Uropathogenic Escherichia coli Releases Extracellular Vesicles That Are Associated with RNA.

BACKGROUND: Bacterium-to-host signalling during infection is a complex process involving proteins, lipids and other diffusible signals that manipulate host cell biology for pathogen survival. Bacteria also release membrane vesicles (MV) that can carry a cargo of effector molecules directly into host cells. Supported by recent publications, we hypothesised that these MVs also associate with RNA, which may be directly involved in the modulation of the host response to infection. METHODS AND RESULTS: Using the uropathogenic Escherichia coli (UPEC) strain 536, we have isolated MVs and found they carry a range of RNA species. Density gradient centrifugation further fractionated and characterised the MV preparation and confirmed that the isolated RNA was associated with the highest particle and protein containing fractions. Using a new approach, RNA-sequencing of libraries derived from three different 'size' RNA populations (<50nt, 50-200nt and 200nt+) isolated from MVs has enabled us to now report the first example of a complete bacterial MV-RNA profile. These data show that MVs carry rRNA, tRNAs, other small RNAs as well as full-length protein coding mRNAs. Confocal microscopy visualised the delivery of lipid labelled MVs into cultured bladder epithelial cells and showed their RNA cargo labelled with 5-EU (5-ethynyl uridine), was transported into the host cell cytoplasm and nucleus. MV RNA uptake by the cells was confirmed by droplet digital RT-PCR of csrC. It was estimated that 1% of MV RNA cargo is delivered into cultured cells. CONCLUSIONS: These data add to the growing evidence of pathogenic bacterial MV being associated a wide range of RNAs. It further raises the plausibility for MV-RNA-mediated cross-kingdom communication whereby they influence host cell function during the infection process.

Characterization of Vibrio parahaemolyticus strains isolated in Chile in 2005 and in 2007.

INTRODUCTION: Vibrio (V.) parahaemolyticus has endemically established in Chilean sea shores, causing outbreaks every year, with an important number of cases. In order to know the genetic relationship, genotype dominance and antibiotic resistance of isolates obtained from two outbreaks, this study characterized 110 strains isolated from environmental and clinical samples in years 2005 and 2007 in Chile. METHODOLOGY: Genotyping was performed by determination of PFGE profiles, and pandemic group and integrons were screened by PCR. Antimicrobial susceptibility was studied by the disk diffusion method. RESULTS: High antibiotic susceptibility frequency was found, mainly among 2007 isolates, except to ampicillin, cephalothin, cefoxitin, cefpodoxime, amikacin, streptomycin and kanamycin. Strains belonging to the pandemic group in clinical isolates account for 88% in 2005, decreasing to 66% in 2007 and among environmental isolates were detected in 20% of the strains from 2005, rising to 36% in 2007. In 2005, nine different PFGE profiles were identified, with 78% of the strains corresponding to a single clone. In 2007, sixteen different PFGE profiles were detected, with 61% of the strains included into a sole clone. The same clone was prevalent in both years. None of class 1, 2, 3 and SXT integrases genes was detected; however, the superintegron integrase gene (intIA) was present in almost all strains. CONCLUSIONS: These results suggest the persistence and dominance of a unique PFGE clone of V. parahaemolyticus during 2005 and 2007, and the absence of genetic elements that capture antibiotic resistance genes described in other species of Vibrio.