Biology subjects

Turton, J. F.

Publications and source records attributed to Turton, J. F..

4 recordsLinked to original sources

bioRxiv2026

Two IncHI1 megaplasmids in Klebsiella species reveal transposable-element-mediated blaIMP-1 mobilisation

Introduction. Carbapenemase-producing Enterobacterales (CPE) represent a major threat to hospitalised patients worldwide. The dissemination of carbapenemase genes, such as blaIMP, is frequently mediated by mobile genetic elements including plasmids. During a previously described multispecies, healthcare-associated outbreak of blaIMP-positive CPE in North West London, two unusual isolates, IMP47 (Klebsiella grimontii) and IMP76 (K. pneumoniae), recovered in 2019 from routinely collected rectal swabs of inpatients, were predicted to harbour blaIMP-1-carrying IncHI1 megaplasmids. Aims. This study aimed to determine complete genomic sequences of IMP47 and IMP76, resolve the genetic context of blaIMP-1, and assess the conjugative mobility of blaIMP-1-carrying megaplasmids. Methods. Genomic sequences of both isolates were recovered through hybrid assembly of Oxford Nanopore and Illumina sequencing reads. Complete plasmid sequences were characterised to determine replicons, conjugation machinery, and genes encoding resistance to antimicrobials or other stress factors. Integrons and transposable elements (TEs) within flanking regions of blaIMP-1 were resolved through genome annotation and search against public databases. Liquid-mating experiments were performed to assess the mobility of blaIMP-1-carrying plasmids. Results. Completed genome assemblies were generated from both isolates, confirming two blaIMP-1-carrying megaplasmids, pIMP47 (391 kbp) and pIMP76_1 (519 kbp), of the replicon type IncHI1A(pNDM-CIT)/IncHI1B(pNDM-CIT). The blaIMP-1 locus was carried by nearly identical class 1 integrons in both plasmids and a closely related IncHI1 megaplasmid pEB3_IMP1 (361 kbp) previously identified in South West England. Comparative analysis revealed conserved genetic structures linking blaIMP-1 to mercury-resistance genes and TEs Tn6025, TnAs3, IS26, and IS5075, suggesting a history of recombination and potential for TE-mediated mobilisation. Conjugation experiments confirmed transfer of pIMP76_1 into a recipient K. pneumoniae strain, resulting in acquisition of ertapenem resistance, whereas transfer of pIMP47 was not observed under the tested conditions. Conclusion. Two IMP-producing IncHI1 megaplasmids in gut-colonising Klebsiella species revealed TE-mediated blaIMP-1 mobilisation. The co-localisation of blaIMP-1 and metal-resistance genes in both plasmids highlights the potential for co-selection of blaIMP-1 in environments enriched with metal ions. Our findings underscore the importance of longitudinal genomic surveillance of carbapenemase-encoding megaplasmids in healthcare settings.

microbiology↗

bioRxiv2025

Genomic and molecular characterisation of a Klebsiella pneumoniae clinical isolate resistant to meropenem-vaborbactam, imipenem-relebactam, and ceftazidime-avibactam

This article reports an unusual Klebsiella pneumoniae clinical isolate, KpMVR1, resistant to meropenem-vaborbactam, imipenem-relebactam, and ceftazidime-avibactam, and investigates the underlying genetic alterations using comparative genomics and molecular experiments. Resistance to carbapenems and third-generation cephalosporins is increasing in K. pneumoniae globally, restricting therapeutic options. The {beta}-lactam/{beta}-lactamase inhibitor combinations are widely used to circumvent {beta}-lactamase-mediated resistance. In 2021, isolate KpMVR1 was recovered from a hospitalised patient in England. Two additional isolates with the same variable-number tandem-repeat profile--KpMVS1, collected from the same patient 42 days before KpMVR1, and KpMVS2, from another patient in the same hospital--were susceptible to meropenem-vaborbactam, imipenem-relebactam, and ceftazidime-avibactam. Illumina and nanopore whole-genome sequencing and hybrid genome assembly were conducted for these three isolates. Annotated genome assemblies were compared to identify genetic variation, and mutagenesis experiments were performed to verify predicted functional alterations. All isolates belonged to a novel clone ST8134 and carried blaKPC-2-like alleles (KpMVR1: blaKPC-157; KpMVS1 and KpMVS2: blaKPC-2) in presumptively conjugative plasmids. ISEc68 caused a frameshift mutation in KpMVR1s ompK36 gene, reducing the meropenem-vaborbactam and imipenem-relebactam susceptibility. KPC-157 demonstrated decreased hydrolysis of imipenem and ceftazidime when compared with KPC-2. KpMVR1 also encoded a disrupted transcriptional repressor MarR and a destabilising mutation in AcrB, a component of the AcrAB-TolC multidrug efflux pump. In conclusion, KpMVR1 harboured complex resistance-associated genetic alterations, with evidence for in vivo emergence of antimicrobial resistance. Our study underlines routine screening for resistant pathogens in vulnerable patients to guide antimicrobial chemotherapy as well as the need to characterise underlying resistance mechanisms to help assess the potential for onward transmission. Data summaryIllumina and nanopore sequencing reads, hybrid genome assemblies, and anonymised metadata of isolates KpMVS1, KpMVR1, and KpMVS2 have been deposited in databases of the National Center for Biotechnology Information (www.ncbi.nlm.nih.gov) under BioProject accession PRJNA1084250, with BioSample accessions SAMN46778009 (KpMVS1), SAMN46778010 (KpMVR1), and SAMN46778011 (KpMVS2). The genome assemblies of these isolates have also been deposited in Pasteur Institutes database for K. pneumoniae species complex (bigsdb.pasteur.fr/klebsiella/) under ids 75608 (KpMVS1), 75609 (KpMVR1), and 75610 (KpMVS2). Impact statementThis is the first blaKPC-positive K. pneumoniae isolate referred to the UKs national reference laboratory with resistance to three last-resort {beta}-lactam/{beta}-lactamase inhibitor combinations meropenem-vaborbactam, imipenem-relebactam, and ceftazidime-avibactam, implicating in vivo emergence of this unusual resistance profile during prolonged antimicrobial chemotherapy. This isolate belonged to a novel clone ST8134 and harboured a plasmid-borne blaKPC-2-like allele blaKPC-157. We identified complex genetic alterations in this isolate: chromosomal large deletions, point mutations, and an ISEc68-induced loss-of-function truncation of the ompK36 porin gene. We determined the impact of KPC-2, KPC-157, and the ompK36 truncation on the susceptibility of K. pneumoniae to meropenem, meropenem-vaborbactam, imipenem, imipenem-relebactam, imipenem-avibactam, aztreonam, aztreonam-avibactam, ceftazidime, ceftazidime-avibactam, and cefiderocol. Our work underscores the need to monitor emerging resistance to beta-lactam/beta-lactamase inhibitor combinations in healthcare and to understand underlying resistance mechanisms for assessing the potential of resistance transmission.

microbiology↗

bioRxiv2024

Long read sequencing provides an insight into plasmids found among carbapenemase producing Enterobacterales from hospitals in the United Kingdom during 2021 to 2023

107 isolates of Enterobacterales consisting of Klebsiella pneumoniae (n=90), Escherichia coli (n=7), Enterobacter cloacae complex (n=6), Klebsiella oxytoca complex (n=3) and Citrobacter freundi (n=1) and additionally an isolate of Acinetobacter baumannii carrying genes encoding NDM (NDM-1, NDM-5 and NDM-14), KPC (KPC-2 and KPC-3), OXA-48-like (OXA-48, OXA-181 and OXA-232) and IMP (IMP-1 and IMP-4) carbapenemases were sequenced using q20 nanopore chemistry to provide complete/near-complete assemblies and relevant plasmids compared. Investigation of potential plasmid outbreaks in individual hospitals among isolates of different types and species revealed a mixed situation with some isolates carrying similar plasmids, but with segments missing/added and some plasmids that were clearly distinct. While most plasmids carrying blaOXA-48 were typical IncL plasmids of approximately 60 kb that are widely described, there was some variation among these. One isolate carried an IncR plasmid that had only limited homology with the others. Identical 51,479 bp ColKP3/IncX3 plasmids carrying blaOXA-181 were found from isolates from different hospitals that exactly matched those on GenBank from other countries. In other isolates blaOXA-181 was carried on IncFII plasmids. blaOXA-232 was found in highly conserved small ColKP3 plasmids that matched those found in other countries and continents. These observations highlight the importance of understanding the wider distribution of plasmids of concern. IncHI2/IncHI2A plasmids were important vehicles for carbapenemase genes and were found with blaKPC-2, blaIMP-1, blaIMP-4 or blaNDM-1, sometimes with the colistin resistance gene mcr-9 in addition. Representatives of K. pneumoniae sequence type (ST) 147 from seven hospitals carried IncFIB(pNDM-Mar)/IncHI1B(pNDM-MAR) hybrid virulence resistance plasmids of 325 to 352 kb that combined blaNDM-5 and other resistance genes with genes found in virulence plasmids. A similar plasmid was also found in an isolate of K. pneumoniae ST1558 and has been described in representatives of ST383. Nanopore sequencing has been instrumental in improving our knowledge of plasmids carrying carbapenemase genes leading to a better understanding of their epidemiology.

microbiology↗

bioRxiv2022

Virulence factors among isolates of extraintestinal Esherichia coli (ExPEC) from hospitals in the United Kingdom.

182 genes associated with virulence were sought from the whole genome sequences of all non-duplicate isolates of Escherichia coli received by the UK Health Security Agencys Antimicrobial Resistance and HealthCare Associated Infections laboratory for typing between June 2019 and March 2021 from hospitals in the United Kingdom and Republic of Ireland (n=593). These were from healthcare associated investigations and were not associated with diarrhoeal disease. Genes that were very common or very rare were excluded from further analysis. The frequency of detection of genes was compared among isolates from invasive infection, screening, urine samples and from neonates. cnf1 (coding for cytotoxic necrotising factor), clbK (coding for colibactin), focCDF (coding for F1 fimbriae), kpsM_K1, neuBD (associated with the K1 capsule), mchBC, mcmA (coding for microcins), papG_alleleIII (part of a cluster encoding P fimbriae), pic (protein involved in intestinal colonization), sfaE/sfafoCDE (coding for S fimbriae), tcpC (encoding TLR domain containing-protein C) and vat (encoding toxin vacuolating autotransporter) were 4 to 28 times more prevalent among isolates from invasive infections than among those from carriage. Representatives of sequence types (ST) 12, 73, 998 and 127 carried multiple of these virulence factors, no matter whether they were from screening swabs or from blood or other infection sites. Isolates carrying multiple virulence factors were more prevalent from neonatal screens than those from general screens. Genes associated with the K1 capsule (ibeA, neuBD, kpsM_K1) were particularly found in STs 1193, 10, 998, 538, 80 and 141. Nanopore sequencing of 14 isolates representing 10 different STs showed that the virulence elements sought were largely carried in chromosomal genomic islands, which were mosaic in nature. Some 9 % of isolates carried more than six of the main virulence genes/gene sets sought, highlighting the potential for significant numbers of carriage isolates to cause extraintestinal infections.

microbiology↗