Plasmid-mediated colistin resistance among human clinical Enterobacterales isolates: national surveillance in the Czech Republic

The occurrence of colistin resistance has increased rapidly among Enterobacterales around the world. We performed a national survey of plasmid-mediated colistin resistance in human clinical isolates through a retrospective analysis of samples from 2009 to 2017 and a prospective sampling in 2018–2020. The aim of this study was to identify and characterize isolates with mcr genes from various regions of the Czech Republic using whole genome sequencing (WGS). Of all 1932 colistin-resistant isolates analyzed, 73 (3.8%) were positive for mcr genes. Most isolates carried mcr-1 (48/73) and were identified as Escherichia coli (n = 44) and Klebsiella pneumoniae (n = 4) of various sequence types (ST). Twenty-five isolates, including Enterobacter spp. (n = 24) and Citrobacter freundii (n = 1) carrying the mcr-9 gene were detected; three of them (Enterobacter kobei ST54) co-harbored the mcr-4 and mcr-9 genes. Multi-drug resistance phenotype was a common feature of mcr isolates and 14% (10/73) isolates also co-harbored clinically important beta-lactamases, including two isolates with carbapenemases KPC-2 and OXA-48. Phylogenetic analysis of E. coli ST744, the dominant genotype in this study, with the global collection showed Czech isolates belonged to two major clades, one containing isolates from Europe, while the second composed of isolates from diverse geographical areas. The mcr-1 gene was carried by IncX4 (34/73, 47%), IncHI2/ST4 (6/73, 8%) and IncI2 (8/73, 11%) plasmid groups. Small plasmids belonging to the ColE10 group were associated with mcr-4 in three isolates, while mcr-9 was carried by IncHI2/ST1 plasmids (4/73, 5%) or the chromosome (18/73, 25%). We showed an overall low level of occurrence of mcr genes in colistin-resistant bacteria from human clinical samples in the Czech Republic

CTX-M15-producing Escherichia coli clone B2-O25b-ST131 and Klebsiella spp. isolates in municipal wastewater treatment plant effluents

Objectives: The global occurrence of antibiotic resistance genes in bacteria in water environments is an increasing concern. Treated wastewater was sampled daily over a 45 day period from the outflow of a municipal wastewater treatment plant in Brno, Czech Republic, and examined for extended-spectrum b-lactamase (ESBL)-producing bacteria. Methods: Water samples were cultivated on MacConkey agar with cefotaxime (2 mg/L) and individual colonies were examined for ESBL production. Phenotypic ESBL-positive bacteria identified as Escherichia coli or Klebsiella spp. were tested for the presence of antibiotic resistance genes, the virulence gene afa/dra and the bla CTX-M upstream region. Genetic relatedness was analysed by PFGE, multilocus sequence typing and plasmid analysis. Results: A total of 68 ESBL-producing Enterobacteriaceae isolates were detected in 34 out of 45 wastewater samples. ESBL-producing isolates included 26 E. coli isolates, 4 Klebsiella pneumoniae isolates and 1 Klebsiella oxytoca isolate. The pandemic and multiresistant B2-O25b-ST131 clone was predominant, being detected among 19 E. coli isolates, and 17 of the B2-O25b-ST131 isolates were positive for the FIA replicon and the afa/dra operon and had an IS26 element flanking bla CTX-M-15 . Seventeen of the B2-O25b-ST131 isolates showed closely related PFGE profiles (defined by 84% band similarity) and belonged to identical clonal groups. Conclusions: The results highlight the inadequacy of the treatment process in removing multiresistant bacteria from municipal wastewater and point to a risk of transmission of clinically important multiresistant strains, such as the pandemic ST131 clone, to the environment. This is the first study demonstrating the pandemic ST131 clone in wastewater

Phylogenomic analysis of a global collection of Escherichia coli ST38: evidence of interspecies and environmental transmission?

We performed a comprehensive phylogenomic analysis of 925 extraintestinal pathogenic Escherichia coli (ExPEC) ST38 genomes from 38 countries and diverse hosts and sources. The phylogeny resolved two broad clades: A (593 strains; 91% human) and B (332 isolates; 42% human), each with distinct ST38 clusters linked to the carriage of specific bla CTX-M alleles, often in association with other antibiotic resistance genes, class 1 integrons and specific plasmid replicon types. Co-carriage of fyuA and irp2 virulence genes, a reliable proxy for carriage of the Yersinia high-pathogenicity island, featured in 580 (62.7%) genomes. ST38 lineages carrying combinations of ExPEC and intestinal pathogenic Escherichia coli virulence factors were also identified. The F plasmid replicon was identified in 536 (58%) genomes, and 112 of these (21%) carry cjrABC-senB, a virulence operon frequently identified in pandemic ExPEC sequence types. Most (108; 96.4%) cjrABC-senB+ ST38 isolates were from human and other sources, except food animals, and were associated with F5:A-:B10 (41 isolates), F1:A2:B20 (20 isolates), and F24:A-:B1 (15 isolates) F replicon types. ST38 genomes that were inferred to carry a ColV-F virulence plasmid (69; 7.4%) were mostly from human (12; 17.4%), avian (26; 37.7%), or poultry (10; 6.9%) sources. We identified multiple examples of putative inter-host and host-environment transmission events, where genomes differed by <35 SNPs. This work emphasizes the importance of adopting a One Health approach for phylogenomic studies that seek to improve understanding of antimicrobial resistance and pathogen evolution

The potential of using E. coli as an indicator for the surveillance of antimicrobial resistance (AMR) in the environment

To understand the dynamics of antimicrobial resistance (AMR), in a One-Health perspective, surveillance play an important role. Monitoring systems already exist in the human health and livestock sectors, but there are no environmental monitoring programs. Therefore there is an urgent need to initiate environmental AMR monitoring programs nationally and globally, which will complement existing systems in different sectors. However, environmental programs should not only identify anthropogenic influences and levels of AMR, but they should also allow for identification of transmissions to and from human and animal populations. In the current review we therefore propose using antimicrobial resistant Escherichia coli as indicators for monitoring occurrence and levels of AMR in the environment, including wildlife.publishedVersio

Genomic Analysis of an I1 Plasmid Hosting a sul3-Class 1 Integron and blaSHV-12 within an Unusual Escherichia coli ST297 from Urban Wildlife

Wild birds, particularly silver gulls (Chroicocephalus novaehollandiae) that nest near anthropogenic sites, often harbour bacteria resistant to multiple antibiotics, including those considered of clinical importance. Here, we describe the whole genome sequence of Escherichia coli isolate CE1867 from a silver gull chick sampled in 2012 that hosted an I1 pST25 plasmid with blaSHV-12, a &beta;-lactamase gene that encodes the ability to hydrolyze oxyimino &beta;-lactams, and other antibiotic resistance genes. Isolate CE1867 is an ST297 isolate, a phylogroup B1 lineage, and clustered with a large ST297 O130:H11 clade, which carry Shiga toxin genes. The I1 plasmid belongs to plasmid sequence type 25 and is notable for its carriage of an atypical sul3-class 1 integron with mefB&#8710;260, a structure most frequently reported in Australia from swine. This integron is a typical example of a Tn21-derived element that captured sul3 in place of the standard sul1 structure. Interestingly, the mercury resistance (mer) module of Tn21 is missing and has been replaced with Tn2-blaTEM-1 and a blaSHV-12 encoding module flanked by direct copies of IS26. Comparisons to similar plasmids, however, demonstrate a closely related family of ARG-carrying plasmids that all host variants of the sul3-associated integron with conserved Tn21 insertion points and a variable presence of both mer and mefB truncations, but predominantly mefB&#8710;260

Antimicrobial Resistance in Fecal Escherichia coli Isolates from Healthy Urban Children of Two Age Groups in Relation to Their Antibiotic Therapy▿

The study was performed in the Czech Republic during 2007 to 2009. Of Escherichia coli isolates from 275 children aged 6 weeks, 36% (n = 177) were resistant to 1 to 7 antibiotics. Of isolates from 253 children aged 6 to 17 years, 24% (n = 205) were resistant to 1 to 5 antibiotics. There was no significant difference in the prevalences of antibiotic-resistant E. coli isolates between these groups of children, even though the consumptions of antibiotics were quite different

Fitness effects of blaCTX-M-15-harbouring F2:A1:B− plasmids on their native Escherichia coli ST131 H30Rx hosts

Objectives: To investigate the fitness effects of large blaCTX-M-15-harbouring F2:A1:B− plasmids on their native Escherichia coli ST131 H30Rx hosts. Methods: We selected five E. coli ST131 H30Rx isolates of diverse origin, each carrying an F2:A1:B− plasmid with the blaCTX-M-15 gene. The plasmid was eliminated from each isolate by displacement using an incompatible curing plasmid, pMDP5_cureEC958. WGS was performed to obtain complete chromosome and plasmid sequences of original isolates and to detect chromosomal mutations in ‘cured’ clones. High-throughput competition assays were conducted to determine the relative fitness of cured clones compared with the corresponding original isolates. Results: We were able to successfully eliminate the F2:A1:B− plasmids from all five original isolates using pMDP5_cureEC958. The F2:A1:B− plasmids produced non-significant fitness effects in three isolates and moderate reductions in relative fitness (3%–4%) in the two remaining isolates. Conclusions: We conclude that F2:A1:B− plasmids pose low fitness costs in their E. coli ST131 H30Rx hosts. This plasmid-host fitness compatibility is likely to promote the maintenance of antibiotic resistance in this clinically important E. coli lineage

Low Rates of Antimicrobial-Resistant Enterobacteriaceae in Wildlife in Taï National Park, Côte d’Ivoire, Surrounded by Villages with High Prevalence of Multiresistant ESBL-Producing Escherichia coli in People and Domestic Animals

Antimicrobial resistance genes can be found in all ecosystems, including those where antibiotic selective pressure has never been exerted. We investigated resistance genes in a collection of faecal samples of wildlife (non-human primates, mice), people and domestic animals (dogs, cats) in Côte d’Ivoire; in the chimpanzee research area of Taï National Park (TNP) and adjacent villages. Single bacteria isolates were collected from antibiotic-containing agar plates and subjected to molecular analysis to detect Enterobacteriaceae isolates with plasmid-mediated genes of extended-spectrum beta-lactamases (ESBLs) and plasmid-mediated quinolone resistance (PMQR). While the prevalence of ESBL-producing E. coli in the villages was 27% in people (n = 77) and 32% in dogs (n = 38), no ESBL-producer was found in wildlife of TNP (n = 75). PMQR genes, mainly represented by qnrS1, were also present in human- and dog-originating isolates from the villages (36% and 42% in people and dogs, respectively), but no qnrS has been found in the park. In TNP, different variants of qnrB were detected in Citrobacter freundii isolates originating non-human primates and mice. In conclusion, ESBL and PMQR genes frequently found in humans and domestic animals in the villages were rather exceptional in wildlife living in the protected area. Although people enter the park, the strict biosecurity levels they are obliged to follow probably impede transmission of bacteria between them and wildlife