Open access
21 June 2018

Single Circular Chromosome Identified from the Genome Sequence of the Vibrio cholerae O1 bv. El Tor Ogawa Strain V060002


We report here the complete genome sequence of the Vibrio cholerae O1 bv. El Tor Ogawa strain V060002, isolated in 1997. The data demonstrate that this clinical strain has a single chromosome resulting from recombination of two prototypical chromosomes.


Vibrio cholerae is a waterborne pathogen that causes the fatal diarrheal disease cholera. Of the more than 200 serogroups of V. cholerae, O1 and O139 are associated with epidemic and pandemic cholera and with the major virulence determinant cholera toxin (1, 2) produced by the filamentous bacteriophage CTXϕ (3). Serogroup O1 comprises two biotypes, classical and El Tor. The classical biotype caused the sixth and probably earlier cholera pandemics, whereas the El Tor biotype is responsible for the current seventh cholera pandemic (4).
The genome of V. cholerae is split into two circular chromosomes (chr1 and chr2) (5, 6), a feature common in the family Vibrionaceae (7, 8). However, recent genomic studies on V. cholerae isolates have revealed two non-O1/non-O139 strains, each with a single chromosome (911). It has also been reported that V. cholerae O1 strains with single chromosomes can be generated by genome engineering (12) or spontaneously isolated as suppressors of lethal mutations that disrupt the replication of chr2 (13, 14).
The sequenced O1 biovar El Tor Ogawa strain V060002 was isolated in 1997 from a patient who traveled to Indonesia, and the strain has been used in our laboratory as a model for studying regulatory mechanisms of chitin-induced natural transformation (1518). Genomic DNA was extracted with the DNeasy blood and tissue kit (Qiagen) following the manufacturer’s instructions. A 20-kbp library for P6-C4 chemistry was prepared using the RS II SMRTbell template preparation kit version 1.0 (PacBio) and sequenced with the P6 version 2 single-molecule real-time (SMRT) sequencing platform (PacBio). Sequencing reads were assembled de novo using the Hierarchical Genome Assembly Process version 3 (HGAP3) (19) with a mean sequence coverage of 196.55-fold. This assembly was corrected with the Quiver consensus algorithm to obtain a high-accuracy genome assembly (19). The contig was further corrected using Pilon version 1.22 (20), and paired-end short reads (300-mer × 2) were obtained from the MiSeq platform (Illumina).
The generated sequence assembly unexpectedly yielded a single circular chromosome with a genome size of 4,057,041 bp and a GC content of 47.5%. The size and number were verified by pulsed-field gel electrophoresis of the intact chromosome of strain V060002 (data not shown). Comparison of the genome sequences of V060002 and the O1 model strain N16961 (5) revealed that a single chromosome of V060002 was generated by recombination of highly homologous insertion sequence elements shared by chr1 and chr2 (99% identity, corresponding to vc1789 to vc1790 on chr1 and vca0791 to vca0792 on chr2 of N16961). It should be noted that these recombination sites are identical to those of a representative chromosome fusion spontaneously isolated from N16961 with a null mutation of the dam gene (14), which is essential for chr2 replication (21).
Annotation of the V060002 genome using the DDBJ Fast Annotation and Submission Tool (DFAST) (22) identified 3,560 coding sequences, 28 rRNA sequences, and 98 tRNA sequences. Strain V060002 also carried well-known gene clusters associated with pathogenesis (2326), as well as two copies of the CTXφ prophage.
More detailed genomic and phenotypic analyses of this naturally occurring V. cholerae O1 strain with a single chromosome will be presented in future publications.

Accession number(s).

The annotated chromosome has been deposited in DDBJ/GenBank under the accession number AP018677.


This work was supported by the Japan Society for the Promotion of Sciences (JSPS KAKENHI, number 16K08798).
We thank Yasunori Saito for assistance with pulsed-field gel electrophoresis and Yu Takizawa for providing help with bioinformatics analyses.


Singh DV, Matte MH, Matte GR, Jiang S, Sabeena F, Shukla BN, Sanyal SC, Huq A, Colwell RR. 2001. Molecular analysis of Vibrio cholerae O1, O139, non-O1, and non-O139 strains: clonal relationships between clinical and environmental isolates. Appl Environ Microbiol 67:910–921.
Kaper JB, Morris JG, Levine MM. 1995. Cholera. Clin Microbiol Rev 8:48–86.
Waldor MK, Mekalanos JJ. 1996. Lysogenic conversion by a filamentous phage encoding cholera toxin. Science 272:1910–1914.
Harris JB, LaRocque RC, Qadri F, Ryan ET, Calderwood SB. 2012. Cholera. Lancet 379:2466–2476.
Heidelberg JF, Eisen JA, Nelson WC, Clayton RA, Gwinn ML, Dodson RJ, Haft DH, Hickey EK, Peterson JD, Umayam L, Gill SR, Nelson KE, Read TD, Tettelin H, Richardson D, Ermolaeva MD, Vamathevan J, Bass S, Qin H, Dragoi I, Sellers P, McDonald L, Utterback T, Fleishmann RD, Nierman WC, White O, Salzberg SL, Smith HO, Colwell RR, Mekalanos JJ, Venter JC, Fraser CM. 2000. DNA sequence of both chromosomes of the cholera pathogen Vibrio cholerae. Nature 406:477–483.
Trucksis M, Michalski J, Deng YK, Kaper JB. 1998. The Vibrio cholerae genome contains two unique circular chromosomes. Proc Natl Acad Sci U S A 95:14464–14469.
Yamaichi Y, Iida T, Park K-S, Yamamoto K, Honda T. 1999. Physical and genetic map of the genome of Vibrio parahaemolyticus: presence of two chromosomes in Vibrio species. Mol Microbiol 31:1513–1521.
Okada K, Iida T, Kita-Tsukamoto K, Honda T. 2005. Vibrios commonly possess two chromosomes. J Bacteriol 187:752–757.
Johnson SL, Khiani A, Bishop-Lilly KA, Chapman C, Patel M, Verratti K, Teshima H, Munk AC, Bruce DC, Han CS, Xie G, Davenport KW, Chain P, Sozhamannan S. 2015. Complete genome assemblies for two single-chromosome Vibrio cholerae isolates, strains 1154-74 (serogroup O49) and 10432-62 (serogroup O27). Genome Announc 3(3):e00462-15.
Chapman C, Henry M, Bishop-Lilly KA, Awosika J, Briska A, Ptashkin RN, Wagner T, Rajanna C, Tsang H, Johnson SL, Mokashi VP, Chain PSG, Sozhamannan S. 2015. Scanning the landscape of genome architecture of non-O1 and non-O139 Vibrio cholerae by whole genome mapping reveals extensive population genetic diversity. PLoS One 10:e0120311.
Xie G, Johnson SL, Davenport KW, Rajavel M, Waldminghaus T, Detter JC, Chain PS, Sozhamannan S. 2017. Exception to the rule: genomic characterization of naturally occurring unusual Vibrio cholerae strains with a single chromosome. Int J Genomics 2017:8724304.
Val M-E, Skovgaard O, Ducos-Galand M, Bland MJ, Mazel D. 2012. Genome engineering in Vibrio cholerae: a feasible approach to address biological issues. PLoS Genet 8:e1002472.
Val M-E, Marbouty M, de Lemos Martins F, Kennedy SP, Kemble H, Bland MJ, Possoz C, Koszul R, Skovgaard O, Mazel D. 2016. A checkpoint control orchestrates the replication of the two chromosomes of Vibrio cholerae. Sci Adv 2:e1501914.
Val M-E, Kennedy SP, Soler-Bistué AJ, Barbe V, Bouchier C, Ducos-Galand M, Skovgaard O, Mazel D. 2014. Fuse or die: how to survive the loss of Dam in Vibrio cholerae. Mol Microbiol 91:665–678.
Yamamoto S, Morita M, Izumiya H, Watanabe H. 2010. Chitin disaccharide (GlcNAc)2 induces natural competence in Vibrio cholerae through transcriptional and translational activation of a positive regulatory gene tfoXVC. Gene 457:42–49.
Yamamoto S, Izumiya H, Mitobe J, Morita M, Arakawa E, Ohnishi M, Watanabe H. 2011. Identification of a chitin-induced small RNA that regulates translation of the tfoX gene, encoding a positive regulator of natural competence in Vibrio cholerae. J Bacteriol 193:1953–1965.
Yamamoto S, Mitobe J, Ishikawa T, Wai SN, Ohnishi M, Watanabe H, Izumiya H. 2014. Regulation of natural competence by the orphan two-component system sensor kinase ChiS involves a non-canonical transmembrane regulator in Vibrio cholerae. Mol Microbiol 91:326–347.
Yamamoto S, Ohnishi M. 2017. Glucose-specific enzyme IIA of the phosphoenolpyruvate: carbohydrate phosphotransferase system modulates chitin signaling pathways in Vibrio cholerae. J Bacteriol 199:e00127-17.
Chin C-S, Alexander DH, Marks P, Klammer AA, Drake J, Heiner C, Clum A, Copeland A, Huddleston J, Eichler EE, Turner SW, Korlach J. 2013. Nonhybrid, finished microbial genome assemblies from long-read SMRT sequencing data. Nat Methods 10:563–569.
Walker BJ, Abeel T, Shea T, Priest M, Abouelliel A, Sakthikumar S, Cuomo CA, Zeng Q, Wortman J, Young SK, Earl AM. 2014. Pilon: an integrated tool for comprehensive microbial variant detection and genome assembly improvement. PLoS One 9:e112963.
Demarre G, Chattoraj DK. 2010. DNA adenine methylation is required to replicate both Vibrio cholerae chromosomes once per cell cycle. PLoS Genet 6:e1000939.
Tanizawa Y, Fujisawa T, Nakamura Y. 2018. DFAST: a flexible prokaryotic genome annotation pipeline for faster genome publication. Bioinformatics 34:1037–1039.
Karaolis DKR, Johnson JA, Bailey CC, Boedeker EC, Kaper JB, Reeves PR. 1998. A Vibrio cholerae pathogenicity island associated with epidemic and pandemic strains. Proc Natl Acad Sci U S A 95:3134–3139.
Jermyn WS, Boyd EF. 2002. Characterization of a novel Vibrio pathogenicity island (VPI-2) encoding neuraminidase (nanH) among toxigenic Vibrio cholerae isolates. Microbiology 148:3681–3693.
Dziejman M, Balon E, Boyd D, Fraser CM, Heidelberg JF, Mekalanos JJ. 2002. Comparative genomic analysis of Vibrio cholerae: genes that correlate with cholera endemic and pandemic disease. Proc Natl Acad Sci U S A 99:1556–1561.
O’Shea YA, Finnan S, Reen FJ, Morrissey JP, O’Gara F, Boyd EF. 2004. The Vibrio seventh pandemic island-II is a 26.9 kb genomic island present in Vibrio cholerae El Tor and O139 serogroup isolates that shows homology to a 43.4 kb genomic island in V. vulnificus. Microbiology 150:4053–4063.

Information & Contributors


Published In

cover image Genome Announcements
Genome Announcements
Volume 6Number 2521 June 2018
eLocator: 10.1128/genomea.00564-18


Received: 17 May 2018
Accepted: 22 May 2018
Published online: 21 June 2018



Shouji Yamamoto
Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan
Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan
Masatomo Morita
Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan
Eiji Arakawa
Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan
Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan
Makoto Ohnishi
Department of Bacteriology I, National Institute of Infectious Diseases, Tokyo, Japan


Address correspondence to Shouji Yamamoto, [email protected].
S.Y. and K.-I.L. contributed equally to this work.

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