalities. These novel findings provide important insights into the pathophysiology of SFTS.
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http://dx.doi.org/10.1016/j.jiac.2017.10.016DOI Listing
April 2018

Unusual presentation of a severely ill patient having severe fever with thrombocytopenia syndrome: a case report.

J Med Case Rep 2017 Feb 3;11(1):27. Epub 2017 Feb 3.

Department of Virology 1, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo, 208-0011, Japan.

Background: Severe fever with thrombocytopenia syndrome is an emerging infectious disease caused by a novel phlebovirus belonging to the family Bunyaviridate. Emergence of encephalitis/encephalopathy during severe fever with thrombocytopenia syndrome progression has been identified as a major risk factor associated with a poor prognosis. Here we report the case of a severely ill patient with severe fever with thrombocytopenia syndrome virus-associated encephalitis/encephalopathy characterized by a lesion of the splenium, which resolved later.

Case Presentation: A 56-year-old Japanese man presented with fever and diarrhea, followed by dysarthria. Diffusion-weighted magnetic resonance imaging demonstrated high signal intensity in the splenium of the corpus callosum. The severe fever with thrombocytopenia syndrome virus genome was detected in our patient's serum, and the clinical course was characterized by convulsion, stupor, and hemorrhagic manifestations, with disseminated intravascular coagulation and hemophagocytic lymphohistiocytosis. Supportive therapy not including administration of corticosteroids led to gradual improvement of the clinical and laboratory findings, and magnetic resonance imaging demonstrated resolution of the splenial lesion. The serum severe fever with thrombocytopenia syndrome viral copy number, which was determined with the quantitative reverse-transcription polymerase chain reaction, rapidly decreased despite the severe clinical course. Our patient's overall condition improved, allowing him to be eventually discharged.

Conclusions: Patients with encephalitis/encephalopathy due to severe fever with thrombocytopenia syndrome virus infection may have a favorable outcome, even if they exhibit splenial lesions and a severe clinical course; monitoring the serum viral load may be of value for prediction of outcome and potentially enables the avoidance of corticosteroids to intentionally cause opportunistic infection.
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http://dx.doi.org/10.1186/s13256-016-1192-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5290612PMC
February 2017

[Genetic Investigation of bla(CTX-M)-typing in Extended-spectrum β-lactamase-producing Escherichia coli and Klebsiella pneumoniae from Clinical Specimens and Commercially Available Chicken Liver].

Kansenshogaku Zasshi 2016 May;90(3):305-9

A genetic investigation consisting of the bla(CTX-M) typing was performed using 40 extended spectrum β-lactamase (ESBL)-producing Escherichia coli isolates from chicken liver and 43 ESBL-producing E. coli and 42 ESBL-producing Klebsiella pneumoniae isolates from patients. The types were determined using a sequence analysis. In 31 isolates in the bla(CTX-M-1) group, there were 13 with the bla(CTX-M-1) and all were from chicken liver. Nine E. coli isolates from chicken liver and one E. coli isolate from patients were found to be bla(CTX-M-55). In the bla(CTX-M-15), there were 6 E. coli isolates and one K. pneumoniae isolate from patients. All 39 isolates in the bla(CTX-M-2) group had the blac(CTX-M-2). Fifty-five isolates were found in the bla(CTX-M-9) group, the highest detection frequency, with 36 possessing bla(CTX-M-14) : 20 E. coli and 13 K. pneumoniae from patients and 3 E. coli from chicken liver. There were 17 bla(CTX-M-27) isolates, including 10 E. coli and 7 K. pneumoniae from patients. One bla(CTX-M-90) K. pneumoniae isolate and one bla(CTX-M-9) E. coli isolate were also obtained from patients. These results indicate that the E. coli isolates from chicken liver consist of bla(CTX-M-1), bla(CTX-M-2) and bla(CTX-M-55) ; the E. coli isolates from patients consist of bla(CTX-M-14) and bla(CTX-M-27) ; and the K. pneumoniae isolates from patients consist of bla(CTX-M-2), bla(CTX-M-14) and bla(CTX-M-27). Therefore, the bla(CTX-M) type differs in isolates from chicken liver and those from humans. These results suggest that it is unlikely that ESBL-producing E. coli from chicken liver are firmly established in the human intestinal tract.
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May 2016

Molecular evolution of the capsid gene in human norovirus genogroup II.

Sci Rep 2016 07 7;6:29400. Epub 2016 Jul 7.

Department of Microbiology, Yokohama City University Graduate School of Medicine, Yokohama-shi, Kanagawa 236-0027, Japan.

Capsid protein of norovirus genogroup II (GII) plays crucial roles in host infection. Although studies on capsid gene evolution have been conducted for a few genotypes of norovirus, the molecular evolution of norovirus GII is not well understood. Here we report the molecular evolution of all GII genotypes, using various bioinformatics techniques. The time-scaled phylogenetic tree showed that the present GII strains diverged from GIV around 1630CE at a high evolutionary rate (around 10(-3) substitutions/site/year), resulting in three lineages. The GII capsid gene had large pairwise distances (maximum > 0.39). The effective population sizes of the present GII strains were large (>10(2)) for about 400 years. Positive (20) and negative (over 450) selection sites were estimated. Moreover, some linear and conformational B-cell epitopes were found in the deduced GII capsid protein. These results suggested that norovirus GII strains rapidly evolved with high divergence and adaptation to humans.
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http://dx.doi.org/10.1038/srep29400DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4935990PMC
July 2016

Predicting genotype compositions in norovirus seasons in Japan.

Microbiol Immunol 2016 Jun;60(6):418-26

Department of Virology II.

Noroviruses cause acute gastroenteritis. Since multiple genotypes of norovirus co-circulate in humans, changing the genotype composition and eluding host immunity, development of a polyvalent vaccine against norovirus in which the genotypes of vaccine strains match the major strains in circulation in the target season is desirable. However, this would require prediction of changes in the genotype composition of circulating strains. A fitness model that predicts the proportion of a strain in the next season from that in the current season has been developed for influenza A virus. Here, such a fitness model that takes into account the fitness effect of herd immunity was used to predict genotype compositions in norovirus seasons in Japan. In the current study, a model that assumes a decline in the magnitude of cross immunity between norovirus strains according to an increase in the divergence of the major antigenic protein VP1 was found to be appropriate for predicting genotype composition. Although it is difficult to predict the proportions of genotypes accurately, the model is effective in predicting the direction of change in the proportions of genotypes. The model predicted that GII.3 and GII.4 may contract, whereas GII.17 may expand and predominate in the 2015-2016 season. The procedure of predicting genotype compositions in norovirus seasons described in the present study has been implemented in the norovirus forecasting system (NOROCAST).
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http://dx.doi.org/10.1111/1348-0421.12384DOI Listing
June 2016

Genetic analysis of human rotavirus C: The appearance of Indian-Bangladeshi strain in Far East Asian countries.

Infect Genet Evol 2016 07 9;41:160-173. Epub 2016 Apr 9.

Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan. Electronic address:

Rotaviruses C (RVCs) circulate worldwide as an enteric pathogen in both humans and animals. Most studies of their genetic diversity focus on the VP7 and VP4 genes, but the complete genomes of 18 human RVCs have been described in independent studies. The genetic background of the Far East Asian RVCs is different than other human RVCs that were found in India and Bangladesh. Recently, a RVC detected in 2010 in South Korea had genetic background similar to the Indian-Bangladeshi RVCs. This study was undertaken to determine the whole genome of eight Japanese RVCs detected in 2005-2012, and to compare them with other human and animal global RVCs to better understand the genetic background of contemporary Far East Asian RVC. By phylogenetic analysis, the human RVCs appeared to be distinct from animal RVCs. Among human RVCs, three lineage constellations had prolonged circulation. The genetic background of the Far East Asian RVC was distinguished from Indian-Bangladeshi RVC as reported earlier. However, we found one Japanese RVC in 2012 that carried the genetic background of Indian-Bangladeshi RVC, whereas the remaining seven Japanese RVCs carried the typical genetic background of Far East Asian RVC. This is the first report of the Indian-Bangladeshi RVC in Japan. With that observation and the reassortment event of human RVCs in Hungary, our study indicates that the RVCs are spreading from one region to another.
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http://dx.doi.org/10.1016/j.meegid.2016.03.027DOI Listing
July 2016

[A Fatal Case of Severe Fever with Thrombocytopenia Syndrome Complicated by Hemophagocytic Lymphohistiocytosis].

Kansenshogaku Zasshi 2015 Sep;89(5):592-6

Severe fever with thrombocytopenia syndrome (SFTS) is a recently identified emerging viral infectious disease in China that is caused by a novel phlebovirus in the family Bunyaviridae, SFTS virus, with an average case fatality rate of 12-30%. A cytokine storm with abnormally expressed cytokine profiles is associated with the disease severity. Hemophagocytic lymphohistiocytosis (HLH) is an aggressive and lifethreatening syndrome associated with excessive immune activation. We report herein on a fatal case of SFTS complicated by HLH. Consecutive plasma exchange and immunomodulatory therapy was ineffective in our case. The pathognomonic histological feature was necrotizing lymphadenitis with massive hemophagocytosis of systemic lymphoid tissues with SFTS viruses and SFTS-RNA copies. No specific treatment of SFTS is available, and an effective treatment strategy for patients with rapidly progressing SFTS has not been established. Appropriate immunomodulatory therapy is necessary for SFTS patients complicated by HLH.
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http://dx.doi.org/10.11150/kansenshogakuzasshi.89.592DOI Listing
September 2015

Molecular Evolution of the Capsid Gene in Norovirus Genogroup I.

Sci Rep 2015 Sep 4;5:13806. Epub 2015 Sep 4.

Department of Molecular Biodefence Research, Yokohama City University Graduate School of Medicine, Yokohama-shi, Kanagawa 236-0004, Japan.

We studied the molecular evolution of the capsid gene in all genotypes (genotypes 1-9) of human norovirus (NoV) genogroup I. The evolutionary time scale and rate were estimated by the Bayesian Markov chain Monte Carlo (MCMC) method. We also performed selective pressure analysis and B-cell linear epitope prediction in the deduced NoV GI capsid protein. Furthermore, we analysed the effective population size of the virus using Bayesian skyline plot (BSP) analysis. A phylogenetic tree by MCMC showed that NoV GI diverged from the common ancestor of NoV GII, GIII, and GIV approximately 2,800 years ago with rapid evolution (about 10(-3) substitutions/site/year). Some positive selection sites and over 400 negative selection sites were estimated in the deduced capsid protein. Many epitopes were estimated in the deduced virus capsid proteins. An epitope of GI.1 may be associated with histo-blood group antigen binding sites (Ser377, Pro378, and Ser380). Moreover, BSP suggested that the adaptation of NoV GI strains to humans was affected by natural selection. The results suggested that NoV GI strains evolved rapidly and date back to many years ago. Additionally, the virus may have undergone locally affected natural selection in the host resulting in its adaptation to humans.
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http://dx.doi.org/10.1038/srep13806DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4559769PMC
September 2015

[Current status of bacteriological studies at prefectural and municipal public health institutes in Japan].

Nihon Saikingaku Zasshi 2015 ;70(2):309-18

Ehime Prefectural Institute of Public Health and Environmental Science.

Prefectural and municipal public health institutes are located in prefectures and ordinance-designated cities in Japan, and play a vital role in the regional surveillance of infectious diseases and foodborne illnesses. These institutes, in close cooperation with national institutes such as the National Institute of Infectious Diseases and the National Institute of Health Sciences, construct the national surveillance network for infectious diseases and their causative agents. Bacteriological examinations and studies on a variety of infectious diseases and foodborne illnesses are core activities of prefectural and municipal public health institutes, through which novel and important bacteriological findings have been acquired. In this article, we report the latest findings regarding bacteriological examinations/studies and interesting cases at these institutes, especially concerning foodborne illnesses, tuberculosis, and antimicrobial resistances.
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http://dx.doi.org/10.3412/jsb.70.309DOI Listing
August 2016

Listeria arpJ gene modifies T helper type 2 subset differentiation.

J Infect Dis 2015 Jul 13;212(2):223-33. Epub 2015 Jan 13.

Department of Immunology and Host Defenses, Ehime University Graduate School of Medicine, Toon, Japan.

Background: Although the T-cell subset differentiation pathway has been characterized extensively from the view of host gene regulation, the effects of genes of the pathogen on T-cell subset differentiation during infection have yet to be elucidated. Especially, the bacterial genes that are responsible for this shift have not yet been determined.

Methods: Utilizing a single-gene-mutation Listeria panel, we investigated genes involved in the host-pathogen interaction that are required for the initiation of T-cell subset differentiation in the early phase of pathogen infection.

Results: We demonstrate that the induction of T helper types 1 and 2 (Th1 and Th2) subsets are separate phenomena and are mediated by distinct Listeria genes. We identified several candidate Listeria genes that appear to be involved in the host-Listeria interaction. Among them, arpJ is the strongest candidate gene for inhibiting Th2 subset induction. Furthermore, the analysis utilizing arpJ-deficient Listeria monocytogenes (Lm) revealed that the tumor necrosis factor (TNF) superfamily (Tnfsf) 9-TNF receptor superfamily (Tnfrsf) 9 interaction inhibits the Th2 response during Lm infection.

Conclusions: arpJ is the candidate gene for inhibiting Th2 T-cell subset induction. The arpJ gene product influences the expression of Tnfsf/Tnfrsf on antigen-presenting cells and inhibits the Th2 T-cell subset differentiation during Listeria infection.
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http://dx.doi.org/10.1093/infdis/jiv021DOI Listing
July 2015

Genotypic analysis of Pseudomonas aeruginosa isolated from ocular infection.

J Infect Chemother 2014 Jul 18;20(7):407-11. Epub 2014 Apr 18.

Department of Ophthalmology, Ehime University, Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.

Pseudomonas aeruginosa is the causative pathogen of keratitis, conjunctivitis, and dacryocystitis. However little is known about their clinical epidemiology in Japan. In this study we investigated the genotypic characterization and serotype of P. aeruginosa isolates from ocular infections. Thirty-four clinical P. aeruginosa isolates were characterized according to infection type, the type III secretion system (TTSS), serotype, and multilocus sequence typing (MLST). We divided the isolates into four clinical infection types as follows: Contact lens (CL)-related keratitis (CL-keratitis; 15 isolates), non CL-related keratitis (non CL-keratitis; 8 isolates), conjunctivitis (7 isolates), and dacryocystitis (4 isolates). Regarding the TTSS classification and serotyping classification, no significant differences were found among the infection types. Two clusters (I, II) and three subclusters (A, B, C) were classified according to MLST. CL-keratitis isolates with exoU positivity were clustered in II-B, and conjunctivitis was clustered in cluster I. Some linkage was found between the genetic background and CL-keratitis or conjunctivitis.
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http://dx.doi.org/10.1016/j.jiac.2014.02.007DOI Listing
July 2014

Characterization of the Escherichia coli O157:H7 outbreak strain whose Shiga toxin 2 gene is inactivated by IS1203v insertion.

Jpn J Infect Dis 2013 ;66(3):201-6

Ehime Prefectural Institute of Public Health and Environmental Science, Ehime, Japan.

A total of 12 enterohemorrhagic Escherichia coli (EHEC) O157:H7 strains were isolated during a recent outbreak in a nursery school in Ehime Prefecture, Japan. These isolates were considered to be derived from a common strain when analyzed using an IS-printing method and pulsed-field gel electrophoresis. PCR analysis revealed that the isolates harbor stx1, stx2, eae, and hlyA. However, assessment of the production of the Stx proteins revealed that these isolates produced Stx1 but not Stx2. We determined their stx2 variants such as stx2c and found that the size of the PCR product was much larger than the expected size. Sequencing of the entire stx2 gene revealed that a 1310-bp fragment was inserted into the coding region of the Stx2A subunit and that the sequences of the insert were identical to those of IS1203v. According to the sequences around the insertion site, additional amino acid residues should be attached at the C-terminus of the A subunit, which may hamper the Stx2 complex formation. Finally, this study also suggested that such an insertion may lead to the misinterpretation of results when screening EHEC isolates for virulence genes by PCR.
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http://dx.doi.org/10.7883/yoken.66.201DOI Listing
December 2013

Different Ca²⁺-sensitivities between the EF-hands of T- and L-plastins.

Biochem Biophys Res Commun 2012 Dec 7;429(3-4):137-41. Epub 2012 Nov 7.

Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan.

Plastins are Ca(2+)-regulated actin-bundling proteins, and essential for developing and stabilizing actin cytoskeletons. T-plastin is expressed in epithelial and mesenchymal cells of solid tissues, whereas L-plastin is expressed in mobile cells such as hemopoietic cell lineages and cancer cells. Using various spectroscopic methods, gel-filtration chromatography, and isothermal titration calorimetry, we here demonstrate that the EF-hand motifs of both T- and L-plastin change their structures in response to Ca(2+), but the sensitivity to Ca(2+) is lower in T-plastin than in L-plastin. These results suggest that T-plastin is suitable for maintaining static cytoskeletons, whereas L-plastin is suitable for dynamic rearrangement of cytoskeletons.
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http://dx.doi.org/10.1016/j.bbrc.2012.10.126DOI Listing
December 2012

Identification and characterization of D-hydroxyproline dehydrogenase and Delta1-pyrroline-4-hydroxy-2-carboxylate deaminase involved in novel L-hydroxyproline metabolism of bacteria: metabolic convergent evolution.

J Biol Chem 2012 Sep 25;287(39):32674-88. Epub 2012 Jul 25.

Faculty of Agriculture, Ehime University, 3-5-7 Tarumi, Matsuyama, Ehime 790-8566, Japan.

L-hydroxyproline (4-hydroxyproline) mainly exists in collagen, and most bacteria cannot metabolize this hydroxyamino acid. Pseudomonas putida and Pseudomonas aeruginosa convert L-hydroxyproline to α-ketoglutarate via four hypothetical enzymatic steps different from known mammalian pathways, but the molecular background is rather unclear. Here, we identified and characterized for the first time two novel enzymes, D-hydroxyproline dehydrogenase and Δ(1)-pyrroline-4-hydroxy-2-carboxylate (Pyr4H2C) deaminase, involved in this hypothetical pathway. These genes were clustered together with genes encoding other catalytic enzymes on the bacterial genomes. D-hydroxyproline dehydrogenases from P. putida and P. aeruginosa were completely different from known bacterial proline dehydrogenases and showed similar high specificity for substrate (D-hydroxyproline) and some artificial electron acceptor(s). On the other hand, the former is a homomeric enzyme only containing FAD as a prosthetic group, whereas the latter is a novel heterododecameric structure consisting of three different subunits (α(4)β(4)γ(4)), and two FADs, FMN, and [2Fe-2S] iron-sulfur cluster were contained in αβγ of the heterotrimeric unit. These results suggested that the L-hydroxyproline pathway clearly evolved convergently in P. putida and P. aeruginosa. Pyr4H2C deaminase is a unique member of the dihydrodipicolinate synthase/N-acetylneuraminate lyase protein family, and its activity was competitively inhibited by pyruvate, a common substrate for other dihydrodipicolinate synthase/N-acetylneuraminate lyase proteins. Furthermore, disruption of Pyr4H2C deaminase genes led to loss of growth on L-hydroxyproline (as well as D-hydroxyproline) but not L- and D-proline, indicating that this pathway is related only to L-hydroxyproline degradation, which is not linked to proline metabolism.
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http://dx.doi.org/10.1074/jbc.M112.374272DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3463351PMC
September 2012

Plastin family of actin-bundling proteins: its functions in leukocytes, neurons, intestines, and cancer.

Authors:
Hiroto Shinomiya

Int J Cell Biol 2012 4;2012:213492. Epub 2012 Jan 4.

Department of Immunology and Host Defenses, Ehime University Graduate School of Medicine, Toon, Ehime 791-0295, Japan.

Sophisticated regulation of the actin cytoskeleton by a variety of actin-binding proteins is essential for eukaryotic cells to perform their diverse functions. The plastin (also know, as fimbrin) protein family belongs to actin-bundling proteins, and the protein family is evolutionarily conserved and expressed in yeast, plant, and animal cells. Plastins are characterized by EF-hand Ca(2+)-binding domains and actin-binding domains and can cross-link actin filaments into higher-order assemblies like bundles. Three isoforms have been identified in mammals. T-plastin is expressed in cells from solid tissues, such as neurons in the brain. I-plastin expression is restricted to intestine and kidney; the isoform plays a vital role in the function of absorptive epithelia in these organs. L-plastin is expressed in hematopoietic cell lineages and in many types of cancer cells; the isoform is thus considered to be a useful biomarker for cancer.
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http://dx.doi.org/10.1155/2012/213492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3259490PMC
August 2012

Interaction of activated Rab5 with actin-bundling proteins, L- and T-plastin and its relevance to endocytic functions in mammalian cells.

Biochem Biophys Res Commun 2011 Apr 21;407(3):615-9. Epub 2011 Mar 21.

Department of Agricultural Chemistry, Graduate School of Agriculture, Tokyo University of Agriculture, 1-1-1 Sakuragaoka, Setagaya-ku, Tokyo 156-8502, Japan.

Rab5 is a GTP-binding protein that is crucial for endocytic machinery functions. We previously identified L-plastin as a binding protein for Rab5, using an affinity column with constitutively active Rab5. L- and T-plastin are isoforms of a plastin protein family belonging to actin-bundling proteins that are implicated in the regulation of cell morphology, lamellipodium protrusion, bacterial invasion and tumor progression. However, the physiological relevance of Rab5 binding to plastin has remained unclear. Here, we show that L- and T-plastin interacted only with activated Rab5 and that they co-localized with Rab5 on the plasma membrane and endosome. Rab5 activity was also higher in both L- and T-plastin over-expressing Cos-1 cells. Furthermore, expression of L- and T-plastin increased the rate of fluid-phase endocytosis. These findings imply that the Rab5 is either activated or the activity is sustained by interaction with plastin, and that this interaction influences endocytic activity.
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http://dx.doi.org/10.1016/j.bbrc.2011.03.082DOI Listing
April 2011

Differences of genetic diversity and antibiotics susceptibility of Pseudomonas aeruginosa isolated from hospital, river and coastal seawater.

Environ Microbiol Rep 2010 Jun;2(3):465-72

Dokkyo Medical University, School of Medicine, Mibu, Tochigi 321-0293, Japan. Center for Marine Environmental Studies, Ehime University, Matsuyama, Ehime 790-8577, Japan. Department of Immunology and Host Defenses, Ehime University Graduate School of Medicine, Toon, Ehime 791-0295, Japan. Division of Medical Technology, Ehime University Hospital, Toon, Ehime 791-0295, Japan.

Pseudomonas aeruginosa is an opportunistic pathogen, and ubiquitously found in natural environments. However, details on difference between clinical and environmental isolates have not been reported enough. In this study, we defined existence of marine specific genogroup and different drug susceptibility among isolates from clinical, river and coastal seawaters. Pseudomonas aeruginosa were isolated by using cetrimide kanamycin nalidixic acid agar media and incubation at 42°C, which was specific selection method of this bacterium from the natural aquatic samples. Pulse field gel electrophoresis analysis showed that the levels of genetic variation within P. aeruginosa were different among environmental sites. Pulse field gel electrophoresis also showed a lower diversity within P. aeruginosa in the coastal waters; and coastal strains isolated different sampling points were positioned closely in the same cluster. Most of the aquatic isolates were sensitive to most of the drugs tested and 'intermediate' to panipenem on the contrast to the clinical isolates, suggesting that the clinical use of antibiotics affect significantly to the emergence of the drug-resistant P. aeruginosa.
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http://dx.doi.org/10.1111/j.1758-2229.2010.00178.xDOI Listing
June 2010

Conformational analysis of the leukocyte-specific EF-hand protein p65/L-plastin by X-ray scattering in solution.

Biophys Chem 2007 Dec 11;131(1-3):36-42. Epub 2007 Sep 11.

Department of Immunology and Host Defenses, Graduate School of Medicine of Ehime University, Ehime, Japan.

p65/L-Plastin is a leukocyte-specific EF-hand protein which plays a vital role in organizing the actin cytoskeleton. Since its overall structural information has been largely unknown, we employed the X-ray scattering technique to elucidate the structure. Kratky plots of p65/L-plastin showed one peak, indicating that the protein takes compact globular conformations. The radii of gyration (Rg) of the monomer p65/L-plastin estimated from Guinier plots were 27.5 +/- 0.5 A and 28.6 A in the absence and presence of Ca(2+), respectively. The distance distribution function P(r) gave single peaks at 31.5-32.3 A and 33 A in the absence and presence of Ca(2+), respectively. These indicate that p65/L-plastin becomes somewhat larger in the presence of Ca(2+). The molecular shape of p65/L-plastin reconstructed from X-ray scattering data using the DAMMIN program has provided the first view of the overall structure of full-length plastin/fimbrin family proteins: a compact horseshoe-like shape with a small projection, which also exhibits Ca(2+) -induced conformational changes.
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http://dx.doi.org/10.1016/j.bpc.2007.09.001DOI Listing
December 2007

Origins and unconventional behavior of neutrophils in developing zebrafish.

Blood 2008 Jan 17;111(1):132-41. Epub 2007 Sep 17.

Unité Macrophages et Développement de l'Immunité, Centre National de la Recherche Scientifique-Unité de Recherche Associée 2578, Institut Pasteur, Paris, France.

The first leukocytes that arise in the development of vertebrate embryos are the primitive macrophages, which differentiate in the yolk sac and then quickly invade embryonic tissues. These macrophages have been considered to constitute a separate lineage, giving rise to no other cell type. Using an in vivo photoactivatable cell tracer in the transparent zebrafish (Danio rerio) embryo, we demonstrated that this lineage also gave rise to an equal or higher number of neutrophilic granulocytes. We were surprised to find that the differentiation of these primitive neutrophils occurs only after primitive myeloid progenitors have dispersed in the tissues. By 2 days after fertilization, these neutrophils have become the major leukocyte type found wandering in the epidermis and mesenchyme. Like the primitive macrophages, all primitive and larval neutrophils express PU.1 and L-plastin and they are highly attracted to local infections, yet only a small fraction of them phagocytose microbes, and to a much lesser extent per cell than the macrophages. They are also attracted to variously stressed or malformed tissues, suggesting a wider role than antimicrobial defense.
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http://dx.doi.org/10.1182/blood-2007-06-095398DOI Listing
January 2008