Molecular Analyses of a Putative CTXφ Precursor and Evidence for Independent Acquisition of Distinct CTXφs by Toxigenic Vibrio cholerae

ABSTRACT The genes encoding cholera toxin (ctxA andctxB) are encoded in the genome of CTXφ, a filamentous phage that infects Vibrio cholerae. To study the evolutionary history of CTXφ, we examined genome diversity in CTXφs derived from a variety of epidemic and nonepidemic Vibriosp. natural isolates. Among these were three V. choleraestrains that contained CTX prophage sequences but not thectxA and ctxB genes. These prophages each gave rise to a plasmid form whose genomic organization was very similar to that of the CTXφ replicative form, with the exception of missingctxAB. Sequence analysis of these three plasmids revealed that they lacked the upstream control region normally found 5′ ofctxA, as well as the ctxAB promoter region and coding sequences. These findings are consistent with the hypothesis that a CTXφ precursor that lacked ctxABsimultaneously acquired the toxin genes and their regulatory sequences. To assess the evolutionary relationships among additional CTXφs, two CTXφ-encoded genes, orfU and zot, were sequenced from 13 V. cholerae and 4 V. mimicusisolates. Comparative nucleotide sequence analyses revealed that the CTXφs derived from classical and El Tor V. choleraeisolates comprise two distinct lineages within otherwise nearly identical chromosomal backgrounds (based on mdh sequences). These findings suggest that nontoxigenic precursors of the twoV. cholerae O1 biotypes independently acquired distinct CTXφs.

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RS1 Element of Vibrio cholerae Can Propagate Horizontally as a Filamentous Phage Exploiting the Morphogenesis Genes of CTXΦ

Infection and Immunity ◽

10.1128/iai.70.1.163-170.2002 ◽

2002 ◽

Vol 70 (1) ◽

pp. 163-170 ◽

Cited By ~ 49

Author(s):

Shah M. Faruque ◽

Asadulghani ◽

M. Kamruzzaman ◽

Ranjan K. Nandi ◽

A. N. Ghosh ◽

...

Keyword(s):

Vibrio Cholerae ◽

Kanamycin Resistance ◽

Open Reading Frames ◽

Filamentous Phage ◽

Pcr Analysis ◽

Recombination System ◽

Genes Encoding ◽

Ctx Prophage ◽

A Site ◽

El Tor

ABSTRACT In toxigenic Vibrio cholerae, cholera toxin is encoded by the CTX prophage, which consists of a core region carrying ctxAB genes and genes required for CTXΦ morphogenesis, and an RS2 region encoding regulation, replication, and integration functions. Integrated CTXΦ is often flanked by another genetic element known as RS1 which carries all open reading frames (ORFs) found in RS2 and an additional ORF designated rstC. We identified a single-stranded circularized form of the RS1 element, in addition to the CTXΦ genome, in nucleic acids extracted from phage preparations of 32 out of 83 (38.5%) RS1-positive toxigenic V. cholerae strains analyzed. Subsequently, the corresponding double-stranded replicative form (RF) of the RS1 element was isolated from a representative strain and marked with a kanamycin resistance (Kmr) marker in an intergenic site to construct pRS1-Km. Restriction and PCR analysis of pRS1-Km and sequencing of a 300-bp region confirmed that this RF DNA was the excised RS1 element which formed a novel junction between ig1 and rstC. Introduction of pRS1-Km into a V. cholerae O1 classical biotype strain, O395, led to the production of extracellular Kmr transducing particles, which carried a single-stranded form of pRS1-Km, thus resembling the genome of a filamentous phage (RS1-KmΦ). Analysis of V. cholerae strains for susceptibility to RS1-KmΦ showed that classical biotype strains were more susceptible to the phage compared to El Tor and O139 strains. Nontoxigenic (CTX−) O1 and O139 strains which carried genes encoding the CTXΦ receptor toxin-coregulated pilus (TCP) were also more susceptible (>1,000-fold) to the phage compared to toxigenic El Tor or O139 strains. Like CTXΦ, the RS1Φ genome also integrated into the host chromosomes by using the attRS sequence. However, only transductants of RS1-KmΦ which also harbored the CTXΦ genome produced a detectable level of extracellular RS1-KmΦ. This suggested that the core genes of CTXΦ are also required for the morphogenesis of RS1Φ. The results of this study showed for the first time that RS1 element, which encodes a site-specific recombination system in V. cholerae, can propagate horizontally as a filamentous phage, exploiting the morphogenesis genes of CTXΦ.

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Expression of Cholera Toxin under Non-AKI Conditions in Vibrio cholerae El Tor Induced by Increasing the Exposed Surface of Cultures

Journal of Bacteriology ◽

10.1128/jb.186.5.1355-1361.2004 ◽

2004 ◽

Vol 186 (5) ◽

pp. 1355-1361 ◽

Cited By ~ 14

Author(s):

Joaquín Sánchez ◽

Gerardo Medina ◽

Thomas Buhse ◽

Jan Holmgren ◽

Gloria Soberón-Chavez

Keyword(s):

Cholera Toxin ◽

Vibrio Cholerae ◽

Expression Pattern ◽

Phase 1 ◽

Inert Atmosphere ◽

Surface Area Ratio ◽

Genes Encoding ◽

El Tor

ABSTRACT The regulatory systems controlling expression of the ctxAB genes encoding cholera toxin (CT) in the classical and El Tor biotypes of pathogenic Vibrio cholerae have been characterized and found to be almost identical. Notwithstanding this, special in vitro conditions, called AKI conditions, are required for El Tor bacteria to produce CT. The AKI conditions involve biphasic cultures. In phase 1 the organism is grown in a still tube for 4 h. In phase 2 the medium is poured into a flask to continue growth with shaking. Virtually no expression of CT occurs if this protocol is not followed. Here we demonstrated that CT expression takes place in single-phase still cultures if the volume-to-surface-area ratio is decreased, both under air and under an inert atmosphere. The expression of key genes involved in the regulation of CT production was analyzed, and we found that the expression pattern closely resembles the in vivo expression pattern.

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Evolutionary history of phycoerythrin pigmentation in the water bloom-forming cyanobacteriumMicrocystis aeruginosa

10.1101/485508 ◽

2018 ◽

Cited By ~ 2

Author(s):

Yuuhiko Tanabe ◽

Haruyo Yamaguchi

Keyword(s):

Evolutionary History ◽

Core Genome ◽

Genomic Organization ◽

Phylogenetic Analyses ◽

Green Light ◽

Water Bloom ◽

Phylogenetic Groups ◽

Genome Phylogeny ◽

History Of ◽

Genomic Basis

AbstractMicrocystis aeruginosais a bloom-forming cyanobacterium found in eutrophic fresh-and brackish water bodies worldwide. As typical for cyanobacteria, mostM. aeruginosastrains are blue-green in color owing to the concomitance of two photosynthetic pigments, phycocyanin (PC) and chlorophylla. Although less common,M. aeruginosastrains that are brownish in color owing to the presence of another pigment phycoerythrin (PE) have been documented. However, the genomic basis, phylogeny, and evolutionary origin of PE pigmentation inM. aeruginosahave only been poorly characterized until date. In the present study, we sequenced and characterized the genomes of five PE-containingM. aeruginosastrains. Putative PE synthesis and regulation genes (thecpecluster) were identified in all five sequenced genomes as well as in three previously publishedM. aeruginosagenomes. Of note, Absorption spectra indicated that the PE content, but not PC content, was markedly altered in response to availability of red/green light in all PE-containing strains. This was consistent with the presence ofccaS/ccaR, a hallmark of type II chromatic adapter, in thecpecluster. Phylogenetic analyses of core genome genes indicated that PE-containing genotypes were located in three different phylogenetic groups. In contrast, the genomic organization of thecpecluster was mostly conserved regardless of genomic background. Additionally, the phylogenies of PE genes were found to be congruent, consistent with the core genome phylogeny. A comparison of core genome and PE genes showed a similar level of genetic divergence between two PE-containing groups. These results suggest that genes responsible for PE pigmentation were introduced intoM. aeruginosaearly during evolution and were repeatedly lost thereafter possibly due to ecological adaptation. Additional horizontal gene transfer (HGT) later during evolution also contributed to the present phylogenetic distribution of PE inM. aeruginosa.

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Structural-Functional Analysis of the Genes, Encoding Biosynthesis of Mannose-Sensitive Hemagglutinating Pili of Adhesion in Different Vibrio cholerae El Tor Strains

Problems of Particularly Dangerous Infections ◽

10.21055/0370-1069-2016-4-75-78 ◽

2016 ◽

pp. 75-78 ◽

Cited By ~ 1

Author(s):

N. A. Plekhanov ◽

S. P. Zadnova

Keyword(s):

Functional Analysis ◽

Vibrio Cholerae ◽

Genes Encoding ◽

El Tor

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RS1 Satellite Phage Promotes Diversity of Toxigenic Vibrio cholerae by Driving CTX Prophage Loss and Elimination of Lysogenic Immunity

Infection and Immunity ◽

10.1128/iai.01699-14 ◽

2014 ◽

Vol 82 (9) ◽

pp. 3636-3643 ◽

Cited By ~ 14

Author(s):

M. Kamruzzaman ◽

William Paul Robins ◽

S. M. Nayeemul Bari ◽

Shamsun Nahar ◽

John J. Mekalanos ◽

...

Keyword(s):

Vibrio Cholerae ◽

Sub Saharan Africa ◽

Host Cells ◽

Adult Rabbit ◽

Content Type ◽

Rabbit Intestine ◽

Sub Saharan ◽

Toxigenic Strains ◽

Ctx Prophage ◽

El Tor

ABSTRACTIn El Tor biotype strains of toxigenicVibrio cholerae, the CTXϕ prophage often resides adjacent to a chromosomally integrated satellite phage genome, RS1, which produces RS1ϕ particles by using CTX prophage-encoded morphogenesis proteins. RS1 encodes RstC, an antirepressor against the CTXϕ repressor RstR, which cooperates with the host-encoded LexA protein to maintain CTXϕ lysogeny. We found that superinfection of toxigenic El Tor strains with RS1ϕ, followed by inoculation of the transductants into the adult rabbit intestine, caused elimination of the resident CTX prophage-producing nontoxigenic derivatives at a high frequency. Further studies usingrecAdeletion mutants and a clonedrstCgene showed that the excision event wasrecAdependent and that introduction of additional copies of the clonedrstCgene instead of infection with RS1ϕ was sufficient to enhance CTXϕ elimination. Our data suggest that once it is excised from the chromosome, the elimination of CTX prophage from host cells is driven by the inability to reestablish CTXϕ lysogeny while RstC is overexpressed. However, with eventual loss of the additional copies ofrstC, the nontoxigenic derivatives can act as precursors of new toxigenic strains by acquiring the CTX prophage either through reinfection with CTXϕ or by chitin-induced transformation. These results provide new insights into the role of RS1ϕ inV. choleraeevolution and the emergence of highly pathogenic clones, such as the variant strains associated with recent devastating epidemics of cholera in Asia, sub-Saharan Africa, and Haiti.

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Multilocus variable-number tandem repeat analysis of Vibrio cholerae O1 El Tor strains harbouring classical toxin B

Journal of Medical Microbiology ◽

10.1099/jmm.0.017939-0 ◽

2010 ◽

Vol 59 (7) ◽

pp. 763-769 ◽

Cited By ~ 31

Author(s):

Seon Young Choi ◽

Je Hee Lee ◽

Yoon-Seong Jeon ◽

Hye Ri Lee ◽

Eun Jin Kim ◽

...

Keyword(s):

Cholera Toxin ◽

Vibrio Cholerae ◽

Tandem Repeat ◽

Variable Number Tandem Repeat ◽

Variable Number ◽

Group Iii ◽

Repeat Analysis ◽

Group I ◽

Ctx Prophage ◽

El Tor

Atypical Vibrio cholerae O1 strains – hybrid strains (strains that cannot be classified either as El Tor or classical biotype) and altered strains (El Tor biotype strains that produce classical cholera toxin) – are currently prevalent in Asia and Africa. A total of 74 hybrid and altered strains that harboured classical cholera toxin were investigated by multilocus variable-number tandem repeat analysis (MLVA). The results showed that the hybrid/altered strains could be categorized into three groups and that they were distant from the El Tor strain responsible for the seventh cholera pandemic. Hybrid/altered strains with a tandem repeat of the classical CTX prophage on the small chromosome were divided into two MLVA groups (group I: Mozambique/Bangladesh group; group III: Vietnam group), and altered strains with the RS1–CTX prophage containing the El Tor type rstR and classical ctxB on the large chromosome were placed in two MLVA groups (group II: India/Bangladesh group; group III: India/Vietnam group).

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Activation of the Vibrio cholerae SOS Response Is Not Required for Intestinal Cholera Toxin Production or Colonization

Infection and Immunity ◽

10.1128/iai.74.2.927-930.2006 ◽

2006 ◽

Vol 74 (2) ◽

pp. 927-930 ◽

Cited By ~ 11

Author(s):

Mariam Quinones ◽

Brigid M. Davis ◽

Matthew K. Waldor

Keyword(s):

Cholera Toxin ◽

Vibrio Cholerae ◽

Sos Response ◽

Toxin Production ◽

Mrna Levels ◽

Suckling Mouse ◽

Potent Inducer ◽

Genes Encoding ◽

Mouse Intestine ◽

Ctx Prophage

ABSTRACT Cholera toxin, one of the main virulence factors of Vibrio cholerae, is encoded in the genome of CTXφ, a V. cholerae-specific lysogenic filamentous bacteriophage. Although the genes encoding cholera toxin, ctxAB, are known to have their own promoter, the toxin genes can also be transcribed from an upstream CTXφ promoter, PrstA . The V. cholerae SOS response to DNA damage induces the CTX prophage by stimulating gene expression initiating from PrstA . Here, we investigated whether ctxA mRNA levels increase along with the levels of the transcripts for the other CTXφ genes following stimulation of the V. cholerae SOS response. Treatment of V. cholerae with the SOS-inducing agent mitomycin C increased the level of ctxA mRNA approximately sevenfold, apparently by augmenting the activity of PrstA . However, using suckling mice as a model host, we found that intraintestinal ctxA transcription does not depend on PrstA . In fact, the suckling mouse intestine does not appear to be a potent inducer of the V. cholerae SOS response. Furthermore, alleviation of LexA-mediated repression of the V. cholerae SOS regulon was not required for V. cholerae growth in the suckling mouse intestine. Our observations suggest that pathogenicity of V. cholerae does not depend on its SOS response.

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Transcriptome and Phenotypic Responses of Vibrio cholerae to Increased Cyclic di-GMP Level

Journal of Bacteriology ◽

10.1128/jb.188.10.3600-3613.2006 ◽

2006 ◽

Vol 188 (10) ◽

pp. 3600-3613 ◽

Cited By ~ 137

Author(s):

Sinem Beyhan ◽

Anna D. Tischler ◽

Andrew Camilli ◽

Fitnat H. Yildiz

Keyword(s):

Vibrio Cholerae ◽

Elevated Level ◽

Life Cycles ◽

Cell Physiology ◽

Human Pathogen ◽

Aquatic Life ◽

Cellular Processes ◽

Genes Encoding ◽

El Tor ◽

Phenotypic Responses

ABSTRACT Vibrio cholerae, the causative agent of cholera, is a facultative human pathogen with intestinal and aquatic life cycles. The capacity of V. cholerae to recognize and respond to fluctuating parameters in its environment is critical to its survival. In many microorganisms, the second messenger, 3′,5′-cyclic diguanylic acid (c-di-GMP), is believed to be important for integrating environmental stimuli that affect cell physiology. Sequence analysis of the V. cholerae genome has revealed an abundance of genes encoding proteins with either GGDEF domains, EAL domains, or both, which are predicted to modulate cellular c-di-GMP concentrations. To elucidate the cellular processes controlled by c-di-GMP, whole-genome transcriptome responses of the El Tor and classical V. cholerae biotypes to increased c-di-GMP concentrations were determined. The results suggest that V. cholerae responds to an elevated level of c-di-GMP by increasing the transcription of the vps, eps, and msh genes and decreasing that of flagellar genes. The functions of other c-di-GMP-regulated genes in V. cholerae are yet to be identified.

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Plasmidic Extended-Spectrum β-Lactamases in Vibrio cholerae O1 El Tor Isolates in Argentina

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.46.5.1462-1468.2002 ◽

2002 ◽

Vol 46 (5) ◽

pp. 1462-1468 ◽

Cited By ~ 53

Author(s):

Alejandro Petroni ◽

Alejandra Corso ◽

Roberto Melano ◽

María Luisa Cacace ◽

Ana María Bru ◽

...

Keyword(s):

Vibrio Cholerae ◽

Profile Analysis ◽

Plasmid Profile ◽

Polymorphism Analysis ◽

Susceptibility Analysis ◽

Extended Spectrum ◽

Genes Encoding ◽

Vibrio Cholerae O1 ◽

El Tor ◽

Susceptibility Profiles

ABSTRACT Since 1992 there have been seven major outbreaks of cholera in Argentina. Susceptibility analysis of 1,947 isolates (40% of reported cases) of Vibrio cholerae O1 biotype El Tor suggested the presence of extended-spectrum β-lactamases (ESBLs) in 28 isolates. Because of their different susceptibility profiles, V. cholerae isolates M1502, M1516, M1573, and M3030 (all of which are of the Ogawa serotype) were selected for the present study. By susceptibility analysis, isoelectric focusing, and PCR-based restriction fragment length polymorphism analysis, CTX-M-type enzymes were identified in three isolates, whereas a PER-2-type enzyme, in addition to a TEM-1-like enzyme, was identified in the other isolate. The presence of these ESBLs in V. cholerae isolates resulted in MICs well below those commonly observed for members of the family Enterobacteriaceae. Genes that encode both ESBLs were transferred to Escherichia coli by conjugation, together with all determinants of resistance to non-β-lactam antibiotics (gentamicin, kanamycin, and sulfamethoxazole for all isolates; amikacin and streptomycin for three isolates; trimethoprim, tetracycline, and chloramphenicol for two isolates). Plasmid profile analysis and Southern blotting revealed the presence of single plasmids of about 150 kb in the four V. cholerae isolates and their respective transconjugants and revealed that the plasmids harbored genes encoding CTX-M-type or PER-2-type ESBLs. These results strongly suggest the broad spread of these ESBLs among genera belong to families other than the Enterobacteriaceae.

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