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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2016

Lipopolysaccharide composition determines the preferred route and entry kinetics of bacterial outer membrane vesicles into host cells

Outer membrane vesicles are microvesicles shed by Gram-negative bacteria and play important roles in immune priming and disease pathogenesis. However, our current mechanistic understanding of vesicle - host cell interactions is limited by a lack of methods to study the kinetics of vesicle entry and cargo delivery to host cells in real-time. Here, we describe a highly sensitive method to study the kinetics of vesicle entry into host cells in real-time using a genetically encoded probe targeted to vesicles. We found that route of vesicular uptake, and thus entry kinetics and efficiency of cargo release, are determined by the chemical composition of the bacterial lipopolysaccharide. The presence of O-antigen facilitates receptor-independent entry, which enhances both rate and efficiency of cargo uptake by host cells. Collectively, our findings highlight the chemical composition of the bacterial cell wall as a major determinant of secretion-independent delivery of virulence factors during Gram-negative infections.

microbiology↗
bioRxiv2016

A Rhizobiales-specific unipolar polysaccharide adhesin contributes to Rhodopseudomonas palustris biofilm formation across diverse photoheterotrophic conditions

Bacteria predominantly exist as members of surfaced-attached communities known as biofilms. Many bacterial species initiate biofilms and adhere to each other using cell surface adhesins. This is the case for numerous ecologically diverse -proteobacteria, which use polar exopolysaccharide adhesins for cell-cell adhesion and surface attachment. Here, we show that Rhodopseudomonas palustris, a metabolically versatile member of the -proteobacterial order Rhizobiales, encodes a functional unipolar polysaccharide (UPP) biosynthesis gene cluster. Deletion of genes predicted to be critical for UPP biosynthesis and export abolished UPP production. We also found that R. palustris uses UPP to mediate biofilm formation across diverse photoheterotrophic growth conditions, wherein light and organic substrates are used to support growth. However, UPP was less important for biofilm formation during photoautotrophy, where light and CO2 support growth, and during aerobic respiration with organic compounds. Expanding our analysis beyond R. palustris, we examined the phylogenetic distribution and genomic organization of UPP gene clusters among Rhizobiales species that inhabit diverse niches. Our analysis suggests that UPP is a conserved ancestral trait of the Rhizobiales but that it has been independently lost multiple times during the evolution of this clade, twice coinciding with adaptation to intracellular lifestyles within animal hosts.\n\nIMPORTANCEBacteria are ubiquitously found as surface-attached communities and cellular aggregates in nature. Here, we address how bacterial adhesion is coordinated in response to diverse environments using two complementary approaches. First, we examined how Rhodopseudomonas palustris, one of the most metabolically versatile organisms ever described, varies its adhesion to surfaces in response to different environmental conditions. We identified 2 critical genes for the production of a unipolar polysaccharide (UPP) and showed that UPP is important for adhesion when light and organic substrates are used for growth. Looking beyond R. palustris, we performed the most comprehensive survey to date on the conservation of UPP biosynthesis genes among a group of closely related bacteria that occupy diverse niches. Our findings suggest that UPP is important for free-living and plant-associated lifestyles but dispensable for animal pathogens. Additionally, we propose guidelines for classifying the adhesins produced by various -proteobacteria, facilitating future functional and comparative studies.

microbiology↗
bioRxiv2016

A novel restraint device for injection of Galleria mellonella larvae that minimises the risk of accidental operator needle stick injury

Larvae of the insect Galleria mellonella are increasingly being used for studying pathogenic microbes and their virulence mechanisms, and as a rapid model for screening novel antimicrobial agents. The larvae (waxworms) are most frequently infected by injection of pathogenic organisms into the haemocoel through the insects prolegs. The mostly widely used method for restraining the waxworms for injection is by grasping them between the operators fingers, which puts the operator at risk of needle stick injury, an important consideration when working with highly pathogenic and/or drug-resistant microorganisms. While use of a stab proof glove can reduce this risk of injury, it does so at the loss of manual dexterity and speed, resulting in a more labour-intensive and cumbersome assay. We describe a simple cost effective device (the so-called Galleria Grabber) for restraining waxworms for injection that keeps the operators fingers clear of the needle thus reducing the risk of injury.

microbiology↗
bioRxiv2016

Standard Issue: Copy number heterogeneity of JC virus standards discovered through next-generation sequencing

Quantitative PCR is a diagnostic mainstay of clinical virology, and accurate quantitation of viral load among labs requires the use of international standards. However, the use of multiple passages of viral isolates to obtain sufficient material for international standards may result in genomic changes that complicate their use as quantitative standards. We performed next-generation sequencing to gain single-nucleotide resolution and relative copy number of JC virus (JCV) clinical standards. Strikingly, the WHO international standard and ExactTM v1/v2 prototype standards for JCV showed 8-fold and 4-fold variation in genomic coverage between different loci in the viral genome, respectively, due to large deletions in the large T-antigen region. No such variation was seen for a clinical sample with high copy-number of JCV nor a plasmid control. Intriguingly, several of the JCV standards sequenced in this study with large T-antigen deletions were cultured in cell lines immortalized using SV40 T-antigen, suggesting the possibility of trans-complementation in cell culture. Using a cut-off of 2% variant allele fraction for junctional reads to define the presence of a strain, 11 different strains were present in the WHO standard. In summary, targeting of different regions of the same international standard could result in up to an 8-fold difference in quantitation. We recommend the use of next-generation sequencing to validate standards in clinical virology.

microbiology↗
bioRxiv2016

Genome-Wide Fitness Analyses of the Foodborne Pathogen Campylobacter jejuni in In Vitro and In Vivo Models

Infection by Campylobacter is recognised as the most common cause of foodborne bacterial illness worldwide. Faecal contamination of meat, especially chicken, during processing represents a key route of transmission to humans. There is currently no licenced vaccine and no Campylobacter-resistant chickens. In addition, preventative measures aimed at reducing environmental contamination and exposure of chickens to Campylobacter jejuni (biosecurity) have been ineffective. There is much interest in the factors/mechanisms that drive C. jejuni colonisation and infection of animals, and survival in the environment. It is anticipated that understanding these mechanisms will guide the development of effective intervention strategies to reduce the burden of C. jejuni infection. Here we present a comprehensive analysis of C. jejuni fitness during growth and survival within and outside hosts. A comparative analysis of transposon (Tn) gene inactivation libraries in three C. jejuni strains by Tn-seq demonstrated that a large proportion, 331 genes, of the C. jejuni genome is dedicated to (in vitro) growth. An extensive Tn library in C. jejuni M1cam (~10,000 mutants) was screened for the colonisation of commercial broiler chickens, survival in houseflies and under nutrient-rich and-poor conditions at low temperature, and infection of human gut epithelial cells. We report C. jejuni factors essential throughout its life cycle and we have identified genes that fulfil important roles across multiple conditions, including maf3, fliW, fliD, pflB and capM, as well as novel genes uniquely implicated in survival outside hosts. Taking a comprehensive screening approach has confirmed previous studies, that the flagella are central to the ability of C. jejuni to interact with its hosts. Future efforts should focus on how to exploit this knowledge to effectively control infections caused by C. jejuni.\n\nAuthor SummaryCampylobacter jejuni is the leading bacterial cause of human diarrhoeal disease. C. jejuni encounters and has to overcome a wide range of \"stress\" conditions whilst passing through the gastrointestinal tract of humans and other animals, during processing of food products, on/in food and in the environment. We have taken a comprehensive approach to understand the basis of C. jejuni growth and within/outside host survival, with the aim to inform future development of intervention strategies. Using a genome-wide transposon gene inactivation approach we identified genes core to the growth of C. jejuni. We also determined genes that were required during the colonisation of chickens, survival in the housefly and under nutrient-rich and -poor conditions at low temperature, and during interaction with human gut epithelial tissue culture cells. This study provides a comprehensive dataset linking C. jejuni genes to growth and survival in models relevant to its life cycle. Genes important across multiple models were identified as well as genes only required under specific conditions. We identified that a large proportion of the C. jejuni genome is dedicated to growth and that the flagella fulfil a prominent role in the interaction with hosts. Our data will aid development of effective control strategies.

microbiology↗
bioRxiv2016

Structure of the Legionella Dot/Icm type IV secretion system in situ by electron cryotomography

Type IV secretion systems (T4SSs) are large macromolecular machines that translocate protein and DNA and are involved in the pathogenesis of multiple human diseases. Here, using electron cryotomography (ECT), we report the in situ structure of the Dot/Icm type IVB secretion system (T4BSS) utilized by the human pathogen Legionella pneumophila. This is the first structure of a type IVB secretion system, and also the first structure of any T4SS in situ. While the Dot/Icm system shares almost no sequence homology with type IVA secretion systems (T4ASSs), its overall structure shows remarkable similarities to two previously imaged T4ASSs, suggesting shared aspects of mechanism. However, compared to one of these, the negative-stain reconstruction of the purified T4ASS from the R388 plasmid, it is approximately twice as long and wide and exhibits several additional large densities, reflecting type-specific elaborations and potentially better structural preservation in situ.

microbiology↗
bioRxiv2016

Vaccination has minimal impact on the intrahost diversity of H3N2 influenza viruses

While influenza virus diversity and antigenic drift have been well characterized on a global scale, the factors that influence the virus rapid evolution within and between human hosts are less clear. Given the modest effectiveness of seasonal vaccination, vaccine-induced antibody responses could serve as a potent selective pressure for novel influenza variants at the individual or community level. We used next generation sequencing of patient-derived viruses from a randomized, placebo-controlled trial of vaccine efficacy to characterize the diversity of influenza A virus and to define the impact of vaccine-induced immunity on within-host populations. Importantly, this study design allowed us to isolate the impact of vaccination while still studying natural infection. We used pre-season hemagglutination inhibition and neuraminidase inhibition titers to quantify vaccine-induced immunity directly and to assess its impact on intrahost populations. We identified 166 cases of H3N2 influenza over 3 seasons and 5119 person-years. We obtained whole genome sequence data for 119 samples and used a stringent and empirically validated analysis pipeline to identify intrahost single nucleotide variants at [≥]1% frequency. Phylogenetic analysis of consensus hemagglutinin and neuraminidase sequences showed no stratification by pre-season HAI and NAI titer, respectively. In our study population, we found that the vast majority of intrahost single nucleotide variants were rare and that very few were found in more than one individual. Most samples had fewer than 15 single nucleotide variants across the entire genome, and the level of diversity did not significantly vary with day of sampling, vaccination status, or pre-season antibody titer. Contrary to what has been suggested in experimental systems, our data indicate that seasonal influenza vaccination has little impact on intrahost diversity in natural infection and that vaccine-induced immunity may be only a minor contributor to antigenic drift at local scales.

microbiology↗
bioRxiv2016

Histone deacetylase inhibitors reduce the number of herpes simplex virus-1 genomes initiating expression in individual cells.

Although many viral particles can enter a single cell, the number of viral genomes per cell that establish infection is limited. However, mechanisms underlying this restriction were not explored in depth. For herpesviruses, one of the possible mechanisms suggested is chromatinization and silencing of the incoming genomes. To test this hypothesis, we followed infection with three herpes simplex virus 1 (HSV-1) fluorescence-expressing recombinants in the presence or absence of histone deacetylases inhibitors (HDACis). Unexpectedly, a lower number of viral genomes initiated expression in the presence of these inhibitors. This phenomenon was observed using several HDACi: Trichostatin A (TSA), Suberohydroxamic Acid (SBX), Valporic Acid (VPA) and Suberoylanilide Hydoxamic Acid (SAHA). We found that HDACi presence did not change the progeny outcome from the infected cells but did alter the kinetic of the infection. Different cell types (HFF, Vero and U2OS), which vary in their capability to activate intrinsic and innate immunity, show a cell specific basal average number of viral genomes establishing infection. Importantly, in all cell types, treatment with TSA reduced the number of viral genomes. ND10 nuclear bodies are known to interact with the incoming herpes genomes and repress viral replication. The viral immediate early protein, ICP0, is known to disassemble the ND10 bodies and to induce degradation of some of the host proteins in these domains. HDACi treated cells expressed higher levels of some of the host ND10 proteins (PML and ATRX), which may down regulate the number of viral genomes initiating expression per cell. Corroborating this hypothesis, infection with three HSV-1 recombinants carrying a deletion in the gene coding for ICP0, show a reduction in the number of genomes being expressed in U2OS cells. We suggest that alterations in the levels of host proteins involved in intrinsic antiviral defense may result in differences in the number of genomes that initiate expression.

microbiology↗
bioRxiv2016

Theory for the spatiotemporal interaction between lytic phages and biofilm-dwelling bacteria

Many bacteria are adapted for attaching to surfaces and for building complex communities, termed biofilms. The biofilm mode of life is predominant in bacterial ecology. So, too, is exposure of bacteria to ubiquitous viral pathogens, termed bacteriophages. Although biofilmphage encounters are likely to be very common in nature, little is known about how phages might interact with biofilm-dwelling bacteria. It is also unclear how the ecological dynamics of phages and their hosts depend on the biological and physical properties of the biofilm environment. To make headway in this area, here we develop the first biofilm simulation framework that captures key mechanistic features of biofilm growth and phage infection. Using these simulations, we find that the equilibrium state of interaction between biofilms and phages is governed largely by nutrient availability to biofilms, infection likelihood per host encounter, and the ability of phages to diffuse through biofilm populations. Interactions between the biofilm matrix and phage particles are thus likely to be of fundamental importance, controlling the extent to which bacteria and phages can coexist in natural contexts. Our results open avenues to new questions of host-parasite coevolution and horizontal gene transfer in spatially structured biofilm contexts.

microbiology↗
bioRxiv2016

Complete mapping of viral escape from neutralizing antibodies

Identifying viral mutations that confer escape from antibodies is crucial for understanding the interplay between immunity and viral evolution. Here we quantify how every amino-acid mutation to influenza hemagglutinin affects neutralization by monoclonal antibodies targeting several antigenic regions. Our approach involves creating all replication-competent protein variants of the virus, selecting these variants with antibody, and using deep sequencing to identify enriched mutations. These high-throughput measurements are predictive of the effects of individual mutations in traditional neutralization assays. At many residues, only some of the possible mutations escape from an antibody. For instance, at a single residue targeted by two different antibodies, we identify some mutations that escape both antibodies and other mutations that escape only one or the other. Therefore, our approach maps how viruses can escape antibodies with mutation-level sensitivity, and shows that only some mutations at antigenic residues actually alter antigenicity.

microbiology↗
bioRxiv2016

On the (im)possibility to reconstruct plasmids from whole genome short-read sequencing data

Plasmids are autonomous extra-chromosomal elements in bacterial cells that can carry genes that are important for bacterial survival. To benchmark algorithms for automated plasmid sequence reconstruction from short read sequencing data, we selected 42 publicly available complete bacterial genome sequences which were assembled by a combination of long- and short-read data. The selected bacterial genome sequence projects span 12 genera, containing 148 plasmids. We predicted plasmids from short-read data with four different programs (PlasmidSPAdes, Recycler, cBar and PlasmidFinder) and compared the outcome to the reference sequences.\n\nPlasmidSPAdes reconstructs plasmids based on coverage differences in the assembly graph. It reconstructed most of the reference plasmids (recall = 0.82) but approximately a quarter of the predicted plasmid contigs were false positives (precision = 0.76). PlasmidSPAdes merged 83 % of the predictions from genomes with multiple plasmids in a single bin. Recycler searches the assembly graph for sub-graphs corresponding to circular sequences and correctly predicted small plasmids but failed with long plasmids (recall = 0.12, precision = 0.30). cBar, which applies pentamer frequency composition analysis to detect plasmid-derived contigs, showed an overall recall and precision of 0.78 and 0.64. However, cBar only categorizes contigs as plasmid-derived and does not bin the different plasmids correctly within a bacterial isolate. PlasmidFinder, which searches for matches in a replicon database, had the highest precision (1.0) but was restricted by the contents of its database and the contig length obtained from de novo assembly (recall = 0.36).\n\nSurprisingly, PlasmidSPAdes and Recycler detected single isolated components corresponding to putative novel small plasmids (<10 kbp) which were also predicted as plasmids by cBar.\n\nThis study shows that it is possible to automatically predict plasmid sequences, but only for small plasmids. The reconstruction of large plasmids (>50 kbp) containing repeated sequences remains challenging and limits the high-throughput analysis of WGS data.\n\nAuthor SummaryShort read sequencing of the DNA of bacteria is often used to understand characteristics such as antibiotic resistance. However the assembly of short read sequencing data with the goal of reconstructing a complete genome is often fragmented and leaves gaps. Therefore independently replicating DNA fragments called plasmids cannot easily be identified from an assembly. Lately a number of programs have been developed to enable the automated prediction of the sequences of plasmids. Here we tested these programs by comparing their outcomes with complete genome sequences. None of the tested programs were able to fully and unambiguously predict distinct plasmid sequences. All programs performed best with the prediction of plasmids smaller than 50 kbp. Larger plasmids were only correctly predicted if they were present as a single contig in the assembly. While predictions by PlasmidSPAdes and cBar contained most of the plasmids, they were merged with or indistinguishable from other plasmids and sometimes chromosome sequences. PlasmidFinder missed most plasmids but all its predictions were correct. Without manual steps or long-read sequencing information, plasmid reconstruction from short read sequencing data remains challenging.

microbiology↗
bioRxiv2017

Modelling Competition for Nutrients between MicrobialPopulations Growing on a Solid Agar Surface

MotivationGrowth rate is a major component of the evolutionary fitness of microbial organisms and an excellent surrogate for cell health. In the high-throughput procedures QFA and SGA, microbial cultures are inoculated in an array on solid agar, and growth is measured to obtain quantitative fitness estimates. Neighbouring cultures, which are often different genetic strains, consume nutrients at different rates, creating gradients in nutrient density. We believe that diffusion of nutrients between cultures is affecting growth in these experiments. Current analysis does not account for this; instead, it is assumed that cultures grow independently. I use a network model of nutrient diffusion and nutrient-dependent growth, to correct for competition, to try to improve the accuracy and precision of fitness estimates. I test the model against QFA data from studies on telomere function in Saccharomyces cerevisiae. Ultimately, the model might be used to improve the reliability of screens for genetic interaction and drug sensitivity.\n\nResultsI fit the competition model to a QFA plate from Addinall et al. (2011). Using far fewer parameters (387 vs 1152), the new model fits timecourses with similar closeness to the previous model. Fitness estimates are less precise for the fastest growing strains, but more precise for the majority of strains (36 out of 50). Fitness rankings agree in the positions of the fastest and slowest growing strains, but disagree in the middle positions. In a cross-plate validation experiment, the competition model overestimates timecourses to a similar degree that the previous model underestimates. A different method of fitting is required to find globally optimal solutions which might improve reliability.\n\nAvailability and ImplementationCANS, a Python package developed for the analysis in this paper, is freely available at https://github.com/lwlss/CANS.

microbiology↗
bioRxiv2016

Giardia alters commensal microbial diversity throughout the murine gut

Giardia lamblia is the most frequently identified protozoan cause of intestinal infection. Over one billion people are estimated to have acute or chronic giardiasis, with infection rates approaching 90% in endemic areas. Despite its significance in global health, the mechanisms of pathogenesis associated with giardiasis remain unclear as the parasite neither produces a known toxin nor induces a robust inflammatory response. Giardia colonization and proliferation in the small intestine of the host may, however, disrupt the ecological homeostasis of gastrointestinal commensal microbes and contribute to diarrheal disease associated with giardiasis. To evaluate the impact of Giardia infection on the host microbiota, we use culture-independent methods to quantify shifts in the diversity of commensal microbes throughout the entire gastrointestinal tract in mice infected with Giardia. We discovered that Giardias colonization of the small intestine causes a systemic dysbiosis of aerobic and anaerobic bacterial taxa. Specifically, giardiasis is typified by both expansions in aerobic Proteobacteria and decreases in anaerobic Firmicutes and Melainabacteria in the murine foregut and hindgut. Based on these shifts, we created a quantitative index of murine Giardia-induced microbial dysbiosis. This index increased at all gut regions during the duration of infection, including both the proximal small intestine and the colon. Thus giardiasis could be an ecological disease, and the observed dysbiosis may be mediated directly via the parasites unique anaerobic fermentative metabolism or indirectly via parasite induction of gut inflammation. This systemic alteration of murine gut commensal diversity may be the cause or the consequence of inflammatory and metabolic changes throughout the gut. Shifts in the commensal microbiota may explain observed variation in giardiasis between hosts with respect to host pathology, degree of parasite colonization, infection initiation, and eventual clearance.

microbiology↗
bioRxiv2016

EMDUnifrac: Exact linear time computation of the Unifrac metric and identification of differentially abundant organisms

Both the weighted and unweighted Unifrac distances have been very successfully employed to assess if two communities differ, but do not give any information about how two communities differ. We take advantage of recent observations that the Unifrac metric is equivalent to the so-called earth movers distance (also known as the Kantorovich-Rubinstein metric) to develop an algorithm that not only computes the Unifrac distance in linear time and space, but also simultaneously finds which operational taxonomic units are responsible for the observed differences between samples. This allows the algorithm, called EMDUnifrac, to determine why given samples are different, not just if they are different, and with no added computational burden. EMDUnifrac can be utilized on any distribution on a tree, and so is particularly suitable to analyzing both operational taxonomic units derived from amplicon sequencing, as well as community profiles resulting from classifying whole genome shotgun metagenomes. The EMDUnifrac source code (written in python) is freely available at: https://github.com/dkoslicki/EMDUnifrac.

microbiology↗
bioRxiv2016

Crosstalk and eavesdropping among quorum sensing peptide signals that regulate bacteriocin production in Streptococcus pneumoniae

Quorum sensing (QS), where bacteria secrete and respond to chemical signals to coordinate population-wide behaviors, has revealed that bacteria are highly social. Here, we investigate how diversity in QS signals and receptors can modify social interactions controlled by the QS system regulating bacteriocin secretion in Streptococcus pneumoniae, encoded by the blp operon (bacteriocin-like peptide). Analysis of 4 096 pneumococcal genomes detected nine blp QS signals (BlpC) and five QS receptor groups (BlpH). Imperfect concordance between signals and receptors suggested widespread social interactions between cells, specifically eavesdropping (where cells respond to signals that they do not produce) and crosstalk (where cells produce signals that non-clones detect). This was confirmed in vitro by measuring the response of reporter strains containing each of six different blp QS receptors to cognate and non-cognate peptides. Assays between pneumococcal colonies grown adjacent to one another provided further evidence that crosstalk and eavesdropping occur at endogenous levels of signal secretion. Finally, simulations of QS strains producing bacteriocins revealed that eavesdropping can be evolutionarily beneficial even when the affinity for non-cognate signals is very weak. Our results highlight that social interactions can mediate intraspecific competition among bacteria and reveal that competitive interactions can be modified by polymorphic QS systems.

microbiology↗
bioRxiv2016

Proteotype profiling unmasks a viral signaling network essential for poxvirus assembly and transcriptional competence

To orchestrate context-dependent signaling programs poxviruses encode two dual-specificity enzymes, the F10 kinase and the H1 phosphatase. These signaling mediators are essential for poxvirus production, yet their substrate profiles and systems level functions remain enigmatic. Using a phosphoproteomic screen of cells infected with wildtype, F10, and H1 mutant viruses we systematically defined the viral signaling network controlled by these enzymes. Quantitative cross-comparison revealed 33 F10 and/or H1 phosphosites within 17 viral proteins. Using this proteotype dataset to inform genotype-phenotype relationships we found that H1-deficient virions harbor a hidden hyper-cleavage phenotype driven by reversible phosphorylation of the virus protease I7 (S134). Quantitative phospho-proteotyping further revealed that the phosphorylation-dependent activity of the viral early transcription factor, A7 (Y367), underlies the transcription-deficient phenotype of H1 mutant virions. Together these results highlight the utility of combining quantitative proteotype screens with mutant viruses to uncover novel proteotype-phenotype-genotype relationships that are masked by classical genetic studies.

microbiology↗
bioRxiv2016

Cobalt enrichment in anaerobic microbial cocultures revealed by synchrotron X-ray fluorescence imaging

Metal cofactors are required for many enzymes in anaerobic microbial respiration. This study examined iron, cobalt, nickel, copper, and zinc in cellular and abiotic phases at the single-cell scale for a sulfate-reducing bacterium (Desulfococcus multivorans) and a methanogenic archaeon (Methanosarcina acetivorans) using synchrotron x-ray fluorescence microscopy. Relative abundances of cellular metals were also measured by inductively coupled plasma mass spectrometry. For both species, zinc and iron were consistently the most abundant cellular metals. M. acetivorans contained higher nickel and cobalt content than D. multivorans, likely due to elevated metal requirements for methylotrophic methanogenesis. Cocultures contained spheroid zinc sulfides and cobalt/copper-sulfides.

microbiology↗
bioRxiv2016

Chromosome segregation drives division site selection in Streptococcus pneumoniae

Accurate spatial and temporal positioning of the tubulin-like protein FtsZ is key for proper bacterial cell division. Streptococcus pneumoniae (pneumococcus) is an oval-shaped, symmetrically dividing human pathogen lacking the canonical systems for division site control (nucleoid occlusion and the Min-system). Recently, the early division protein MapZ was identified and implicated in pneumococcal division site selection. We show that MapZ is important for proper division plane selection; thus the question remains what drives pneumococcal division site selection. By mapping the cell cycle in detail, we show that directly after replication both chromosomal origin regions localize to the future cell division sites, prior to FtsZ. Perturbing the longitudinal chromosomal organization by mutating the condensin SMC, by CRISPR/Cas9-mediated chromosome cutting or by poisoning DNA decatenation resulted in mistiming of MapZ and FtsZ positioning and subsequent cell elongation. Together, we demonstrate an intimate relationship between DNA replication, chromosome segregation and division site selection in the pneumococcus, providing a simple way to ensure equally sized daughter cells without the necessity for additional protein factors.

microbiology↗