Moraxella Species as Potential Sources of MCR-Like Polymyxin Resistance Determinants

Abstract

Plasmid-mediated resistance to polymyxins mediated by the MCR-1/2 determinants has been reported in Enterobacteriaceae worldwide. Using PCR-based and cloning strategies, a series of Moraxella spp. were screened for mcr-like genes. Moraxella spp. that are mainly animal pathogens but may also be human pathogens were identified as potential reservoirs of mcr-like genes.KEYWORDS polymyxin, plasmid, resistance, reservoir, animal, MCR R esistance to polymyxins in Enterobacteriaceae results mostly from chromosomal mutations in genes involved in modification of lipopolysaccharide (LPS) (1, 2). In particular, it has been shown that mutations, truncations, or insertions into genes encoding LPS-modifying enzymes may be responsible for acquired resistance to polymyxins in Enterobacteriaceae (1, 3, 4), Acinetobacter baumannii, and Pseudomonas aeruginosa (1, 5). However, the plasmid-mediated polymyxin resistance determinant MCR-1 was recently found in Escherichia coli and Klebsiella pneumoniae isolates recovered from humans and animals in China (6). The mcr-1 gene has been reported worldwide in other enterobacterial genera, including Enterobacter, Salmonella, and Shigella (7-11). This resistance was identified in Enterobacteriaceae isolated from various animal species, including cattle, chicken, and pigs (12-14), and from river samples and vegetables (15).MCR-1 is a 541-amino-acid phosphoethanolamine transferase that adds phosphoethanolamine to the lipid A moiety of LPS, leading to a more cationic LPS structure and consequently to resistance to polymyxins (6). Recently, the MCR-2 protein (538 amino acids long, 81% amino acid identity with MCR-1) was identified in Belgium from colistin-resistant E. coli (16). Additionally, the MCR-1.2 and MCR-1.3 variants (both exhibiting single amino acid substitutions versus MCR-1) were identified in Italy and China, respectively (17,18). Several plasmid types carrying the mcr-1 gene, belonging to the IncI2, IncHI2, or IncX4 incompatibility groups, have been identified (8,10,12). In addition, the mcr-1.2 and mcr-2 genes were found on IncX4 plasmid scaffolds (16,17).It is often suggested that animals treated with polymyxins represent a significant reservoir of polymyxin-resistant Gram-negative isolates, particularly MCR-producing isolates (19). In fact, polymyxins are heavily used in veterinary medicine, and Europeans, particularly in Spain and Italy, have a high consumption of those antibiotics for food-producing animals (20). The frequent identification of MCR-1-producing isolates from animals further supports the hypothesis that the animal world might be the primary source of this resistance determinant.

MCR-2-mediated plasmid-borne polymyxin resistance most likely originates from Moraxella pluranimalium

“…1 We recently showed that some Moraxella species might constitute putative reservoirs of MCR-like proteins, with corresponding genes located on the chromosome of these species. 6 Hence, MCRlike proteins were identified in Moraxella catarrhalis, Moraxella lincolnii, Moraxella porci and Moraxella osloensis. They all share significant amino acid identities with MCR-1 and MCR-2, ranging from 59% to 64%.…”

“…Internal outward primers were then designed and used for an inverse PCR strategy, as previously performed. 6 The sequence of the entire mcr gene was thus obtained and it revealed that this new enzyme (termed MCR-2.2) was almost identical to MCR-2 (99% amino acid identity), with only 8 amino acid differences out of the 538 constituting the MCR-2 enzyme, and shared 82% amino acid identity with MCR-1.…”

“…They all share significant amino acid identities with MCR-1 and MCR-2, ranging from 59% to 64%. 6 Even though these Moraxella species have been shown to carry intrinsic mcr-like genes, these genes remain quite distantly related to the plasmid-borne mcr-1 and mcr-2.…”

“…6 Then MICs of colistin were determined for the recombinant strains expressing mcr-1, mcr-2 and mcr-2.2 by broth microdilution (BMD), 1 and this showed that MCR-2.2 conferred exactly the same level of resistance as MCR-1 and MCR-2 (MIC 4 mg/L compared with 0.03 mg/L for the E. coli WT recipient strain), thus showing that the few differences in terms of amino acid sequence did not have an impact on resistance to colistin in E. coli.…”

In Vitro Study of IS Apl1 -Mediated Mobilization of the Colistin Resistance Gene mcr-1

“…This work confirms previous hypotheses (12) that the mcr-1 gene had been initially mobilized by two copies of ISApl1 from an unknown progenitor, targeting broad-host range plasmid(s) that subsequently transferred this resistance gene into Enterobacteriaceae. Interestingly, our very recent work showed that Moraxella species are natural sources of mcr-like genes, and may harbor ISApl1 elements (21). Therefore, mobilization of the mcr-1 gene might have occurred into a Moraxella species (still to be precisely identified) through an ISApl1-mediated transposition process.…”