Wastewaters belong among the most important sources of environmental pollution, including antibiotic-resistant bacteria. The aim of the study was to evaluate treated wastewaters as a possible transmission pathway for bacterial colonisation of gulls occupying the receiving river. A collection of antibiotic-resistant Escherichia coli originating both from treated municipal wastewaters discharged to the river Svratka (Czech Republic) and nestlings of Black-headed Gull (Chroicocephalus ridibundus) living 35 km downstream of the outlet was obtained using selective cultivation. Isolates were further characterised by various phenotyping and genotyping methods. From a total of 670 E. coli isolates (450 from effluents, 220 from gulls), 86 isolates (41 from effluents, 45 from gulls) showed identical antibiotic resistance phenotype and genotype and were further analysed for clonal relatedness using pulsed-field gel electrophoresis (PFGE). Despite the overall high diversity of the isolates, 21 isolates from both sources showed similar PFGE profiles. Isolates belonging to epidemiologically important sequence types (ST131, 15 isolates; ST23, three isolates) were subjected to whole-genome sequencing. Subsequent phylogenetic analysis did not reveal any close clonal relationship between the isolates from the effluents and gulls' nestlings with the closest strains showing 90 SNPs difference. Although our study did not provide direct evidence of transmission of antibiotic-resistant E. coli to wild gulls via treated wastewaters, we observed gull chicks as carriers of diverse multi-resistant E. coli, including high-risk clones, posing risk of further bacterial contamination of the surrounding environment.

• Keywords
• Enterobacterales, Environment, Whole-genome sequencing, Wild birds,
• Publication type
• Journal Article MeSH

BACKGROUND: Wastewaters are considered as important players in the spread of antimicrobial resistance, thus affecting the health of humans and animals. Here, we focused on wastewaters as a possible source of carbapenemase-producing Enterobacterales for the environment. METHODS: A total of 180 presumptive coliforms from hospital and municipal wastewaters, and a river in the Czech Republic were obtained by selective cultivation on meropenem-supplemented media and tested for presence of carbapenemase-encoding genes by PCR. Strains carrying genes of interest were characterized by testing antimicrobial susceptibility, carbapenemase production and combination of short- and long- read whole-genome sequencing. The phylogenetic tree including publicly available genomes of Enterobacter asburiae was conducted using Prokka, Roary and RAxML. RESULTS: Three VIM-producing Enterobacter asburiae isolates, members of the Enterobacter cloacae complex, were detected from hospital and municipal wastewaters, and the river. The blaVIM-1 gene was located within a class 1 integron that was carried by different F-type plasmids and one non-typeable plasmid. Furthermore, one of the isolates carried plasmid-borne colistin-resistance gene mcr-10, while in another isolate chromosomally located mcr-9 without colistin resistance phenotype was detected. In addition, the analysis of 685 publicly available E. asburiae genomes showed they frequently carry carbapenemase genes, highlighting the importance of this species in the emergence of resistance to last-line antibiotics. CONCLUSION: Our findings pointed out the important contribution of hospital and community wastewaters in transmission of multi-drug resistant pathogens.

• Keywords
• mcr, Antimicrobial resistance, Carbapenemase, Environment,
• MeSH
• Anti-Bacterial Agents pharmacology   MeSH
• Drug Resistance, Bacterial genetics   MeSH
• Bacterial Proteins genetics   MeSH
• beta-Lactamases * genetics   MeSH
• Enterobacter * genetics   drug effects   isolation & purification   MeSH
• Phylogeny MeSH
• Colistin * pharmacology   MeSH
• Humans MeSH
• Microbial Sensitivity Tests MeSH
• Wastewater * microbiology   MeSH
• Plasmids genetics   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Anti-Bacterial Agents MeSH
• Bacterial Proteins MeSH
• beta-Lactamases * MeSH
• carbapenemase MeSH Browser
• Colistin * MeSH
• Wastewater * MeSH
• VIM-1 metallo-beta-lactamase MeSH Browser

Extraintestinal pathogenic Escherichia coli (ExPEC) sequence type (ST) 38 is one of the top 10 human pandemic lineages. Although a major cause of urinary tract and blood stream infections, ST38 has been poorly characterized from a global phylogenomic perspective. A comprehensive genome-scale analysis of 925 ST38 isolate genomes identified two broad ancestral clades and linkage of discrete ST38 clusters with specific bla CTX-M variants. In addition, the clades and clusters carry important virulence genes, with diverse but poorly characterized plasmids. Numerous putative interhost and environment transmission events were identified here by the presence of ST38 clones (defined as isolates with ≤35 SNPs) within humans, companion animals, food sources, urban birds, wildlife, and the environment. A small cluster of international ST38 clones from diverse sources, likely representing progenitors of a hospital outbreak that occurred in Brisbane, Australia, in 2017, was also identified. Our study emphasizes the importance of characterizing isolate genomes derived from nonhuman sources and geographical locations, without any selection bias.

• Keywords
• EAEC, One Health, ST38, bla CTX-M, drug resistance, enteroaggregative E. coli, genomic surveillance, phylogenomics, β-lactamase genes,
• MeSH
• Escherichia coli genetics   MeSH
• Extraintestinal Pathogenic Escherichia coli * MeSH
• Phylogeny MeSH
• Escherichia coli Infections * epidemiology   MeSH
• Humans MeSH
• Plasmids MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

INTRODUCTION: Hospitals and wastewater are recognized hot spots for the selection and dissemination of antibiotic-resistant bacteria to the environment, but the total participation of hospitals in the spread of nosocomial pathogens to municipal wastewater treatment plants (WWTPs) and adjacent rivers had not previously been revealed. METHODS: We used a combination of culturing and whole-genome sequencing to explore the transmission routes of Escherichia coli from hospitalized patients suffering from urinary tract infections (UTI) via wastewater to the environment. Samples were collected in two periods in three locations (A, B, and C) and cultured on selective antibiotic-enhanced plates. RESULTS: In total, 408 E. coli isolates were obtained from patients with UTI (n=81), raw hospital sewage (n=73), WWTPs inflow (n=96)/outflow (n=106), and river upstream (n=21)/downstream (n=31) of WWTPs. The majority of the isolates produced extended-spectrum beta-lactamase (ESBL), mainly CTX-M-15, and showed multidrug resistance (MDR) profiles. Seven carbapenemase-producing isolates with GES-5 or OXA-244 were obtained in two locations from wastewater and river samples. Isolates were assigned to 74 different sequence types (ST), with the predominance of ST131 (n=80) found in all sources including rivers. Extraintestinal pathogenic lineages frequently found in hospital sewage (ST10, ST38, and ST69) were also found in river water. Despite generally high genetic diversity, phylogenetic analysis of ST10, ST295, and ST744 showed highly related isolates (SNP 0-18) from different sources, providing the evidence for the transmission of resistant strains through WWTPs to surface waters. DISCUSSION: Results of this study suggest that 1) UTI share a minor participation in hospitals wastewaters; 2) a high diversity of STs and phylogenetic groups in municipal wastewaters derive from the urban influence rather than hospitals; and 3) pathogenic lineages and bacteria with emerging resistance genotypes associated with hospitals spread into surface waters. Our study highlights the contribution of hospital and municipal wastewater to the transmission of ESBL- and carbapenemase-producing E. coli with MDR profiles to the environment.

• Keywords
• Escherichia coli, antibiotic resistance, beta-lactamases, wastewater, whole-genome sequencing,
• MeSH
• Anti-Bacterial Agents pharmacology   MeSH
• beta-Lactamases genetics   MeSH
• Escherichia coli genetics   MeSH
• Phylogeny MeSH
• Urinary Tract Infections * microbiology   MeSH
• Escherichia coli Infections * microbiology   MeSH
• Humans MeSH
• Microbial Sensitivity Tests MeSH
• Multilocus Sequence Typing MeSH
• Hospitals MeSH
• Wastewater MeSH
• Sewage microbiology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Anti-Bacterial Agents MeSH
• beta-Lactamases MeSH
• Wastewater MeSH
• Sewage MeSH

Escherichia coli sequence type 963 (ST963) is a neglected lineage closely related to ST38, a globally widespread extraintestinal pathogenic ST causing urinary tract infections (UTI) as well as sepsis in humans. Our current study aimed to improve the knowledge of this understudied ST by carrying out a comprehensive comparative analysis of whole-genome sequencing data consisting of 31 isolates from silver gulls in Australia along with another 80 genomes from public resources originating from geographically scattered regions. ST963 was notable for carriage of cephalosporinase gene blaCMY-2, which was identified in 99 isolates and was generally chromosomally encoded. ST963 isolates showed otherwise low carriage of antibiotic resistance genes, in contrast with the closely related E. coli ST38. We found considerable phylogenetic variability among international ST963 isolates (up to 11,273 single nucleotide polymorphisms [SNPs]), forming three separate clades. A major clade that often differed by 20 SNPs or less consisted of Australian isolates of both human and animal origin, providing evidence of zoonotic or zooanthropogenic transmission. There was a high prevalence of virulence F29:A-:B10 pUTI89-like plasmids within E. coli ST963 (n = 88), carried especially by less variable isolates exhibiting ≤1,154 SNPs. We characterized a novel 115,443-bp pUTI89-like plasmid, pCE2050_A, that carried a traS:IS5 insertion absent from pUTI89. Since IS5 was also present in a transposition unit bearing blaCMY-2 on chromosomes of ST963 strains, IS5 insertion into pUTI89 may enable mobilization of the blaCMY-2 gene from the chromosome/transposition unit to pUTI89 via homologous recombination. IMPORTANCE We have provided the first comprehensive genomic study of E. coli ST963 by analyzing various genomic and phenotypic data sets of isolates from Australian silver gulls and comparison with genomes from geographically dispersed regions of human and animal origin. Our study suggests the emergence of a specific blaCMY-2-carrying E. coli ST963 clone in Australia that is widely spread across the continent by humans and birds. Genomic analysis has revealed that ST963 is a globally dispersed lineage with a remarkable set of virulence genes and virulence plasmids described in uropathogenic E. coli. While ST963 separated into three clusters, a unique specific clade of Australian ST963 isolates harboring a chromosomal copy of AmpC β-lactamase encoding the gene blaCMY-2 and originating from both humans and wild birds was identified. This phylogenetically close cluster comprised isolates of both animal and human origin, thus providing evidence of interspecies zoonotic transmission. The analysis of the genetic environment of the AmpC β-lactamase-encoding gene highlighted ongoing evolutionary events that shape the carriage of this gene in ST963.

• Keywords
• Australia, Escherichia coli, ST963, WGS, comparative genomics, humans, phylogenetic analysis, silver gulls, transmission, β-lactamases,
• MeSH
• Charadriiformes * microbiology   MeSH
• Escherichia coli * genetics   MeSH
• Phylogeny MeSH
• Escherichia coli Infections * transmission   veterinary   MeSH
• Humans MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Australia MeSH

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 β-lactamase gene that encodes the ability to hydrolyze oxyimino β-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∆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∆260.

• Keywords
• AMR, Escherichia coli, IS26, Tn21, antibiotic, extended spectrum β-lactamase,
• Publication type
• Journal Article MeSH

The Australian silver gull is an urban-adapted species that frequents anthropogenic waste sites. The enterobacterial flora of synanthropic birds often carries antibiotic resistance genes. Whole-genome sequence analyses of 425 Escherichia coli isolates from cloacal swabs of chicks inhabiting three coastal sites in New South Wales, Australia, cultured on media supplemented with meropenem, cefotaxime, or ciprofloxacin are reported. Phylogenetically, over 170 antibiotic-resistant lineages from 96 sequence types (STs) representing all major phylogroups were identified. Remarkably, 25 STs hosted the carbapenemase gene blaIMP-4, sourced only from Five Islands. Class 1 integrons carrying blaIMP and blaOXA alongside blaCTX-M and qnrS were notable. Multiple plasmid types mobilized blaIMP-4 and blaOXA-1, and 121 isolates (28%) carried either a ColV-like (18%) or a pUTI89-like (10%) F virulence plasmid. Phylogenetic comparisons to human isolates provided evidence of interspecies transmission. Our study underscores the importance of bystander species in the transmission of antibiotic-resistant and pathogenic E. coli. IMPORTANCE By compiling various genomic and phenotypic data sets, we have provided one of the most comprehensive genomic studies of Escherichia coli isolates from the Australian silver gull, on media containing clinically relevant antibiotics. The analysis of genetic structures capturing antimicrobial resistance genes across three gull breeding colonies in New South Wales, Australia, and comparisons to clinical data have revealed a range of trackable genetic signatures that highlight the broad distribution of clinical antimicrobial resistance in more than 170 different lineages of E. coli. Conserved truncation sizes of the class 1 integrase gene, a key component of multiple-drug resistance structures in the Enterobacteriaceae, represent unique deletion events that are helping to link seemingly disparate isolates and highlight epidemiologically relevant data between wildlife and clinical sources. Notably, only the most anthropogenically affected of the three sites (Five Islands) was observed to host carbapenem resistance, indicating a potential reservoir among the sites sampled.

• Keywords
• AMR, Escherichia coli, genomics, wildlife,
• MeSH
• Anti-Bacterial Agents pharmacology   MeSH
• Drug Resistance, Microbial MeSH
• Anti-Infective Agents * MeSH
• Charadriiformes * microbiology   MeSH
• Animals, Wild MeSH
• Enterobacteriaceae MeSH
• Escherichia coli genetics   MeSH
• Phylogeny MeSH
• Humans MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Australia epidemiology   MeSH
• Names of Substances
• Anti-Bacterial Agents MeSH
• Anti-Infective Agents * MeSH

Escherichia coli ST216, including those that carry blaKPC-2, blaFOX-5, blaCTX-M-15 and mcr-1, have been linked to wild and urban-adapted birds and the colonisation of hospital environments causing recalcitrant, carbapenem-resistant human infections. Here we sequenced 22 multiple-drug resistant ST216 isolates from Australian silver gull chicks sampled from Five Islands, of which 21 carried nine or more antibiotic resistance genes including blaIMP-4 (n = 21), blaTEM-1b (n = 21), aac(3)-IId (n = 20), mph(A) (n = 20), catB3 (n = 20), sul1 (n = 20), aph(3")-Ib (n = 18) and aph(6)-Id (n = 18) on FIB(K) (n = 20), HI2-ST1 (n = 11) and HI2-ST3 (n = 10) plasmids. We show that (i) all HI2 plasmids harbour blaIMP-4 in resistance regions containing In809 flanked by IS26 (HI2-ST1) or IS15DI (HI2-ST3) and diverse metal resistance genes; (ii) HI2-ST1 plasmids are highly related to plasmids reported in diverse Enterobacteriaceae sourced from humans, companion animals and wildlife; (iii) HI2 were a feature of the Australian gull isolates and were not observed in international ST216 isolates. Phylogenetic analyses identified close relationships between ST216 from Australian gull and clinical isolates from overseas. E. coli ST216 from Australian gulls harbour HI2 plasmids encoding resistance to clinically important antibiotics and metals. Our studies underscore the importance of adopting a one health approach to AMR and pathogen surveillance.

• Keywords
• Australian silver gull, Chroicocephalus novaehollandiae, ST216, anthropogenic pollution, urban birds, whole genome sequencing, wildlife,
• Publication type
• Journal Article MeSH

Wild corvids were examined for the presence of carbapenemase-producing Gram-negative bacteria in the United States. A total of 13 isolates were detected among 590 fecal samples of American crow; 11 Providencia rettgeri isolates harboring blaIMP-27 on the chromosome as a class 2 integron gene cassette within the Tn7 transposon, 1 Klebsiella pneumoniae ST258 isolate carrying blaKPC-2 on a pKpQIL-like plasmid as a part of Tn4401a, and 1 Enterobacter bugandensis isolate with blaIMI-1 located within EcloIMEX-2.

• Keywords
• IMI, IMP-27, KPC, Providencia rettgeri, carbapenemase, wild birds,
• MeSH
• Anti-Bacterial Agents pharmacology   MeSH
• Bacterial Proteins genetics   MeSH
• beta-Lactamases genetics   MeSH
• Enterobacter MeSH
• Klebsiella Infections * MeSH
• Klebsiella pneumoniae genetics   MeSH
• Microbial Sensitivity Tests MeSH
• Plasmids genetics   MeSH
• Providencia MeSH
• Crows * MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• United States MeSH
• Names of Substances
• Anti-Bacterial Agents MeSH
• Bacterial Proteins MeSH
• beta-Lactamases MeSH
• carbapenemase MeSH Browser

Silver gulls carry phylogenetically diverse Escherichia coli, including globally dominant extraintestinal pathogenic E. coli (ExPEC) sequence types and pandemic ExPEC-ST131 clades; however, our large-scale study (504 samples) on silver gulls nesting off the coast of New South Wales identified E. coli ST457 as the most prevalent. A phylogenetic analysis of whole-genome sequences (WGS) of 138 ST457 samples comprising 42 from gulls, 2 from humans (Australia), and 14 from poultry farmed in Paraguay were compared with 80 WGS deposited in public databases from diverse sources and countries. E. coli ST457 strains are phylogenetic group F, carry fimH145, and partition into five main clades in accordance to predominant flagella H-antigen carriage. Although we identified considerable phylogenetic diversity among the 138 ST457 strains, closely related subclades (<100 SNPs) suggested zoonotic or zooanthroponosis transmission between humans, wild birds, and food-producing animals. Australian human clinical and gull strains in two of the clades were closely related (≤80 SNPs). Regarding plasmid content, country, or country/source, specific connections were observed, including I1/ST23, I1/ST314, and I1/ST315 disseminating blaCMY-2 in Australia, I1/ST113 carrying blaCTX-M-8 and mcr-5 in Paraguayan poultry, and F2:A-:B1 plasmids of Dutch origin being detected across multiple ST457 clades. We identified a high prevalence of nearly identical I1/ST23 plasmids carrying blaCMY-2 among Australian gull and clinical human strains. In summary, ST457 is a broad host range, geographically diverse E. coli lineage that can cause human extraintestinal disease, including urinary tract infection, and displays a remarkable ability to capture mobile elements that carry and transmit genes encoding resistance to critically important antibiotics.

• Keywords
• AmpC, ESBL, ExPEC, I1 plasmids, ST457,
• MeSH
• Anti-Bacterial Agents pharmacology   MeSH
• beta-Lactamases genetics   MeSH
• beta-Lactams MeSH
• Animals, Wild MeSH
• Escherichia coli * genetics   MeSH
• Phylogeny MeSH
• Escherichia coli Infections * veterinary   MeSH
• Humans MeSH
• Plasmids genetics   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Australia MeSH
• Names of Substances
• Anti-Bacterial Agents MeSH
• beta-Lactamases MeSH
• beta-Lactams MeSH