July 19, 2019  |  

Evolutionary dynamics of Vibrio cholerae O1 following a single-source introduction to Haiti.

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.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.


July 19, 2019  |  

Evolution of hypervirulence by a MRSA clone through acquisition of a transposable element.

Staphylococcus aureus has evolved as a pathogen that causes a range of diseases in humans. There are two dominant modes of evolution thought to explain most of the virulence differences between strains. First, virulence genes may be acquired from other organisms. Second, mutations may cause changes in the regulation and expression of genes. Here we describe an evolutionary event in which transposition of an IS element has a direct impact on virulence gene regulation resulting in hypervirulence. Whole-genome analysis of a methicillin-resistant S. aureus (MRSA) strain USA500 revealed acquisition of a transposable element (IS256) that is absent from close relatives of this strain. Of the multiple copies of IS256 found in the USA500 genome, one was inserted in the promoter sequence of repressor of toxins (Rot), a master transcriptional regulator responsible for the expression of virulence factors in S. aureus. We show that insertion into the rot promoter by IS256 results in the derepression of cytotoxin expression and increased virulence. Taken together, this work provides new insight into evolutionary strategies by which S. aureus is able to modify its virulence properties and demonstrates a novel mechanism by which horizontal gene transfer directly impacts virulence through altering toxin regulation. © 2014 John Wiley & Sons Ltd.


July 19, 2019  |  

Insertion sequence IS26 reorganizes plasmids in clinically isolated multidrug-resistant bacteria by replicative transposition.

Carbapenemase-producing Enterobacteriaceae (CPE), which are resistant to most or all known antibiotics, constitute a global threat to public health. Transposable elements are often associated with antibiotic resistance determinants, suggesting a role in the emergence of resistance. One insertion sequence, IS26, is frequently associated with resistance determinants, but its role remains unclear. We have analyzed the genomic contexts of 70 IS26 copies in several clinical and surveillance CPE isolates from the National Institutes of Health Clinical Center. We used target site duplications and their patterns as guides and found that a large fraction of plasmid reorganizations result from IS26 replicative transpositions, including replicon fusions, DNA inversions, and deletions. Replicative transposition could also be inferred for transposon Tn4401, which harbors the carbapenemase blaKPC gene. Thus, replicative transposition is important in the ongoing reorganization of plasmids carrying multidrug-resistant determinants, an observation that carries substantial clinical and epidemiological implications for understanding how such extreme drug resistance phenotypes evolve.Although IS26 is frequently reported to reside in resistance plasmids of clinical isolates, the characteristic hallmark of transposition, target site duplication (TSD), is generally not observed, raising questions about the mode of transposition for IS26. The previous observation of cointegrate formation during transposition implies that IS26 transposes via a replicative mechanism. The other possible outcome of replicative transposition is DNA inversion or deletion, when transposition occurs intramolecularly, and this would also generate a specific TSD pattern that might also serve as supporting evidence for the transposition mechanism. The numerous examples we present here demonstrate that replicative transposition, used by many mobile elements (including IS26 and Tn4401), is prevalent in the plasmids of clinical isolates and results in significant plasmid reorganization. This study also provides a method to trace the evolution of resistance plasmids based on TSD patterns. Copyright © 2015 He et al.


July 7, 2019  |  

Emergence of scarlet fever Streptococcus pyogenes emm12 clones in Hong Kong is associated with toxin acquisition and multidrug resistance.

A scarlet fever outbreak began in mainland China and Hong Kong in 2011 (refs. 1-6). Macrolide- and tetracycline-resistant Streptococcus pyogenes emm12 isolates represent the majority of clinical cases. Recently, we identified two mobile genetic elements that were closely associated with emm12 outbreak isolates: the integrative and conjugative element ICE-emm12, encoding genes for tetracycline and macrolide resistance, and prophage FHKU.vir, encoding the superantigens SSA and SpeC, as well as the DNase Spd1 (ref. 4). Here we sequenced the genomes of 141 emm12 isolates, including 132 isolated in Hong Kong between 2005 and 2011. We found that the introduction of several ICE-emm12 variants, FHKU.vir and a new prophage, FHKU.ssa, occurred in three distinct emm12 lineages late in the twentieth century. Acquisition of ssa and transposable elements encoding multidrug resistance genes triggered the expansion of scarlet fever-associated emm12 lineages in Hong Kong. The occurrence of multidrug-resistant ssa-harboring scarlet fever strains should prompt heightened surveillance within China and abroad for the dissemination of these mobile genetic elements.


July 7, 2019  |  

Sequence type 1 group B Streptococcus, an emerging cause of invasive disease in adults, evolves by small genetic changes.

The molecular mechanisms underlying pathogen emergence in humans is a critical but poorly understood area of microbiologic investigation. Serotype V group B Streptococcus (GBS) was first isolated from humans in 1975, and rates of invasive serotype V GBS disease significantly increased starting in the early 1990s. We found that 210 of 229 serotype V GBS strains (92%) isolated from the bloodstream of nonpregnant adults in the United States and Canada between 1992 and 2013 were multilocus sequence type (ST) 1. Elucidation of the complete genome of a 1992 ST-1 strain revealed that this strain had the highest homology with a GBS strain causing cow mastitis and that the 1992 ST-1 strain differed from serotype V strains isolated in the late 1970s by acquisition of cell surface proteins and antimicrobial resistance determinants. Whole-genome comparison of 202 invasive ST-1 strains detected significant recombination in only eight strains. The remaining 194 strains differed by an average of 97 SNPs. Phylogenetic analysis revealed a temporally dependent mode of genetic diversification consistent with the emergence in the 1990s of ST-1 GBS as major agents of human disease. Thirty-one loci were identified as being under positive selective pressure, and mutations at loci encoding polysaccharide capsule production proteins, regulators of pilus expression, and two-component gene regulatory systems were shown to affect the bacterial phenotype. These data reveal that phenotypic diversity among ST-1 GBS is mainly driven by small genetic changes rather than extensive recombination, thereby extending knowledge into how pathogens adapt to humans.


July 7, 2019  |  

Novel recA-independent horizontal gene transfer in Escherichia coli K-12.

In bacteria, mechanisms that incorporate DNA into a genome without strand-transfer proteins such as RecA play a major role in generating novelty by horizontal gene transfer. We describe a new illegitimate recombination event in Escherichia coli K-12: RecA-independent homologous replacements, with very large (megabase-length) donor patches replacing recipient DNA. A previously uncharacterized gene (yjiP) increases the frequency of RecA-independent replacement recombination. To show this, we used conjugal DNA transfer, combining a classical conjugation donor, HfrH, with modern genome engineering methods and whole genome sequencing analysis to enable interrogation of genetic dependence of integration mechanisms and characterization of recombination products. As in classical experiments, genomic DNA transfer begins at a unique position in the donor, entering the recipient via conjugation; antibiotic resistance markers are then used to select recombinant progeny. Different configurations of this system were used to compare known mechanisms for stable DNA incorporation, including homologous recombination, F’-plasmid formation, and genome duplication. A genome island of interest known as the immigration control region was specifically replaced in a minority of recombinants, at a frequency of 3 X 10-12 CFU/recipient per hour.


July 7, 2019  |  

Common cell shape evolution of two nasopharyngeal pathogens.

Respiratory infectious diseases are the third cause of worldwide death. The nasopharynx is the portal of entry and the ecological niche of many microorganisms, of which some are pathogenic to humans, such as Neisseria meningitidis and Moraxella catarrhalis. These microbes possess several surface structures that interact with the actors of the innate immune system. In our attempt to understand the past evolution of these bacteria and their adaption to the nasopharynx, we first studied differences in cell wall structure, one of the strongest immune-modulators. We were able to show that a modification of peptidoglycan (PG) composition (increased proportion of pentapeptides) and a cell shape change from rod to cocci had been selected for along the past evolution of N. meningitidis. Using genomic comparison across species, we correlated the emergence of the new cell shape (cocci) with the deletion, from the genome of N. meningitidis ancestor, of only one gene: yacF. Moreover, the reconstruction of this genetic deletion in a bacterium harboring the ancestral version of the locus together with the analysis of the PG structure, suggest that this gene is coordinating the transition from cell elongation to cell division. Accompanying the loss of yacF, the elongation machinery was also lost by several of the descendants leading to the change in the PG structure observed in N. meningitidis. Finally, the same evolution was observed for the ancestor of M. catarrhalis. This suggests a strong selection of these genetic events during the colonization of the nasopharynx. This selection may have been forced by the requirement of evolving permissive interaction with the immune system, the need to reduce the cellular surface exposed to immune attacks without reducing the intracellular storage capacity, or the necessity to better compete for adhesion to target cells.


July 7, 2019  |  

Genomic reconnaissance of clinical isolates of emerging human pathogen Mycobacterium abscessus reveals high evolutionary potential.

Mycobacterium abscessus (Ma) is an emerging human pathogen that causes both soft tissue infections and systemic disease. We present the first comparative whole-genome study of Ma strains isolated from patients of wide geographical origin. We found a high proportion of accessory strain-specific genes indicating an open, non-conservative pan-genome structure, and clear evidence of rapid phage-mediated evolution. Although we found fewer virulence factors in Ma compared to M. tuberculosis, our data indicated that Ma evolves rapidly and therefore should be monitored closely for the acquisition of more pathogenic traits. This comparative study provides a better understanding of Ma and forms the basis for future functional work on this important pathogen.


July 7, 2019  |  

Genetic diversity of O-antigens in Hafnia alvei and the development of a suspension array for serotype detection.

Hafnia alvei is a facultative and rod-shaped gram-negative bacterium that belongs to the Enterobacteriaceae family. Although it has been more than 50 years since the genus was identified, very little is known about variations among Hafnia species. Diversity in O-antigens (O-polysaccharide, OPS) is thought to be a major factor in bacterial adaptation to different hosts and situations and variability in the environment. Antigenic variation is also an important factor in pathogenicity that has been used to define clones within a number of species. The genes that are required to synthesize OPS are always clustered within the bacterial chromosome. A serotyping scheme including 39 O-serotypes has been proposed for H. alvei, but it has not been correlated with known OPS structures, and no previous report has described the genetic features of OPS. In this study, we obtained the genome sequences of 21 H. alvei strains (as defined by previous immunochemical studies) with different lipopolysaccharides. This is the first study to show that the O-antigen gene cluster in H. alvei is located between mpo and gnd in the chromosome. All 21 of the OPS gene clusters contain both the wzx gene and the wzy gene and display a large number of polymorphisms. We developed an O serotype-specific wzy-based suspension array to detect all 21 of the distinct OPS forms we identified in H. alvei. To the best of our knowledge, this is the first report to identify the genetic features of H. alvei antigenic variation and to develop a molecular technique to identify and classify different serotypes.


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