By 2030, the global population will be 8.5 billion, placing pressure on international poultry production, of which China is a key producer. From April 2017, China will implement the withdrawal of colistin as a growth promoter, removing over 8,000 tonnes per year from the Chinese farming sector. To understand the impact of banning colistin and the epidemiology of multi-drug-resistant (MDR) Escherichia coli (using bla and mcr-1 as marker genes), we sampled poultry, dogs, sewage, wild birds and flies. Here, we show that mcr-1, but not bla, is prevalent in hatcheries, but bla quickly contaminates flocks through dogs, flies and wild birds. We also screened samples directly for resistance genes to understand the true breadth and depth of the environmental and animal resistome. Direct sample testing for bla and mcr-1 in hatcheries, commercial farms, a slaughterhouse and supermarkets revealed considerably higher levels of positive samples than the bla- and mcr-1-positive E. coli, indicating a substantial segment of unseen resistome-a phenomenon we have termed the 'phantom resistome'. Whole-genome sequencing identified common bla-positive E. coli shared among farms, flies, dogs and farmers, providing direct evidence of carbapenem-resistant E. coli transmission and environmental contamination.

Two adjacent colistin resistance gene variants, termed and-like, were identified in the chromosome of an isolate obtained from retail chicken meat. The variants showed 95.20% and 84.19% nucleotide sequence identity, respectively, to from porcine Functional cloning indicated that only conferred polymyxin resistance in both and The -like segment was also observed in other species, including ,, and .

This study investigated the characteristics of Escherichia coli isolates carrying mcr-1-bla NDM from a chicken farm in China. Of the 78 E. coli isolates, 21 clonally unrelated isolates carried mcr-1-bla NDM . Diverse IncI2 plasmids disseminated mcr-1, while the dissemination of bla NDM was mediated by diverse IncB/O plasmids. More striking was the colocalization of resistance genes mcr-1 and bla in an IncHI2/ST3 plasmid, which might pose a great challenge for public health.KEYWORDS high incidence, mcr-1, bla NDM , colocalization, Escherichia coli C arbapenems have been reliable and potent agents against Gram-negative bacteria. The rapid increase of carbapenem-resistant Enterobacteriaceae poses a great threat to public health and has prompted the reconsideration of colistin as a last-resort therapeutic option (1). Recently, a plasmid-borne colistin resistance gene (mcr-1) was identified in Escherichia coli and Klebsiella pneumoniae strains from animals and humans in China (2). This finding foreshadowed the inevitable dissemination of colistin resistance worldwide and was confirmed by the presence of mcr-1 in other countries (3-6).The mcr-1 gene is often associated with the extended-spectrum ␤-lactamase gene and has been found with bla CTX-M in the same plasmid (4,7,8). The mcr-1 gene has also emerged in carbapenem-resistant isolates (4, 9, 10), and the cotransfer of mcr-1 and carbapenem resistance genes is obviously of great clinical concern. Recently, mcr-1 was even found with bla NDM in the same IncX3-X4 hybrid plasmid in E. coli from pets in China (11). However, only a few isolates resistant to both carbapenem and colistin were characterized in those previous studies.Food-producing animals, especially chickens, serve as resistance gene "reservoirs," so it is crucial to identify the origins of multidrug-resistant plasmids in these animals. In this study, we investigated the genetically diverse E. coli isolates carrying mcr-1 and bla NDM from a chicken farm and characterized the plasmids harboring mcr-1 or bla NDM .Seventy-eight E. coli isolates were collected from diseased chickens in four separate barns on a large chicken farm (100,000 animals) in Shandong Province, China in October 2015. Feces from chickens that showed signs of diarrhea were randomly collected and streaked onto MacConkey agar. After incubating at 37°C for 20 h, one colony with typical E. coli morphology was selected from each chicken sample. Although detailed information on antibiotic usage was not available, ceftiofur and colistin were often used for prophylaxis and treatment of bacterial infections on this farm.Susceptibilities to 18 antimicrobials were determined for these 78 isolates by the agar dilution method using the recommended breakpoints (12, 13). The colistin breakpoint (Ն2 g/ml) was used according to recommendations by the European Committee for Antimicrobial Susceptibility Testing.

T he rapid rise and dissemination of multidrug-resistant (MDR) bacteria are a major threat to public health worldwide and have narrowed the treatment options for infections caused by these bacteria (1). Colistin is one of the last-resort drugs for the treatment of infections caused by MDR Gram-negative bacteria. However, within the past 2 years, shortly after the report of the first plasmid-mediated colistin resistance gene, mcr-1, in 2015, four other mobile colistin resistance genes (mcr-2, mcr-3, mcr-4, and mcr-5) and multiple variants have been reported (2-6). Compared to mcr-2, mcr-4, and mcr-5, which have been detected solely in Europe so far, both mcr-1 and mcr-3 have been identified globally. Here, we report a novel mcr-3 variant in Aeromonas caviae, Proteus mirabilis, and Escherichia coli from a single domestic duck. A total of 15 cloacal samples were obtained from free-range domestic ducks near a river in a suburban area of Qingdao City, Shandong Province, China, in March 2017. Direct sample testing was then performed on all 15 samples to detect the mcr-3 gene, and only one sample was identified as mcr-3 positive. The sample positive for mcr-3 was further isolated on CHROMagar orientation agar plates (bioMérieux, Lyon, France) containing 2 mg/liter colistin. Three mcr-3-positive strains, A. caviae 17AC, P. mirabilis 17PM, and E. coli 17EC, were obtained, and their sequences were confirmed by using 16S rRNA gene sequencing and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry analysis. A. caviae 17AC, P. mirabilis 17PM, and E. coli 17EC were then subjected to 150-bp paired-end whole-genome sequencing (WGS) using the Illumina HiSeq 2500 platform (Annoroad, Beijing, China). The draft genomes were assembled using CLC Genomics Workbench 9.0 (CLC Bio, Aarhus, Denmark), and all contigs were searched for mcr-3 by standalone BLAST analysis. WGS analysis identified mcr-3-carrying fragments in each genome, including a 26.2-kb contig from 17AC, a 17.8-kb contig from 17PM, and a 13.0-kb contig from 17EC. An mcr-3 variant gene in three contigs showed 98.83% nucleotide sequence identity to mcr-3 from porcine E. coli. The deduced protein sequence differed from MCR-3.1 by seven amino acid substitutions, one of which (V122G) was located in a putative transmembrane region, while the remaining six (R297L, I313V, E337K, H341Y, D358E, and Q468K) were in the catalytic domain. This novel mcr-3 variant was designated mcr-3.10. A. caviae 17AC and E. coli 17EC belong to novel sequence types (STs), ST513 and ST457, respectively. E. coli of ST457 carrying mcr-1 has been reported from humans in the United States and Vietnam (7, 8). Plasmid replicon typing revealed that the mcr-3.10-carrying plasmid in both 17EC and its transconjugant T-17EC belongs to the IncI2 replicon.

No abstract