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HD-GYP is a protein domain of unknown biochemical function implicated in bacterial signaling and regulation. In the plant pathogen Xanthomonas campestris pv. campestris, the synthesis of virulence factors and dispersal of biofilms are positively controlled by a two-component signal transduction system comprising the HD-GYP domain regulatory protein RpfG and cognate sensor RpfC and by cell-cell signaling mediated by the diffusible signal molecule DSF (diffusible signal factor). The RpfG/RpfC two-component system has been implicated in DSF perception and signal transduction. Here we show that the role of RpfG is to degrade the unusual nucleotide cyclic di-GMP, an activity associated with the HD-GYP domain. Mutation of the conserved H and D residues of the isolated HD-GYP domain resulted in loss of both the enzymatic activity against cyclic di-GMP and the regulatory activity in virulence factor synthesis. Two other protein domains, GGDEF and EAL, are already implicated in the synthesis and degradation respectively of cyclic di-GMP. As with GGDEF and EAL domains, the HD-GYP domain is widely distributed in free-living bacteria and occurs in plant and animal pathogens, as well as beneficial symbionts and organisms associated with a range of environmental niches. Identification of the role of the HD-GYP domain thus increases our understanding of a signaling network whose importance to the lifestyle of diverse bacteria is now emerging.

Background Lactobacillus reuteri is a gut symbiont of a wide variety of vertebrate species that has diversified into distinct phylogenetic clades which are to a large degree host-specific. Previous work demonstrated host specificity in mice and begun to determine the mechanisms by which gut colonisation and host restriction is achieved. However, how L. reuteri strains colonise the gastrointestinal (GI) tract of pigs is unknown. Results To gain insight into the ecology of L. reuteri in the pig gut, the genome sequence of the porcine small intestinal isolate L. reuteri ATCC 53608 was completed and consisted of a chromosome of 1.94 Mbp and two plasmids of 138.5 kbp and 9.09 kbp, respectively. Furthermore, we generated draft genomes of four additional L. reuteri strains isolated from pig faeces or lower GI tract, lp167-67, pg-3b, 20-2 and 3c6, and subjected all five genomes to a comparative genomic analysis together with the previously completed genome of strain I5007. A phylogenetic analysis based on whole genomes showed that porcine L. reuteri strains fall into two distinct clades, as previously suggested by multi-locus sequence analysis. These six pig L. reuteri genomes contained a core set of 1364 orthologous gene clusters, as determined by OrthoMCL analysis, that contributed to a pan-genome totalling 3373 gene clusters. Genome comparisons of the six pig L. reuteri strains with 14  L. reuteri strains from other host origins gave a total pan-genome of 5225 gene clusters that included a core genome of 851 gene clusters but revealed that there were no pig-specific genes per se . However, genes specific for and conserved among strains of the two pig phylogenetic lineages were detected, some of which encoded cell surface proteins that could contribute to the diversification of the two lineages and their observed host specificity. Conclusions This study extends the phylogenetic analysis of L. reuteri strains at a genome-wide level, pointing to distinct evolutionary trajectories of porcine L. reuteri lineages, and providing new insights into the genomic events in L. reuteri that occurred during specialisation to their hosts. The occurrence of two distinct pig-derived clades may reflect differences in host genotype, environmental factors such as dietary components or to evolution from ancestral strains of human and rodent origin following contact with pig populations.

Bacteriovorax marinus SJ is a predatory delta-proteobacterium isolated from a marine environment. The genome sequence of this strain provides an interesting contrast to that of the terrestrial predatory bacterium Bdellovibrio bacteriovorus HD100. Based on their predatory lifestyle, Bacteriovorax were originally designated as members of the genus Bdellovibrio but subsequently were re-assigned to a new genus and family based on genetic and phenotypic differences. B. marinus attaches to Gram-negative bacteria, penetrates through the cell wall to form a bdelloplast, in which it replicates, as shown using microscopy. Bacteriovorax is distinct, as it shares only 30% of its gene products with its closest sequenced relatives. Remarkably, 34% of predicted genes over 500 nt in length were completely unique with no significant matches in the databases. As expected, Bacteriovorax shares several characteristic loci with the other delta-proteobacteria. A geneset shared between Bacteriovorax and Bdellovibrio that is not conserved among other delta-proteobacteria such as Myxobacteria (which destroy prey bacteria externally via lysis), or the non-predatory Desulfo-bacteria and Geobacter species was identified. These 291 gene orthologues common to both Bacteriovorax and Bdellovibrio may be the key indicators of host-interaction predatory-specific processes required for prey entry. The locus from Bdellovibrio bacteriovorus is implicated in the switch from predatory to prey/host-independent growth. Although the locus is conserved in B. marinus , the sequence has only limited similarity. The results of this study advance understanding of both the similarities and differences between Bdellovibrio and Bacteriovorax and confirm the distant relationship between the two and their separation into different families.

This work centres on the genomic comparisons of two closely-related nitrogen-fixing symbiotic bacteria, Rhizobium leguminosarum biovar viciae 3841 and Rhizobium etli CFN42. These strains maintain a stable genomic core that is also common to other rhizobia species plus a very variable and significant accessory component. The chromosomes are highly syntenic, whereas plasmids are related by fewer syntenic blocks and have mosaic structures. The pairs of plasmids p42f-pRL12, p42e-pRL11 and p42b-pRL9 as well large parts of p42c with pRL10 are shown to be similar, whereas the symbiotic plasmids (p42d and pRL10) are structurally unrelated and seem to follow distinct evolutionary paths. Even though purifying selection is acting on the whole genome, the accessory component is evolving more rapidly. This component is constituted largely for proteins for transport of diverse metabolites and elements of external origin. The present analysis allows us to conclude that a heterogeneous and quickly diversifying group of plasmids co-exists in a common genomic framework.

Enteroaggregative E. coli (EAEC) are a major cause of diarrhoea worldwide. Due to their heterogeneity and carriage in healthy individuals, identification of diagnostic virulence markers for pathogenic strains has been difficult. In this study, we have determined phenotypic and genotypic differences between EAEC strains of sequence types (STs) epidemiologically associated with asymptomatic carriage (ST31) and diarrhoeal disease (ST40). ST40 strains demonstrated significantly enhanced intestinal adherence, biofilm formation, and pro-inflammatory interleukin-8 secretion compared with ST31 isolates. This was independent of whether strains were derived from diarrhoea patients or healthy controls. Whole genome sequencing revealed differences in putative virulence genes encoding aggregative adherence fimbriae, E. coli common pilus, flagellin and EAEC heat-stable enterotoxin 1. Our results indicate that ST40 strains have a higher intrinsic potential of human pathogenesis due to a specific combination of virulence-related factors which promote host cell colonization and inflammation. These findings may contribute to the development of genotypic and/or phenotypic markers for EAEC strains of high virulence.

Non-aureus staphylococci (NAS) are implicated in many healthcare-acquired infections and an understanding of the genetics of antimicrobial resistance is important in relation to both clinical intervention and the role of NAS as a reservoir of resistance genes. Gap statement: The burden of antimicrobial resistance in NAS, particularly to clinically relevant antimicrobials, is under-recognised. We sourced 394 NAS isolates from clinical samples, healthy human volunteers, animals and type cultures and subjected them to minimum inhibitory concentration (MIC) testing by agar dilution using eight antimicrobials. Cefoxitin was used to screen for methicillin resistance, as it stimulates the expression of mecA in S. aureus. We performed whole genome sequencing on 366 isolates and analysed these genotypically for the presence of genetic mechanisms responsible for the phenotypic levels of reduced antimicrobial susceptibility. We observed 175 sequenced isolates with a MIC ≥ 4 µg/ml to cefoxitin, of which 50% did not harbour a known mec homologue. Eight clinical NAS isolates displayed high daptomycin MICs (>4 µg/ml), with no known mechanism identified. Differences in MICs against erythromycin were attributable to the presence of different resistance genes (msrA and ermC). In total, 49% of isolates displayed reduced susceptibility to three or more of the antimicrobials tested. The widespread presence of reduced antimicrobial susceptibility in NAS is concerning. An increased likelihood of harder-to-treat infections caused directly by NAS with acquired resistance genes has clinical implications for AMR detection, the horizontal resistance gene pool and the management of patients.

Background Clostridium botulinum is a group of four physiologically and phylogenetically distinct bacteria that produce botulinum neurotoxin. While studies have characterised variability between strains of Group I (proteolytic) C. botulinum , the genetic and physiological variability and relationships between strains within Group II (non-proteolytic) C. botulinum are not well understood. In this study the genome of Group II strain C. botulinum Eklund 17B (NRP) was sequenced and used to construct a whole genome DNA microarray. This was used in a comparative genomic indexing study to compare the relatedness of 43 strains of Group II C. botulinum (14 type B, 24 type E and 5 type F). These results were compared with characteristics determined from physiological tests. Results Whole genome indexing showed that strains of Group II C. botulinum isolated from a wide variety of environments over more than 75 years clustered together indicating the genetic background of Group II C. botulinum is stable. Further analysis showed that strains forming type B or type F toxin are closely related with only toxin cluster genes targets being unique to either type. Strains producing type E toxin formed a separate subset. Carbohydrate fermentation tests supported the observation that type B and F strains form a separate subset to type E strains. All the type F strains and most of type B strains produced acid from amylopectin, amylose and glycogen whereas type E strains did not. However, these two subsets did not differ strongly in minimum growth temperature or maximum NaCl concentration for growth. No relationship was found between tellurite resistance and toxin type despite all the tested type B and type F strains carrying tehB , while the sequence was absent or diverged in all type E strains. Conclusions Although Group II C. botulinum form a tight genetic group, genomic and physiological analysis indicates there are two distinct subsets within this group. All type B strains and type F strains are in one subset and all type E strains in the other.

The obligately anaerobic bacterium Bacteroides fragilis, an opportunistic pathogen and inhabitant of the normal human colonic microbiota, exhibits considerable within-strain phase and antigenic variation of surface components. The complete genome sequence has revealed an unusual breadth (in number and in effect) of DNA inversion events that potentially control expression of many different components, including surface and secreted components, regulatory molecules, and restriction-modification proteins. Invertible promoters of two different types (12 group 1 and 11 group 2) were identified. One group has inversion crossover (fix) sites similar to the hix sites of Salmonella typhimurium. There are also four independent intergenic shufflons that potentially alter the expression and function of varied genes. The composition of the 10 different polysaccharide biosynthesis gene clusters identified (7 with associated invertible promoters) suggests a mechanism of synthesis similar to the O-antigen capsules of Escherichia coli.

Complex relationships between bacteria and multicellular eukaryotes have evolved in several environments but science has traditionally focused on interactions that are pathogenic. Recently there has been increased recognition of symbiotic interactions that benefit both the microorganism and the host. The bacterial partner commonly produces essential nutrients that are not synthesized by the host. In return, bacteria benefit from an ample supply of growth materials and environmental stability inside the host. An array of recent genome papers examining microbial symbionts in hosts ranging from insects and worms to plants, has shed light on the genetics that underpin these metabolic partnerships.