Evidence of a Dominant Lineage of Vibrio cholerae-Specific Lytic Bacteriophages Shed by Cholera Patients over a 10-Year Period in Dhaka, Bangladesh

Seed, Kimberley D.; Bodi, Kip; Kropinski, Andrew M.; Ackermann, Hans-Wolfgang; Calderwood, Stephen B.; Qadri, Firdausi; Camilli, Andrew

doi:10.1128/mbio.00334-10

Cited by 126 publications

(203 citation statements)

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“…The conversion of smooth2012Env-9 to rugose2012Env-9 phenotype, creation of in-frame vpsA mutation in 2012Env-9 smooth background, and complementation of vpsA mutant was performed5. Vibriophages infection assay (plaque assay) was performed as described previously to determine whether Vibriophages can infect 2012Env-9 and 2012Env-390, and a 2012Env-9 ΔvpsA mutant26.…”

Section: Methodsmentioning

confidence: 99%

Non-toxigenic environmental Vibrio cholerae O1 strain from Haiti provides evidence of pre-pandemic cholera in Hispaniola

Azarian

Ali

Johnson

et al. 2016

Sci Rep

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Vibrio cholerae is ubiquitous in aquatic environments, with environmental toxigenic V. cholerae O1 strains serving as a source for recurrent cholera epidemics and pandemic disease. However, a number of questions remain about long-term survival and evolution of V. cholerae strains within these aquatic environmental reservoirs. Through monitoring of the Haitian aquatic environment following the 2010 cholera epidemic, we isolated two novel non-toxigenic (ctxA/B-negative) Vibrio cholerae O1. These two isolates underwent whole-genome sequencing and were investigated through comparative genomics and Bayesian coalescent analysis. These isolates cluster in the evolutionary tree with strains responsible for clinical cholera, possessing genomic components of 6th and 7th pandemic lineages, and diverge from “modern” cholera strains around 1548 C.E. [95% HPD: 1532–1555]. Vibrio Pathogenicity Island (VPI)-1 was present; however, SXT/R391-family ICE and VPI-2 were absent. Rugose phenotype conversion and vibriophage resistance evidenced adaption for persistence in aquatic environments. The identification of V. cholerae O1 strains in the Haitian environment, which predate the first reported cholera pandemic in 1817, broadens our understanding of the history of pandemics. It also raises the possibility that these and similar environmental strains could acquire virulence genes from the 2010 Haitian epidemic clone, including the cholera toxin producing CTXϕ.

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Section: Methodsmentioning

confidence: 99%

Non-toxigenic environmental Vibrio cholerae O1 strain from Haiti provides evidence of pre-pandemic cholera in Hispaniola

Azarian

Ali

Johnson

et al. 2016

Sci Rep

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“…We successfully generated a small deletion (33 bp) and a large deletion (Ͼ2.6 kb), as well as a gene replacement, using editing templates with arms of homology ranging from ϳ250 to 575 bp. Novel genes without a predictable biological function are often very abundant in phage genomes; for example, 75% of the 230 predicted ORFs in the ICP1 genome lack homology to known proteins (44). In order to understand this largely unexplored reservoir of novel genetic information, efficient genetic tools are required.…”

Section: Discussionmentioning

confidence: 99%

“…To evaluate the sequence requirements of CRISPR-Cas immunity in V. cholerae, we took advantage of our collection of phages whose whole genomes had been sequenced (44) and in which the targets of several of the spacers are heterogeneous be- (Fig. 5).…”

Section: Methodsmentioning

confidence: 99%

Functional Analysis of Bacteriophage Immunity through a Type I-E CRISPR-Cas System in Vibrio cholerae and Its Application in Bacteriophage Genome Engineering

et al. 2016

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The classical and El Tor biotypes of Vibrio cholerae serogroup O1, the etiological agent of cholera, are responsible for the sixth and seventh (current) pandemics, respectively. A genomic island (GI), GI-24, previously identified in a classical biotype strain of V. cholerae, is predicted to encode clustered regularly interspaced short palindromic repeat (CRISPR)-associated proteins (Cas proteins); however, experimental evidence in support of CRISPR activity in V. cholerae has not been documented. Here, we show that CRISPR-Cas is ubiquitous in strains of the classical biotype but excluded from strains of the El Tor biotype. We also provide in silico evidence to suggest that CRISPR-Cas actively contributes to phage resistance in classical strains. We demonstrate that transfer of GI-24 to V. cholerae El Tor via natural transformation enables CRISPR-Cas-mediated resistance to bacteriophage CP-T1 under laboratory conditions. To elucidate the sequence requirements of this type I-E CRISPR-Cas system, we engineered a plasmid-based system allowing the directed targeting of a region of interest. Through screening for phage mutants that escape CRISPR-Cas-mediated resistance, we show that CRISPR targets must be accompanied by a 3= TT protospacer-adjacent motif (PAM) for efficient interference. Finally, we demonstrate that efficient editing of V. cholerae lytic phage genomes can be performed by simultaneously introducing an editing template that allows homologous recombination and escape from CRISPR-Cas targeting. IMPORTANCECholera, caused by the facultative pathogen Vibrio cholerae, remains a serious public health threat. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) provide prokaryotes with sequence-specific protection from invading nucleic acids, including bacteriophages. In this work, we show that one genomic feature differentiating sixth pandemic (classical biotype) strains from seventh pandemic (El Tor biotype) strains is the presence of a CRISPR-Cas system in the classical biotype. We demonstrate that the CRISPR-Cas system from a classical biotype strain can be transferred to a V. cholerae El Tor biotype strain and that it is functional in providing resistance to phage infection. Finally, we show that this CRISPR-Cas system can be used as an efficient tool for the editing of V. cholerae lytic phage genomes. Vibrio cholerae is a Gram-negative facultative pathogen that causes the acute diarrheal disease cholera. The current cholera pandemic, the seventh in recorded history, began in 1961 and is caused by V. cholerae O1 of the El Tor biotype (1). This pandemic has affected much of the developing world, including many countries in Asia and Africa and most recently in the Caribbean (2). The sixth cholera pandemic, however, was caused by V. cholerae O1 of the classical biotype. Classical biotype strains declined following the emergence of El Tor strains and are now believed to be extinct, after last being seen in 1990 in Bangladesh (3). The mechanisms underpinn...

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“…Both clinical data and mathematical studies support a model where lytic phages may be important in ending outbreaks (Faruque et al 2005a, b; Jensen et al 2006). Recent work has evaluated genomic characterization of phages year-to-year in cholera prone areas using coupled-genomic approaches (Seed et al 2011) and host-pathogen interactions using microarray and RNA-seq (Larocque et al 2005; Mandlik et al 2011). …”

Section: Application Of Genomicsmentioning

confidence: 99%

Genomic Science in Understanding Cholera Outbreaks and Evolution of Vibrio cholerae as a Human Pathogen

Robins

Mekalanos

2014

Cholera Outbreaks

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Modern genomic and bioinformatic approaches have been applied to interrogate the V. cholerae genome, the role of genomic elements in cholera disease, and the origin, relatedness, and dissemination of epidemic strains. A universal attribute of choleragenic strains includes a repertoire of pathogenicity islands and virulence genes, namely the CTX–ϕ prophage and Toxin Co-regulated Pilus (TCP) in addition to other virulent genetic elements including those referred to as Seventh Pandemic Islands. During the last decade, the advent of Next Generation Sequencing (NGS) has provided highly resolved and often complete genomic sequences of epidemic isolates in addition to both clinical and environmental strains isolated from geographically unconnected regions. Genomic comparisons of these strains, as was completed during and following the Haitian outbreak in 2010, reveals that most epidemic strains appear closely related, regardless of region of origin. Non-O1 clinical or environmental strains may also possess some virulence islands, but phylogenic analysis of the core genome suggests they are more diverse and distantly related than those isolated during epidemics. Like Haiti, genomic studies that examine both the Vibrio core- and pan-genome in addition to Single Nucleotide Polymorphisms (SNPs) conclude that a number of epidemics are caused by strains that closely resemble those in Asia, and often appear to originate there and then spread globally. The accumulation of SNPs in the epidemic strains over time can then be applied to better understand the evolution of the V. cholerae genome as an etiological agent.

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Evidence of a Dominant Lineage of Vibrio cholerae-Specific Lytic Bacteriophages Shed by Cholera Patients over a 10-Year Period in Dhaka, Bangladesh

Cited by 126 publications

References 56 publications

Non-toxigenic environmental Vibrio cholerae O1 strain from Haiti provides evidence of pre-pandemic cholera in Hispaniola

Non-toxigenic environmental Vibrio cholerae O1 strain from Haiti provides evidence of pre-pandemic cholera in Hispaniola

Functional Analysis of Bacteriophage Immunity through a Type I-E CRISPR-Cas System in Vibrio cholerae and Its Application in Bacteriophage Genome Engineering

Genomic Science in Understanding Cholera Outbreaks and Evolution of Vibrio cholerae as a Human Pathogen

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