Are bacteriophage defence and virulence two sides of the same coin in Campylobacter jejuni?

Abstract The continuous battle for survival in the environment has led to the development or acquisition of sophisticated defence systems in bacteria. These defence systems have contributed to the survival of the bacterial species in the environment for millions of years. Some systems appear to have evolved in a number of pathogenic bacteria towards a role in virulence and host immune evasion. Recently, different bacterial cell envelope components from diverse bacterial species have been linked not only to bacteriophage defence, but also to virulence features. In the present review we focus specifically on the bacterial cell envelopeexpressed sialic-acid-containing LOS (lipo-oligosaccharide) structures and Type II CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR-associated) genes that both occur in specific Gramnegative pathogens. In Campylobacter jejuni circumstantial evidence points at a potential intertwined dual function between sialylated LOS structures and subtype II-C CRISPR-Cas, i.e. in phage defence and virulence. In the present review we discuss whether a dual functionality of sialylated LOS and subtype II-C CRISPR-Cas is exclusive to C. jejuni only or could be more widespread within the group of Type II CRISPR-Cas-harbouring bacteria. We conclude from the literature that, at least in C. jejuni, circumstantial evidence exists for a complex intertwined dual functionality between sialylated LOS and Type II CRISPR-Cas, and that other bacteria show similar genomic signatures

KREAP: An automated Galaxy platform to quantify in vitro re-epithelialization kinetics

Background: In vitro scratch assays have been widely used to study the influence of bioactive substances on the processes of cell migration and proliferation that are involved in re-epithelialization

Active Human and Porcine Serum Induce Competence for Genetic Transformation in the Emerging Zoonotic Pathogen Streptococcus suis.

The acquisition of novel genetic traits through natural competence is a strategy used by bacteria in microbe-rich environments where microbial competition, antibiotics, and host immune defenses threaten their survival. Here, we show that virulent strains of Streptococcus suis, an important zoonotic agent and porcine pathogen, become competent for genetic transformation with plasmid or linear DNA when cultured in active porcine and human serum. Competence was not induced in active fetal bovine serum, which contains less complement factors and immunoglobulins than adult serum and was strongly reduced in heat-treated or low-molecular weight fractions of active porcine serum. Late competence genes, encoding the uptake machinery for environmental DNA, were upregulated in the active serum. Competence development was independent of the early competence regulatory switch involving XIP and ComR, as well as sigma factor ComX, suggesting the presence of an alternative stress-induced pathway for regulation of the late competence genes required for DNA uptake

Transcriptomics in serum and culture medium reveal shared and differential gene regulation in pathogenic and commensal Streptococcus suis

Streptococcus suis colonizes the upper respiratory tract of healthy pigs at high abundance but can also cause opportunistic respiratory and systemic disease. Disease-associated S. suis reference strains are well studied, but less is known about commensal lineages. It is not known what mechanisms enable some S. suis lineages to cause disease while others persist as commensal colonizers, or to what extent gene expression in disease-associated and commensal lineages diverge. In this study we compared the transcriptomes of 21S. suis strains grown in active porcine serum and Todd–Hewitt yeast broth. These strains included both commensal and pathogenic strains, including several strains of sequence type (ST) 1, which is responsible for most cases of human disease and is considered to be the most pathogenic S. suis lineage. We sampled the strains during their exponential growth phase and mapped RNA sequencing reads to the corresponding strain genomes. We found that the transcriptomes of pathogenic and commensal strains with large genomic divergence were unexpectedly conserved when grown in active porcine serum, but that regulation and expression of key pathways varied. Notably, we observed strong variation of expression across media of genes involved in capsule production in pathogens, and of the agmatine deiminase system in commensals. ST1 strains displayed large differences in gene expression between the two media compared to strains from other clades. Their capacity to regulate gene expression across different environmental conditions may be key to their success as zoonotic pathogens

Identification of the transcriptional response of human intestinal mucosa to Lactobacillus plantarum WCFS1 in vivo

Contains fulltext : 69887.pdf ( ) (Open Access)BACKGROUND: There is limited knowledge on the extent and dynamics of the mucosal response to commensal and probiotic species in the human intestinal lumen. This study aimed to identify the acute, time-dependent responses of intestinal mucosa to commensal Lactobacillus plantarum WCFS1 in vivo in two placebo-controlled human intervention studies in healthy volunteers. Transcriptional changes in duodenal mucosa upon continuous intraduodenal infusion of L. plantarum WCFS1 for one- and six h, respectively, were studied using oro- and nasogastric intubations with dedicated orogastric catheters and tissue sampling by standard flexible gastroduodenoscopy. RESULTS: One- and six-h exposure of small intestinal mucosa to L. plantarum WCFS1 induced differential expression of 669 and 424 gene reporters, respectively. While short-term exposure to L. plantarum WCFS1 inhibited fatty acid metabolism and cell cycle progression, cells switched to a more proliferative phase after prolonged exposure with an overall expression profile characterized by upregulation of genes involved in lipid metabolism, cellular growth and development. Cell death and immune responses were triggered, but cell death-executing genes or inflammatory signals were not expressed. Proteome analysis showed differential expression of several proteins. Only the microsomal protein 'microsomal triglyceride transfer protein' was regulated on both the transcriptional and the protein level in all subjects. CONCLUSION: Overall, this study showed that intestinal exposure to L. plantarum WCFS1 induced consistent, time-dependent transcriptional responses in healthy intestinal mucosa. This extensive exploration of the human response to L. plantarum WCFS1 could eventually provide molecular support for specific or probiotic activity of this strain or species, and exemplifies the strength of the applied technology to identify the potential bio-activity of microbes in the human intestine

Gene expression analysis of peripheral cells for subclassification of pediatric inflammatory bowel disease in remission

Objective: In current clinical practice, optimal treatment of inflammatory bowel disease (IBD) aims at the induction and maintenance of clinical remission. Clinical remission is apparent when laboratory markers of inflammation are normal and clinical symptoms are absent. However, sub-clinical inflammation can still be present. A detailed analysis of the immune status during this inactive state of disease may provide a useful tool to categorize patients with clinical remission into subsets with variable states of immune activation. Design: By using Affymetrix GeneChips, we analysed RNA gene expression profiles of peripheral blood leukocytes from pediatric IBD patients in clinical remission and controls. We performed (un)supervised clustering analysis of IBD-associated genes and applied Ingenuity® pathway software to identify specific molecular profiles between patients. Results: Pediatric IBD patients with disease in clinical remission display heterogeneously distributed gene expression profiles that are significantly distinct from controls. We identified three clusters of IBD patients, each displaying specific expression profiles of IBD-associated genes. Conclusion: The expression of immune- and IBD-associated genes in peripheral blood leukocytes from pediatric IBD patients in clinical remission was different from healthy controls, indicating that sub-clinical immune mechanisms are still active during remission. As such, RNA profiling of peripheral blood may allow for non-invasive patient subclassification and new perspectives in treatment regimes of IBD patients in the future

Identification of plasminogen-binding sites in Streptococcus suis enolase that contribute to bacterial translocation across the blood-brain barrier

Streptococcus suis is an emerging zoonotic pathogen that can cause invasive disease commonly associated with meningitis in pigs and humans. To cause meningitis, S. suis must cross the blood-brain barrier (BBB) comprising blood vessels that vascularize the central nervous system (CNS). The BBB is highly selective due to interactions with other cell types in the brain and the composition of the extracellular matrix (ECM). Purified streptococcal surface enolase, an essential enzyme participating in glycolysis, can bind human plasminogen (Plg) and plasmin (Pln). Plg has been proposed to increase bacterial traversal across the BBB via conversion to Pln, a protease which cleaves host proteins in the ECM and monocyte chemoattractant protein 1 (MCP1) to disrupt tight junctions. The essentiality of enolase has made it challenging to unequivocally demonstrate its role in binding Plg/Pln on the bacterial surface and confirm its predicted role in facilitating translocation of the BBB. Here, we report on the CRISPR/Cas9 engineering of S. suis enolase mutants eno261, eno252/253/255, eno252/261, and eno434/435 possessing amino acid substitutions at in silico predicted binding sites for Plg. As expected, amino acid substitutions in the predicted Plg binding sites reduced Plg and Pln binding to S. suis but did not affect bacterial growth in vitro compared to the wild-type strain. The binding of Plg to wild-type S. suis enhanced translocation across the human cerebral microvascular endothelial cell line hCMEC/D3 but not for the eno mutant strains tested. To our knowledge, this is the first study where predicted Plg-binding sites of enolase have been mutated to show altered Plg and Pln binding to the surface of S. suis and attenuation of translocation across an endothelial cell monolayer in vitro

Campylobacter jejuni Cas9 Modulates the Transcriptome in Caco-2 Intestinal Epithelial Cells

The zoonotic human pathogen Campylobacter jejuni is known for its ability to induce DNA-damage and cell death pathology in humans. The molecular mechanism behind this phenomenon involves nuclear translocation by Cas9, a nuclease in C. jejuni (CjeCas9) that is the molecular marker of the Type II CRISPR-Cas system. However, it is unknown via which cellular pathways CjeCas9 drives human intestinal epithelial cells into cell death. Here, we show that CjeCas9 released by C. jejuni during the infection of Caco-2 human intestinal epithelial cells directly modulates Caco-2 transcriptomes during the first four hours of infection. Specifically, our results reveal that CjeCas9 activates DNA damage (p53, ATM (Ataxia Telangiectasia Mutated Protein)), pro-inflammatory (NF-κB (Nuclear factor-κB)) signaling and cell death pathways, driving Caco-2 cell

Vectorial secretion of interleukin-8 mediates autocrine signalling in intestinal epithelial cells via apically located CXCR1

BACKGROUND: In the intestinal mucosa, several adaptations of TLR signalling have evolved to avoid chronic inflammatory responses to the presence of commensal microbes. Here we investigated whether polarized monolayers of intestinal epithelial cells might regulate inflammatory responses by secreting IL-8 in a vectorial fashion (i.e. apical versus basolateral) depending on the location of the TLR stimulus. RESULTS: In the Caco-2 BBE model of polarized villus-like epithelium, apical stimulation with TLR2 and TLR5 ligands resulted in the apical secretion of IL-8. The CXCR1 receptor for IL-8 was expressed only on the apical membrane of Caco-2 BBE cells and differentiated epithelial cells in the human small intestine and colon. Transcriptome analyses revealed that Caco-2 BBE cells respond to stimulation with IL-8 supporting the hypothesis that IL-8 induces G protein-coupled receptor signalling. CONCLUSIONS: These results show that IL-8 induces autocrine signalling via an apical CXCR1 in Caco-2 BBE intestinal epithelial cells and that this receptor is also expressed on the apical surface of differentiated human intestinal epithelial cells in vivo, suggesting an autocrine function for IL-8 secreted in the lumen