Horizontal DNA transfer mechanisms of bacteria as weapons of intragenomic conflict

Horizontal DNA transfer (HDT) is a pervasive mechanism of diversification in many microbial species, but its primary evolutionary role remains controversial. Much recent research has emphasised the adaptive benefit of acquiring novel DNA, but here we argue instead that intragenomic conflict provides a coherent framework for understanding the evolutionary origins of HDT. To test this hypothesis, we developed a mathematical model of a clonally descended bacterial population undergoing HDT through transmission of mobile genetic elements (MGEs) and genetic transformation. Including the known bias of transformation toward the acquisition of shorter alleles into the model suggested it could be an effective means of counteracting the spread of MGEs. Both constitutive and transient competence for transformation were found to provide an effective defence against parasitic MGEs; transient competence could also be effective at permitting the selective spread of MGEs conferring a benefit on their host bacterium. The coordination of transient competence with cell-cell killing, observed in multiple species, was found to result in synergistic blocking of MGE transmission through releasing genomic DNA for homologous recombination while simultaneously reducing horizontal MGE spread by lowering the local cell density. To evaluate the feasibility of the functions suggested by the modelling analysis, we analysed genomic data from longitudinal sampling of individuals carrying Streptococcus pneumoniae. This revealed the frequent within-host coexistence of clonally descended cells that differed in their MGE infection status, a necessary condition for the proposed mechanism to operate. Additionally, we found multiple examples of MGEs inhibiting transformation through integrative disruption of genes encoding the competence machinery across many species, providing evidence of an ongoing "arms race." Reduced rates of transformation have also been observed in cells infected by MGEs that reduce the concentration of extracellular DNA through secretion of DNases. Simulations predicted that either mechanism of limiting transformation would benefit individual MGEs, but also that this tactic's effectiveness was limited by competition with other MGEs coinfecting the same cell. A further observed behaviour we hypothesised to reduce elimination by transformation was MGE activation when cells become competent. Our model predicted that this response was effective at counteracting transformation independently of competing MGEs. Therefore, this framework is able to explain both common properties of MGEs, and the seemingly paradoxical bacterial behaviours of transformation and cell-cell killing within clonally related populations, as the consequences of intragenomic conflict between self-replicating chromosomes and parasitic MGEs. The antagonistic nature of the different mechanisms of HDT over short timescales means their contribution to bacterial evolution is likely to be substantially greater than previously appreciated

Dominant Role of Nucleotide Substitution in the Diversification of Serotype 3 Pneumococci over Decades and during a Single Infection

Streptococcus pneumoniae of serotype 3 possess a mucoid capsule and cause disease associated with high mortality rates relative to other pneumococci. Phylogenetic analysis of a complete reference genome and 81 draft sequences from clonal complex 180, the predominant serotype 3 clone in much of the world, found most sampled isolates belonged to a clade affected by few diversifying recombinations. However, other isolates indicate significant genetic variation has accumulated over the clonal complex’s entire history. Two closely related genomes, one from the blood and another from the cerebrospinal fluid, were obtained from a patient with meningitis. The pair differed in their behaviour in a mouse model of disease and in their susceptibility to antimicrobials, with at least some of these changes attributable to a mutation that upregulated the patAB efflux pump. This indicates clinically important phenotypic variation can accumulate rapidly through small alterations to the genotype

Transformation asymmetry and the evolution of the bacterial accessory genome

Bacterial transformation can insert or delete genomic islands (GIs), depending on the donor and recipient genotypes, if an homologous recombination spans the GI’s integration site and includes sufficiently long flanking homologous arms. Combining mathematical models of recombination with experiments using pneumococci found GI insertion rates declined geometrically with the GI’s size. The decrease in acquisition frequency with length (1.08x10−3 bp−1) was higher than a previous estimate of the analogous rate at which core genome recombinations terminated. Although most efficient for shorter GIs, transformation-mediated deletion frequencies did not vary consistently with GI length, with removal of 10 kb GIs approximately 50% as efficient as acquisition of base substitutions. Fragments of two kilobases, typical of transformation event sizes, could drive all these deletions independent of island length. The strong asymmetry of transformation, and its capacity to efficiently remove GIs, suggests non-mobile accessory loci will decline in frequency without preservation by selection

Diverse regulatory pathways modulate bet hedging of competence induction in epigenetically-differentiated phase variants of Streptococcus pneumoniae

Despite enabling Streptococcus pneumoniae to acquire antibiotic resistance and evade vaccine-induced immunity, transformation occurs at variable rates across pneumococci. Phase variants of isolate RMV7, distinguished by altered methylation patterns driven by the translocating variable restriction-modification (tvr) locus, differed significantly in their transformation efficiencies and biofilm thicknesses. These differences were replicated when the corresponding tvr alleles were introduced into an RMV7 derivative lacking the locus. RNA-seq identified differential expression of the type 1 pilus, causing the variation in biofilm formation, and inhibition of competence induction in the less transformable variant, RMV7domi. This was partly attributable to RMV7domi's lower expression of ManLMN, which promoted competence induction through importing N-acetylglucosamine. This effect was potentiated by analogues of some proteobacterial competence regulatory machinery. Additionally, one of RMV7domi's phage-related chromosomal island was relatively active, which inhibited transformation by increasing expression of the stress response proteins ClpP and HrcA. However, HrcA increased competence induction in the other variant, with its effects depending on Ca2+ supplementation and heat shock. Hence the heterogeneity in transformation efficiency likely reflects the diverse signalling pathways by which it is affected. This regulatory complexity will modulate population-wide responses to synchronising quorum sensing signals to produce co-ordinated yet stochastic bet hedging behaviour.Graphical Abstrac

Bayesian inference of ancestral dates on bacterial phylogenetic trees

The sequencing and comparative analysis of a collection of bacterial genomes from a single species or lineage of interest can lead to key insights into its evolution, ecology or epidemiology. The tool of choice for such a study is often to build a phylogenetic tree, and more specifically when possible a dated phylogeny, in which the dates of all common ancestors are estimated. Here, we propose a new Bayesian methodology to construct dated phylogenies which is specifically designed for bacterial genomics. Unlike previous Bayesian methods aimed at building dated phylogenies, we consider that the phylogenetic relationships between the genomes have been previously evaluated using a standard phylogenetic method, which makes our methodology much faster and scalable. This two-step approach also allows us to directly exploit existing phylogenetic methods that detect bacterial recombination, and therefore to account for the effect of recombination in the construction of a dated phylogeny. We analysed many simulated datasets in order to benchmark the performance of our approach in a wide range of situations. Furthermore, we present applications to three different real datasets from recent bacterial genomic studies. Our methodology is implemented in a R package called BactDating which is freely available for download at https://github.com/xavierdidelot/BactDating

Bacterial microevolution and the Pangenome

The comparison of multiple genome sequences sampled from a bacterial population reveals considerable diversity in both the core and the accessory parts of the pangenome. This diversity can be analysed in terms of microevolutionary events that took place since the genomes shared a common ancestor, especially deletion, duplication, and recombination. We review the basic modelling ingredients used implicitly or explicitly when performing such a pangenome analysis. In particular, we describe a basic neutral phylogenetic framework of bacterial pangenome microevolution, which is not incompatible with evaluating the role of natural selection. We survey the different ways in which pangenome data is summarised in order to be included in microevolutionary models, as well as the main methodological approaches that have been proposed to reconstruct pangenome microevolutionary history

Genomic epidemiology of penicillin-non-susceptible Streptococcus pneumoniae

Penicillin-non-susceptible Streptococcus pneumoniae (PNSP) were first detected in the 1960s, and are now common worldwide, predominantly through the international spread of a limited number of strains. Extant PNSP are characterized by mosaic pbp2x, pbp2b and pbp1a genes generated by interspecies recombinations, with the extent of these alterations determining the range and concentrations of β-lactams to which the genotype is non-susceptible. The complexity of the genetics underlying these phenotypes has been the subject of both molecular microbiology and genome-wide association and epistasis analyses. Such studies can aid our understanding of PNSP evolution and help improve the already highly-performing bioinformatic methods capable of identifying PNSP from genomic surveillance data

Differences in genotype and virulence among four multidrug-resistant <i>Streptococcus pneumoniae</i> isolates belonging to the PMEN1 clone

We report on the comparative genomics and characterization of the virulence phenotypes of four &lt;i&gt;S. pneumoniae&lt;/i&gt; strains that belong to the multidrug resistant clone PMEN1 (Spain&lt;sup&gt;23F&lt;/sup&gt; ST81). Strains SV35-T23 and SV36-T3 were recovered in 1996 from the nasopharynx of patients at an AIDS hospice in New York. Strain SV36-T3 expressed capsule type 3 which is unusual for this clone and represents the product of an in vivo capsular switch event. A third PMEN1 isolate - PN4595-T23 - was recovered in 1996 from the nasopharynx of a child attending day care in Portugal, and a fourth strain - ATCC700669 - was originally isolated from a patient with pneumococcal disease in Spain in 1984. We compared the genomes among four PMEN1 strains and 47 previously sequenced pneumococcal isolates for gene possession differences and allelic variations within core genes. In contrast to the 47 strains - representing a variety of clonal types - the four PMEN1 strains grouped closely together, demonstrating high genomic conservation within this lineage relative to the rest of the species. In the four PMEN1 strains allelic and gene possession differences were clustered into 18 genomic regions including the capsule, the blp bacteriocins, erythromycin resistance, the MM1-2008 prophage and multiple cell wall anchored proteins. In spite of their genomic similarity, the high resolution chinchilla model was able to detect variations in virulence properties of the PMEN1 strains highlighting how small genic or allelic variation can lead to significant changes in pathogenicity and making this set of strains ideal for the identification of novel virulence determinant

Ethnic differences in oral health and use of dental services:cross-sectional study using the 2009 Adult Dental Health Survey

Background Oral health impacts on general health and quality of life, and oral diseases are the most common non-communicable diseases worldwide. Non-White ethnic groups account for an increasing proportion of the UK population. This study explores whether there are ethnic differences in oral health and whether these are explained by differences in sociodemographic or lifestyle factors, or use of dental services. Methods We used the Adult Dental Health Survey 2009 to conduct a cross-sectional study of the adult general population in England, Wales and Northern Ireland. Ethnic groups were compared in terms of oral health, lifestyle and use of dental services. Logistic regression analyses were used to determine whether ethnic differences in fillings, extractions and missing teeth persisted after adjustment for potential sociodemographic confounders and whether they were explained by lifestyle or dental service mediators. Results The study comprised 10,435 (94.6 %) White, 272 (2.5 %) Indian, 165 (1.5 %) Pakistani/Bangladeshi and 187 (1.7 %) Black participants. After adjusting for confounders, South Asian participants were significantly less likely, than White, to have fillings (Indian adjusted OR 0.25, 95 % CI 0.17-0.37; Pakistani/Bangladeshi adjusted OR 0.43, 95 % CI 0.26-0.69), dental extractions (Indian adjusted OR 0.33, 95 % CI 0.23-0.47; Pakistani/Bangladeshi adjusted OR 0.41, 95 % CI 0.26-0.63), and &lt;20 teeth (Indian adjusted OR 0.31, 95 % CI 0.16-0.59; Pakistani/Bangladeshi adjusted OR 0.22, 95 % CI 0.08-0.57). They attended the dentist less frequently and were more likely to add sugar to hot drinks, but were significantly less likely to consume sweets and cakes. Adjustment for these attenuated the differences but they remained significant. Black participants had reduced risk of all outcomes but after adjustment for lifestyle the difference in fillings was attenuated, and extractions and tooth loss became non-significant. Conclusions Contrary to most health inequalities, oral health was better among non-White groups, in spite of lower use of dental services. The differences could be partially explained by reported differences in dietary sugar