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Results for “Microbiology”

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At least 199 records · Page 11Linked to original sources

bioRxiv2015

Identification of protein secretion systems in bacterial genomes

Bacteria with two cell membranes (diderms) have evolved complex systems for protein secretion. These systems were extensively studied in some model bacteria, but the characterisation of their diversity has lagged behind due to lack of standard annotation tools. We built models for accurate identification of protein secretion systems and related appendages in bacteria with LPS-containing outer membranes. They can be used with MacSyFinder (standalone program) or online (http://mobyle.pasteur.fr/cgi-bin/portal.py#forms::txsscan). They include protein profiles and information on the systems composition and genetic organisation. They can be used to search for T1SS-T6SS, T9SS, and accessorily for flagella, Type IV and Tad pili. We identified ~10,000 systems in bacterial genomes, where T1SS and T5SS were by far the most abundant and widespread. The recently described T6SSiii and T9SS were restricted to Bacteroidetes, and T6SSii to Francisella. T2SS, T3SS, and T4SS were frequently encoded in single-copy in one locus, whereas most T1SS were encoded in two loci. The secretion systems of diderm Firmicutes were similar to those found in other diderms. Novel systems may remain to be discovered, since some clades of environmental bacteria lacked all known protein secretion systems. Our models can be fully customized, which should facilitate the identification of novel systems.Introduction

Microbiology↗

bioRxiv2015

Scan-o-matic: high-resolution microbial phenomics at a massive scale

The capacity to map traits over large cohorts of individuals - phenomics - lags far behind the explosive development in genomics. For microbes the estimation of growth is the key phenotype. We introduce an automated microbial phenomics framework that delivers accurate and highly resolved growth phenotypes at an unprecedented scale. Advancements were achieved through introduction of transmissive scanning hardware and software technology, frequent acquisition of precise colony population size measurements, extraction of population growth rates from growth curves and removal of spatial bias by reference-surface normalization. Our prototype arrangement automatically records and analyses 100,000 experiments in parallel. We demonstrate the power of the approach by extending and nuancing the known salt defence biology in bakers yeast. The introduced framework will have a transformative impact by providing high-quality microbial phenomics data for extensive cohorts of individuals and generating well-populated and standardized phenomics databases.

Microbiology↗

bioRxiv2015

EcOH: In silico serotyping of E. coli from short read data

The lipopolysaccharide (O) and flagellar (H) surface antigens of Escherichia coli are targets for serotyping that have traditionally been used to identify pathogenic lineages of E. coli. As serotyping has several limitations, public health reference laboratories are increasingly moving towards whole genome sequencing (WGS) for the rapid characterisation of bacterial isolates. Here we present a method to rapidly and accurately serotype E. coli isolates from raw, short read sequence data, leveraging the known genetic basis for the biosynthesis of O- and H-antigens. Our approach bypasses the need for de novo genome assembly by directly screening WGS reads against a curated database of alleles linked to known E. coli O-groups and H-types (the EcOH database) using the software package SRST2. We validated our approach by comparing in silico results with those obtained via serological phenotyping of 197 enteropathogenic (EPEC) isolates. We also demonstrated the utility of our method to characterise enterotoxigenic E. coli (ETEC) and the uropathogenic E. coli (UPEC) epidemic clone ST131, and for in silico serotyping of foodborne outbreak-related isolates in the public GenomeTrakr database.

Microbiology↗

bioRxiv2015

Latitude delineates patterns of biogeography in terrestrial Streptomyces

We examined the biogeography of Streptomyces at regional spatial scales to identify factors that govern patterns of microbial diversity. Streptomyces are spore forming filamentous bacteria which are widespread in soil. Streptomyces strains were isolated from perennial grass habitats sampled across a spatial scale of more than 6,000 km. Previous analysis of this geographically explicit culture collection provided evidence for a latitudinal diversity gradient in Streptomyces species. Here we evaluate the hypothesis that this latitudinal diversity gradient is a result of evolutionary dynamics associated with historical demographic processes. Historical demographic phenomena have genetic consequences that can be evaluated through analysis of population genetics. We applied population genetic approaches to analyze population structure in six of the most numerically abundant and geographically widespread Streptomyces phylogroups from our culture collection. Streptomyces population structure varied at regional spatial scales and allelic diversity correlated with geographic distance. In addition, allelic diversity and gene flow are partitioned by latitude. Finally, we found that nucleotide diversity within phylogroups is negatively correlated with latitude. These results indicate that phylogroup diversification is constrained by dispersal limitation at regional spatial scales and they are consistent with the hypothesis that historical demographic processes have influenced the contemporary biogeography of Streptomyces.\n\nOriginality-Significance StatementWe provide the first population genetic evidence that patterns of Streptomyces biogeography, which manifest in geographically explicit patterns of gene flow and a latitudinal gradient of nucleotide diversity, result from dispersal limitation and regional diversification due to drift. This contribution elucidates evolutionary processes that underlie patterns of microbial biogeography.

Microbiology↗

bioRxiv2015

Exopolysaccharide biosynthetic glycoside hydrolases can be utilized to disrupt and prevent Pseudomonas aeruginosa biofilms

Bacterial biofilms are a significant medical challenge as they are recalcitrant to current therapeutic regimes. A key component of biofilm formation in the opportunistic human pathogen Pseudomonas aeruginosa is the biosynthesis of the exopolysaccharides Pel and Psl, which are involved in the formation and maintenance of the structural biofilm scaffold and protection against antimicrobials and host defenses. Given that the glycoside hydrolases - PelAh and PslGh - encoded in the pel and psl biosynthetic operons, respectively, are utilized for in vivo exopolysaccharide processing, we reasoned that these would provide specificity to target P. aeruginosa biofilms. Evaluating these enzymes as potential therapeutics, we demonstrate that these glycoside hydrolases selectively target and degrade the exopolysaccharide component of the biofilm matrix and that nanomolar concentrations of these enzymes can both prevent biofilm formation as well as rapidly disrupt preexisting biofilms in vitro. This treatment was effective against clinical and environmental P. aeruginosa isolates and reduced biofilm biomass by 58-94%. These non-cytotoxic enzymes potentiated antibiotics as the addition of either enzyme to a sub-lethal concentration of colistin reduced viable bacterial counts by 2.5 orders of magnitude. Additionally, PelAh was able to increase neutrophil killing by ~50%. This work illustrates the feasibility and benefits of using bacterial exopolysaccharide biosynthetic glycoside hydrolases and synthetic biology to develop novel anti-biofilm therapeutics.

Microbiology↗

bioRxiv2015

Spread of the pandemic Zika virus lineage is associated with NS1 codon usage adaptation in humans

Zika virus (ZIKV) infections were more common in the zoonotic cycle until the end of the 20th century with few human cases in Africa and Southeastern Asia. Recently, the Asian lineage of ZIKV is spreading along human-to-human chains of transmission in the Pacific Islands and in South America. To better understand its recent urban expansion, we compared genetic differences among the lineages. Herein we show that the recent Asian lineage spread is associated with significant NS1 codon usage adaptation to human housekeeping genes, which could facilitate viral replication and increase viral titers. These findings were supported by a significant correlation with growth in Malthusian fitness. Furthermore, we predicted several epitopes in the NS1 protein that are shared between ZIKV and Dengue. Our results imply in a significant dependence of the recent human ZIKV spread on NS1 translational selection.

Microbiology↗

bioRxiv2015

The limitations of in vitro experimentation in understanding biofilms and chronic infection

We have become increasingly aware that during infection, pathogenic bacteria often grow in multicellular biofilms which are often highly resistant to antibacterial strategies. In order to understand how biofilms form and contribute to infection, in vitro biofilm systems such as microtitre plate assays and flow cells, have been heavily used by many research groups around the world. Whilst these methods have greatly increased our understanding of the biology of biofilms, it is becoming increasingly apparent that many of our in vitro methods do not accurately represent in vivo conditions. Here we present a systematic review of the most widely used in vitro biofilm systems, and we discuss why they are not always representative of the in vivo biofilms found in chronic infections. We present examples of methods that will help us to bridge the gap between in vitro and in vivo biofilm work, so that our bench-side data can ultimately be used to improve bedside treatment.

Microbiology↗

bioRxiv2015

Phylogenetic and functional substrate specificity for endolithic microbial communities in hyper-arid environments

Under extreme water deficit, endolithic (inside rock) microbial ecosystems are considered environmental refuges for life in cold and hot deserts, yet their diversity and functional adaptations remain vastly unexplored. The metagenomic analyses of the communities from two rock substrates, calcite and ignimbrite, revealed that they were dominated by Cyanobacteria, Actinobacteria, and Chloroflexi. The relative distribution of major phyla was significantly different between the two substrates and biodiversity estimates, from 16S rRNA gene sequences and from the metagenome data, all pointed to a higher taxonomic diversity in the calcite community. While both endolithic communities showed adaptations to extreme aridity and to the rock habitat, their functional capabilities revealed significant differences. ABC transporters and pathways for osmoregulation were more diverse in the calcite chasmoendolithic community. In contrast, the ignimbrite cryptoendolithic community was enriched in pathways for secondary metabolites, such as non-ribosomal peptides (NRPS) and polyketides (PKS). Assemblies of the metagenome data produced population genomes for the major phyla found in both communities and revealed a greater diversity of Cyanobacteria population genomes for the calcite substrate. Draft genomes of the dominant Cyanobacteria in each community were constructed with more than 93% estimated completeness. The two annotated proteomes shared 64% amino acid identity and a significantly higher number of genes involved in iron update, and NRPS gene clusters, were found in the draft genomes from the ignimbrite. Both the community-wide and genome-specific differences may be related to higher water availability and the colonization of large fissures and cracks in the calcite in contrast to a harsh competition for colonization space and nutrient resources in the narrow pores of the ignimbrite. Together, these results indicated that the habitable architecture of both lithic substrates-chasmoendolithic versus cryptoendolithic - might be an essential element in determining the colonization and the diversity of the microbial communities in endolithic substrates at the dry limit for life.

Microbiology↗

bioRxiv2015

A new Defective Helper RNA to produce Sindbis virus that infects neurons but does not propagate

Recombinant Sindbis viruses are important tools in neuroscience because they combine rapid and high transgene expression with a capacity to carry large transgenes. Currently, two packaging systems based on the DH(26S)5SIN and the DH-BB(tRNA;TE12) Defective Helper (DH) RNAs are available for making recombinant Sindbis virus that is neurotropic (able to infect neurons and potentially other cells). Both systems produce a fraction of viral particles that can propagate beyond the primary infected neuron. When injected into mouse brains, viruses produced using these DH RNAs label neurons at the injection site, but also elsewhere in the brain. Such ectopic labeling caused recombinant Sindbis viruses to be classified as anterograde viruses with limited retrograde spread, and can complicate the interpretation of neuroanatomical and other experiments.\n\nHere we describe a new DH RNA, DH-BB(5SIN;TE12ORF), that can be used to produce virus that is both neurotropic and propagation-incompetent. We show in mice that DH-BB(5SIN;TE12ORF)- packaged virus eliminates infection of cells outside the injection site. We also provide evidence that ectopically labeled cells observed in previous experiments with recombinant Sindbis virus resulted from secondary infection by propagation-competent virus, rather than from inefficient retrograde spread.\n\nVirus produced with our new packaging system retains all the advantages of previous recombinant Sindbis viruses, but minimizes the risks of confounding results with unwanted ectopic labeling. It should therefore be considered in future studies in which a neurotropic, recombinant Sindbis virus is needed.

Microbiology↗

bioRxiv2015

Dynamics of microbial community composition and soil organic carbon mineralization in soil following addition of pyrogenic and fresh organic matter

Pyrogenic organic matter (PyOM) additions to soils can have large impacts on soil organic C (SOC) cycling. Because the soil microbial community drives SOC fluxes, understanding how PyOM additions affect soil microbes is essential to understanding how PyOM affects SOC. We studied SOC dynamics and surveyed soil microbial communities after OM additions in a field experiment. We produced and applied either 350{degrees}C corn stover PyOM or an equivalent amount of dried corn stover to a Typic Fragiudept soil. Stover increased SOC-derived and total CO2 fluxes (up to 6x), and caused rapid and persistent changes in bacterial community composition over 82 days. In contrast, PyOM only temporarily increased total soil CO2 fluxes (up to 2x) and caused fewer changes in bacterial community composition. 70% of the OTUs that increased in response to PyOM additions also responded to stover additions. These OTUs likely thrive on easily-mineralizable C that is found both in stover and, to a lesser extent, in PyOM. In contrast, we also identified unique PyOM-responders, which may respond to substrates such as polyaromatic C. In particular, members of Gemmatimonadetes tended to increase in relative abundance in response to PyOM but not to fresh organic matter. We identify taxa to target for future investigations of the mechanistic underpinnings of ecological phenomena associated with PyOM additions to soil.

Microbiology↗

bioRxiv2015

Changes in the relative abundance of two Saccharomyces species from oak forests to wine fermentations

Saccharomyces cerevisiae and its sibling species S. paradoxus are known to inhabit temperate arboreal habitats across the globe. Despite their sympatric distribution in the wild, S. cerevisiae is predominantly associated with human fermentations. The apparent ecological differentiation of these species is particularly striking in Europe where S. paradoxus is abundant in forests and S. cerevisiae is abundant in vineyards. However, ecological differences may be confounded with geographic differences in species abundance. To compare the distribution and abundance of these two species we isolated Saccharomyces strains from over 1,200 samples taken from vineyard and forest habitats in Slovenia. We isolated numerous strains of S. cerevisiae and S. paradoxus as well as small number of S. kudriavzevii strains from both vineyard and forest environments. We find S. cerevisiae less abundant than S. paradoxus on oak trees within and outside the vineyard, but more abundant on grapevines and associated substrates. Analysis of the uncultured microbiome shows that both S. cerevisiae and S. paradoxus are rare species in soil and bark samples, but can be much more common in grape must. In contrast to S. paradoxus, European strains of S. cerevisiae have acquired multiple traits thought to be important for life in the vineyard and dominance of wine fermentations. We conclude that S. cerevisiae and S. paradoxus currently share both vineyard and non-vineyard habitats in Slovenia and we discuss factors relevant to their global distribution and relative abundance.

Microbiology↗

bioRxiv2015

Retroviruses integrate into a shared, non-palindromic motif

Palindromic consensus nucleotide sequences are found at the genomic integration sites of retroviruses and other transposable elements. It has been suggested that the palindromic consensus arises as a consequence of structural symmetry in the integrase complex, but the precise mechanism has yet to be elucidated. Here we perform a statistical analysis of large datasets of HTLV-1 and HIV-1 integration sites. The results show that the palindromic consensus sequence is not present in individual integration sites, but appears to arise in the population average as a consequence of the existence of a non-palindromic nucleotide motif that occurs in approximately equal proportions on the plus-strand and the minus-strand of the host genome. We demonstrate that palindromic probability position matrices are characteristic of such situations. We develop a generally applicable algorithm to sort the individual integration site sequences into plus-strand and minus-strand subpopulations. We apply this algorithm to identify the respective integration site nucleotide motifs of five retroviruses of different genera: HTLV-1, HIV-1, MLV, ASLV, and PFV. The results reveal a non-palindromic motif that is shared between these retroviruses.

Microbiology↗

bioRxiv2015

Diversity of Pico- to Mesoplankton Along the 2000 km Salinity Gradient of the Baltic Sea

Microscopic plankton form the productive base of both marine and freshwater ecosystems and are key drivers of global biogeochemical cycles of carbon and nutrients. Plankton diversity is immense with representations from all major phyla within the three domains of life. So far, plankton monitoring has mainly been based on microscopic identification, which has limited sensitivity and reproducibility, not least because of the numerical majority of plankton being unidentifiable under the light microscope. High-throughput sequencing of taxonomic marker genes offers a means to identify taxa inaccessible by traditional methods; thus, recent studies have unveiled an extensive previously unknown diversity of plankton. Here, we conducted ultra-deep Illumina sequencing (average 105 sequences/sample) of rRNA gene amplicons of surface water eukaryotic and bacterial plankton communities along a 2000 km transect following the salinity gradient of the Baltic Sea. Community composition was strongly correlated with salinity for both bacterial and eukaryotic plankton assemblages, highlighting the importance of salinity for structuring the biodiversity within this ecosystem. The distribution of major planktonic taxa followed expected patterns as observed in monitoring programs, but also novel groups to the Baltic were identified, such as relatives to the coccolithophore Emiliana huxleyi in the northern Baltic Sea. The deep sequencing also enabled accurate enumeration of highly resolved (> 99% identity) operational taxonomic units, which revealed contrasting distribution profiles among closely related populations, reflecting niche partitioning into ecotypes. This study provides the first ultra-deep sequencing-based survey on eukaryotic and bacterial plankton biogeography in the Baltic Sea.

Microbiology↗

bioRxiv2016

Comparative evaluation of the genomes of common bacterial members of the Drosophila intestinal community

Drosophila melanogaster is an excellent model to explore the molecular exchanges that occur between an animal intestine and their microbial passengers. For example, groundbreaking studies in flies uncovered a sophisticated web of host responses to intestinal bacteria. The outcomes of these responses define critical events in the host, such as the establishment of immune responses, access to nutrients, and the rate of larval development. Despite our steady march towards illuminating the host machinery that responds to bacterial presence in the gut, we know remarkably little about the microbial products that influence bacterial association with a fly host. To address this deficiency, we sequenced and characterized the genomes of three common Drosophila-associated microbes: Lactobacillus plantarum, Lactobacillus brevis and Acetobacter pasteurianus. In each case, we compared the genomes of Drosophila-associated strains to the genomes of strains isolated from alternative sources. This approach allowed us to identify molecular functions common to Drosophila-associated microbes, and, in the case of A. pasteurianus, to identify genes that are essential for association with the host. Of note, many of the gene products unique to fly-associated strains have established roles in the stabilization of host-microbe interactions. We believe that these data provide a valuable starting point for a more thorough examination of the microbial perspective on host-microbe relationships.

Microbiology↗

bioRxiv2016

Eighteenth century Yersinia pestis genomes reveal the long-term persistence of an historical plague focus

The 14th-18th century pandemic of Yersinia pestis caused devastating disease outbreaks in Europe for almost 400 years. The reasons for plagues persistence and abrupt disappearance in Europe are poorly understood, but could have been due to either the presence of now-extinct plague foci in Europe itself, or successive disease introductions from other locations. Here we present five Y. pestis genomes from one of the last European outbreaks of plague, from 1722 in Marseille, France. The lineage identified has not been found in any extant Y. pestis foci sampled to date, and has its ancestry in strains obtained from victims of the 14th century Black Death. These data suggest the existence of a previously uncharacterized historical plague focus that persisted for at least three centuries. We propose that this disease source may have been responsible for the many resurgences of plague in Europe following the Black Death.

Microbiology↗

bioRxiv2016

Real-time predictions of reservoir size and rebound time during antiretroviral therapy interruption trials for HIV

Monitoring the efficacy of novel reservoir-reducing treatments for HIV is challenging. The limited ability to sample and quantify latent infection means that supervised antiretroviral therapy (ART) interruption studies are generally required. Here we introduce a set of mathematical and statistical modeling tools to aid in the design and interpretation of ART-interruption trials. We show how the likely size of the remaining reservoir can be updated in real-time as patients continue off treatment, by combining the output of laboratory assays with insights from models of reservoir dynamics and rebound. We design an optimal schedule for viral load sampling during interruption, whereby the frequency of follow-up can be decreased as patients continue off ART without rebound. While this scheme can minimize costs when the chance of rebound between visits is low, we find that the reservoir will be almost completely reseeded before rebound is detected unless sampling occurs at least every two weeks and the most sensitive viral load assays are used. We use simulated data to predict the clinical trial size needed to estimate treatment effects in the face of highly variable patient outcomes and imperfect reservoir assays. Our findings suggest that large numbers of patients - between 40 and 150 - will be necessary to reliably estimate the reservoir-reducing potential of a new therapy and to compare this across interventions. As an example, we apply these methods to the two \"Boston patients\", recipients of allogeneic hematopoietic stem cell transplants who experienced large reductions in latent infection and underwent ART-interruption. We argue that the timing of viral rebound was not particularly surprising given the information available before treatment cessation. Additionally, we show how other clinical data can be used to estimate the relative contribution that remaining HIV+ cells in the recipient versus newly infected cells from the donor made to the residual reservoir that eventually caused rebound. Together, these tools will aid HIV researchers in the evaluating new potentially-curative strategies that target the latent reservoir.

Microbiology↗

bioRxiv2016

The status of the microbial census: an update

A census is typically carried out for people at a national level; however, microbial ecologists have implemented a molecular census of bacteria and archaea by sequencing their 16S rRNA genes. We assessed how well the microbial census of full-length 16S rRNA gene sequences is proceeding in the context of recent advances in high throughput sequencing technologies. Among the 1,411,234 and 53,546 full-length bacterial and archaeal sequences sequences, 94.5% and 95.1% of the bacterial and archeaeal sequences, respectively, belonged to operational taxonomic units (OTUs) that have been observed more than once. Although these metrics suggest that the census is approaching completion, 29.2% of the bacterial and 38.5% of the archaeal OTUs have been observed more than once. Thus, there is still considerable microbial diversity to be explored. Unfortunately, the rate of new full-length sequences has been declining and new sequences are primarily being deposited by a small number of studies. Furthermore, sequences from soil and aquatic environments, which are known to be rich in bacterial diversity, only represent 7.8 and 16.5% of the census while sequences associated with zoonotic environments represent 55.0% of the census. Continued use of traditional approaches and new technologies such as single cell genomics and short read assembly are likely to improve our ability to sample rare OTUs if it is possible to overcome this sampling bias. The success of ongoing efforts to use short read sequencing to characterize microbial communities requires that researchers strive to expand the depth and breadth of the microbial census.\n\nImportanceThe biodiversity contained within the bacterial and archaeal domains dwarfs that of the eukaryotes and the services these organisms provide to the biosphere are critical. Surprisingly, we have done a relatively poor job of keeping track of the ongoing effort to characterize the biodiversity as represented in full-length 16S rRNA genes. By understanding how this census is proceeding, it is possible to suggest the best allocation of resources for advancing the census. We found that the ongoing effort has done an excellent job of sampling the most abundant organisms, but struggles to sample the more rare organisms. Through the use of new sequencing technologies we should be able to obtain full-length sequences from these rare organisms. Furthermore, we suggest that by allocating more resources to sampling environments known to have the greatest biodiversity we will be able to make significant advances in our characterization of microbial diversity.

Microbiology↗

bioRxiv2016

Screening of anti-mycobacterial compounds in a naturally infected zebrafish embryo model

Synopsis Synopsis Introduction Materials and Methods Results Discussion Funding Transparency statement References Mycobacterium tuberculosis is a deadly human pathogen that latently infects a third of the world's population, resulting in approximately 1.5 million deaths per year. Due to the difficulties and expense of carrying out animal drug trials using M. tuberculosis and rodents, infections of the zebrafish Danio rerio with M. marinum have been used as a surrogate. However the methods so far described require specialised equipment and a high level of operator expertise.\n\nWe investigated a natural infection model where zebrafish embryos are infected through incubation in media containing M. marinum. Using bioluminescently labelled ...

Microbiology↗