Objective: The aim of this study was to detect potential animal reservoirs of Escherichia coli carrying the mcr-1 gene in an Ecuadorian household. Methods: The mobile colistin-resistance gene, mcr-1, was first detected in Ecuador in a commensal E. coli isolate from a boy. A cross-sectional study was performed to detect the possible source of colistin-resistant E. coli in the boy's household. Faecal swabs and soil faecal samples were collected from companion animals. Samples were plated on selective media to isolate colistin-resistant E. coli and isolates were submitted to PCR detection of mcr-1, pulsed field gel electrophoresis (PFGE), and multi-locus sequences typing (MLST). Moreover, the genomes of all the isolates were sequenced. Results: Three different colistin-resistant E. coli sequence types (ST3941,1630 and 2170), corresponding to three PFGE patterns, were obtained from a chicken and two dogs; these isolates were different from the human isolate (ST609). By whole-genome sequencing, the mcr-1.1 gene was found on Incl2 plasmids with very high nucleotide identity. Conclusions: Our results indicate a polyclonal dissemination of mcr-1.1 in the environment surrounding the first MCR-producing E. coli strain reported in Ecuador. Our findings support the idea of lateral dissemination of mcr-1.1 gene between unrelated E. coli isolates. Crown Copyright (C) 2019 Published by Elsevier Ltd on behalf of International Society for Antimicrobial Chemotherapy.

To assess the molecular events exerted by Helicobacter pylori interacting directly with gastric epithelial cells, an improved procedure for microbial DNA isolation from stained hematoxilin-eosin gastric biopsies was developed based on laser micro-dissection (LM) [1]. Few articles have described the use of LM to select and detect H. pylori genome from formalin-fixed paraffin embedded gastric tissue [2]. To improve the yield and quality of DNA isolated from H. pylori contacting intestinal epithelial cells, the following conditions were established after modification of the QIAamp DNA Micro kit. • Use of at least 25 cut sections of 10–20 μm of diameter and 3 μm thick with more than 10 bacteria in each cut. • Lysis with 30 μL of tissue lysis buffer and 20 μL of proteinase K (PK) with the tube in an upside-down position. • The use of thin purification columns with 35 μL of elution buffer. The mean of DNA concentration obtained from 25 LM cut sections was 1.94± 0 .16 ng/μL, and it was efficiently amplified with qPCR in a Bio Rad iCycler instrument. The LM can improve the sample selection and DNA extraction for molecular analysis of H. pylori associated with human gastric epithelium.

The aim of this study was to investigate the presence of Escherichia coli carrying mcr-1 gene in domestic animals close to a child who suffered a peritoneal infection by a mcr-1 positive E. coli . Rectal or cloacal swabs and fecal samples from domestic animals were plated on selective media to isolate colistin-resistant E. coli and isolates were submitted to detection of mcr-1 gene, pulsed field gel electrophoresis (PFGE), multi-locus sequence typing (MLST), replicon typing and S1-PFGE. Four mcr-1 positive E. coli isolates (from chicken, turkey and dog) were recovered. No shared PFGE pattern or MLST sequence type were observed among isolates. A 60Kb IncI1 γ mcr -1-carrying plasmid was detected in all isolates. Our results suggest that mcr-1 gene was horizontally disseminated amongst different lineages of E. coli from domestic animals in the child’s household. Importance: Horizontally transferable colistin resistance ( mcr -1 gene) is thought to have originated in domestic animals and transferred to humans through meat and dairy products. In the present report we show evidence that the mcr -1 gene could be transferred to different E. coli strains colonizing different hosts (humans and pets) in the same household.

The exponential increase in the numbers of isolates of Carbapenem-Resistant Enterobacteriaceae (CRE) creates the need for using novel therapeutic approaches to save the lives of patients. Fosfomycin has long been considered a rational option for the treatment of CRE to be used as part of a combined therapy scheme. However, the assessment of fosfomycin susceptibility in the laboratory presents a great challenge due to the discrepancies found between different methodologies. Thus, our goal was to evaluate fosfomycin susceptibility in a group of 150 Enterobacteriaceae bacterial isolates using agar dilution as the gold standard technique to compare the results with those obtained by disk diffusion. We found a fosfomycin susceptibility of 79.3% in general terms. By comparing both methodologies, we reported a categorical agreement of 96% without Very Major Errors (VMEs) or Major Errors (MEs) and 4% of minor Errors (mEs). Our results suggest that fosfomycin could provide a rational alternative treatment for those patients that are infected by a Multidrug-Resistant (MDR) microorganism that is currently untreatable and that the disk diffusion and classical agar dilution techniques are adequate to assess the resistance profile of CRE to fosfomycin.