GERMAN FEDERAL INSTITUTE FOR RISK ASSESSMENT Federal Institute for Risk Assessment • Max-Dohrn-Str. 8-10 • 10589 Berlin, GERMANY • Phone + 49 30 - 184 12 - 0 • Fax + 49 30 - 184 12 – 99 0 99 • bfr@bfr.bund.de • www.bfr.bund.de Isolation and characterization of a novel mcr-5 carrying Escherichia coli plasmid from chicken feces in Germany Colistin is considered as an important antibiotic of the last resort against human infections with multidrug-resistant (MDR) Gram- negative bacteria. The emergence and spread of mobilizable colistin resistance in Enterobacteriaceae is associated with major public health concerns. Since 2015, several mobile colistin resistance genes were described coding for enzymes of the phosphoethanolamine-transferase family. Nowadays, eight different mcr-genes (mcr-1 to -8) were identified, mediating resistances to colistin in different bacterial genera (especially in Enterobacteriaceae). In this study, a novel mcr-5 carrying plasmid of a commensal E. coli recovered from chicken feces is described. Introduction Here we describe a novel mcr-5 plasmid-prototype from an E. coli isolate of the German national monitoring of zoonoses in food and livestock in 2013/2014. The isolate exhibited a MIC against colistin of 4 mg/L as determined by using broth microdilution according to the EUCAST guidelines. Basis genetic features of the Whole Genome Sequencing (WGS) analysis of the isolate are summarized in Table 1. S1-PFGE (Fig. 1) analysis and WGS revealed that the genome of the plasmid pEC1897-13 was 38 kb in size and exhibited 44 putative open reading frames (ORFs; Table 2), respectively. Basic properties of the E. coli isolate and its plasmid Transmissibility of the mcr-5 carrying plasmid K. Juraschek1,†, M. Borowiak1,†, D. Shamoun1, S. Schmoger1, A. Irrgang1, N. Pauly1, M. Grobbel1, A. Käsbohrer1,2, B. Malorny1 & J.A. Hammerl1 Bioinformatic analysis indicated that the plasmid belongs to the IncFII group, but represent a novel pMLST-allele that is closely relate to the allele FII-82. Beside mcr-5, no further resistance determinant was detected on the plasmid. Interestingly, the plasmid genome of pEC1897-13 obviously comprises all necessary components of a functional IncF conjugative- transfer system (Table 2). However, up to now no self-transmission of the plasmid was observed in E. coli by filter mating studies at 37°C under different conditions. Correspondence to Dr. Jens A. Hammerl, National Reference Laboratory for Antibiotic Resistance, Department of Biological Safety, German Federal Institute for Risk Assessment, Diedersdorfer Weg 1, 12277 Berlin, Germany; E-mail: jens-andre.hammerl@bfr.bund.de Here we report on a novel IncFII mcr-5 carrying plasmid-prototype recovered from chicken feces in 2013. The impact of the plasmid for the transmission of colistin resistances in unknown. However, despite of most of the described mcr-5 carrying plasmids pEC1897-13 carry a complex IncF-like transfer system that may be potentially functional under some specific circumstances. Conclusion 1German Federal Institute for Risk Assessment, Department of Biological Safety, Berlin, Germany; 2Institute for Veterinary Public Health, University of Veterinary Medicine, Vienna, Austria; † both author contributed equally to this work Isolate 13-AB01897 Year of isolation 2013 Source Chicken, feces Genome features Genome size (bp) 5,256,358 GC content (%) 50.44 Genes (total) 5,758 Genes (coding) 5,317 CDS (total) 5,653 CDS (coding) 5,317 RNA genes (total) 105 rRNAs (total) 5S, 16S, 23S 6, 7, 5 tRNAs 77 ncRNAs 10 Pseudo genes (total) 336 Aquired antimicrobial resistances Aminoglycoside aph(3'')-Ib (100%) aph(6)-Id (100%) aadA1 (100%) Beta-lactam blaTEM-1B (100%) Colistin mcr-5 (100%) MLS mph(B) (100%) Phenicol catA1 (99.85%) Sulphonamide sul1 (100%) sul2 (100%) Tetracycline tet(34) (84.75%, partial) tet(A) (100%) Trimethoprim dfrA1 (100%) Chromosomal point muations gyrA S83L PlasmidFinder (inc group) Col ColRNAI (91.74%, partial) Col(MG828)