The order Methylococcales constitutes the methanotrophs – bacteria that can metabolize methane, a potent greenhouse gas, as their sole source of energy. These bacteria are significant players in the global carbon cycle and can produce value-added products from methane, such as biopolymers, biofuels, and single-cell proteins for animal feed, among others. Previous studies using single-gene phylogenies have shown inconsistencies in the currently established taxonomic structure of this group. This study aimed to determine and resolve these issues by using whole-genome sequence analyses. Phylogenomic analysis and the use of similarity indexes for genomic comparisons – average amino acid identity, digital DNA–DNA hybridization (dDDH), and average nucleotide identity (ANI) – were performed on 91 Methylococcales genomes. Results suggest the reclassification of members at the genus and species levels. Firstly, to resolve polyphyly of the genus Methylomicrobium, Methylomicrobium alcaliphilum, “Methylomicrobium buryatense,” Methylomicrobium japanense, Methylomicrobium kenyense, and Methylomicrobium pelagicum are reclassified to a newly proposed genus, Methylotuvimicrobium gen. nov.; they are therefore renamed to Methylotuvimicrobium alcaliphilum comb. nov., “Methylotuvimicrobium buryatense” comb. nov., Methylotuvimicrobium japanense comb. nov., Methylotuvimicrobium kenyense comb. nov., and Methylotuvimicrobium pelagicum comb. nov., respectively. Secondly, due to the phylogenetic affinity and phenotypic similarities of Methylosarcina lacus with Methylomicrobium agile and Methylomicrobium album, the reclassification of the former species to Methylomicrobium lacus comb. nov. is proposed. Thirdly, using established same-species delineation thresholds (70% dDDH and 95% ANI), Methylobacter whittenburyi is proposed to be a later heterotypic synonym of Methylobacter marinus (89% dDDH and 99% ANI). Also, the effectively but not validly published “Methylomonas denitrificans” was identified as Methylomonas methanica (92% dDDH and 100% ANI), indicating that the former is a later heterotypic synonym of the latter. Lastly, strains MC09, R-45363, and R-45371, currently identified as M. methanica, each represent a putative novel species of the genus Methylomonas (21–35% dDDH and 74–88% ANI against M. methanica) and were reclassified as Methylomonas sp. strains. It is imperative to resolve taxonomic inconsistencies within this group, first and foremost, to avoid confusion with ecological and evolutionary interpretations in subsequent studies.

Prior to the epidemic that emerged in Haiti in October of 2010, cholera had not been documented in this country. After its introduction, a strain of Vibrio cholerae O1 spread rapidly throughout Haiti, where it caused over 600,000 cases of disease and >7,500 deaths in the first two years of the epidemic. We applied whole-genome sequencing to a temporal series of V. cholerae isolates from Haiti to gain insight into the mode and tempo of evolution in this isolated population of V. cholerae O1. Phylogenetic and Bayesian analyses supported the hypothesis that all isolates in the sample set diverged from a common ancestor within a time frame that is consistent with epidemiological observations. A pangenome analysis showed nearly homogeneous genomic content, with no evidence of gene acquisition among Haiti isolates. Nine nearly closed genomes assembled from continuous-long-read data showed evidence of genome rearrangements and supported the observation of no gene acquisition among isolates. Thus, intrinsic mutational processes can account for virtually all of the observed genetic polymorphism, with no demonstrable contribution from horizontal gene transfer (HGT). Consistent with this, the 12 Haiti isolates tested by laboratory HGT assays were severely impaired for transformation, although unlike previously characterized noncompetent V. cholerae isolates, each expressed hapR and possessed a functional quorum-sensing system. Continued monitoring of V. cholerae in Haiti will illuminate the processes influencing the origin and fate of genome variants, which will facilitate interpretation of genetic variation in future epidemics. IMPORTANCE Vibrio cholerae is the cause of substantial morbidity and mortality worldwide, with over three million cases of disease each year. An understanding of the mode and rate of evolutionary change is critical for proper interpretation of genome sequence data and attribution of outbreak sources. The Haiti epidemic provides an unprecedented opportunity to study an isolated, single-source outbreak of Vibrio cholerae O1 over an established time frame. By using multiple approaches to assay genetic variation, we found no evidence that the Haiti strain has acquired any genes by horizontal gene transfer, an observation that led us to discover that it is also poorly transformable. We have found no evidence that environmental strains have played a role in the evolution of the outbreak strain.

Cholera is a devastating diarrhoeal disease caused by certain strains of serogroup O1/O139 Vibrio cholerae. Mobile genetic elements such as genomic islands (GIs) have been pivotal in the evolution of O1/O139 V. cholerae. Perhaps the most important GI involved in cholera disease is the V. cholerae pathogenicity island 1 (VPI-1). This GI contains the toxin-coregulated pilus (TCP) gene cluster that is necessary for colonization of the human intestine as well as being the receptor for infection by the cholera-toxin bearing CTX phage. In this study, we report a GI (designated GIVchS12) from a non-O1/O139 strain of V. cholerae that is present in the same chromosomal location as VPI-1, contains an integrase gene with 94% nucleotide and 100% protein identity to the VPI-1 integrase, and attachment (att) sites 100% identical to those found in VPI-1. However, instead of TCP and the other accessory genes present in VPI-1, GIVchS12 contains a CRISPR-Cas element and a type VI secretion system (T6SS). GIs similar to GIVchS12 were identified in other V. cholerae genomes, also containing CRISPR-Cas elements and/or T6SS’s. This study highlights the diversity of GIs circulating in natural V. cholerae populations and identifies GIs with VPI-1 recombination characteristics as a propagator of CRISPR-Cas and T6SS modules.

Vibrio metoecus is the closest relative of Vibrio cholerae , the causative agent of the potent diarrheal disease cholera. Although the pathogenic potential of this new species is yet to be studied in depth, it has been co-isolated with V. cholerae in coastal waters and found in clinical specimens in the United States. We used these two organisms to investigate the genetic interaction between closely related species in their natural environment. The genomes of 20 V. cholerae and 4 V. metoecus strains isolated from a brackish coastal pond on the US east coast, as well as 4 clinical V. metoecus strains were sequenced and compared with reference strains. Whole genome comparison shows 86–87% average nucleotide identity (ANI) in their core genes between the two species. On the other hand, the chromosomal integron, which occupies approximately 3% of their genomes, shows higher conservation in ANI between species than any other region of their genomes. The ANI of 93–94% observed in this region is not significantly greater within than between species, meaning that it does not follow species boundaries. Vibrio metoecus does not encode toxigenic V. cholerae major virulence factors, the cholera toxin and toxin-coregulated pilus. However, some of the pathogenicity islands found in pandemic V. cholerae were either present in the common ancestor it shares with V. metoecus , or acquired by clinical and environmental V. metoecus in partial fragments. The virulence factors of V. cholerae are therefore both more ancient and more widespread than previously believed. There is high interspecies recombination in the core genome, which has been detected in 24% of the single-copy core genes, including genes involved in pathogenicity. Vibrio metoecus was six times more often the recipient of DNA from V. cholerae as it was the donor, indicating a strong bias in the direction of gene transfer in the environment.

Marine microbes form host-associated biofilm communities that are shaped by complex interactions between bacteria and their host. The roseobacter Phaeobacter inhibens exploits both symbiotic and pathogenic niches while interacting with its microalgal host Emiliania huxleyi. During co-cultivation over extended periods with E. huxleyi, we show that P. inhibens selectively kills two host cell types, the diploid calcifying strain and the haploid flagellated strain. Meanwhile, various non-calcifying diploid strains are resistant to this pathogen or the pathogen is avirulent to this cell type. This differential pathogenesis has the potential of dramatically altering the composition of E. huxleyi blooms, which are typically dominated by the roseobacter-susceptible calcifying strain. This cell type makes calcite plates, which are an important sink in the marine carbon cycle and forms part of the marine paleobotanic record. P. inhibens kills the haploid cells, which have been proposed as critical to the survival of the algae, as they readily escape both eukaryotic predation and viral infection. Consequently, bacteria such as P. inhibens could influence E. huxleyi’s life history by selective pathogenesis, thereby altering the composition of cell types within its populations and its bloom-bust lifestyle.