23The aim of this study was to investigate the presence of Escherichia coli carrying mcr-1 gene 24 in domestic animals close to a child who suffered a peritoneal infection by a mcr-1 positive 25 E. coli. Rectal or cloacal swabs and fecal samples from domestic animals were plated on 26 selective media to isolate colistin-resistant E. coli and isolates were submitted to detection of 27 mcr-1 gene, pulsed field gel electrophoresis (PFGE), multi-locus sequence typing (MLST), 28 replicon typing and S1-PFGE. Four mcr-1 positive E. coli isolates (from chicken, turkey and 29 dog) were recovered. No shared PFGE pattern or MLST sequence type were observed among 30 isolates. A 60Kb IncI1 mcr-1-carrying plasmid was detected in all isolates. Our results 31 suggest that mcr-1 gene was horizontally disseminated amongst different lineages of E. coli 32 from domestic animals in the child's household. 33 Importance 34Horizontally transferable colistin resistance (mcr-1 gene) is thought to have originated in 35 domestic animals and transferred to humans through meat and dairy products. In the present 36 report we show evidence that the mcr-1 gene could be transferred to different E. coli strains 37 colonizing different hosts (humans and pets) in the same household. 38 42 different bacterial genera colonizing different animal species (2, 3).43 3The first report of a colistin resistance (CR) gene carried by plasmids (mcr-1) came from China in 44 2015 (4). This gene codes for a phosphoethanolamine transferase (MCR-1), which modifies the 45 lipid A moiety in the outer membrane of Gram negative bacteria and confers resistance to 46 polymyxins (4, 5). Among Enterobacteriacea different mcr gen groups (1-5) could be transferred by 47 PCR was performed to detect mcr gene (4); amplicons were sequence and aligned using mcr-1 130 (NG_055582.1), mcr-2 (NG_051171.1), mcr-3 ( NG_056184.1), mcr-4( MG822665.1), mcr-5 131 (MG384740.1) accession numbers with Geneius software. Pulsed field gel electrophoresis (PFGE) 132 (25) and multilocus sequence typing (MLST) was performed on seven housekeeping genes to 133 define clonal relatedness (26). Replicon typing was performed using a commercial kit (PBRT KIT, 134 DIATHE, Fano, Italy) (20, 27, 28). -lactamase genes (blaCTX-M-1, blaTEM, blaSHV) were detected 135 using primers previously described (29). Briefly, 12,5mL of GoTaq ® Green Master Mix 136 (Promega, Madison, USA) were mixed with 1L of upstream primer, 10M, 1L of downstream 137 primer, 10 L of DNA template and 9,5 L of Nuclease-free water to complete a 25L of 138 reaction volume. The reaction mix were amplified using 2-minute of initial denaturation at 94°C 139 followed by 40 cycles of DNA denaturation at 94°C (40 sec), annealing at 60°C (40sec) and 140 extension at 72°C (1min). The final elongation step at 72°C for 5 min. Amplicons were detected by 141 electrophoresis in a 2% agarose gel. For complete amplification and sequencing of the detected 142resistance genes, primers and conditions previously described were used (30). 143 S1-PFG...
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