Oral microbiomes from hunter-gatherers and traditional farmers reveal shifts in commensal balance and pathogen load linked to diet.

Maladaptation to modern diets has been implicated in several chronic disorders. Given the higher prevalence of disease such as dental caries and chronic gum diseases in industrialized societies, we sought to investigate the impact of different subsistence strategies on oral health and physiology, as documented by the oral microbiome. To control for confounding variables such as environment and host genetics, we sampled saliva from three pairs of populations of hunter-gatherers and traditional farmers living in close proximity in the Philippines. Deep shotgun sequencing of salivary DNA generated high-coverage microbiomes along with human genomes. Comparing these microbiomes with publicly available data from individuals living on a Western diet revealed that abundance ratios of core species were significantly correlated with subsistence strategy, with hunter-gatherers and Westerners occupying either end of a gradient of Neisseria against Haemophilus, and traditional farmers falling in between. Species found preferentially in hunter-gatherers included microbes often considered as oral pathogens, despite their hosts' apparent good oral health. Discriminant analysis of gene functions revealed vitamin B5 autotrophy and urease-mediated pH regulation as candidate adaptations of the microbiome to the hunter-gatherer and Western diets, respectively. These results suggest that major transitions in diet selected for different communities of commensals and likely played a role in the emergence of modern oral pathogens.

Presence of pathogenic Vibrio parahaemolyticus in waters and seafood from the Tunisian Sea.

The occurrence of the hemolysin genes, tdh and trh, in Vibrio parahaemolyticus strains isolated from environmental samples collected from various exported seafood products comprising of fishes and shellfish (Mytilus edulis and Crassostrea gigas) or seawater, was studied. Eight strains were confirmed as V. parahaemolyticus by toxR -based polymerase chain reaction and only one strain out of these 8 strains was positive for tdh and trh genes. Toxigenic V. parahaemolyticus isolates are present in Tunisian coastal areas and they may also be present in Tunisian exported seafood products.

Molecular characterization of enterotoxigenic and borderline oxacillin resistant Staphylococcus strains from ovine milk.

Staphylococcal enterotoxins (SEs) produced by Staphylococcus spp. are superantigens responsible for food-poisoning and are associated to mobile genetic elements such as Staphylococcus aureus pathogenicity islands (SaPI). The presence of 13 enterotoxin genes (sea, seb, sec, sed, see, seg, seh, sei, sej, sel, sek, seq, and tst) was tested in 15 S. aureus and 24 coagulase-negative Staphylococcus (CNS) multi-resistant strains isolated from ovine milk in Sardinia. All CNS isolates were enterotoxin-negative, whereas co-presence of sec, sel and tst was observed in most of the S. aureus strains. One isolate of S. aureus was characterized by tst alone. A multiplex PCR assay aimed at discriminating between the integrase genes of pathogenicity islands SaPI2, SaPIbov1, and SaPIMW2 was developed. We demonstrated that strains harboring sec, sel and tst were associated with SaPIbov1, whereas the strain positive for tst was associated with SaPI2. Borderline oxacillin resistant S. aureus strains were also detected. RAPD analysis of the Staphylococcus strains showed that clonal relationships were correlated with pathogenic profiles.

Comparative ICE genomics: insights into the evolution of the SXT/R391 family of ICEs.

Integrating and conjugative elements (ICEs) are one of the three principal types of self-transmissible mobile genetic elements in bacteria. ICEs, like plasmids, transfer via conjugation; but unlike plasmids and similar to many phages, these elements integrate into and replicate along with the host chromosome. Members of the SXT/R391 family of ICEs have been isolated from several species of gram-negative bacteria, including Vibrio cholerae, the cause of cholera, where they have been important vectors for disseminating genes conferring resistance to antibiotics. Here we developed a plasmid-based system to capture and isolate SXT/R391 ICEs for sequencing. Comparative analyses of the genomes of 13 SXT/R391 ICEs derived from diverse hosts and locations revealed that they contain 52 perfectly syntenic and nearly identical core genes that serve as a scaffold capable of mobilizing an array of variable DNA. Furthermore, selection pressure to maintain ICE mobility appears to have restricted insertions of variable DNA into intergenic sites that do not interrupt core functions. The variable genes confer diverse element-specific phenotypes, such as resistance to antibiotics. Functional analysis of a set of deletion mutants revealed that less than half of the conserved core genes are required for ICE mobility; the functions of most of the dispensable core genes are unknown. Several lines of evidence suggest that there has been extensive recombination between SXT/R391 ICEs, resulting in re-assortment of their respective variable gene content. Furthermore, our analyses suggest that there may be a network of phylogenetic relationships among sequences found in all types of mobile genetic elements.

ICEVchInd5 is prevalent in epidemic Vibrio cholerae O1 El Tor strains isolated in India.

Integrative conjugative elements (ICEs) of the SXT/R391 family are self-transmissible mobile elements mainly involved in antibiotic resistance spread among γ-Proteobacteria, including Vibrio cholerae. We demonstrated that the recently described ICEVchInd5 is prevailing in V. cholerae O1 clinical strains isolated in Wardha province (Maharashtra, India) from 1994 to 2005. Genetic characterization by ribotyping and multiple-locus SSR analysis proved the same clonal origin for V. cholerae O1 isolates in Wardha province over an 11-year period and was used to assess the correlation between strain and ICE content among ours and different Indian reference strains. In silico analysis showed the existence of at least 3 sibling ICEs of ICEVchInd5 in V. cholerae O1 El Tor reference strains, isolated in the Indian subcontinent after 1992.

New V. cholerae atypical El Tor variant emerged during the 2006 epidemic outbreak in Angola.

BACKGROUND: V. cholerae is the etiological agent of cholera, a major public health concern in most developing countries. Virulence of V. cholerae relies on the powerful cholera toxin, encoded by the CTX prophage. The emergence of new pathogenic variants in the recent years has been mostly associated with new CTX prophage rearrangements. RESULTS: In this retrospective study, we show that the epidemic V. cholerae O1 El Tor strain responsible for the 2006 outbreak in Angola is clonally and genetically different from El Tor strains circulating in the 1990s in the same area. Strains from 2006 carry ICEVchAng3 of the SXT/R391 family. This ICE is associated with a narrower multidrug resistance profile compared to the one conferred by plasmid p3iANG to strains of the 1990s. The CTX prophage carried by 2006 El Tor strains is characterized by rstR(ET) and ctxB(Cla) alleles organized in a RS1-RS2-Core array on chromosome I. Interestingly, the newly emerging atypical strain belongs to a clade previously known to comprise only clinical isolates from the Indian subcontinent that also contain the same ICE of the SXT/R391 family. CONCLUSIONS: Our findings remark the appearance of a novel V. cholerae epidemic variant in Africa with a new CTXΦ arrangement previously described only in the Indian Subcontinent.

Recent Asian origin of chytrid fungi causing global amphibian declines.

Globalized infectious diseases are causing species declines worldwide, but their source often remains elusive. We used whole-genome sequencing to solve the spatiotemporal origins of the most devastating panzootic to date, caused by the fungus Batrachochytrium dendrobatidis, a proximate driver of global amphibian declines. We traced the source of B. dendrobatidis to the Korean peninsula, where one lineage, BdASIA-1, exhibits the genetic hallmarks of an ancestral population that seeded the panzootic. We date the emergence of this pathogen to the early 20th century, coinciding with the global expansion of commercial trade in amphibians, and we show that intercontinental transmission is ongoing. Our findings point to East Asia as a geographic hotspot for B. dendrobatidis biodiversity and the original source of these lineages that now parasitize amphibians worldwide.

Acquisition and evolution of SXT-R391 integrative conjugative elements in the seventh-pandemic Vibrio cholerae lineage.

UNLABELLED: SXT-R391 Integrative conjugative elements (ICEs) are self-transmissible mobile genetic elements able to confer multidrug resistance and other adaptive features to bacterial hosts, including Vibrio cholerae, the causative agent of cholera. ICEs are arranged in a mosaic genetic structure composed of a conserved backbone interspersed with variable DNA clusters located in conserved hot spots. In this study, we investigated ICE acquisition and subsequent microevolution in pandemic V. cholerae. Ninety-six ICEs were retrieved from publicly available sequence databases from V. cholerae clinical strains and were compared to a set of reference ICEs. Comparative genomics highlighted the existence of five main ICE groups with a distinct genetic makeup, exemplified by ICEVchInd5, ICEVchMoz10, SXT, ICEVchInd6, and ICEVchBan11. ICEVchInd5 (the most frequent element, represented by 70 of 96 elements analyzed) displayed no sequence rearrangements and was characterized by 46 single nucleotide polymorphisms (SNPs). SNP analysis revealed that recent inter-ICE homologous recombination between ICEVchInd5 and other ICEs circulating in gammaproteobacteria generated ICEVchMoz10, ICEVchInd6, and ICEVchBan11. Bayesian phylogenetic analyses indicated that ICEVchInd5 and SXT were independently acquired by the current pandemic V. cholerae O1 and O139 lineages, respectively, within a period of only a few years. IMPORTANCE: SXT-R391 ICEs have been recognized as key vectors of antibiotic resistance in the seventh-pandemic lineage of V. cholerae, which remains a major cause of mortality and morbidity on a global scale. ICEs were acquired only recently in this clade and are acknowledged to be major contributors to horizontal gene transfer and the acquisition of new traits in bacterial species. We have reconstructed the temporal dynamics of SXT-R391 ICE acquisition and spread and have identified subsequent recombination events generating significant diversity in ICEs currently circulating among V. cholerae clinical strains. Our results showed that acquisition of SXT-R391 ICEs provided the V. cholerae seventh-pandemic lineage not only with a multidrug resistance phenotype but also with a powerful molecular tool for rapidly accessing the pan-genome of a large number of gammaproteobacteria.

A new integrative conjugative element detected in Haitian isolates of Vibrio cholerae non-O1/non-O139.

The presence of SXT/R391-related integrating conjugative elements (ICEs) in Vibrio cholerae O1 and non-O1/non-O139 isolated from clinical and environmental samples in Haiti in 2010 was studied. The main finding of this work was the identification of the novel ICEVchHai2 among closely related V. cholerae non-O1/non-O139 clinical strains. The mosaic structure of this element confirms the role of ICEs as efficient recombination systems whereby new genetic material can be acquired and exchanged, according V. cholerae strains new accessory functions.

Vibrio cholerae O1 epidemic variants in Angola: a retrospective study between 1992 and 2006.

Cholera is still a major public health concern in many African countries. In Angola, after a decade of absence, cholera reemerged in 1987, spreading throughout the country until 1996, with outbreaks recurring in a seasonal pattern. In 2006 Angola was hit by one of the most severe outbreaks of the last decade, with ca. 240,000 cases reported. We analyzed 21 clinical strains isolated between 1992 and 2006 from several provinces throughout the country: Benguela, Bengo, Luanda, Cuando Cubango, and Cabinda. We used two multiplex PCR assays to investigate discriminatory mobile genetic elements (MGE) [Integrative Conjugative Elements (ICEs), VSP-II, GI12, GI14, GI15, K, and TLC phages] and we compared the profiles obtained with those of different reference V. cholerae O1 variants (prototypical, altered, and hybrid), responsible for the ongoing 7th pandemic. We also tested the strains for the presence of specific VSP-II variants and for the presence of a genomic island (GI) (WASA-1), correlated with the transmission of seventh pandemic cholera from Africa to South America. Based on the presence/absence of the analyzed genetic elements, five novel profiles were detected in the epidemic strains circulating in the 1990s. The most frequent profiles, F and G, were characterized by the absence of ICEs and the three GIs tested, and the presence of GI WASA-1 and the WASA variant of the VSP-II island. Our results identified unexpected variability within the 1990s epidemic, showing different rearrangements in a dynamic part of the genome not present in the prototypical V. cholerae O1 N16961. Moreover the 2006 strains differed from the current pandemic V. cholerae O1 strain. Taken together, our results highlight the role of horizontal gene transfer (HGT) in diversifying the genetic background of V. cholerae within a single epidemic.

Rapid detection by multiplex PCR of Genomic Islands, prophages and Integrative Conjugative Elements in V. cholerae 7th pandemic variants.

Vibrio cholerae poses a threat to human health, and new epidemic variants have been reported so far. Seventh pandemic V. cholerae strains are characterized by highly related genomic sequences but can be discriminated by a large set of Genomic Islands, phages and Integrative Conjugative Elements. Classical serotyping and biotyping methods do not easily discriminate among new variants arising worldwide, therefore the establishment of new methods for their identification is required. We developed a multiplex PCR assay for the rapid detection of the major 7th pandemic variants of V. cholerae O1 and O139. Three specific genomic islands (GI-12, GI-14 and GI-15), two phages (Kappa and TLC), Vibrio Seventh Pandemic Island 2 (VSP-II), and the ICEs of the SXT/R391 family were selected as targets of our multiplex PCR based on a comparative genomic approach. The optimization and specificity of the multiplex PCR was assessed on 5 V. cholerae 7th pandemic reference strains, and other 34 V. cholerae strains from various epidemic events were analyzed to validate the reliability of our method. This assay had sufficient specificity to identify twelve different V. cholerae genetic profiles, and therefore has the potential to be used as a rapid screening method.

Genomic analysis of ICEVchBan8: An atypical genetic element in Vibrio cholerae.

Genomic islands (GIs) and integrative conjugative elements (ICEs) are major players in bacterial evolution since they encode genes involved in adaptive functions of medical or environmental importance. Here we performed the genomic analysis of ICEVchBan8, an unusual ICE found in the genome of a clinical non-toxigenic Vibrio cholerae O37 isolate. ICEVchBan8 shares most of its genetic structure with SXT/R391 ICEs. However, this ICE codes for a different integration/excision module is located at a different insertion site, and part of its genetic cargo shows homology to other pathogenicity islands of V. cholerae.