Seventh Pandemic Research Articles

Development of an INDEL typing system for <i>ctx</i>+ strains of <i>Vibrio cholerae</i> from the seventh pandemic

BACKGROUND: The seventh cholera pandemic is accompanied by the formation of Vibrio cholerae clones with new genetic properties, including those with the ability to spread pandemically and cause diseases with a more severe clinical course. The widespread distribution of such genetic variants of Vibrio cholerae and the possibility of their introduction into the territory of the Russian Federation necessitate constant comprehensive monitoring using modern molecular genetic technologies. AIM: To improve INDEL typing of ctx+ strains of V. cholerae of the seventh pandemic by using additional INDEL loci. Materials and methods: A bioinformatic analysis of 2105 full-genome sequences of toxigenic ctxAB+tcpA+ strains of Vibrio cholerae O1 El Tor from open databases was carried out in order to search for INDEL loci for molecular typing. Based on the convenience criterion for allele size identification, eight INDEL loci were selected. Three loci have been described previously, and five were identified as a result of this work. The designed primers formed amplicons ranging in size from 67 to 390 base pairs, which made it possible to confidently identify them during gel electrophoresis. Results: The distribution of alleles formed 11 unique INDEL clusters, which we designated A-K. Based on the number of strains within the clusters, three types of clusters were identified: major (A, B and C) made up 89% of the total number of sequences studied, intermediate (D, E, F, G and H) 10.5% of the genomes. Three minor clusters (I, J and K) were represented by single strains. Four clusters united strains isolated in the 20th century (A — in 1941, F — in 1957, G — in 1993, E — in 1999), and seven clusters — in the 21st century in the period from 2003 to 2016. In the period from 2019 to 2023, representatives of INDEL clusters were active: A, B, D and E. ConclusionS: The study of the timing of circulation suggested that representatives of different clusters have different epidemic potential, which was manifested in the absence of isolation of strains of some clusters in recent years. A comparative study of INDEL typing with SNP typing in the in silico analysis of 378 genomes of strains isolated on the African continent indicates that the proposed INDEL typing method is not inferior to SNP typing in terms of resolution.

Genomic evidence of two-staged transmission of the early seventh cholera pandemic

The seventh cholera pandemic started in 1961 in Indonesia and spread across the world in three waves in the decades that followed. Here, we utilised genomic evidence to detail the first wave of the seventh pandemic. Genomes of 22 seventh pandemic Vibrio cholerae isolates from 1961 to 1979 were completely sequenced. Together with 152 publicly available genomes from the same period, they fell into seven phylogenetic clusters (CL1–CL7). By multilevel genome typing (MGT), all were assigned to MGT2 ST1 (Wave 1) except three isolates in CL7 which were typed as MGT2 ST2 (Wave 2). The Wave 1 seventh pandemic expanded in two stages, with Stage 1 (CL1–CL5) spread across Asia and Stage 2 (CL6 and CL7) spread to the Middle East and Africa. Three non-synonymous mutations, one each, in three regulatory genes, csrD (global regulator), acfB (chemotaxis), and luxO (quorum sensing) may have critically contributed to its pandemicity. The three MGT2 ST2 isolates in CL7 were the progenitors of Wave 2 and evolved from within Wave 1 with acquisition of a novel IncA/C plasmid. Our findings provide new insight into the evolution and transmission of the early seventh pandemic, which may aid future cholera prevention and control.

Diversity and Complexity of CTXΦ and Pre-CTXΦ Families in Vibrio cholerae from Seventh Pandemic.

CTXΦ is a lysogenic filamentous phage that carries the genes encoding cholera toxin (ctxAB), the main virulence factor of Vibrio cholerae. The toxigenic conversion of environmental V. cholerae strains through CTXΦ lysogenic infection is crucial for the emergence of new pathogenic clones. A special allelic form of CTXΦ, called pre-CTXΦ, is a precursor of CTXΦ and without ctxAB. Different members of the pre-CTXΦ and CTXΦ families are distinguished by the sequence of the transcriptional repressor-coding gene rstR. Multiple rstR alleles can coexist within a single strain, demonstrating the diverse structure and complex genomic integration patterns of CTXΦ/pre-CTXΦ prophage on the chromosome. Exploration of the diversity and co-integration patterns of CTXΦ/pre-CTXΦ prophages in V. cholerae can help to understand the evolution of this phage family. In this study, 21 V. cholerae strains, which were shown to carry the CTXΦ/pre-CTXΦ prophages as opposed to typical CTXETΦ-RS1 structure, were selected from approximately 1000 strains with diverse genomes. We identified two CTXΦ members and six pre-CTXΦ members with distinct rstR alleles, revealing complex chromosomal DNA integration patterns and arrangements of different prophages in these strains. Promoter activity assays showed that the transcriptional repressor RstR protected against CTXΦ superinfection by preventing the replication and integration of CTXΦ/pre-CTXΦ phages containing the same rstR allele, supporting the co-integration of the diverse CTXΦ/pre-CTXΦ members observed. The numbers and types of prophages and their co-integration arrangements in serogroup O139 strains were more complex than those in serogroup O1 strains. Also, these CTXΦ/pre-CTXΦ members were shown to present the bloom period of the CTXΦ/pre-CTXΦ family during wave 2 of the seventh cholera pandemic. Together, these analyses deepen our comprehension of the genetic variation of CTXΦ and pre-CTXΦ and provide insights into the evolution of the CTXΦ/pre-CTXΦ family in the seventh cholera pandemic.

Comparative analysis of the structure of regulatory genes of Vibrio cholerae serotype О1 biotype El Tor strains

Introduction. The expression of the ctxAB and tcpA-F genes encoding the main pathogenicity factors of the Vibrio cholerae is controlled by regulatory genes. The structure of these genes has not been fully studied in the pathogen strains isolated during different periods of the current pandemic. The aim of the study was a comparative analysis of the structure of regulatory genes of V. cholerae O1 biovar El Tor strains isolated on the territory of the Russian Federation and neighboring countries during the seventh cholera pandemic. Materials and methods. The nucleotide sequences of the complete genomes of 29 toxigenic strains isolated from 1970 to 2023 were analyzed. The analysis was carried out using BioEdit v7.2.6.1 software and Blast tool. Results. The analysis of ten regulatory genes (toxT, aphA, aphB, hns, hapR, vieA, luxO, luxT, carS, carR) was carried out. Almost all strains were found to have a thymine insertion in the hapR gene at position 219. The exception was V. cholerae strain M3208 (Tambov, 2023), which had an insertion of five nucleotides in this gene. Mutations of the luxO gene with an unknown effect were detected in 44.8% of the studied strains. In 46.7% and 33.3% of the studied genetic variants carrying the ctxB1 allele, non-synonymous substitutions were detected in the hns (G319A) and vieA (C235T) genes, respectively. All genetic variants with the ctxB7 allele have mutations in both the hns and vieA genes. Three genetic variants with the ctxB7 allele, imported to the Russian Federation in recent years, contain an altered structure of the carR gene (G265A). Conclusion. The structure of genes (toxT, aphA, aphB, carS, luxT, hapR) of V. cholerae O1 El Tor strains remains unchanged for the majority of the studied isolates. At the same time, variability in the hns (G319A), vieA (C235T) and carR (G256A) genes was detected. Mutations in these genes can be used as genetic markers of modern V. cholerae O1 El Tor genetic variants.

Genomic Analysis of <I>Vibrio cholerae</I> Strains Isolated in Siberia and the Far East during the Seventh Cholera Pandemic: Determining the Appurtenance to Global Phylogenetic Lines

Genomic Analysis of <I>Vibrio cholerae</I> Strains Isolated in Siberia and the Far East during the Seventh Cholera Pandemic: Determining the Appurtenance to Global Phylogenetic Lines

Plasmid targeting and destruction by the DdmDE bacterial defence system.

Although eukaryotic Argonautes have a pivotal role in post-transcriptional gene regulation through nucleic acid cleavage, some short prokaryotic Argonaute variants (pAgos) rely on auxiliary nuclease factors for efficient foreign DNA degradation1. Here we reveal the activation pathway of the DNA defence module DdmDE system, which rapidly eliminates small, multicopy plasmids from the Vibrio cholerae seventh pandemic strain (7PET)2. Through a combination of cryo-electron microscopy, biochemistry and in vivo plasmid clearance assays, we demonstrate that DdmE is a catalytically inactive, DNA-guided, DNA-targeting pAgo with a distinctive insertion domain. We observe thatthe helicase-nuclease DdmD transitions from an autoinhibited, dimeric complex to amonomeric state upon loading of single-stranded DNA targets. Furthermore, the complete structure of the DdmDE-guide-target handover complex provides a comprehensive view into how DNA recognition triggers processive plasmid destruction. Our work establishes a mechanistic foundation for how pAgos utilize ancillary factors to achieve plasmid clearance, and provides insights into anti-plasmid immunity in bacteria.

Spatiotemporal dynamics of cholera epidemics in Ethiopia: 2015–2021

Since the onset of the seventh cholera pandemic, Ethiopia has been affected by recurrent epidemics. However, the epidemiology of cholera in this country remains poorly understood. This study aimed to describe cholera outbreak characteristics in Ethiopia from 2015 to 2021. During this period, Ethiopia experienced four epidemic waves. The first wave involved nationwide outbreaks during the second half of 2016 followed by outbreaks predominantly affecting Somali Region in 2017. The second wave primarily affected Tigray and Afar Regions. During the third wave, multiple smaller-scale outbreaks occurred during 2019. The fourth wave was limited to Bale Zone (Oromia Region) in 2021. Overall, a north to south shift was observed over the course of the study period. Major cholera transmission factors included limited access to safe water and sanitation facilities. Severe weather events (drought and flooding) appear to aggravate cholera diffusion. Cholera transmission between Ethiopia and nearby countries (Kenya and Somalia), likely plays a major role in regional cholera dynamics. Overall, this study provides the first understanding of recent spatiotemporal cholera dynamics in Ethiopia to inform cholera control and elimination strategies.

Clinical surveillance systems obscure the true cholera infection burden in an endemic region

Our understanding of cholera transmission and burden largely relies on clinic-based surveillance, which can obscure trends, bias burden estimates and limit the impact of targeted cholera-prevention measures. Serological surveillance provides a complementary approach to monitoring infections, although the link between serologically derived infections and medically attended disease incidence—shaped by immunological, behavioral and clinical factors—remains poorly understood. We unravel this cascade in a cholera-endemic Bangladeshi community by integrating clinic-based surveillance, healthcare-seeking and longitudinal serological data through statistical modeling. Combining the serological trajectories with a reconstructed incidence timeline of symptomatic cholera, we estimated an annual Vibrio cholerae O1 infection incidence rate of 535 per 1,000 population (95% credible interval 514–556), with incidence increasing by age group. Clinic-based surveillance alone underestimated the number of infections and reported cases were not consistently correlated with infection timing. Of the infections, 4 in 3,280 resulted in symptoms, only 1 of which was reported through the surveillance system. These results impart insights into cholera transmission dynamics and burden in the epicenter of the seventh cholera pandemic, where >50% of our study population had an annual V. cholerae O1 infection, and emphasize the potential for a biased view of disease burden and infection risk when depending solely on clinical surveillance data.

Genome sequences of Vibrio cholerae strains isolated in the DRC between 2009 and 2012.

Vibrio cholerae has caused seven cholera pandemics in the past two centuries. The seventh and ongoing pandemic has been particularly severe on the African continent. Here, we report long read-based genome sequences of six V. cholerae strains isolated in the Democratic Republic of the Congo between 2009 and 2012.

EPIDEMIOLOGICAL SITUATION OF CHOLERA IN THE WORLD AND IN KAZAKHSTAN FOR 2022-2024

This paper provides an overview of the monitoring of the spread of cholera in the world, the CIS countries and the Republic of Kazakhstan with an assessment of the risks of possible introduction and further spread of this particularly dangerous disease among the population of the country. The ways of introduction, measures to prevent and contain the spread are considered in order to ensure the epidemiological well-being and biological safety of the population of the Republic of Kazakhstan for 2022-2023 and for six months of 2024.Despite the large-scale vaccination of the population of Asia and Africa, there is an upward trend in the dynamics of the incidence of cholera in the world. In 2022-2023 and in the five months of 2024, 1,135,601 cholera patients were registered in 44 countries worldwide, of which 9297 were fatal. Including by country: 147 people fell ill in Europe, 2 died; in Asia - 666 350/,620; in Africa - 458 316/7357; in North and Central America - 10788/1318. WHO has reported imported cholera cases to countries in Asia, the Americas, including the Caribbean, Europe and Australia with Oceania. Administrative territories endemic to cholera have been identified in 24 countries. According to WHO, the increasing increase in the incidence of cholera is associated with social and natural risks caused by emergencies of various origins, the presence of endemic foci, imported infections and other risk factors. The forecast for cholera in the world for 2024-2025, taking into account the established high degree of activation of the epidemic process, is unfavorable. For the Republic of Kazakhstan, the cholera forecast is determined by the presence of external risks caused by the continuation of the seventh cholera pandemic, possible imports of infection to the regions of the country

Vibrio cholerae's ToxRS bile sensing system.

The seventh pandemic of the diarrheal cholera disease, which began in 1960, is caused by the Gram-negative bacterium Vibrio cholerae. Its environmental persistence provoking recurring sudden outbreaks is enabled by V. cholerae's rapid adaption to changing environments involving sensory proteins like ToxR and ToxS. Located at the inner membrane, ToxR and ToxS react to environmental stimuli like bile acid, thereby inducing survival strategies e.g. bile resistance and virulence regulation. The presented crystal structure of the sensory domains of ToxR and ToxS in combination with multiple bile acid interaction studies, reveals that a bile binding pocket of ToxS is only properly folded upon binding to ToxR. Our data proposes an interdependent functionality between ToxR transcriptional activity and ToxS sensory function. These findings support the previously suggested link between ToxRS and VtrAC-like co-component systems. Besides VtrAC, ToxRS is now the only experimentally determined structure within this recently defined superfamily, further emphasizing its significance. In-depth analysis of the ToxRS complex reveals its remarkable conservation across various Vibrio species, underlining the significance of conserved residues in the ToxS barrel and the more diverse ToxR sensory domain. Unravelling the intricate mechanisms governing ToxRS's environmental sensing capabilities, provides a promising tool for disruption of this vital interaction, ultimately inhibiting Vibrio's survival and virulence. Our findings hold far-reaching implications for all Vibrio strains that rely on the ToxRS system as a shared sensory cornerstone for adapting to their surroundings.

Genetic characterization of non-O1/non-O139 <i>Vibrio cholerae</i> mobilome: a strategy for understanding and discriminating emerging environmental bacterial strains

Acute diarrhea and cholera (AWD/C) result in more than 21000 to 143000 global mortality annually and are associated with Vibrio cholerae. The pathogen has shown increasing evolutionary/emerging dynamics linked with mobilome or ubiquitous nature of mobile integrative genetic and conjugative elements (MIGCE), however, such dynamics are rarely reported amongst somatic-antigen non-agglutinating Type-1/-139 V. cholerae (SA-NAG-T-1/139Vc). The study reports the genetic detection of mobilome-associated indices in SA-NAG-T-1/139Vc as a potential strategy for differentiating/discriminating emerging environmental bacteria. Presumptive V. cholerae isolates were retrieved from five water sources, while strains were characterized/serogrouped and confirmed using simplex and comparative-genomic-multiplex Polymerase Chain Reaction (PCR). Genomic island (GI-12det, GI-14det, GI-15det); Phages (TLC-phagedet, Kappa-phagedet) and ICEs of the SXT/R391 family genes (SXT/R391-ICEs integrase, SXT-Hotspot-IV, ICEVchInd5Hotspot-IV, ICEVchMoz10Hotspot-IV) were detected. Other rare ICE members such as the ICEVcBan8att gene and Vibrio Seventh Pandemic island detection (VSP-II Integrase, Prototypical VSP-II) were also detected. Results revealed that the 8.22% (61/742) SA-NAG-T-1/139Vc serogroup observed harbors the Vibrio Seventh Pandemic island integrase (34/61; 55.7%) and other rare genetic traits including; attB/attP (29/61; 47.5%, 14/61; 23%), integrative genetic elements (4/61; 6.56%), phage types (TLC-phagedet: 2/61; 3.28% and Kappa-phagedet: 7/61; 11.48%) as well as the integrase genes (INT1, Sul1, Sul2) (29/61: 47.5%; 21/61: 34.4%; 25/61: 41%). Such genetic detection of mobilome determinants/MIGCE suggests potential discriminatory tendencies amongst SA-NAG-T-1/139Vcwhich may be applied in mobilome typing of evolving/emerging environmental bacteria. The need to encourage the application of such mobilome typing indices and continuous study of these strains is suggestive of interest in controlling future potential emerging environmental strains.

Recent surge in cholera outbreaks globally during the COVID-19 pandemic era: a potential threat to the African continent and salient counteracting strategies.

Recent surge in cholera outbreaks globally during the COVID-19 pandemic era: a potential threat to the African continent and salient counteracting strategies.

New Insights into Vibrio cholerae Biofilms from Molecular Biophysics to Microbial Ecology.

With the discovery that 48% of cholera infections in rural Bangladesh villages could be prevented by simple filtration of unpurified waters and the detection of Vibrio cholerae aggregates in stools from cholera patients it was realized V. cholerae biofilms had a central function in cholera pathogenesis. We are currently in the seventh cholera pandemic, caused by O1 serotypes of the El Tor biotypes strains, which initiated in 1961. It is estimated that V. cholerae annually causes millions of infections and over 100,000 deaths. Given the continued emergence of cholera in areas that lack access to clean water, such as Haiti after the 2010 earthquake or the ongoing Yemen civil war, increasing our understanding of cholera disease remains a worldwide public health priority. The surveillance and treatment of cholera is also affected as the world is impacted by the COVID-19 pandemic, raising significant concerns in Africa. In addition to the importance of biofilm formation in its life cycle, V. cholerae has become a key model system for understanding bacterial signal transduction networks that regulate biofilm formation and discovering fundamental principles about bacterial surface attachment and biofilm maturation. This chapter will highlight recent insights into V. cholerae biofilms including their structure, ecological role in environmental survival and infection, regulatory systems that control them, and biomechanical insights into the nature of V. cholerae biofilms.

Genomes of Vibrio cholerae O1 Serotype Ogawa Associated with Current Cholera Activity in Pakistan.

Here, this report presents two genomes of Vibrio cholerae O1 serotype Ogawa, recovered from cholera cases in Australia linked to travel to Pakistan in 2022. Their multidrug-resistant genotype represents the current activity of cholera within the seventh pandemic. One of the genome sequences was assembled using both short- and long-read sequences.

Vibrio cholerae O1 El Tor strains linked to global cholera show region-specific patterns by pulsed-field gel electrophoresis.

Vibrio cholerae O1 El Tor, causative agent of the ongoing seventh cholera pandemic, is native to the aquatic environment of the Ganges Delta, Bay of Bengal (GDBB). Recent studies traced pandemic strains to the GDBB and proposed global spread of cholera had occurred via intercontinental transmission. In the research presented here, NotI-digested genomic DNA extracted from V. cholerae O1 clinical and environmental strains isolated in Bangladesh during 20042014 was analyzed by pulsed-field gel electrophoresis (PFGE). Results of cluster analysis showed 94.67% of the V. cholerae strains belonged to clade A and included the majority of clinical strains of spatio-temporal origin and representing different cholera endemic foci. The rest of the strains were estuarine, all environmental strains from Mathbaria, Bangladesh, and occurred as singletons, clustered in clades B and C, or in the small clades D and E. Cluster analysis of the Bangladeshi strains and including 157 El Tor strains from thirteen countries in Asia, Africa, and the Americas revealed 85% of the total set of strains belonged to clade A, indicating all were related, yet did not form an homogeneous cluster. Overall, 15% of the global strains comprised multiple small clades or segregated as singletons. Three sub-clades could be discerned within the major clade A, reflecting distinct lineages of V. cholerae O1 El Tor associated with cholera in Asia, Africa, and the Americas. The presence in Asia and the Americas of non-pandemic V. cholerae O1 El Tor populations differing by PFGE and from strains associated with cholera globally suggests different ecotypes are resident in distant geographies.

The Vibrio cholerae Seventh Pandemic Islands act in tandem to defend against a circulating phage.

The current circulating pandemic El Tor biotype of Vibrio cholerae has persisted for over sixty years and is characterized by its acquisition of two unique genomic islands called the Vibrio Seventh Pandemic Islands 1 and 2 (VSP-I and VSP-II). However, the functions of most of the genes on VSP-I and VSP-II are unknown and the advantages realized by El Tor through these two islands are not clear. Recent studies have broadly implicated these two mobile genetic elements with phage defense. Still, protection against phage infection through these islands has not been observed directly in any V. cholerae El Tor biotype. Here we report the isolation of a circulating phage from a cholera patient stool sample and demonstrate that propagation of this phage in its native host is inhibited by elements in both VSP-I and VSP-II, providing direct evidence for the role of these genomic islands in phage defense. Moreover, we show that these defense systems are regulated by quorum sensing and active only at certain cell densities. Finally, we have isolated a naturally occurring phage variant that is resistant to the defense conferred by the VSP islands, illustrating the countermeasures used by phages to evade these defense mechanisms. Together, this work demonstrates a functional role for the VSPs in V. cholerae and highlights the key regulatory and mechanistic insights that can be gained by studying anti-phage systems in their native contexts.

Elaboration of a Toxigenic Vibrio cholerae Typing Scheme Based on Bioinformatiсs Analysis Data

Introduction: The current stage of the seventh cholera pandemic is characterized by the emergence of novel Vibrio cholerae gene variants, gradually replacing their predecessors and occupying a dominant position in the etiology of the disease. Determining their epidemic potential by identifying the number of genetic markers is unsuitable for operational analysis. Thus, the development of a method for differentiating pathogens based on PCR detection of a limited number of markers seems relevant. Objective: To create a database of whole genome V. cholerae sequences containing different alleles of cheA3 (VCA1095) and rtxA genes based on bioinformatics analysis data and to elaborate a simple and informative toxigenic vibrio typing scheme. Materials and methods: The NCBI database-extracted results of whole genome sequencing of 3,309 strains of Vibrio cholerae isolated in 1962–2021 were used for the analysis. The software was developed in Java. Results: The bioinformatics analysis of the database of whole genome V. cholerae sequences, including 3,309 genomes of third wave strains, enabled us to divide them into three groups: “pre-Haitian”, “Haitian”, and “post-Haitian”. All of them contained alleles of the genes of toxin-co-regulated tcpACIRS101 pili and the MARTX rtxA4 cytotoxin with a null mutation that caused a premature stop codon. However, in the “pre-Haitian” strains, the gene of the cholera toxin subunit B of the classical ctxB1 type and the prototype gene of histidine kinase cheA3 (VCA1095) were always detected, which in PCR formed a 95 bp long amplicon and was designated as VCA1095-95. In the “Haitian” strains, a deletion of 8 bp occurred in this gene, and the PCR amplicon was shortened to 87 bp (VCA1095-87). Its mandatory combination with the ctxB7 allele was revealed. The “post-Haitian” strains contained an even shorter rtxA4a allele due to the deletion of 60 bp in the proximal part. Conclusion: Since the analysis of a large number of genomes revealed strict correlations between certain alleles in each group, we consider it possible to use only two markers for operational analysis, i.e. alleles of the cheA3 and rtxA genes. The typing scheme based on their PCR detection can be used to facilitate determination of the epidemic potential of newly isolated cultures.

Renewed Interest in Cholera Control

Cholera is an ancient disease of significant public health importance accounting for approximately 1.3-4 million cases worldwide and about 21000–143,000 deaths. Cholera is endemic in many developing countries where sanitation is poor and lack in safe water. The disease has the potential to cause large-scale epidemics and pandemics. The ongoing seventh pandemic has spread to more than 100 countries since its origin in 1961 in Indonesia. Travellers from developed countries travelling to cholera endemic areas are at higher risk of getting cholera. Oral rehydration therapy (ORT) has revolutionized the treatment of cholera and causes reduction of mortality. ORT has saved the lives of millions of children all over the world and is considered the single most significant discovery in medicine of the century. Effective oral cholera vaccines have been developed and are being used but not to its full potential to control cholera. Since the disease burden due to cholera is huge and we have the tools to combat cholera, particularly reduce mortality, it is imperative that the global communities should embark on cholera control as an urgent priority of human health. Cholera control should be viewed as a poverty alleviation programme for the marginalised population. The current pandemic of COVID-19 overwhelmed the economy and healthcare systems so much so that the health authorities undermined the cholera control measures even during ongoing epidemics of cholera. The World Health Organization (WHO) has reframed the strategy with renewed interest on cholera control with a target of reducing the mortality from cholera by 90 percent by 2030. Member countries should mobilize enough funds for the programme and actively implement it. Improvement of sanitation and safe water for all should continue as long-term solution for combating the menace of cholera.

Identifying long-term colonization factors of Vibrio cholerae O1 El Tor in the zebrafish natural host model

The human disease cholera, marked by acute, voluminous watery diarrhea, is caused by the gram-negative, aquatic bacterium Vibrio cholerae. All seven cholera pandemics since 1817 were identified as being caused by just 2 of over 155 known V. choleraeserogroups: O1 and O139. The O1 serogroup is divided into two biotypes: classical and El Tor. Classical biotype is associated with pandemics 1 through 6, but the El Tor biotype has since displaced classical as the causative agent of the ongoing 7th cholera pandemic over the past 60 years. The El Tor genome resembles that of classical but has acquired two unique pathogenicity islands known as Vibrio Seventh Pandemic (VSP) -1 and -2. El Tor biotype has been associated with prolonged colonization, infection, and disease both in humans and in the zebrafish natural host model. The zebrafish model allows for complete observation of Vibrio cholerae infection in a system undisrupted by antibiotic use or immune suppression. El Tor strains colonize the zebrafish intestine for up to 10 days longer than classical strains. Preliminary studies demonstrate VSP-2 is required to observe this phenotype, but VSP-1 is not. By creating targeted regional knockouts of the VSP-2 island, the specific gene(s) essential for enabling prolonged colonization will be identified and applied to the understanding of how El Tor interacts with a natural host. By identifying the genes used by El Tor to colonize a natural host for prolonged periods, we gain insight into how this pathogen may persist in the environment and perpetuate disease.