Diagnostic tool for surveillance, detection and monitoring of the high-risk clone K. pneumoniae ST15.

BACKGROUND: The global spread of Klebsiella pneumoniae ST15, causing multi-continental outbreaks, contributes to the movement of resistance genes between clones increasing the antimicrobial resistance crisis. The genomic traits providing it with the ability to outcompete other bacteria and cause epidemics remain unclear. AIM: To identify the specific genomic traits of K. pneumoniae ST15 to develop a diagnostic test. METHODS: An outbreak caused by K. pneumoniae occurred in Hospital A Coruña, Spain. Antimicrobial susceptibility analysis and molecular typing (PGFE and MLST) were performed. One isolate of each sequence type was selected for whole-genome sequencing analysis. Comparative analysis of genomes was performed using RAST. BLASTn was used to evaluate the presence of the fhaC and kpiD genes. Two hundred and ninety-four K. pneumoniae from a Spanish nationwide collection were analysed by PCR. FINDINGS: Genotyping showed that 87.5% of the isolates tested belonged to a clone with a unique PFGE pattern which corresponded to ST15. Comparative genomic analysis of the different STs enabled us to determine the specific genomic traits of K. pneumoniae ST15. Two adherence-related systems (Kpi and KpFhaB/FhaC) were specific markers of this clone. Multiplex-PCR analysis with kpiD and fhaC oligonucleotides revealed that K. pneumoniae ST15 is specifically detected with a sensitivity of 100% and a specificity of 97.76%. The PCR results showed 100% concordance with the MLST and whole-genome sequencing data. CONCLUSION: K. pneumoniae ST15 possesses specific genomic traits that could favour its dissemination. They could be used as targets to detect K. pneumoniae ST15 with high sensitivity and specificity.

Change of concept about the regulation of angiotensin II-induced monocyte chemoattractant protein-1 production in human endothelial cells.

AIMS: Some intriguing clinical observations about the anti-inflammatory effects of angiotensin type 1 (AT1) receptor blockers and angiotensin converting enzyme inhibitors in cardiovascular patients brought us to study the signalling pathways which lead to angiotensin II (ANG)-induced monocyte chemoattractant protein-1 (MCP-1) production in human endothelial cells. METHODS: MCP-1 production in human umbilical vein endothelial cells (HUVECs) under treatments with ANG, AT1 and angiotensin type 2 (AT2) receptor blockers and pravastatin was measured by ELISA. The expression of AT1 and AT2 receptors and NADPH oxidase catalytic subunits (NOX 1-5) was analysed at mRNA and protein levels. Nuclear factor-kappa B (NF-κB) activation was studied by p65 subunit translocation to the cellular nucleus. Cell viability was tested by the MTT method. Nox4 subcellular distribution was analysed by subcellular protein fractionation and by immunoprecipitation followed by matrix-assisted laser desorption/ionization mass spectrometry analysis. RESULTS: ANG-induced MCP-1 production was mediated by AT2 receptor, but not AT1 receptor in HUVECs in culture, which in turn activated NF-κB, promoting p65 subunit translocation to the nucleus. Reactive oxygen species produced by NADPH oxidase participated in this activation, mainly by the Nox4 subunit, ubiquitously expressed in all the compartments of HUVECs. Pravastatin inhibited ANG-induced MCP-1 production. CONCLUSIONS: Our results support that ANG-induced MCP-1 production in HUVECs is mediated by AT2 instead AT1 receptor activation, which in turn activates NF-κB involving reactive oxygen species produced by the NADPH oxidase complex. Statins can also block ANG-induced MCP-1 production, probably by their inhibitory effects on NADPH oxidase activity.

The joint power of sex and stress to modulate brain-gut-microbiota axis and intestinal barrier homeostasis: implications for irritable bowel syndrome.

BACKGROUND: Intestinal homeostasis is a dynamic process that takes place at the interface between the lumen and the mucosa of the gastrointestinal tract, where a constant scrutiny for antigens and toxins derived from food and microorganisms is carried out by the vast gut-associated immune system. Intestinal homeostasis is preserved by the ability of the mucus layer and the mucosal barrier to keep the passage of small-sized and antigenic molecules across the epithelium highly selective. When combined and preserved, immune surveillance and barrier's selective permeability, the host capacity of preventing the development of intestinal inflammation is optimized, and viceversa. In addition, the brain-gut-microbiome axis, a multidirectional communication system that integrates distant and local regulatory networks through neural, immunological, metabolic, and hormonal signaling pathways, also regulates intestinal function. Dysfunction of the brain-gut-microbiome axis may induce the loss of gut mucosal homeostasis, leading to uncontrolled permeation of toxins and immunogenic particles, increasing the risk of appearance of intestinal inflammation, mucosal damage, and gut disorders. Irritable bowel syndrome is prevalent stress-sensitive gastrointestinal disorder that shows a female predominance. Interestingly, the role of stress, sex and gonadal hormones in the regulation of intestinal mucosal and the brain-gut-microbiome axis functioning is being increasingly recognized. PURPOSE: We aim to critically review the evidence linking sex, and stress to intestinal barrier and brain-gut-microbiome axis dysfunction and the implications for irritable bowel syndrome.

Receptor for advanced glycation end-products expression in subcutaneous adipose tissue is related to coronary artery disease.

OBJECTIVE: Obesity, a risk factor for coronary artery disease (CAD), is associated with inflammation and reactive oxygen species (ROS) production, while advanced glycation end-products, through their receptor (AGER or RAGE), play an important role on these processes. The aim of this study was to analyze the expression levels of RAGE, NADPH oxidase subunits, and catalase in adipose tissue in relation with CAD. DESIGN AND METHODS: Patients undergoing heart surgery were included in two groups: with and without CAD. Epicardial adipose tissue (EAT) and subcutaneous adipose tissue (SAT) biopsies were analyzed for gene expression by RT-quantitative PCR, immunohistochemistry, or western blot. RESULTS: RAGE mRNA and protein expression in SAT from patients with CAD was lower than in patients without CAD. However, there was no change in EAT from patients with or without CAD. P22-PHOX and RAGE gene expression were higher in EAT than in SAT, whereas catalase mRNA levels were lower. NADPH oxidase subunits and catalase mRNA expression were not influenced by CAD. Whereas NADPH oxidase-dependent oxidative response of SAT and EAT to lipid circulating levels could be different; glycemic levels were not related with the analyzed genes expression. CONCLUSIONS: This study demonstrates that RAGE expression in SAT, but not in EAT, is down-regulated in patients with CAD with respect to those without CAD. Although changes were not observed for NADPH oxidase subunits or catalase expression between CAD and non-CAD patients, a possible relationship between ROS production and RAGE expression in adipose tissues cannot be ruled out.