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

Genome Dynamics of Short Oligonucleotides: The Example of Bacterial DNA Uptake Enhancing Sequences

Among the many bacteria naturally competent for transformation by DNA uptake—a phenomenon with significant clinical and financial implications— Pasteurellaceae and Neisseriaceae species preferentially take up DNA containing specific short sequences. The genomic overrepresentation of these DNA uptake enhancing sequences (DUES) causes preferential uptake of conspecific DNA, but the function(s) behind this overrepresentation and its evolution are still a matter for discovery. Here I analyze DUES genome dynamics and evolution and test the validity of the results to other selectively constrained oligonucleotides. I use statistical methods and computer simulations to examine DUESs accumulation in Haemophilus influenzae and Neisseria gonorrhoeae genomes. I analyze DUESs sequence and nucleotide frequencies, as well as those of all their mismatched forms, and prove the dependence of DUESs genomic overrepresentation on their preferential uptake by quantifying and correlating both characteristics. I then argue that mutation, uptake bias, and weak selection against DUESs in less constrained parts of the genome combined are sufficient enough to cause DUESs accumulation in susceptible parts of the genome with no need for other DUES function. The distribution of overrepresentation values across sequences with different mismatch loads compared to the DUES suggests a gradual yet not linear molecular drive of DNA sequences depending on their similarity to the DUES. Other genomically overrepresented sequences, both pro- and eukaryotic, show similar distribution of frequencies suggesting that the molecular drive reported above applies to other frequent oligonucleotides. Rare oligonucleotides, however, seem to be gradually drawn to genomic underrepresentation, thus, suggesting a molecular drag. To my knowledge this work provides the first clear evidence of the gradual evolution of selectively constrained oligonucleotides, including repeated, palindromic and protein/transcription factor-binding DNAs

Neisseria meningitidis Differentially Controls Host Cell Motility through PilC1 and PilC2 Components of Type IV Pili

Neisseria meningitidis is a strictly human pathogen that has two facets since asymptomatic carriage can unpredictably turn into fulminant forms of infection. Meningococcal pathogenesis relies on the ability of the bacteria to break host epithelial or endothelial cellular barriers. Highly restrictive, yet poorly understood, mechanisms allow meningococcal adhesion to cells of only human origin. Adhesion of encapsulated and virulent meningococci to human cells relies on the expression of bacterial type four pili (T4P) that trigger intense host cell signalling. Among the components of the meningococcal T4P, the concomitantly expressed PilC1 and PilC2 proteins regulate pili exposure at the bacterial surface, and until now, PilC1 was believed to be specifically responsible for T4P-mediated meningococcal adhesion to human cells. Contrary to previous reports, we show that, like PilC1, the meningococcal PilC2 component is capable of mediating adhesion to human ME180 epithelial cells, with cortical plaque formation and F-actin condensation. However, PilC1 and PilC2 promote different effects on infected cells. Cellular tracking analysis revealed that PilC1-expressing meningococci caused a severe reduction in the motility of infected cells, which was not the case when cells were infected with PilC2-expressing strains. The amount of both total and phosphorylated forms of EGFR was dramatically reduced in cells upon PilC1-mediated infection. In contrast, PilC2-mediated infection did not notably affect the EGFR pathway, and these specificities were shared among unrelated meningococcal strains. These results suggest that meningococci have evolved a highly discriminative tool for differential adhesion in specific microenvironments where different cell types are present. Moreover, the fine-tuning of cellular control through the combined action of two concomitantly expressed, but distinctly regulated, T4P-associated variants of the same molecule (i.e. PilC1 and PilC2) brings a new model to light for the analysis of the interplay between pathogenic bacteria and human host cells

Whence the Interstellar Magnetic Field Shaping the Heliosphere?

Measurements of starlight polarized by aligned interstellar dust grains are used to probe the relation between the orientation of the ambient interstellar magnetic field (ISMF) and the ISMF traced by the ribbons of energetic neutral atoms discovered by the Interstellar Boundary Explorer spacecraft. We utilize polarization data, many acquired specifically for this study, to trace the configuration of the ISMF within 40 pc. A statistical analysis yields a best-fit ISMF orientation, B (magpol), aligned with Galactic coordinates l = 42 degrees, b = 49 degrees. Further analysis shows the ISMF is more orderly for "downfield" stars located over 90 degrees from B (magpol). The data subset of downfield stars yields an orientation for the nearby ISMF at ecliptic coordinates lambda, beta approximate to 219 degrees +/- 15 degrees, 43 degrees +/- 9 degrees (Galactic coordinates l, b approximate to 40 degrees, 56 degrees, +/- 17 degrees). This best-fit ISMF orientation from polarization data is close to the field direction obtained from ribbon models. This agreement suggests that the ISMF shaping the heliosphere belongs to an extended ordered magnetic field. Extended filamentary structures are found throughout the sky. A previously discovered filament traversing the heliosphere nose region, "Filament A," extends over 300 degrees of the sky, and crosses the upwind direction of interstellar dust flowing into the heliosphere. Filament A overlaps the locations of the Voyager kilohertz emissions, three quasar intraday variables, cosmic microwave background (CMB) components, and the inflow direction of interstellar grains sampled by Ulysses and Galileo. These features are likely located in the upstream outer heliosheath where ISMF drapes over the heliosphere, suggesting Filament A coincides with a dusty magnetized plasma. A filament 55 degrees long is aligned with a possible shock interface between local interstellar clouds. A dark spot in the CMB is seen within 5 degrees of the filament and within 10 degrees of the downfield ISMF direction. Two large magnetic arcs are centered on the directions of the heliotail. The overlap between CMB components and the aligned dust grains forming Filament A indicates the configuration of dust entrained in the ISMF interacting with the heliosphere provides a measurable foreground to the CMB

A Cell-Based Model for Quorum Sensing in Heterogeneous Bacterial Colonies

Although bacteria are unicellular organisms, they have the ability to act in concert by synthesizing and detecting small diffusing autoinducer molecules. The phenomenon, known as quorum sensing, has mainly been proposed to serve as a means for cell-density measurement. Here, we use a cell-based model of growing bacterial microcolonies to investigate a quorum-sensing mechanism at a single cell level. We show that the model indeed predicts a density-dependent behavior, highly dependent on local cell-clustering and the geometry of the space where the colony is evolving. We analyze the molecular network with two positive feedback loops to find the multistability regions and show how the quorum-sensing mechanism depends on different model parameters. Specifically, we show that the switching capability of the network leads to more constraints on parameters in a natural environment where the bacteria themselves produce autoinducer than compared to situations where autoinducer is introduced externally. The cell-based model also allows us to investigate mixed populations, where non-producing cheater cells are shown to have a fitness advantage, but still cannot completely outcompete producer cells. Simulations, therefore, are able to predict the relative fitness of cheater cells from experiments and can also display and account for the paradoxical phenomenon seen in experiments; even though the cheater cells have a fitness advantage in each of the investigated groups, the overall effect is an increase in the fraction of producer cells. The cell-based type of model presented here together with high-resolution experiments will play an integral role in a more explicit and precise comparison of models and experiments, addressing quorum sensing at a cellular resolution

Heterogeneous Response to a Quorum-Sensing Signal in the Luminescence of Individual Vibrio fischeri

The marine bacterium Vibrio fischeri regulates its bioluminescence through a quorum sensing mechanism: the bacterium releases diffusible small molecules (autoinducers) that accumulate in the environment as the population density increases. This accumulation of autoinducer (AI) eventually activates transcriptional regulators for bioluminescence as well as host colonization behaviors. Although V.fischeri quorum sensing has been extensively characterized in bulk populations, far less is known about how it performs at the level of the individual cell, where biochemical noise is likely to limit the precision of luminescence regulation. We have measured the time-dependence and AI-dependence of light production by individual V.fischeri cells that are immobilized in a perfusion chamber and supplied with a defined concentration of exogenous AI. We use low-light level microscopy to record and quantify the photon emission from the cells over periods of several hours as they respond to the introduction of AI. We observe an extremely heterogeneous response to the AI signal. Individual cells differ widely in the onset time for their luminescence and in their resulting brightness, even in the presence of high AI concentrations that saturate the light output from a bulk population. The observed heterogeneity shows that although a given concentration of quorum signal may determine the average light output from a population of cells, it provides far weaker control over the luminescence output of each individual cell

Structure and dynamics of the pan-genome of Streptococcus pneumoniae and closely related species

Background Streptococcus pneumoniae is one of the most important causes of microbial diseases in humans. The genomes of 44 diverse strains of S. pneumoniae were analyzed and compared with strains of non-pathogenic streptococci of the Mitis group. Results Despite evidence of extensive recombination, the S. pneumoniae phylogenetic tree revealed six major lineages. With the exception of serotype 1, the tree correlated poorly with capsular serotype, geographical site of isolation and disease outcome. The distribution of dispensable genes, genes present in not all, but more than one strain, was consistent with phylogeny, although horizontal gene transfer events attenuated this correlation in the case of ancient lineages. Homologous recombination, involving short stretches of DNA, was the dominant 13 evolutionary process of the core genome of S. pneumoniae. Genetic exchange occurred both within and across the borders of the species, and S. mitis was the main reservoir of genetic diversity of S. pneumoniae. The pan-genome size of S. pneumoniae increased logarithmically with the number of strains and linearly with the number of polymorphic sites of the sampled genomes, suggesting that acquired genes accumulate proportionately to the age of clones. Most genes associated with pathogenicity were shared by all S. pneumoniae strains, but were also present in S. mitis, S. oralis and S. infantis, indicating that these genes are not sufficient to determine virulence. Conclusion Genetic exchange with related species sharing the same ecological niche is the main mechanism of evolution of S. pneumoniae. The open pan genome guarantees the species a quick and economical response to diverse environments

Bacteria clustering by polymers induces the expression of quorum sense controlled phenotypes

Bacteria deploy a range of chemistries to regulate their behaviour and respond to their environment. Quorum sensing is one mean by which bacteria use chemical reactions to modulate pre-infection behaviour such as surface attachment. Polymers that can interfere with bacterial adhesion or the chemical reactions used for quorum sensing are thus a potential means to control bacterial population responses. Here we report how polymeric "bacteria sequestrants", designed to bind to bacteria through electrostatic interactions and thus inhibit bacterial adhesion to surfaces, induce the expression of quorum sensing controlled phenotypes as a consequence of cell clustering. A combination of polymer and analytical chemistry, biological assays and computational modelling has been used to characterise the feedback between bacteria clustering and quorum sensing signaling. We have also derived design principles and chemical strategies for controlling bacterial behaviour at the population leve

Identification and characterization of a direct activator of a gene transfer agent

Gene transfer agents (GTAs) are thought to be ancient bacteriophages that have been co-opted into serving their host and can now transfer any gene between bacteria. Production of GTAs is controlled by several global regulators through unclear mechanisms. In Rhodobacter capsulatus, gene rcc01865 encodes a putative regulatory protein that is essential for GTA production. Here, I show that rcc01865 (hereafter gafA) encodes a transcriptional regulator that binds to the GTA promoter to initiate production of structural and DNA packaging components. Expression of gafA is in turn controlled by the pleiotropic regulator protein CtrA and the quorum-sensing regulator GtaR. GafA and CtrA work together to promote GTA maturation and eventual release through cell lysis. Identification of GafA as a direct GTA regulator allows the first integrated regulatory model to be proposed and paves the way for discovery of GTAs in other species that possess gafA homologues