Genomic Characterization of VIM and MCR Co-Producers: The First Two Clinical Cases, in Italy

Abstract

Background: the co-production of carbapenemases and mcr-genes represents a worrisome event in the treatment of Enterobacteriaceae infections. The aim of the study was to characterize the genomic features of two clinical Enterobacter cloacae complex (ECC) isolates, co-producing VIM and MCR enzymes, in Italy. Methods: species identification and antibiotic susceptibility profiling were performed using MALDI-TOF and broth microdilution methods, respectively. Transferability of the blaVIM- and mcr- type genes was verified through conjugation experiment. Extracted DNA was sequenced using long reads sequencing technology on the Sequel I platform (PacBio). Results: the first isolate showed clinical resistance against ertapenem yet was colistin susceptible (EUCAST 2020 breakpoints). The mcr-9.2 gene was harbored on a conjugative IncHI2 plasmid, while the blaVIM-1 determinant was harbored on a conjugative IncN plasmid. The second isolate, resistant to both carbapenems and colistin, harbored: mcr-9 gene and its two component regulatory genes for increased expression on the chromosome, mcr-4.3 on non-conjugative (yet co-transferable) ColE plasmid, and blaVIM-1 on a non-conjugative IncA plasmid. Conclusions: to our knowledge, this is the first report of co-production of VIM and MCR in ECC isolates in Italy.

Emergence of IncHI2 Plasmids With Mobilized Colistin Resistance (mcr)-9 Gene in ESBL-Producing, Multidrug-Resistant Salmonella Typhimurium and Its Monophasic Variant ST34 From Food-Producing Animals in Italy

“…In our three selected isolates, mcr-9 was demonstrated to be located on IncHI2/IncHI2A plasmids by the hybrid (Illumina-Oxford Nanopore) approach, together with other AMR genes conferring resistance to different antimicrobial classes, including the bla SHV −12 ESBL gene type detected in the two plasmids of pig origin, and different genes involved in heavy metals metabolism. Similarly, previous findings detected IncHI2-or IncHI2A-type plasmids in mcr-9 positive Enterobacteriaceae, often together with other resistance determinants (including HPCIAs) and to heavy metals mainly from human patients (Carroll et al, 2019;Chavda et al, 2019;Kieffer et al, 2019;Bitar et al, 2020;Faccone et al, 2020;Kananizadeh et al, 2020;Lin et al, 2020;Osei Sekyere et al, 2020;Soliman et al, 2020;Tsui et al, 2020;Umeda et al, 2020;Elbediwi et al, 2021;Marchetti et al, 2021;Sun et al, 2021), and also from animal (Börjesson et al, 2020;Yuan et al, 2019;Borowiak et al, 2020;Haenni et al, 2020;Leite et al, 2020), food (Borowiak et al, 2020;Sadek et al, 2020;Tyson et al, 2020), and environmental (Kamathewatta et al, 2020) sources. Few reports also described mcr-9 integrated into the chromosome of Citrobacter (Ribeiro et al, 2021) and Salmonella (Pan et al, 2020;Tyson et al, 2020) isolates, with a genetic context similar to the structure observed in mcr-9-harboring plasmid sequences, suggesting a possible mcr-9 transfer as a gene cassette between plasmids and chromosomes (Pan et al, 2020).…”

“…The results obtained with BLAST analysis with the two previously published IncHI2 mcr-9-carrying plasmids revealed that our three IncHI2 mcr-9-carrying plasmids were closely related (88% coverage and 99.9% identity) to the IncHI2 ∼293 kb plasmid (p3846) described in Italy by Marchetti et al (2021) in E. cloacae. As for the mcr-9/bla VIM −1 -carrying IncHI2 plasmid (pRH-R27, size ∼299 kb) described in Germany in S. Infantis from a fattening pig farm, it shared with our three mcr-9-carrying plasmids a coverage range of 80-84% and an identity of 99.9%.…”

“…The mcr-9-carrying plasmids obtained were also compared using BLAST (Altschul et al, 1990) with two previously published complete mcr-9-carrying plasmid sequences from Enterobacteriacae: (a) an mcr-9/bla VIM −1 -carrying plasmid named p3846_IncHI2_mcr (accession number CP052871) with a size of 293,138 bp, isolated from Enterobacter cloacae firstly identified in Italy from a human clinical case (Marchetti et al, 2021); (b) an mcr-9/bla VIM −1 -carrying plasmid named pRH-R27 (accession number LN555650.1) with a size of 299,305 bp from a Salmonella infantis isolated from a fattening pig farm in Germany (Falgenhauer et al, 2017).…”

Screening of mcr-1 among Gram-Negative Bacteria from Different Clinical Samples from ICU Patients in Alexandria, Egypt: One-Year Study

“… Khalifa et al (2020b) found mcr-9 in colistin-susceptible foodborne K. pneumoniae . The mcr-9 subtype was also reported by Marchetti et al (2021) who reported mcr-9.2 in a colistin susceptible E. cloacae . The mcr-10 was described by Wang et al (2020a) in Enterobacter roggenkampii that was susceptible to colistin.…”

Resistance Patterns, mcr-4 and OXA-48 Genes, and Virulence Factors of Escherichia coli from Apennine Chamois Living in Sympatry with Domestic Species, Italy

“…In addition to the phenotypic resistance, it was reported the occurrence of mcr-4 gene. In Italy, mcr-4 first characterization was in Salmonella enterica serovar Typhimurium in a pig [ 12 ], afterward, mcr-4 was also reported in Salmonella enterica serovar Typhimurium from human patients with gastroenteritis [ 40 ], in E. coli from various stages of the broiler production pyramid (breeder, manure and soil), [ 15 ] and in Enterobacter cloacae from a hospitalized elderly woman [ 41 ]. Based on the above research, our study represents the first report of mcr-4 occurrence in wildlife with particular regard to red deer and Apennine chamois species.…”