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  1. Article ; Online: Genetic incorporation of non-canonical amino acid photocrosslinkers in Neisseria meningitidis

    Hideyuki Takahashi / Naoshi Dohmae / Kwang Sik Kim / Ken Shimuta / Makoto Ohnishi / Shigeyuki Yokoyama / Tatsuo Yanagisawa

    PLoS ONE, Vol 15, Iss 8, p e

    New method provides insights into the physiological function of the function-unknown NMB1345 protein.

    2020  Volume 0237883

    Abstract: Although whole-genome sequencing has provided novel insights into Neisseria meningitidis, many open reading frames have only been annotated as hypothetical proteins with unknown biological functions. Our previous genetic analyses revealed that the ... ...

    Abstract Although whole-genome sequencing has provided novel insights into Neisseria meningitidis, many open reading frames have only been annotated as hypothetical proteins with unknown biological functions. Our previous genetic analyses revealed that the hypothetical protein, NMB1345, plays a crucial role in meningococcal infection in human brain microvascular endothelial cells; however, NMB1345 has no homology to any identified protein in databases and its physiological function could not be elucidated using pre-existing methods. Among the many biological technologies to examine transient protein-protein interaction in vivo, one of the developed methods is genetic code expansion with non-canonical amino acids (ncAAs) utilizing a pyrrolysyl-tRNA synthetase/tRNAPyl pair from Methanosarcina species: However, this method has never been applied to assign function-unknown proteins in pathogenic bacteria. In the present study, we developed a new method to genetically incorporate ncAAs-encoded photocrosslinking probes into N. meningitidis by utilizing a pyrrolysyl-tRNA synthetase/tRNAPyl pair and elucidated the biological function(s) of the NMB1345 protein. The results revealed that the NMB1345 protein directly interacts with PilE, a major component of meningococcal pili, and further physicochemical and genetic analyses showed that the interaction between the NMB1345 protein and PilE was important for both functional pilus formation and meningococcal infectious ability in N. meningitidis. The present study using this new methodology for N. meningitidis provides novel insights into meningococcal pathogenesis by assigning the function of a hypothetical protein.
    Keywords Medicine ; R ; Science ; Q
    Subject code 612
    Language English
    Publishing date 2020-01-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  2. Article ; Online: Urethritis Caused by Novel Neisseria meningitidis Serogroup W in Man Who Has Sex with Men, Japan

    Kayoko Hayakawa / Ichiro Itoda / Ken Shimuta / Hideyuki Takahashi / Makoto Ohnishi

    Emerging Infectious Diseases, Vol 20, Iss 9, Pp 1585-

    2014  Volume 1587

    Keywords Neisseria meningitidis ; urethritis ; serogroup W ; men who have sex with men ; MSM ; sequence type 11 ; Medicine ; R ; Infectious and parasitic diseases ; RC109-216
    Language English
    Publishing date 2014-09-01T00:00:00Z
    Publisher Centers for Disease Control and Prevention
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  3. Article ; Online: An attenuated Shigella mutant lacking the RNA-binding protein Hfq provides cross-protection against Shigella strains of broad serotype.

    Jiro Mitobe / Ritam Sinha / Soma Mitra / Dhrubajyoti Nag / Noriko Saito / Ken Shimuta / Nobuo Koizumi / Hemanta Koley

    PLoS Neglected Tropical Diseases, Vol 11, Iss 7, p e

    2017  Volume 0005728

    Abstract: Few live attenuated vaccines protect against multiple serotypes of bacterial pathogen because host serotype-specific immune responses are limited to the serotype present in the vaccine strain. Here, immunization with a mutant of Shigella flexneri 2a ... ...

    Abstract Few live attenuated vaccines protect against multiple serotypes of bacterial pathogen because host serotype-specific immune responses are limited to the serotype present in the vaccine strain. Here, immunization with a mutant of Shigella flexneri 2a protected guinea pigs against subsequent infection by S. dysenteriae type 1 and S. sonnei strains. This deletion mutant lacked the RNA-binding protein Hfq leading to increased expression of the type III secretion system via loss of regulation, resulting in attenuation of cell viability through repression of stress response sigma factors. Such increased antigen production and simultaneous attenuation were expected to elicit protective immunity against Shigella strains of heterologous serotypes. Thus, the vaccine potential of this mutant was tested in two guinea pig models of shigellosis. Animals vaccinated in the left eye showed fewer symptoms upon subsequent challenge via the right eye, and even survived subsequent intestinal challenge. In addition, oral vaccination effectively induced production of immunoglobulins without severe side effects, again protecting all animals against subsequent intestinal challenge with S. dysenteriae type 1 or S. sonnei strains. Antibodies against common virulence proteins and the O-antigen of S. flexneri 2a were detected by immunofluorescence microscopy. Reaction of antibodies with various strains, including enteroinvasive Escherichia coli, suggested that common virulence proteins induced protective immunity against a range of serotypes. Therefore, vaccination is expected to cover not only the most prevalent serotypes of S. sonnei and S. flexneri 2a, but also various Shigella strains, including S. dysenteriae type 1, which produces Shiga toxin.
    Keywords Arctic medicine. Tropical medicine ; RC955-962 ; Public aspects of medicine ; RA1-1270
    Subject code 570
    Language English
    Publishing date 2017-07-01T00:00:00Z
    Publisher Public Library of Science (PLoS)
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  4. Article ; Online: Emergence and evolution of antimicrobial resistance genes and mutations in Neisseria gonorrhoeae

    Koji Yahara / Kevin C. Ma / Tatum D. Mortimer / Ken Shimuta / Shu-ichi Nakayama / Aki Hirabayashi / Masato Suzuki / Michio Jinnai / Hitomi Ohya / Toshiro Kuroki / Yuko Watanabe / Mitsuru Yasuda / Takashi Deguchi / Vegard Eldholm / Odile B. Harrison / Martin C. J. Maiden / Yonatan H. Grad / Makoto Ohnishi

    Genome Medicine, Vol 13, Iss 1, Pp 1-

    2021  Volume 12

    Abstract: Abstract Background Antimicrobial resistance in Neisseria gonorrhoeae is a global health concern. Strains from two internationally circulating sequence types, ST-7363 and ST-1901, have acquired resistance to third-generation cephalosporins, mainly due to ...

    Abstract Abstract Background Antimicrobial resistance in Neisseria gonorrhoeae is a global health concern. Strains from two internationally circulating sequence types, ST-7363 and ST-1901, have acquired resistance to third-generation cephalosporins, mainly due to mosaic penA alleles. These two STs were first detected in Japan; however, the timeline, mechanism, and process of emergence and spread of these mosaic penA alleles to other countries remain unknown. Methods We studied the evolution of penA alleles by obtaining the complete genomes from three Japanese ST-1901 clinical isolates harboring mosaic penA allele 34 (penA-34) dating from 2005 and generating a phylogenetic representation of 1075 strains sampled from 35 countries. We also sequenced the genomes of 103 Japanese ST-7363 N. gonorrhoeae isolates from 1996 to 2005 and reconstructed a phylogeny including 88 previously sequenced genomes. Results Based on an estimate of the time-of-emergence of ST-1901 (harboring mosaic penA-34) and ST-7363 (harboring mosaic penA-10), and > 300 additional genome sequences of Japanese strains representing multiple STs isolated in 1996–2015, we suggest that penA-34 in ST-1901 was generated from penA-10 via recombination with another Neisseria species, followed by recombination with a gonococcal strain harboring wildtype penA-1. Following the acquisition of penA-10 in ST-7363, a dominant sub-lineage rapidly acquired fluoroquinolone resistance mutations at GyrA 95 and ParC 87-88, by independent mutations rather than horizontal gene transfer. Data in the literature suggest that the emergence of these resistance determinants may reflect selection from the standard treatment regimens in Japan at that time. Conclusions Our findings highlight how antibiotic use and recombination across and within Neisseria species intersect in driving the emergence and spread of drug-resistant gonorrhea.
    Keywords Recombination ; Horizontal gene transfer ; Genomic epidemiology ; Antimicrobial resistance ; Phylogeny ; Surveillance ; Medicine ; R ; Genetics ; QH426-470
    Subject code 572
    Language English
    Publishing date 2021-03-01T00:00:00Z
    Publisher BMC
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  5. Article ; Online: Identifying antibiotics based on structural differences in the conserved allostery from mitochondrial heme-copper oxidases

    Yuya Nishida / Sachiko Yanagisawa / Rikuri Morita / Hideki Shigematsu / Kyoko Shinzawa-Itoh / Hitomi Yuki / Satoshi Ogasawara / Ken Shimuta / Takashi Iwamoto / Chisa Nakabayashi / Waka Matsumura / Hisakazu Kato / Chai Gopalasingam / Takemasa Nagao / Tasneem Qaqorh / Yusuke Takahashi / Satoru Yamazaki / Katsumasa Kamiya / Ryuhei Harada /
    Nobuhiro Mizuno / Hideyuki Takahashi / Yukihiro Akeda / Makoto Ohnishi / Yoshikazu Ishii / Takashi Kumasaka / Takeshi Murata / Kazumasa Muramoto / Takehiko Tosha / Yoshitsugu Shiro / Teruki Honma / Yasuteru Shigeta / Minoru Kubo / Seiji Takashima / Yasunori Shintani

    Nature Communications, Vol 13, Iss 1, Pp 1-

    2022  Volume 14

    Abstract: Antimicrobial resistance to currently available antibiotics requires innovation of antibiotics. Here, the authors identify a critical inhibitory site in an energy-producing enzyme, which can lead to rational design of antibiotics. ...

    Abstract Antimicrobial resistance to currently available antibiotics requires innovation of antibiotics. Here, the authors identify a critical inhibitory site in an energy-producing enzyme, which can lead to rational design of antibiotics.
    Keywords Science ; Q
    Language English
    Publishing date 2022-12-01T00:00:00Z
    Publisher Nature Portfolio
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  6. Article ; Online: Assessment of SARS-CoV-2 infectivity of upper respiratory specimens from COVID-19 patients by virus isolation using VeroE6/TMPRSS2 cells

    Tadaki Suzuki / Souichi Yamada / Shuetsu Fukushi / Hitomi Kinoshita / Makoto Ohnishi / Tsuguto Fujimoto / Masayuki Saijo / Ken Maeda / Nozomu Hanaoka / Naomi Nojiri / Ai Kawana-Tachikawa / Shigeru Kusagawa / Koichi Ishikawa / Shigeyoshi Harada / Saori Matsuoka / Tadashi Kikuchi / Sayuri Seki / Midori Nakamura-Hoshi / Shoji Miki /
    Lucky Ronald Runtuwene / Nobuo Koizumi / Sunao Iyoda / Hideyuki Takahashi / Hidemasa Izumiya / Jiro Mitobe / Shouji Yamamoto / Masatomo Morita / Ken-ichi Lee / Ken Shimuta / Kyoko Saito / Masayoshi Fukasawa / Yasutaka Hoshino / Ken Miyazawa / Minoru Nagi / Chikako Shimokawa / Yasuyuki Morishima / Takashi Sakudoh / Yoshihiro Kaku / Chang Kweng Lim / Shigeru Tajima / Takahiro Maeki / Eri Nakayama / Satoshi Taniguchi / Motohiko Ogawa / Takanobu Kato / Hussein Hassan Aly / Kousho Wakae / Kento Fukano

    BMJ Open Respiratory Research, Vol 8, Iss

    2021  Volume 1

    Abstract: Background An outbreak of novel coronavirus (SARS-CoV-2)-associated respiratory infectious diseases (COVID-19) emerged in 2019 and has spread rapidly in humans around the world. The demonstration of in vitro infectiousness of respiratory specimens is an ... ...

    Abstract Background An outbreak of novel coronavirus (SARS-CoV-2)-associated respiratory infectious diseases (COVID-19) emerged in 2019 and has spread rapidly in humans around the world. The demonstration of in vitro infectiousness of respiratory specimens is an informative surrogate for SARS-CoV-2 transmission from patients with COVID-19; accordingly, viral isolation assays in cell culture are an important aspect of laboratory diagnostics for COVID-19.Methods We developed a simple and rapid protocol for isolating SARS-CoV-2 from respiratory specimens using VeroE6/TMPRSS2 cells, a cell line that is highly susceptible to the virus. We also investigated a correlation between isolation of SARS-CoV-2 and viral load detected by real-time RT-PCR (rRT-PCR) using N2 primer/probe set that has been developed for testing of COVID-19 in Japan.Results The SARS-CoV-2 isolation protocol did not require blind passage of inoculated cells and yielded the results of viral isolation within 7 days after inoculation. Specimens with cycle threshold (Ct) values of <20.2, determined by rRT-PCR, were predicted to be isolation-positive. On the other hand, 6.9% of specimens with Ct values >35 were virus isolation-positive, indicating that low viral loads (high Ct values) in upper respiratory specimens do not always indicate no risk of containing transmissible virus.Conclusion In combination with rRT-PCR, the SARS-CoV-2 isolation protocol provides a means for assessing the potential risk of transmissible virus in upper respiratory specimens.
    Keywords Medicine ; R ; Diseases of the respiratory system ; RC705-779
    Subject code 610
    Language English
    Publishing date 2021-08-01T00:00:00Z
    Publisher BMJ Publishing Group
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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