Journal of Microbiology

Journal Articles

Repositioning of a mucolytic drug to a selective antibacterial against Vibrio cholerae: In-Young Chung† , Bi-o Kim† , Hye-Jeong Jang† , You-Hee Cho; J. Microbiol. 2020;58(1):61-66. Published online January 2, 2020; DOI: https://doi.org/10.1007/s12275-020-9590-9

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Drug repositioning, the approach to explore existing drugs for use in new therapeutic indications, has emerged as an alternative drug development strategy. In this study, we found that a mucolytic drug, N-acetylcysteine (NAC) showed antibacterial activity against Vibrio cholerae. NAC can provide acid stress that selectively inhibited the growth of V. cholerae among other bacterial pathogens. To address the antibacterial mechanism of NAC against V. cholerae, six acr (acetylcysteine- resistant) mutants were isolated from 3,118 random transposon insertion clones. The transposon insertion sites of the six mutants were mapped at the five genes. All these mutants did not display NAC resistance under acidic conditions, despite their resistance to NAC under alkaline conditions, indicating that the NAC resistance directed by the acr mutations was independent of the unusual pH-sensitivity of V. cholerae. Furthermore, all these mutants displayed attenuated virulence and reduced biofilm formation, suggesting that the acr genes are required for pathogenesis of V. cholerae. This study validates the relevance of drug repositioning for antibacterials with new modes of action and will provide an insight into a novel antibacterial therapy for V. cholerae infections to minimize side effects and resistance emergence.

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Identification of brevinin-1EMa-derived stapled peptides as broad-spectrum virus entry blockers
Mi Il Kim, Thanh K. Pham, Dahee Kim, Minkyung Park, Bi-o Kim, You-Hee Cho, Young-Woo Kim, Choongho Lee
Virology.2021; 561: 6. CrossRef

Guinea pig complement potently measures vibriocidal activity of human antibodies in response to cholera vaccines: Kyoung Whun Kim , Soyoung Jeong , Ki Bum Ahn , Jae Seung Yang , Cheol-Heui Yun , Seung Hyun Han; J. Microbiol. 2017;55(12):973-978. Published online December 7, 2017; DOI: https://doi.org/10.1007/s12275-017-7478-0

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Abstract

The vibriocidal assay using guinea pig complement is widely used for the evaluation of immune responses to cholera vaccines in human clinical trials. However, it is unclear why guinea pig complement has been used over human complement in the measurement of vibriocidal activity of human sera and there have not been comparison studies for the use of guinea pig complement over those from other species. Therefore, we comparatively investigated the effects of complements derived from human, guinea pig, rabbit, and sheep on vibriocidal activity. Complements from guinea pig, rabbit, and human showed concentration-dependent vibriocidal activity in the presence of quality control serum antibodies. Of these complements, guinea pig complement was the most sensitive and effective over a wide concentration range. When the vibriocidal activity of complements was measured in the absence of serum antibodies, human, sheep, and guinea pig complements showed vibriocidal activity up to 40-fold, 20- fold, and 1-fold dilution, respectively. For human pre- and post-vaccination sera, the most potent vibriocidal activity was observed when guinea pig complement was used. In addition, the highest fold-increases between pre- and post- vaccinated sera were obtained with guinea pig complement. Furthermore, human complement contained a higher amount of V. cholerae- and its lipopolysaccharide-specific antibodies than guinea pig complement. Collectively, these results suggest that guinea pig complements are suitable for vibriocidal assays due to their high sensitivity and effectiveness to human sera.

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Immunogenicity and protective efficacy of a live, oral cholera vaccine formulation stored outside-the-cold-chain for 140 days
Tew Hui Xian, Kurunathan Sinniah, Chan Yean Yean, Venkateskumar Krishnamoorthy, Mohd Baidi Bahari, Manickam Ravichandran, Guruswamy Prabhakaran
BMC Immunology.2020;[Epub] CrossRef
A high-throughput, bead-based, antigen-specific assay to assess the ability of antibodies to induce complement activation
Stephanie Fischinger, Jonathan K. Fallon, Ashlin R. Michell, Thomas Broge, Todd J. Suscovich, Hendrik Streeck, Galit Alter
Journal of Immunological Methods.2019; 473: 112630. CrossRef
Characterization of antibody response in patients with acute and chronic chikungunya virus disease
Fatih Anfasa, Stephanie M. Lim, Susan Fekken, Robert Wever, Albert D.M.E. Osterhaus, Byron E.E. Martina
Journal of Clinical Virology.2019; 117: 68. CrossRef

Research Support, Non-U.S. Gov'ts

Crystal structure of the bacterial type VI secretion system component TssL from Vibrio cholerae: Jeong Ho Chang , Yeon-Gil Kim; J. Microbiol. 2015;53(1):32-37. Published online December 4, 2014; DOI: https://doi.org/10.1007/s12275-015-4539-0

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Abstract

The type VI secretion system (T6SS), commonly found in Gram-negative bacteria, is responsible for exporting effector proteins. The T6SS has been reported to be cytotoxic to host cells. While the components and assembly of the T6SS complex have been largely assessed, structural data on T6SS components from virulent bacteria is remarkably insufficient. Here, we report the crystal structure of Vibrio cholerae TssL (VcTssL), a core component of T6SS. In spite of a relatively low sequence identity, the overall structure of VcTssL is largely similar to those from other bacterial homologs except for several differences found in local structural elements. A unique feature attributed to the C-terminal fragment of Vc- TssL is a crystallographic artifact. This incidental feature of VcTssL may provide insights into screening of molecular partners for the cytoplasmic domain of TssL. Additionally, our results may help in the design of molecular probes for a detailed understanding of the functional relationship between TssL and other T6SS components.

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Structural Characterization of TssL from Acinetobacter baumannii: a Key Component of the Type VI Secretion System
Federico M. Ruiz, Juvenal Lopez, C. Gastón Ferrara, Elena Santillana, Yanis R. Espinosa, Mario F. Feldman, Antonio Romero, Ann M. Stock
Journal of Bacteriology.2020;[Epub] CrossRef
In situ and high‐resolution cryo‐ EM structure of a bacterial type VI secretion system membrane complex
Chiara Rapisarda, Yassine Cherrak, Romain Kooger, Victoria Schmidt, Riccardo Pellarin, Laureen Logger, Eric Cascales, Martin Pilhofer, Eric Durand, Rémi Fronzes
The EMBO Journal.2019;[Epub] CrossRef
Crystal Structure of the Type VI Secretion System Accessory Protein TagF from Pseudomonas Aeruginosa
Chang-Kyu Ok, Jeong Ho Chang
Protein & Peptide Letters.2019; 26(3): 204. CrossRef
Structure and Activity of the Type VI Secretion System
Yassine Cherrak, Nicolas Flaugnatti, Eric Durand, Laure Journet, Eric Cascales, Maria Sandkvist, Peter J. Christie
Microbiology Spectrum.2019;[Epub] CrossRef
Crystal structure of the periplasmic domain of TssL, a key membrane component of Type VI secretion system
Xiangbei Wang, Bo Sun, Mengxue Xu, Shenshen Qiu, Dongqing Xu, Tingting Ran, Jianhua He, Weiwu Wang
International Journal of Biological Macromolecules.2018; 120: 1474. CrossRef
Tryptophan-mediated Dimerization of the TssL Transmembrane Anchor Is Required for Type VI Secretion System Activity
Abdelrahim Zoued, Jean-Pierre Duneau, Eric Durand, Alexandre P. España, Laure Journet, Françoise Guerlesquin, Eric Cascales
Journal of Molecular Biology.2018; 430(7): 987. CrossRef
Structure–Function Analysis of the TssL Cytoplasmic Domain Reveals a New Interaction between the Type VI Secretion Baseplate and Membrane Complexes
Abdelrahim Zoued, Chloé J. Cassaro, Eric Durand, Badreddine Douzi, Alexandre P. España, Christian Cambillau, Laure Journet, Eric Cascales
Journal of Molecular Biology.2016; 428(22): 4413. CrossRef
Aim, Load, Fire: The Type VI Secretion System, a Bacterial Nanoweapon
Francesca R. Cianfanelli, Laura Monlezun, Sarah J. Coulthurst
Trends in Microbiology.2016; 24(1): 51. CrossRef
Biogenesis and structure of a type VI secretion membrane core complex
Eric Durand, Van Son Nguyen, Abdelrahim Zoued, Laureen Logger, Gérard Péhau-Arnaudet, Marie-Stéphanie Aschtgen, Silvia Spinelli, Aline Desmyter, Benjamin Bardiaux, Annick Dujeancourt, Alain Roussel, Christian Cambillau, Eric Cascales, Rémi Fronzes
Nature.2015; 523(7562): 555. CrossRef
Type VI secretion system: secretion by a contractile nanomachine
Marek Basler
Philosophical Transactions of the Royal Society B: Biological Sciences.2015; 370(1679): 20150021. CrossRef

Structural and Functional Importance of Outer Membrane Proteins in Vibrio cholerae Flagellum: Wasimul Bari , Kang-Mu Lee , Sang Sun Yoon; J. Microbiol. 2012;50(4):631-637. Published online August 25, 2012; DOI: https://doi.org/10.1007/s12275-012-2116-3

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Abstract: Vibrio cholerae has a sheath-covered monotrichous flagellum that is known to contribute to virulence. Although the structural organization of the V. cholerae flagellum has been extensively studied, the involvement of outer membrane proteins as integral components in the flagellum still remains elusive. Here we show that flagella produced by V. cholerae O1 El Tor strain C6706 were two times thicker than those from two other Gram-negative bacteria. A C6706 mutant strain (SSY11) devoid of two outer membrane proteins (OMPs), OmpU and OmpT, produced thinner flagella. SSY11 showed significant defects in the flagella-mediated motility as compared to its parental strain. Moreover, increased shedding of the flagella-associated proteins was observed in the culture supernatant of SSY11. This finding was also supported by the observation that culture supernatants of the SSY11 strain induced the production of a significantly higher level of IL-8 in human colon carcinoma HT29 and alveolar epithelial A549 cells than those of the wild-type C6706 strain. These results further suggest a definite role of these two OMPs in providing the structural integrity of the V. cholerae flagellum as part of the surrounding sheath.

Molecular Characterization Reveals Involvement of Altered El Tor Biotype Vibrio cholerae O1 Strains in Cholera Outbreak at Hyderabad, India: Ajay Kumar Goel , Meenu Jain , Pramod Kumar , Pennagaram Sarguna , Meera Bai , Neha Ghosh , Natrajan Gopalan; J. Microbiol. 2011;49(2):280-284. Published online May 3, 2011; DOI: https://doi.org/10.1007/s12275-011-0317-9

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Abstract: Thirty-four Vibrio cholerae isolates collected from a cholera outbreak in Hyderabad, South India were found to belong to serogroup O1 biotype El Tor serotype Ogawa. The genotype of all the isolates was confirmed by PCR assays. All the isolates were found PCR positive for ctxAB, ompW, rfbO1, rtxC, and tcpA genes. All the isolates but one harboured rstREl Tor allele. However, one isolate carried both rstREl Tor as well as rstRClassical alleles. Cholera toxin (ctxB) genotyping of the isolates confirmed the presence of altered cholera toxin B of classical biotype in all the isolates. All the isolates except VCH35 harboured an RS1-CTX prophage array on the large chromosome. The isolate VCH35 contained a tandem repeat of classical CTX prophage on the small chromosome. The clonal relationship among the V. cholerae isolates as carried out by enterobacterial repetitive intergenic consensus sequences PCR, BOX PCR and randomly amplified polymorphic DNA, uniformly showed a genetic relationship among the outbreak isolates. The results of this study suggest that altered El Tor biotype V. cholerae with the classical cholera toxin gene are involved in cholera outbreaks in India.

Journal Article

Molecular Characterization of Vibrio cholerae Isolates from Cholera Outbreaks in North India: Joseph J. Kingston , Kuruvilla Zachariah , Urmil Tuteja , Sanjay Kumar , Harsh Vardhan Batra; J. Microbiol. 2009;47(1):110-115. Published online February 20, 2009; DOI: https://doi.org/10.1007/s12275-008-0162-7

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Abstract: Vibrio cholerae isolates recovered from cholera outbreaks in Bhind district of Madhya Pradesh and Delhi, Northern India were characterized. The O1 serogroup isolates from Bhind outbreak were of Inaba serotype whereas both Ogawa and Inaba serotypes were recovered from Delhi. PCR analysis revealed that only O1 serogroup V. cholerae isolates carried the virulence-associated genes like ctxA, tcpA, ace, and zot. Molecular typing by repetitive sequence based ERIC, VCR1, and VC1 PCR’s revealed similar DNA profile for both Inaba and Ogawa serotypes. A discrete VC1-PCR band identified among the El Tor strains had greater similarity (>97%) to the V. cholerae genome sequence and therefore has the potential to be used as a marker for the identification of the V. cholerae strains. Non-O1 strains recovered from Bhind region differed among themselves as well as from that of the O1 isolates. All the O1 serogroup isolates possessed SXT element and were uniformly resistant to the antibiotics nalidixic acid, polymyxin-B, furazolidone, cloxacilin, trimethoprim-sulfamethaxazole, and vibriostatic agent O129. Inaba strains from both Delhi and Bhind differed from Ogawa strains by their resistance to streptomycin despite sharing similar DNA patterns in all the three rep-PCRs. Though Delhi and Bhind are separate geographical regions in Northern India, Inaba strains from both these places appear to be closely related owing to their similarity in antibiogram and genetic profile.

Research Support, Non-U.S. Gov't

pVC, a Small Cryptic Plasmid from the Environmental Isolate of Vibrio cholerae MP-1: Ruifu Zhang , Yanling Wang , Pak Chow Leung , Ji-Dong Gu; J. Microbiol. 2007;45(3):193-198.; DOI: https://doi.org/2543 [pii]

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Abstract: A marine bacterium was isolated from Mai Po Nature Reserve of Hong Kong and identified as Vibrio cholerae MP-1. It contains a small plasmid designated as pVC of 3.8 kb. Four open reading frames (ORFs) are identified on the plasmid, but none of them shows homology to any known protein. Database search indicated that a 440 bp fragment is 96% identical to a fragment found in a small plasmid of another V. cholerae. Further experiments demonstrated that a 2.3 kb EcoRI fragment containing the complete ORF1, partial ORF4 and their intergenic region could self-replicate. Additional analyses revealed that sequence upstream of ORF1 showed the features characteristic of theta type replicons. Protein encoded by ORF1 has two characteristic motifs existed in most replication initiator proteins (Rep): the leucine zipper (LZ) motif located at the N-terminal region and the alpha helix-turn-alpha helix motif (HTH) located at the C-terminal end. The results suggest that pVC replicates via the theta type mechanism and is likely a novel type of theta replicon.

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