Genomic Characterization of Prevalent mcr-1, mcr-4, and mcr-5 Escherichia coli Within Swine Enteric Colibacillosis in Spain

Abstract

Antimicrobial agents are crucial for the treatment of many bacterial diseases in pigs, however, the massive use of critically important antibiotics such as colistin, fluoroquinolones and 3rd–4th-generation cephalosporins often selects for co-resistance. Based on a comprehensive characterization of 35 colistin-resistant Escherichia coli from swine enteric colibacillosis, belonging to prevalent Spanish lineages, the aims of the present study were to investigate the characteristics of E. coli clones successfully spread in swine and to assess the correlation of the in vitro results with in silico predictions from WGS data. The resistome analysis showed six different mcr variants: mcr-1.1; mcr-1.10; mcr-4.1; mcr-4.2; mcr-4.5; and mcr-5.1. Additionally, blaCTX–M–14, blaCTX–M–32 and blaSHV–12 genes were present in seven genomes. PlasmidFinder revealed that mcr-1.1 genes located mainly on IncHI2 and IncX4 types, and mcr-4 on ColE10-like plasmids. Twenty-eight genomes showed a gyrA S83L substitution, and 12 of those 28 harbored double-serine mutations gyrA S83L and parC S80I, correlating with in vitro quinolone-resistances. Notably, 16 of the 35 mcr-bearing genomes showed mutations in the PmrA (S39I) and PmrB (V161G) proteins. The summative presence of mechanisms, associated with high-level of resistance to quinolones/fluoroquinolones and colistin, could be conferring adaptive advantages to prevalent pig E. coli lineages, such as the ST10-A (CH11-24), as presumed for ST131. SerotypeFinder allowed the H-antigen identification of in vitro non-mobile (HNM) isolates, revealing that 15 of the 21 HNM E. coli analyzed were H39. Since the H39 is associated with the most prevalent O antigens worldwide within swine colibacillosis, such as O108 and O157, it would be probably playing a role in porcine colibacillosis to be considered as a valuable subunit antigen in the formulation of a broadly protective Enterotoxigenic E. coli (ETEC) vaccine. Our data show common features with other European countries in relation to a prevalent clonal group (CC10), serotypes (O108:H39, O138:H10, O139:H1, O141:H4), high plasmid content within the isolates and mcr location, which would support global alternatives to the use of antibiotics in pigs. Here, we report for first time a rare finding so far, which is the co-occurrence of double colistin-resistance mechanisms in a significant number of E. coli isolates.

Whole Genome Sequencing and Characteristics of mcr-1–Harboring Plasmids of Porcine Escherichia coli Isolates Belonging to the High-Risk Clone O25b:H4-ST131 Clade B

“…ColE1-like plasmids were the second more prevalent in our collection and the replication protein could not be determined; these results are in accordance with the findings of Lanza et al (2014). Furthermore, ColE plasmids have been reported as mcr-4 and mcr-5 carriers Carattoli et al, 2017;García-Meniño et al, 2019). The ColE1-like plasmids from this study did not carry resistance genes.…”

“…In contrast, F2:A-:B1 plasmids were dominant in the porcine clade B isolates used in this study. This type of plasmid was absent in 35 non-ST131 E. coli porcine isolates from colibacillosis (García-Meniño et al, 2019), suggesting that IncF plasmids vary greatly within the different lineages. Reid et al (2019) reported 23 F-types among 282 isolates from a mixture of origins belonging to the clade B of ST131, and the F2:A-:B1 IncF subtype was one of the commonest (32%).…”

“…However, it has been widely used in veterinary medicine. Livestock, and particularly porcine farming, has been singled out as reservoir for colistin resistance (García-Meniño et al, 2019) and foodborne pathogens (FBPs) that could have devastating health and economic consequences (Sekse et al, 2017).…”

“…et al, 2016;Olaitan et al, 2016;Pham Thanh et al, 2016;Stoesser et al, 2016a;Zeng et al, 2016;Zhao and Zong, 2016;AbuOun et al, 2017;Li et al, 2017c;Luo et al, 2017). Although mcr genes have been found in a large diversity of clones, Matamoros et al (2017) and García-Meniño et al (2019) established the ST10 and ST155 E. coli clones as potential reservoirs of the mcr-1 gene.…”

“…The mcr gene has been detected in many plasmid types, including IncI2, IncHI2, IncP, IncX4, IncY, IncF, and ColE10-like ones (Madec and Haenni, 2018) from different origins (Sun et al, 2018). Nevertheless, Matamoros et al (2017) and García-Meniño et al (2019) found that the majority of the mcr-carrier plasmids belonged mainly to four plasmid incompatibility groups: IncX4, IncI2, IncHI2, and ColE10-like. In contrast, it remains rare to find colistin resistance genes chromosomally encoded, in accordance to studies, where the prevalence of chromosomal mcr carrier isolates was estimated in 4% of the analyzed isolates.…”