The cost of phage resistance in a plant pathogenic bacterium is context-dependent.

Parasites are ubiquitous features of living systems and many parasites severely reduce the fecundity or longevity of their hosts. This parasite-imposed selection on host populations should strongly favor the evolution of host resistance, but hosts typically face a trade-off between investment in reproductive fitness and investment in defense against parasites. The magnitude of such a trade-off is likely to be context-dependent, and accordingly costs that are key in shaping evolution in nature may not be easily observable in an artificial environment. We set out to assess the costs of phage resistance for a plant pathogenic bacterium in its natural plant host versus in a nutrient-rich, artificial medium. We demonstrate that mutants of Pseudomonas syringae that have evolved resistance via a single mutational step pay a substantial cost for this resistance when grown on their tomato plant hosts, but do not realize any measurable growth rate costs in nutrient-rich media. This work demonstrates that resistance to phage can significantly alter bacterial growth within plant hosts, and therefore that phage-mediated selection in nature is likely to be an important component of bacterial pathogenicity

Galaxy tools and workflows for sequence analysis with applications in molecular plant pathology

The Galaxy Project offers the popular web browser-based platform Galaxy for running bioinformatics tools and constructing simple workflows. Here, we present a broad collection of additional Galaxy tools for large scale analysis of gene and protein sequences. The motivating research theme is the identification of specific genes of interest in a range of non-model organisms, and our central example is the identification and prediction of "effector" proteins produced by plant pathogens in order to manipulate their host plant. This functional annotation of a pathogen's predicted capacity for virulence is a key step in translating sequence data into potential applications in plant pathology. This collection includes novel tools, and widely-used third-party tools such as NCBI BLASTC wrapped for use within Galaxy. Individual bioinformatics software tools are typically available separately as standalone packages, or in online browserbased form. The Galaxy framework enables the user to combine these and other tools to automate organism scale analyses as workflows, without demanding familiarity with command line tools and scripting.Workflows created using Galaxy can be saved and are reusable, so may be distributed within and between research groups, facilitating the construction of a set of standardised, reusable bioinformatic protocols. The Galaxy tools and workflows described in this manuscript are open source and freely available from the Galaxy Tool Shed (http://usegalaxy.org/toolshed or http://toolshed.g2.bx.psu.edu)

Genome-wide chromatin mapping with size resolution reveals a dynamic sub-nucleosomal landscape in Arabidopsis

All eukaryotic genomes are packaged as chromatin, with DNA interlaced with both regularly patterned nucleosomes and sub-nucleosomal-sized protein structures such as mobile and labile transcription factors (TF) and initiation complexes, together forming a dynamic chromatin landscape. Whilst details of nucleosome position in Arabidopsis have been previously analysed, there is less understanding of their relationship to more dynamic sub-nucleosomal particles (subNSPs) defined as protected regions shorter than the ~150bp typical of nucleosomes. The genome-wide profile of these subNSPs has not been previously analysed in plants and this study investigates the relationship of dynamic bound particles with transcriptional control. Here we combine differential micrococcal nuclease (MNase) digestion and a modified paired-end sequencing protocol to reveal the chromatin structure landscape of Arabidopsis cells across a wide particle size range. Linking this data to RNAseq expression analysis provides detailed insight into the relationship of identified DNA-bound particles with transcriptional activity. The use of differential digestion reveals sensitive positions, including a labile -1 nucleosome positioned upstream of the transcription start site (TSS) of active genes. We investigated the response of the chromatin landscape to changes in environmental conditions using light and dark growth, given the large transcriptional changes resulting from this simple alteration. The resulting shifts in the suites of expressed and repressed genes show little correspondence to changes in nucleosome positioning, but led to significant alterations in the profile of subNSPs upstream of TSS both globally and locally. We examined previously mapped positions for the TFs PIF3, PIF4 and CCA1, which regulate light responses, and found that changes in subNSPs co-localized with these binding sites. This small particle structure is detected only under low levels of MNase digestion and is lost on more complete digestion of chromatin to nucleosomes. We conclude that wide-spectrum analysis of the Arabidopsis genome by differential MNase digestion allows detection of sensitive features hereto obscured, and the comparisons between genome-wide subNSP profiles reveals dynamic changes in their distribution, particularly at distinct genomic locations (i.e. 5’UTRs). The method here employed allows insight into the complex influence of genetic and extrinsic factors in modifying the sub-nucleosomal landscape in association with transcriptional changes

A pelagic microbiome (viruses to protists) from a small cup of seawater

The aquatic microbiome is composed of a multi-phylotype community of microbes, ranging from the numerically dominant viruses to the phylogenetically diverse unicellular phytoplankton. They influence key biogeochemical processes and form the base of marine food webs, becoming food for secondary consumers. Due to recent advances in next-generation sequencing, this previously overlooked component of our hydrosphere is starting to reveal its true diversity and biological complexity. We report here that 250 mL of seawater is sufficient to provide a comprehensive description of the microbial diversity in an oceanic environment. We found that there was a dominance of the order Caudovirales (59%), with the family Myoviridae being the most prevalent. The families Phycodnaviridae and Mimiviridae made up the remainder of pelagic double-stranded DNA (dsDNA) virome. Consistent with this analysis, the Cyanobacteria dominate (52%) the prokaryotic diversity. While the dinoflagellates and their endosymbionts, the superphylum Alveolata dominates (92%) the microbial eukaryotic diversity. A total of 834 prokaryotic, 346 eukaryotic and 254 unique virus phylotypes were recorded in this relatively small sample of water. We also provide evidence, through a metagenomic-barcoding comparative analysis, that viruses are the likely source of microbial environmental DNA (meDNA). This study opens the door to a more integrated approach to oceanographic sampling and data analysis

SWI/SNF-like chromatin remodeling factor Fun30 supports point centromere function in S. cerevisiae

Budding yeast centromeres are sequence-defined point centromeres and are, unlike in many other organisms, not embedded in heterochromatin. Here we show that Fun30, a poorly understood SWI/SNF-like chromatin remodeling factor conserved in humans, promotes point centromere function through the formation of correct chromatin architecture at centromeres. Our determination of the genome-wide binding and nucleosome positioning properties of Fun30 shows that this enzyme is consistently enriched over centromeres and that a majority of CENs show Fun30-dependent changes in flanking nucleosome position and/or CEN core micrococcal nuclease accessibility. Fun30 deletion leads to defects in histone variant Htz1 occupancy genome-wide, including at and around most centromeres. FUN30 genetically interacts with CSE4, coding for the centromere-specific variant of histone H3, and counteracts the detrimental effect of transcription through centromeres on chromosome segregation and suppresses transcriptional noise over centromere CEN3. Previous work has shown a requirement for fission yeast and mammalian homologs of Fun30 in heterochromatin assembly. As centromeres in budding yeast are not embedded in heterochromatin, our findings indicate a direct role of Fun30 in centromere chromatin by promoting correct chromatin architecture

Genetic import and phenotype specific alleles associated with hyper-invasion in Campylobacter jejuni

Background: Campylobacter jejuni is a major zoonotic pathogen, causing gastroenteritis in humans. Invasion is an important pathogenesis trait by which C. jejuni causes disease. Here we report the genomic analysis of 134 strains to identify traits unique to hyperinvasive isolates. Methods: A total of 134 C. jejuni genomes were used to create a phylogenetic tree to position the hyperinvasive strains. Comparative genomics lead to the identification of mosaic capsule regions. A pan genome approach led to the discovery of unique loci, or loci with unique alleles, to the hyperinvasive strains. Results: Phylogenetic analysis showed that the hyper-invasive phenotype is a generalist trait. Despite the fact that hyperinvasive strains are only distantly related based on the whole genome phylogeny, they all possess genes within the capsule region with high identity to capsule genes from C. jejuni subsp. doylei and C. lari. In addition there were genes unique to the hyper-invasive strains with identity to non-C. jejuni genes, as well as allelic variants of mainly pathogenesis related genes already known in the other C. jejuni. In particular, the sequence of flagella genes, flgD-E and flgL were highly conserved amongst the hyper-invasive strains and divergent from sequences in other C. jejuni. A novel cytolethal distending toxin (cdt) operon was also identified as present in all hyper-invasive strains in addition to the classic cdt operon present in other C. jejuni. Conclusions: Overall, the hyper-invasive phenotype is strongly linked to the presence of orthologous genes from other Campylobacter species in their genomes, notably within the capsule region, in addition to the observed association with unique allelic variants in flagellar genes and the secondary cdt operon which is unlikely under random sharing of accessory alleles in separate lineages.Peer reviewe

The impact of the HIRA histone chaperone upon global nucleosome architecture

HIRA is an evolutionarily conserved histone chaperone that mediates replication-independent nucleosome assembly and is important for a variety of processes such as cell cycle progression, development, and senescence. Here we have used a chromatin sequencing approach to determine the genome-wide contribution of HIRA to nucleosome organization in Schizosaccharomyces pombe. Cells lacking HIRA experience a global reduction in nucleosome occupancy at gene sequences, consistent with the proposed role for HIRA in chromatin reassembly behind elongating RNA polymerase II. In addition, we find that at its target promoters, HIRA commonly maintains the full occupancy of the −1 nucleosome. HIRA does not affect global chromatin structure at replication origins or in rDNA repeats but is required for nucleosome occupancy in silent regions of the genome. Nucleosome organization associated with the heterochromatic (dg-dh) repeats located at the centromere is perturbed by loss of HIRA function and furthermore HIRA is required for normal nucleosome occupancy at Tf2 LTR retrotransposons. Overall, our data indicate that HIRA plays an important role in maintaining nucleosome architecture at both euchromatic and heterochromatic loci

Transcriptional reprogramming underpins enhanced plant growth promotion by the biocontrol fungus Trichoderma hamatum GD12 during antagonistic interactions with Sclerotinia sclerotiorumin soil

The free-living soil fungus Trichoderma hamatum strain GD12 is notable amongst Trichoderma strains in both controlling plant diseases and in stimulating plant growth, a property enhanced during its antagonistic interactions with pathogens in soil. These attributes, alongside its markedly expanded genome and proteome compared to other biocontrol and plant growth promoting Trichoderma strains, imply a rich potential for sustainable alternatives to synthetic pesticides and fertilisers for controlling plant disease and increasing yields. The purpose of this study was to investigate the transcriptional responses of GD12 underpinning its biocontrol and plant growth promotion capabilities during antagonistic interactions with the pathogen Sclerotinia sclerotiorum in soil. Using an extensive mRNA-seq study capturing different time points during the pathogen-antagonist interaction in soil, we show that dynamic and biphasic signatures in the GD12 transcriptome underpin its biocontrol and plant (lettuce) growth promotional activities. Functional predictions of differentially expressed genes demonstrate the enrichment of transcripts encoding proteins involved in transportation and oxidation-reduction reactions during both processes and an over-representation of siderophores. We identify a biphasic response during biocontrol characterised by a significant induction of transcripts encoding small-secreted cysteine rich proteins, secondary metabolite producing gene clusters and genes unique to GD12. These data support the hypothesis that Sclerotinia biocontrol is mediated by the synthesis and secretion of antifungal compounds and that GD12's unique reservoir of uncharacterised genes is actively recruited during effective biological control of a plurivorous plant pathogen. This article is protected by copyright. All rights reserved