Phylodynamic Analysis of Clinical and Environmental Vibrio cholerae Isolates from Haiti Reveals Diversification Driven by Positive Selection

ABSTRACTPhylodynamic analysis of genome-wide single-nucleotide polymorphism (SNP) data is a powerful tool to investigate underlying evolutionary processes of bacterial epidemics. The method was applied to investigate a collection of 65 clinical and environmental isolates ofVibrio choleraefrom Haiti collected between 2010 and 2012. Characterization of isolates recovered from environmental samples identified a total of four toxigenicV. choleraeO1 isolates, four non-O1/O139 isolates, and a novel nontoxigenicV. choleraeO1 isolate with the classicaltcpAgene. Phylogenies of strains were inferred from genome-wide SNPs using coalescent-based demographic models within a Bayesian framework. A close phylogenetic relationship between clinical and environmental toxigenicV. choleraeO1 strains was observed. As cholera spread throughout Haiti between October 2010 and August 2012, the population size initially increased and then fluctuated over time. Selection analysis along internal branches of the phylogeny showed a steady accumulation of synonymous substitutions and a progressive increase of nonsynonymous substitutions over time, suggesting diversification likely was driven by positive selection. Short-term accumulation of nonsynonymous substitutions driven by selection may have significant implications for virulence, transmission dynamics, and even vaccine efficacy.IMPORTANCECholera, a dehydrating diarrheal disease caused by toxigenic strains of the bacteriumVibrio cholerae, emerged in 2010 in Haiti, a country where there were no available records on cholera over the past 100 years. While devastating in terms of morbidity and mortality, the outbreak provided a unique opportunity to study the evolutionary dynamics ofV. choleraeand its environmental presence. The present study expands on previous work and provides an in-depth phylodynamic analysis inferred from genome-wide single nucleotide polymorphisms of clinical and environmental strains from dispersed geographic settings in Haiti over a 2-year period. Our results indicate that even during such a short time scale,V. choleraein Haiti has undergone evolution and diversification driven by positive selection, which may have implications for understanding the global clinical and epidemiological patterns of the disease. Furthermore, the continued presence of the epidemic strain in Haitian aquatic environments has implications for transmission.

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Clonal Lineages of Erysipelothrix rhusiopathiae Responsible for Acute Swine Erysipelas in Japan Identified by Using Genome-Wide Single-Nucleotide Polymorphism Analysis

Applied and Environmental Microbiology ◽

10.1128/aem.00130-17 ◽

2017 ◽

Vol 83 (11) ◽

Cited By ~ 12

Author(s):

Yohsuke Ogawa ◽

Kazumasa Shiraiwa ◽

Yoshitoshi Ogura ◽

Tadasuke Ooka ◽

Sayaka Nishikawa ◽

...

Keyword(s):

Polymorphism Analysis ◽

Whole Genome ◽

Sequencing Analysis ◽

Erysipelothrix Rhusiopathiae ◽

Single Nucleotide ◽

Content Type ◽

Genome Wide ◽

Wide Range ◽

Swine Erysipelas ◽

Lineage Iv

ABSTRACTErysipelothrix rhusiopathiaecauses swine erysipelas, an important infectious disease in the swine industry. In Japan, the incidence of acute swine erysipelas due toE. rhusiopathiaeserovar 1a has recently increased markedly. To study the genetic relatedness of the strains from the recent cases, we analyzed 34E. rhusiopathiaeserovar 1a swine isolates collected between 1990 and 2011 and further investigated the possible association of the live Koganei 65-0.15 vaccine strain (serovar 1a) with the increase in cases. Pulsed-field gel electrophoresis analysis revealed no marked variation among the isolates; however, sequencing analysis of a hypervariable region in the surface-protective antigen A gene (spaA) revealed that the strains isolated after 2007 exhibited the samespaAgenotype and could be differentiated from older strains. Phylogenetic analysis based on genome-wide single-nucleotide polymorphisms (SNPs) revealed that the Japanese strains examined were closely related, showing a relatively small number of SNPs among them. The strains were classified into four major lineages, with Koganei 65-0.15 (lineage III) being phylogenetically separated from the other three lineages. The strains isolated after 2007 and the two older strains constituted one major lineage (lineage IV) with a specificspaAgenotype (M203/I257-SpaA), while the recent isolates were further divided into two geographic groups. The remaining older isolates belonged to either lineage I, with the I203/L257-SpaA type, or lineage II, with the I203/I257-SpaA type. These results indicate that the recent increased incidence of acute swine erysipelas in Japan is associated with two sublineages of lineage IV, which have independently evolved in two different geographic regions.IMPORTANCEUsing large-scale whole-genome sequence data fromErysipelothrix rhusiopathiaeisolates from a wide range of hosts and geographic origins, a recent study clarified the existence of three distinct clades (clades 1, 2, and 3) that are found across multiple continents and host species, representing both livestock and wildlife, and an “intermediate” clade between clade 2 and the dominant clade 3 within the species. In this study, we found that theE. rhusiopathiaeJapanese strains examined exhibited remarkably low levels of genetic diversity and confirmed that all of the Japanese and Chinese swine isolates examined in this study belong to clonal lineages within the intermediate clade. We report thatspaAgenotyping ofE. rhusiopathiaestrains is a practical alternative to whole-genome sequencing analysis of theE. rhusiopathiaeisolates from eastern Asian countries.

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The coral symbiont Candidatus Aquarickettsia is variably abundant in threatened Caribbean acroporids and transmitted horizontally

10.1101/2021.01.28.428674 ◽

2021 ◽

Author(s):

Lydia J. Baker ◽

Hannah G. Reich ◽

Sheila A. Kitchen ◽

J. Grace Klinges ◽

Hanna R. Koch ◽

...

Keyword(s):

Positive Selection ◽

Horizontal Transmission ◽

Sampling Location ◽

Acropora Cervicornis ◽

Single Nucleotide ◽

Algal Symbiont ◽

Genome Wide ◽

The Us ◽

Variant Analysis

AbstractThe aquatic symbiont “Candidatus Aquarickettsia rohweri” infects a diversity of non-bilaterian metazoan phyla. In the threatened coral Acropora cervicornis, Aquarickettsia proliferates in response to increased nutrient exposure, resulting in suppressed growth and increased disease susceptibility and mortality. This study evaluated the extent, as well as the ecology and evolution of Aquarickettsia infecting the Caribbean corals: Ac. cervicornis and Ac. palmata and their hybrid (‘Ac. prolifera’). The bacterial parasite Aquarickettsia was found in all acroporids, with host and sampling location impacting infection magnitude. Phylogenomic and genome-wide single nucleotide variant analysis found Aquarickettsia clustering by region, not by coral taxon. Fixation analysis suggested within coral colonies, Aquarickettsia are genetically isolated to the extent that reinfection is unlikely. Relative to other Rickettsiales, Aquarickettsia is undergoing positive selection, with Florida populations experiencing greater positive selection relative to the other Caribbean locations. This may be due to Aquarickettsia response to increased nutrient stress in Florida, as indicated by greater in situ replication rates in these corals. Aquarickettsia did not significantly codiversify with either coral animal nor algal symbiont, and qPCR analysis of gametes and juveniles from susceptible coral genotypes indicated absence in early life stages. Thus, despite being an obligate parasite, Aquarickettsia must be horizontally transmitted via coral mucocytes, an unidentified secondary host, or a yet unexplored environmentally mediated mechanism. Importantly, the prevalence of Aquarickettsia in Ac. cervicornis and high abundance in Florida populations suggests that disease mitigation efforts in the US and Caribbean should focus on preventing early infection via horizontal transmission.

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Genome-Wide Identification of Host-Segregating Single-Nucleotide Polymorphisms for Source Attribution of Clinical Campylobacter coli Isolates

Applied and Environmental Microbiology ◽

10.1128/aem.01787-20 ◽

2020 ◽

Vol 86 (24) ◽

Author(s):

Quentin Jehanne ◽

Ben Pascoe ◽

Lucie Bénéjat ◽

Astrid Ducournau ◽

Alice Buissonnière ◽

...

Keyword(s):

Single Nucleotide Polymorphisms ◽

Common Species ◽

Frequency Variation ◽

Common Source ◽

Campylobacter Coli ◽

Nucleotide Polymorphisms ◽

Source Attribution ◽

Single Nucleotide ◽

Content Type ◽

Genome Wide

ABSTRACT Campylobacter is among the most common causes of gastroenteritis worldwide. Campylobacter jejuni and Campylobacter coli are the most common species causing human disease. DNA sequence-based methods for strain characterization have focused largely on C. jejuni, responsible for 80 to 90% of infections, meaning that C. coli epidemiology has lagged behind. Here, we have analyzed the genome of 450 C. coli isolates to determine genetic markers that can discriminate isolates sampled from 3 major reservoir hosts (chickens, cattle, and pigs). These markers then were applied to identify the source of infection of 147 C. coli strains from French clinical cases. Using STRUCTURE software, 259 potential host-segregating markers were revealed by probabilistic characterization of single-nucleotide polymorphism (SNP) frequency variation in strain collections from three different hosts. These SNPs were found in 41 genes or intergenic regions, mostly coding for proteins involved in motility and membrane functions. Source attribution of clinical isolates based on the differential presence of these markers confirmed chickens as the most common source of C. coli infection in France. IMPORTANCE Genome-wide and source attribution studies based on Campylobacter species have shown their importance for the understanding of foodborne infections. Although the use of multilocus sequence typing based on 7 genes from C. jejuni is a powerful method to structure populations, when applied to C. coli, results have not clearly demonstrated its robustness. Therefore, we aim to provide more accurate data based on the identification of single-nucleotide polymorphisms. Results from this study reveal an important number of host-segregating SNPs, found in proteins involved in motility, membrane functions, or DNA repair systems. These findings offer new, interesting opportunities for further study of C. coli adaptation to its environment. Additionally, the results demonstrate that poultry is potentially the main reservoir of C. coli in France.

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Population Structure of KPC-Producing Klebsiella pneumoniae Isolates from Midwestern U.S. Hospitals

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00125-14 ◽

2014 ◽

Vol 58 (8) ◽

pp. 4961-4965 ◽

Cited By ~ 56

Author(s):

Meredith S. Wright ◽

Federico Perez ◽

Lauren Brinkac ◽

Michael R. Jacobs ◽

Keith Kaye ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Capsular Polysaccharide ◽

Sequence Type ◽

Single Nucleotide Variants ◽

Single Nucleotide ◽

Content Type ◽

Strain Diversity ◽

Carbapenem Resistant ◽

Genetic Context ◽

Over Time

ABSTRACTGenome sequencing of carbapenem-resistantKlebsiella pneumoniaeisolates from regional U.S. hospitals was used to characterize strain diversity and theblaKPCgenetic context. A phylogeny based on core single-nucleotide variants (SNVs) supports a division of sequence type 258 (ST258) into two distinct groups. The primary differences between the groups are in the capsular polysaccharide locus (cps) and their plasmid contents. A strict association between clade and KPC variant was found. TheblaKPCgene was found on variants of two plasmid backbones. This study indicates that highly similarK. pneumoniaesubpopulations coexist within the same hospitals over time.

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Core Genome Multilocus Sequence Typing Scheme for High-Resolution Typing of Enterococcus faecium

Journal of Clinical Microbiology ◽

10.1128/jcm.01946-15 ◽

2015 ◽

Vol 53 (12) ◽

pp. 3788-3797 ◽

Cited By ~ 130

Author(s):

Mark de Been ◽

Mette Pinholt ◽

Janetta Top ◽

Stefan Bletz ◽

Alexander Mellmann ◽

...

Keyword(s):

High Resolution ◽

Enterococcus Faecium ◽

Multilocus Sequence Typing ◽

Core Genome ◽

Target Genes ◽

Multidrug Resistant ◽

Single Nucleotide ◽

Content Type ◽

Genome Wide ◽

Computationally Intensive

Enterococcus faecium, a common inhabitant of the human gut, has emerged in the last 2 decades as an important multidrug-resistant nosocomial pathogen. Since the start of the 21st century, multilocus sequence typing (MLST) has been used to study the molecular epidemiology ofE. faecium. However, due to the use of a small number of genes, the resolution of MLST is limited. Whole-genome sequencing (WGS) now allows for high-resolution tracing of outbreaks, but current WGS-based approaches lack standardization, rendering them less suitable for interlaboratory prospective surveillance. To overcome this limitation, we developed a core genome MLST (cgMLST) scheme forE. faecium. cgMLST transfers genome-wide single nucleotide polymorphism (SNP) diversity into a standardized and portable allele numbering system that is far less computationally intensive than SNP-based analysis of WGS data. TheE. faeciumcgMLST scheme was built using 40 genome sequences that represented the diversity of the species. The scheme consists of 1,423 cgMLST target genes. To test the performance of the scheme, we performed WGS analysis of 103 outbreak isolates from five different hospitals in the Netherlands, Denmark, and Germany. The cgMLST scheme performed well in distinguishing between epidemiologically related and unrelated isolates, even between those that had the same sequence type (ST), which denotes the higher discriminatory power of this cgMLST scheme over that of conventional MLST. We also show that in terms of resolution, the performance of theE. faeciumcgMLST scheme is equivalent to that of an SNP-based approach. In conclusion, the cgMLST scheme developed in this study facilitates rapid, standardized, and high-resolution tracing ofE. faeciumoutbreaks.

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Shared Signature of Recent Positive Selection on the TSBP1–BTNL2–HLA-DRA Genes in Five Native Populations from North Borneo

Genome Biology and Evolution ◽

10.1093/gbe/evaa207 ◽

2020 ◽

Vol 12 (12) ◽

pp. 2245-2257

Author(s):

Boon-Peng Hoh ◽

Xiaoxi Zhang ◽

Lian Deng ◽

Kai Yuan ◽

Chee-Wei Yew ◽

...

Keyword(s):

Natural Selection ◽

Positive Selection ◽

Shared Environment ◽

Nucleotide Polymorphisms ◽

Single Nucleotide ◽

Genome Wide ◽

Endemic Malaria ◽

North Borneo ◽

Native Populations ◽

History Of

Abstract North Borneo (NB) is home to more than 40 native populations. These natives are believed to have undergone local adaptation in response to environmental challenges such as the mosquito-abundant tropical rainforest. We attempted to trace the footprints of natural selection from the genomic data of NB native populations using a panel of ∼2.2 million genome-wide single nucleotide polymorphisms. As a result, an ∼13-kb haplotype in the Major Histocompatibility Complex Class II region encompassing candidate genes TSBP1–BTNL2–HLA-DRA was identified to be undergoing natural selection. This putative signature of positive selection is shared among the five NB populations and is estimated to have arisen ∼5.5 thousand years (∼220 generations) ago, which coincides with the period of Austronesian expansion. Owing to the long history of endemic malaria in NB, the putative signature of positive selection is postulated to be driven by Plasmodium parasite infection. The findings of this study imply that despite high levels of genetic differentiation, the NB populations might have experienced similar local genetic adaptation resulting from stresses of the shared environment.

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Single Nucleotide Polymorphisms in Regulator-Encoding Genes Have an Additive Effect on Virulence Gene Expression in a Vibrio cholerae Clinical Isolate

mSphere ◽

10.1128/msphere.00253-16 ◽

2016 ◽

Vol 1 (5) ◽

Cited By ~ 5

Author(s):

Bailey M. Carignan ◽

Kyle D. Brumfield ◽

Mike S. Son

Keyword(s):

Infectious Disease ◽

Disease Progression ◽

Vibrio Cholerae ◽

Mortality Rates ◽

Watery Diarrhea ◽

Nucleotide Polymorphisms ◽

Single Nucleotide ◽

Content Type ◽

Virulent Strains ◽

Acute Watery Diarrhea

ABSTRACT Cholera, an infectious disease of the small intestine caused by the aquatic bacterium Vibrio cholerae, often results in vomiting and acute watery diarrhea. If left untreated or if the response is too slow, the symptoms can quickly lead to extreme dehydration and ultimately death of the patient. Recent anecdotal evidence of cholera patients suffering from increasingly severe symptoms and of disease progression at a much higher rate than previously observed has emerged. As recent cholera outbreaks caused by increasingly virulent strains have resulted in higher mortality rates, the need to investigate the mechanism(s) allowing this observed increased virulence is apparent. The significance of our research is in identifying the mechanism for increased virulence capabilities, which will allow the development of a model that will greatly enhance our understanding of cholera disease and V. cholerae pathogenesis, leading to broader biomedical impacts, as cholera serves as a model for other enteric diarrheal diseases. Vibrio cholerae is the etiological agent of the infectious disease cholera, which is characterized by vomiting and severe watery diarrhea. Recently, V. cholerae clinical isolates have demonstrated increased virulence capabilities, causing more severe symptoms with a much higher rate of disease progression than previously observed. We have identified single nucleotide polymorphisms (SNPs) in four virulence-regulatory genes (hapR, hns, luxO, and vieA) of a hypervirulent V. cholerae clinical isolate, MQ1795. Herein, all SNPs and SNP combinations of interest were introduced into the prototypical El Tor reference strain N16961, and the effects on the production of numerous virulence-related factors, including cholera toxin (CT), the toxin-coregulated pilus (TCP), and ToxT, were analyzed. Our data show that triple-SNP (hapR hns luxO and hns luxO vieA) and quadruple-SNP combinations produced the greatest increases in CT, TCP, and ToxT production. The hns and hns luxO SNP combinations were sufficient for increased TCP and ToxT production. Notably, the hns luxO vieA triple-SNP combination strain produced TCP and ToxT levels similar to those of MQ1795. Certain SNP combinations (hapR and hapR vieA) had the opposite effect on CT, TCP, and ToxT expression. Interestingly, the hns vieA double-SNP combination strain increased TCP production while decreasing CT production. Our findings suggest that SNPs identified in the four regulatory genes, in various combinations, are associated with increased virulence capabilities observed in V. cholerae clinical isolates. These studies provide insight into the evolution of highly virulent strains. IMPORTANCE Cholera, an infectious disease of the small intestine caused by the aquatic bacterium Vibrio cholerae, often results in vomiting and acute watery diarrhea. If left untreated or if the response is too slow, the symptoms can quickly lead to extreme dehydration and ultimately death of the patient. Recent anecdotal evidence of cholera patients suffering from increasingly severe symptoms and of disease progression at a much higher rate than previously observed has emerged. As recent cholera outbreaks caused by increasingly virulent strains have resulted in higher mortality rates, the need to investigate the mechanism(s) allowing this observed increased virulence is apparent. The significance of our research is in identifying the mechanism for increased virulence capabilities, which will allow the development of a model that will greatly enhance our understanding of cholera disease and V. cholerae pathogenesis, leading to broader biomedical impacts, as cholera serves as a model for other enteric diarrheal diseases.

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The NorR Regulon Is Critical for Vibrio cholerae Resistance to Nitric Oxide and Sustained Colonization of the Intestines

mBio ◽

10.1128/mbio.00013-12 ◽

2012 ◽

Vol 3 (2) ◽

Cited By ~ 43

Author(s):

Andrew M. Stern ◽

Amanda J. Hay ◽

Zhi Liu ◽

Fiona A. Desland ◽

Juan Zhang ◽

...

Keyword(s):

Nitric Oxide ◽

Mouse Model ◽

Vibrio Cholerae ◽

Critical Role ◽

Regulatory Protein ◽

Global Public Health ◽

Content Type ◽

Time Periods ◽

Microaerobic Conditions ◽

Over Time

ABSTRACT Vibrio cholerae, the cause of an often fatal infectious diarrhea, remains a large global public health threat. Little is known about the challenges V. cholerae encounters during colonization of the intestines, which genes are important for overcoming these challenges, and how these genes are regulated. In this study, we examined the V. cholerae response to nitric oxide (NO), an antibacterial molecule derived during infection from various sources, including host inducible NO synthase (iNOS). We demonstrate that the regulatory protein NorR regulates the expression of NO detoxification genes hmpA and nnrS, and that all three are critical for resisting low levels of NO stress under microaerobic conditions in vitro. We also show that prxA, a gene previously thought to be important for NO detoxification, plays no role in NO resistance under microaerobic conditions and is upregulated by H2O2, not NO. Furthermore, in an adult mouse model of prolonged colonization, hmpA and norR were important for the resistance of both iNOS- and non-iNOS-derived stresses. Our data demonstrate that NO detoxification systems play a critical role in the survival of V. cholerae under microaerobic conditions resembling those of an infectious setting and during colonization of the intestines over time periods similar to that of an actual V. cholerae infection. IMPORTANCE Little is known about what environmental stresses Vibrio cholerae, the etiologic agent of cholera, encounters during infection, and even less is known about how V. cholerae senses and counters these stresses. Most prior studies of V. cholerae infection relied on the 24-h infant mouse model, which does not allow the analysis of survival over time periods comparable to that of an actual V. cholerae infection. In this study, we used a sustained mouse colonization model to identify nitric oxide resistance as a function critical for the survival of V. cholerae in the intestines and further identified the genes responsible for sensing and detoxifying this stress.

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Vibrio cholerae Virulence Activator ToxR Regulates Manganese Transport and Resistance to Reactive Oxygen Species

Infection and Immunity ◽

10.1128/iai.00944-19 ◽

2019 ◽

Vol 88 (3) ◽

Author(s):

Hang-hang Jiang ◽

Yitian Zhou ◽

Ming Liu ◽

Jessie Larios-Valencia ◽

Zachariah Lee ◽

...

Keyword(s):

Reactive Oxygen Species ◽

Vibrio Cholerae ◽

Reactive Oxygen ◽

Oxygen Species ◽

Content Type ◽

Genome Wide ◽

A Genome ◽

Its Gene ◽

Virulence Regulator ◽

Manganese Transport

ABSTRACT Like many other pathogens, Vibrio cholerae, the causative agent of cholera, can modulate its gene expression to combat stresses encountered in both aquatic and host environments, including stress posed by reactive oxygen species (ROS). We previously reported that the virulence activator AphB in V. cholerae is involved in ROS resistance. In this study, we found that another key virulence regulator, ToxR, was important for V. cholerae resistance to hydrogen peroxide. Through a genome-wide transposon screen, we discovered that a deletion in mneA, which encodes a manganese exporter, restored ROS resistance of the toxR mutant. We then showed that ToxR did not affect mneA transcription but that the ToxR-regulated major porin OmpU was critical for ROS resistance. The addition of manganese in culture medium restored ROS resistance in both the toxR and ompU mutants. Furthermore, elemental analysis indicated that the intracellular concentration of manganese in both the toxR and ompU mutants was reduced. This may result in intracellular ROS accumulation in these mutants. Our data suggest that ToxR plays an important role in the resistance to reactive oxygen species through the regulation of manganese transport.

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Vibrio cholerae Represses Polysaccharide Synthesis To Promote Motility in Mucosa

Infection and Immunity ◽

10.1128/iai.02841-14 ◽

2015 ◽

Vol 83 (3) ◽

pp. 1114-1121 ◽

Cited By ~ 18

Author(s):

Zhenyu Liu ◽

Yuning Wang ◽

Shengyan Liu ◽

Ying Sheng ◽

Karl-Gustav Rueggeberg ◽

...

Keyword(s):

Vibrio Cholerae ◽

Epithelial Surface ◽

Semisolid Medium ◽

Content Type ◽

Infant Mouse ◽

Genome Wide ◽

Polysaccharide Synthesis ◽

A Genome ◽

Rapid Spread ◽

Colonization Model

The viscoelastic mucus layer of gastrointestinal tracts is a host defense barrier that a successful enteric pathogen, such asVibrio cholerae, must circumvent.V. cholerae, the causative agent of cholera, is able to penetrate the mucosa and colonize the epithelial surface of the small intestine. In this study, we found that mucin, the major component of mucus, promotedV. choleraemovement on semisolid medium and in liquid medium. A genome-wide screen revealed thatVibriopolysaccharide (VPS) production was inversely correlated with mucin-enhanced motility. Mucin adhesion assays indicated that VPS bound to mucin. Moreover, we found thatvpsexpression was reduced upon exposure to mucin. In an infant mouse colonization model, mutants that overexpressed VPS colonized less effectively than wild-type strains in more distal intestinal regions. These results suggest thatV. choleraeis able to sense mucosal signals and modulatevpsexpression accordingly so as to promote fast motion in mucus, thus allowing for rapid spread throughout the intestines.

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