Identification of a new OmpA-like protein in Neisseria gonorrhoeae involved in the binding to human epithelial cells and in vivo colonization

Outer membrane protein As (OmpAs) are highly conserved proteins within the Enterobacteriaceae family. OmpA contributes to the maintenance of structural membrane integrity and invasion into mammalian cells. In Escherichia coli K1 OmpA also contributes to serum resistance and is involved in the virulence of the bacterium. Here we describe the identification of an OmpA-like protein in Neisseria gonorrhoeae (Ng-OmpA). We show that the gonococcal OmpA-like protein, similarly to E. coli OmpA, plays a significant role in the adhesion and invasion into human cervical carcinoma and endometrial cells and is required for entry into macrophages and intracellular survival. Furthermore, the isogenic knockout ompA mutant demonstrates reduced recovery in a mouse model of infection when compared with the wild-type strain, suggesting that Ng-OmpA plays an important role in the in vivo colonization. All together, these data suggest that the newly identified surface exposed protein Ng-OmpA represents a novel virulence factor of gonococcus.     

The discovery of 6-[2-(5-chloro-2-{[(2,4-difluorophenyl)methyl]oxy}phenyl)-1-cyclopenten-1-yl]-2-pyridinecarboxylic acid, GW848687X, a potent and selective prostaglandin EP1 receptor antagonist for the treatment of inflammatory pain

The discovery of a series of selective EP1 receptor antagonists based on a 1,2-diarylcyclopentene template is described. After defining the structural requirements for EP1 potency and selectivity, heterocyclic rings were incorporated to reduce logD and improve in vitro pharmacokinetic properties. The 2,6-substituted pyridines and pyridazines gave an appropriate balance of potency, in vivo pharmacokinetic properties and a low potential for inhibiting a range of CYP450 enzymes. From this series, GW848687X was shown to have an excellent profile in models of inflammatory pain and was selected as a development candidate.     

Structure-based design of peptidomimetic antagonists of p56(lck) SH2 domain

Starting from the tetrapeptide Ac-pYEEI-NHMe and using a structure-based approach, we have designed and synthesised a peptidomimetic ligand for p56(lck) SH2 domain containing a conformationally restricted replacement for the two glutamate residues. We have explored replacments for the isoleucine residue in the pY+3 pocket and thus identified 1-(R)-amino-3-(S)-indaneacetic acid as the most potent replacement. We also report the X-ray crystal structures of two of the antagonists.     

Evolution of functional polymorphism in the gene coding for the Helicobacter pylori cytotoxin

There are two functionally different alleles of the Helicobacter pylori vacA gene, which code for proteins with different in target cell specificity. The alleles (m1 and m2) differ by approximately 50% in amino acid sequence in a 300 amino acid region, the m-region, which determines specificity. An analysis of partial likelihood anomalies in a set of eight Chinese and six Western vacA genes revealed highly significant phylogenetic deviation of a region of the gene including the m-region. Phylogenetic analysis of the conserved regions of these genes failed to reveal any distinction between m1 alleles and m2 alleles, however clear cut geographic variation was observed. In the m-region, the m1 alleles also show separate clustering of Chinese and Western isolates, however the m-region of the m2 alleles has a phylogenetic structure markedly different from the rest of the gene. The data indicate that the m2 m-region was acquired and spread through the population by horizontal transfer of DNA.     

Pilus operon evolution in Streptococcus pneumoniae is driven by positive selection and recombination

Background

The evolution of bacterial organelles involved in host-pathogen interactions is subject to intense and competing selective pressures due to the need to maintain function while escaping the host immune response. To characterize the interplay of these forces in an important pathogen, we sequenced the rlrA islet, a chromosomal region encoding for a pilus-like structure involved in adherence to lung epithelial cells in vitro and in colonization in a murine model of infection, in 44 clinical isolates of Streptococcus pneumoniae.

Results

We found that the rrgA and rrgB genes, encoding the main structural components of the pilus, are under the action of positive selection. In contrast, the rrgC gene, coding for a component present in low quantities in the assembled pilus, and the srtB, srtC and srtD genes, coding for three sortase enzymes essential for pilus assembly but probably not directly exposed to the host immune system, show no evidence of positive selection. We found several events of homologous recombination in the region containing these genes, identifying 4 major recombination hotspots. An analysis of the most recent recombination events shows a high level of mosaicism of the region coding for the rrgC, srtB, srtC and srtD genes.

Conclusions

In the rlrA islet, the genes coding for proteins directly exposed to the host immune response are under the action of positive selection, and exist in distinct forms in the population of circulating strains. The genes coding for proteins not directly exposed on the surface of the bacterial cell are more conserved probably due to the homogenizing effect of recombination.     

Independent Origins of Vectored Plant Pathogenic Bacteria from Arthropod-Associated <Emphasis Type="Italic">Arsenophonus</Emphasis> Endosymbionts

The genus Arsenophonus (Gammaproteobacteria) is comprised of intracellular symbiotic bacteria that are widespread across the arthropods. These bacteria can significantly influence the ecology and life history of their hosts. For instance, Arsenophonus nasoniae causes an excess of females in the progeny of parasitoid wasps by selectively killing the male embryos. Other Arsenophonus bacteria have been suspected to protect insect hosts from parasitoid wasps or to expand the host plant range of phytophagous sap-sucking insects. In addition, a few reports have also documented some Arsenophonus bacteria as plant pathogens. The adaptation to a plant pathogenic lifestyle seems to be promoted by the infection of sap-sucking insects in the family Cixiidae, which then transmit these bacteria to plants during the feeding process. In this study, we define the specific localization of an Arsenophonus bacterium pathogenic to sugar beet and strawberry plants within the plant hosts and the insect vector, Pentastiridius leporinus (Hemiptera: Cixiidae), using fluorescence in situ hybridization assays. Phylogenetic analysis on 16S rRNA and nucleotide coding sequences, using both maximum likelihood and Bayesian criteria, revealed that this bacterium is not a sister taxon to “Candidatus Phlomobacter fragariae,” a previously characterized Arsenophonus bacterium pathogenic to strawberry plants in France and Japan. Ancestral state reconstruction analysis indicated that the adaptation to a plant pathogenic lifestyle likely evolved from an arthropod-associated lifestyle and showed that within the genus Arsenophonus, the plant pathogenic lifestyle arose independently at least twice. We also propose a novel Candidatus status, “Candidatus Arsenophonus phytopathogenicus” novel species, for the bacterium associated with sugar beet and strawberry diseases and transmitted by the planthopper P. leporinus.