New Vibrio cholerae Core-Genome Multi-Locus Sequence Typing (cgMLST) Website Online

A new core-genome multi-locus sequence typing (cgMLST) scheme for V. cholerae is now publicly available on pubmlst.org/vcholerae. New genomes can now be submitted and analyzed on PubMLST within minutes. Tools such as GrapeTree, Interactive Tree of Life, and PHYLOViZ are all integrated within the website to allow for quick follow-up analyses. This cgMLST scheme, containing 2,443 core genes, has been used to analyze over 1,000 V. cholerae genomes from both private collections and publicly available databases. This dataset includes representative isolates from known outbreaks and endemic countries. Along with epidemiological data, it is possible to use this cgMLST scheme to monitor V. cholerae diversity at both the local and global levels. A 1:1 mapping with a previously established V. cholerae MLST scheme from our lab has been carried out to demonstrate backwards compatibility with prior MLST studies.

The full manuscript entitled "A Vibrio cholerae core genome multilocus sequence typing scheme to facilitate the epidemiological study of cholera," published in the Journal of Bacteriology, details the application of this cgMLST scheme to study cholera outbreaks. This work was done in collaboration with Dr. Cheryl Tarr from the Centers of Disease Control and Prevention and Dr. Munirul Alam from the International Centre for Diarrhoeal Disease Research, Bangladesh.

cgMLST minimum spanning tree of all Yemen isolates and representative 7th pandemic El Tor strains (from Liang et al., 2020)

Insights into an Environmental Vibrio cholerae Population on a Subspecies Scale

This manuscript represents the original Boucher Lab Gang’s (Paul, Fabini and Tania) first paper together! Their publication in Environmental Microbiology, “Culture‐independent tracking of Vibrio cholerae lineages reveals complex spatiotemporal dynamics in a natural population”, combines culture-independent amplicon sequencing and qPCR to study the diversity and abundance of dozens of sympatric V. cholerae strains in a single location. Kirchberger et. al finds that among many co-occurring strains, few manage to grow to large numbers on resource-rich particles, while the rest fight for the scraps (perhaps using their diverse Type VI secretion system toxins?).

Size-fractionated Vibrio cholerae population composition using culture independent techniques from a single, cholera-free environment. From Kirchberger et al., 2020.

Eight Novel Vibrio Genomes Announced

In addition to the nine Vibrio sp. genomes deposited late last November, a new genome announcement with eight additional Vibrio isolates has come out today. Working alongside the US Centers for Disease Control and Prevention again, these newly described Vibrio strains have come from a variety of blood, stool or wound infections, in addition to a previously described environmental strain. All genomes within the article "Draft genome sequences of eight Vibrio sp. clinical isolates from across the United States that form a basal sister clade to Vibrio cholerae" cluster within a single clade, and molecular analyses for species delineation place these isolates at or below the species cutoff values when compared to representative V. cholerae strains. This suggests these strains may form a basal V. cholerae lineage, or a new species related to V. cholerae.

Nine Divergent Vibrio Genomes Announced

The Boucher Lab and the Centres for Disease Control and Prevention in the US have published the genomes of nine Vibrio species suggested to belong to a unique basal lineage of Vibrio cholerae, or a new closely related novel species to V. cholerae. The genome report, "Draft genome sequences of nine Vibrio sp. isolates from across the United States closely related to Vibrio cholerae" was published in Microbiology Resource Announcements today. These new genomes, based on molecular analyses, are at the cusp of the delineating species cutoffs.

Tracking the Pandemic Generating Lineage of Vibrio cholerae

In collaboration with Dr. Munirul Alam (International Centre for Diarrheal Disease Research, Bangladesh), Yan and Tareq's new review outlining the evolution and dissemination of the pandemic lineage of Vibrio cholerae has been published. "Emergence, ecology and dispersal of the pandemic generating Vibrio cholerae lineage" is now available in the open access journal of International Microbiology.

The initial development of pandemic Vibrio cholerae highlights important gene acquisition events required for the Pandemic Generating (PG) lineage to form before being able to infect human hosts. These preliminary gene acquisition stages drive the evolution of pandemic cholera, up to the seventh and current pandemic El Tor strains. Phylogenetic analysis of PG strains indicate a replacement of El Tor predecessors that is now associated with current cholera outbreaks. Pandemic V. cholerae exists in two life stages, as a pathogen in human hosts and in environmental reservoirs, with ecological factors influencing regions that have annual patterns of infection outbreaks, such as water chemistry and host associations in environmental reservoirs. This review provides insight into the evolutionary path of PG V. cholerae, and its dispersal in the environment outside of human hosts. 

Phylogenetic analysis of pandemic V. cholerae evolution (from Islam et al., 2017)

Evolution of the Type VI Secretion System in Vibrio cholerae

Paul and Yan's new paper, "Sequential displacement of type VI secretion system effector genes leads to evolution of diverse immunity gene arrays in Vibrio cholerae," is out!

In collaboration with Dr. Stefan Pukatzki (University of Colorado Denver), Dr. Daniele Provenzano (University of Texas Rio Grande Valley), and Dr. Daniel Unterweger (University of Oxford), Paul and Yan conducted an extensive study on the role of horizontal gene transfer in shaping the diversity of type VI secretion system (T6SS) loci in V. cholerae and closely related species. They discovered the existence of complex arrays of effector and immunity genes in these loci. These arrays differ even between closely related sympatric strains and appear to be formed by successive horizontal gene transfer events. The resulting accumulation of large numbers of immunity genes potentially enhance the recipient's fitness in T6SS-mediated bactericidal interactions. Additionally, they show how the accumulation of T6SS elements through horizontal gene transfer could have contributed to the evolution of some V. cholerae strains from harmless environmental bacteria to pandemic pathogens.

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Whole-genome phylogeny and T6SS effector-immunity module composition of V. cholerae and closely related species (from Kirchberger et al., 2017)

A Genomic Island in Vibrio cholerae with CRISPR-Cas and T6SS Regions

Yan, Fabini, and Paul collaborated with Dr. Maurizio Labbate (University Technology Sydney) and published a paper in Scientific Reports about a ~28-kb genomic island (GI; designated as GIVchS12) in a non-O1/O139 strain of Vibrio cholerae located in the same position where the Vibrio Pathogenicity Island - 1 (VPI-1) would be as it has VPI-1 site-specific recombination characteristics. VPI-1, which has so far only been found in some O1 and O139 V. cholerae strains (with pathogenic strains causing cholera epidemics and pandemics), contains the toxin-coregulated pilus (tcp) cluster. TCP is a precursor for infection of V. cholerae by the CTX phage (that contains the cholera toxin) as it serves as receptor for the phage. It is also important for the colonization of the small intestine during V. cholerae infection of the host. On the other hand, GIVchS12 does not contain the same genes as VPI-1. It contains CRISPR-Cas and type VI secretion system (T6SS) modules. CRISPR-Cas is a defense mechanism by bacteria against unwanted lateral gene transfer by recognizing foreign DNA and cleaving it. T6SS is for inter-cell anatgonism, where T6SS-harbouring bacteria produce a membrane-spanning protein complex used to puncture and kill nearby eukaryotic or prokaryotic cells.

Our survey of representative V. cholerae genomes suggests the presence of genomic islands similar to GIVchS12 containing CRISPR-Cas and T6SS modules from various strains. Natural populations of V. cholerae can serve as reservoir for diverse GIs such as GIVchS12.

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Schematic representation of V. cholerae S12 genomic island GIVchS12 (from Labbate et al., 2016)

Vibrio cidicii, a Novel Vibrio Species Closely Related to Vibrio navarrensis

The official species description paper of the novel Vibrio species, Vibrio cidicii, is now out. The paper is entitled "Characterization of clinical and environmental isolates of Vibrio cidicii sp. nov., a close relative of Vibrio navarrensis" and published in the International Journal of Systematic and Evolutionary Microbiology. Vibrio cidicii was initially identified by the group of Dr. Cheryl Tarr from the Centers for Disease Control and Prevention (CDC) while studying clinical isolates of V. navarrensis. Thus, the new species was named after the CDC.

Characterization of the isolates involved extensive biochemical and genotypic tests, including whole-genome comparisons with close relatives, V. navarrensis and Vibrio vulnificus. Metabolic profiling found one major phenotypic difference between V. cidicii from its closest relatives, the utilization of L-rhamnose, where the V. cidicii isolates are able to utilize the substrate. Genome comparisons and multilocus sequence analysis (MLSA) show that the V. cidicii genomes to belong to a species different from V. navarrensis and V. vulnificus.

This work was in collaboration with the teams of Dr. Cheryl Tarr (CDC), Dr. I. King Jordan (Georgia Institute of Technology), and Dr. Rebecca Case (Department of Biological Sciences). This is the second novel Vibrio species characterized from the Boucher Lab (Vibrio metoecus being the first one).

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MLSA of housekeeping genes show distinct clustering of V. cidicii from its closest relatives (from Orata et al., 2016)

Genome Sequences of Bacteria in Close Association with Emiliania huxleyi

Yan and Fabini, in collaboration with the lab of Dr. Rebecca Case (also from the Department of Biological Sciences), released two Genome Announcements today on the genome sequences of bacterial strains isolated from the polymicrobial cultures of coccolith-bearing and naked (no coccolith) microalga Emiliania huxleyi. The Case Lab isolated multiple bacteria from these cultures, including strains of Balneola sp., Erythrobacter sp., Jannaschia sp., Marinobacter sp., Roseovarius sp., and Sulfitobacter spp. among others, which form a symbiotic relationship with E. huxleyi. The Boucher Lab sequenced the genomes of these isolates. This is part of an ongoing work with both labs. We are currently working on resolving the taxonomic inconsistencies within the Rhodobacteraceae family, specifically within the Rosobeacter clade, by comparative genomic analysis.

Population Structure of Single-Source Vibrio cholerae

Our lab's newest paper is finally out! Bacteria display a stunning array of diversity, from higher order groups all the way down to the subspecies level of endlessly differing strains. Our main organism of study, Vibrio cholerae, is no different. Only by deeply sampling groups of closely related organisms from the same environment can we begin to understand how this diversity arises – both for V. cholerae and bacteria in general. Using multi locus sequence typing and phylogenomics, we found that the scenic Oyster Pond and Lagoon ecosystem in Woods Hole, Massachusetts is dominated by a handful of distantly related V. cholerae strains, with many more found in much lower numbers, perhaps waiting for the right opportunity to rise to the top. The dominating strains show signs of ecological differentiation, but are likely in strong competition with each other, as evidenced by their different repertoires of type VI secretion system effectors that mediate fatal bacterial interactions. Oh, and some are bioluminescent!

The paper is entitled "A small number of phylogenetically distinct clonal complexes dominate a coastal Vibrio cholerae population" and published in Applied and Environmental Microbiology. The work is a carryover from Yan's postdoc at the Massachusetts Institute of Technology (as evidenced by having Dr. Kathryn Kauffman and Dr. Martin Polz from MIT as co-authors), so it has been a long time in the making!

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Clonal backbone of V. cholerae sampled from different locations around Oyster Pond (from Kirchberger et al., 2016)