Biology subjects

Paterson, D. L.

Publications and source records attributed to Paterson, D. L..

13 recordsLinked to original sources

bioRxiv2026

Disruption of cAMP Signalling and RNA Regulation Confers Cefiderocol Resistance in Acinetobacter baumannii

Cefiderocol is a siderophore-conjugated cephalosporin used to treat severe multidrug-resistant Gram-negative infections. Its recent prioritisation for US national preparedness underscores its clinical importance, yet resistance can emerge despite apparent in vitro susceptibility. Here, we identify two regulatory routes to cefiderocol resistance in carbapenem-resistant Acinetobacter baumannii (CRAB): mutation of the cAMP-producing adenylate cyclase CavA and truncation of the glycine-rich RNA-binding domain of the post-transcriptional regulator Hfq. These variants occurred in contemporary clinical isolates, conferred resistance in a murine CRAB lung-infection model and imposed minimal fitness costs. In Escherichia coli, deletion of the CavA orthologue cyaA increased competitive fitness during sub-inhibitory cefiderocol exposure. Hfq-CLASH and integrated omics revealed Hfq-dependent regulation of Fur-associated iron homeostasis in A. baumannii. Under iron limitation, truncated-Hfq induced a defence-primed state that attenuated siderophore uptake and reduced cefiderocol activity. These findings identify bacterial regulatory state and iron availability as determinants of cefiderocol susceptibility, with implications for resistance surveillance and susceptibility testing.

microbiology↗

bioRxiv2026

Cross-species convergence of surface polysaccharides shapes the virulence of an atypical carbapenem resistant uropathogenic Escherichia coli clone

Carbapenem-resistant Escherichia coli (CREC) is an emerging global health threat associated with limited treatment options and increased mortality. Here, analysis of 20,331 E. coli genomes from urinary tract infection (UTI) and urine samples identifies phylogroup A sequence type 361 (ST361) as an emerging CREC clone associated with UTI. Phylogenetic and phylodynamic analyses reveal two dominant ST361 clades that emerged in Asia around 2006 and subsequently disseminated intercontinentally. ST361 exhibits near pan-drug resistance, with blaNDM-5 conferring carbapenem resistance. In silico capsule and O-antigen typing reveals a highly conserved Klebsiella-like surface polysaccharide serotype in 95.9% of ST361 genomes, which we show contributes to virulence in UTI and catheter-associated UTI mouse models. Analysis of the mobile genetic elements harbouring blaNDM-5 in ST361 and other CREC further reveals a dynamic interplay between plasmid transfer and insertion elements that contributes to blaNDM-5 dissemination. Together, these findings identify ST361 as an emerging UTI-associated CREC clone and reveal the convergence of clonal expansion, virulence-associated surface architecture and mobile genetic elements in its spread.

microbiology↗

bioRxiv2024

Induced degradation of SNAP-fusion proteins

Self-labeling protein tags are an efficient means to visualize, manipulate, and isolate engineered fusion proteins with suitable chemical probes. The SNAP-tag, which covalently conjugates to benzyl-guanine and -chloropyrimidine derivatives is used extensively in fluorescence microscopy, given the availability of suitable SNAP-ligand-based probes. Here, we extend the applicability of the SNAP-tag to targeted protein degradation. We developed a set of SNAP PROteolysis TArgeting Chimeras (SNAP-PROTACs), which recruit the VHL or CRBN-ubiquitin E3 ligases to induce the degradation of SNAP-fusion proteins. Endogenous tagging enabled the visualization and the selective depletion of a SNAP-clathrin light chain fusion protein using SNAP-PROTACs. The addition of PROTACs to the SNAP-tag reagent toolbox facilitates the comprehensive analysis of protein function with a single gene tagging event.

cell biology↗

bioRxiv2023

Identification of a novel MCR-1 variant displaying low level of co-resistance to β-lactam antibiotics that uncovers a potential novel antimicrobial peptide

The emerging and global spread of a novel plasmid-mediated colistin resistance gene, mcr-1, threatens human health. It is accepted that MCR-1 affects bacterial fitness, and this fitness cost correlates with bacterial membrane lipid A perturbation. However, the detailed molecular mechanism remains unclear. Here, we screened out a novel MCR-1 variant, named M6, with a two-point mutation that rendered a low level of co-resistance to {beta}-lactam antibiotics. Compared to wild-type (WT) MCR-1, this variant caused severe lipid A perturbation resulting in peptidoglycan layer remodelling and thus resulted in phenotypic co-resistance to {beta}-lactams. Moreover, we found that a lipid A loading cavity is localized at the linker domain of MCR-1 and governs colistin resistance and bacterial membrane permeability, and the mutated pocket of M6 facilitates the binding affinity of lipid A. More importantly, synthetic peptides derived from M6 achieved broad-spectrum antimicrobial activity. These findings provide insights into a potential vulnerability that could be exploited in future antimicrobial drug design.

microbiology↗

bioRxiv2021

MicroPIPE: An end-to-end solution for high-quality complete bacterial genome construction

Oxford Nanopore Technology (ONT) long-read sequencing has become a popular platform for microbial researchers; however, easy and automated construction of high-quality bacterial genomes remains challenging. Here we present MicroPIPE: a reproducible end-to-end bacterial genome assembly pipeline for ONT and Illumina sequencing. To construct MicroPIPE, we evaluated the performance of several tools for genome reconstruction and assessed overall genome accuracy using ONT both natively and with Illumina. Further validation of MicroPIPE was carried out using 11 sequence type (ST)131 Escherichia coli and eight publicly available Gram-negative and Gram-positive bacterial isolates. MicroPIPE uses Singularity containers and the workflow manager Nextflow and is available at https://github.com/BeatsonLab-MicrobialGenomics/micropipe.

bioinformatics↗

bioRxiv2020

Strain and lineage-level methylome heterogeneity in the multi-drug resistant pathogenic Escherichia coli ST101 clone

Escherichia coli Sequence Type (ST)101 is an emerging, multi-drug resistant lineage associated with carbapenem resistance. We recently completed a comprehensive genomics study on mobile genetic elements (MGEs) and their role in blaNDM-1 dissemination within the ST101 lineage. DNA methyltransferases (MTases) are also frequently associated with MGEs, with DNA methylation guiding numerous biological processes including genomic defence against foreign DNA and regulation of gene expression. The availability of Pacific Biosciences Single Molecule Real Time Sequencing data for seven ST101 strains enabled us to investigate the role of DNA methylation on a genome-wide scale (methylome). We defined the methylome of two complete (MS6192 and MS6193) and five draft (MS6194, MS6201, MS6203, MS6204, MS6207) ST101 genomes. Our analysis identified 14 putative MTases and eight N6-methyladenine DNA recognition sites, with one site that has not been described previously. Furthermore, we identified a Type I MTase encoded within a Transposon 7-like Transposon and show its acquisition leads to differences in the methylome between two almost identical isolates. Genomic comparisons with 13 previously published ST101 draft genomes identified variations in MTase distribution, consistent with MGE differences between genomes, highlighting the diversity of active MTases within strains of a single E. coli lineage. It is well established that MGEs can contribute to the evolution of E. coli due to their virulence and resistance gene repertoires. This study emphasises the potential for mobile genetic elements to also enable highly similar bacterial strains to rapidly acquire genome-wide functional differences via changes to the methylome. Impact StatementEscherichia coli ST101 is an emerging human pathogen frequently associated with carbapenem resistance. E. coli ST101 strains carry numerous mobile genetic elements that encode virulence determinants, antimicrobial resistance, and DNA methyltransferases (MTases). In this study we provide the first comprehensive analysis of the genome-wide complement of DNA methylation (methylome) in seven E. coli ST101 genomes. We identified a Transposon carrying a Type I restriction modification system that may lead to functional differences between two almost identical genomes and showed how small recombination events at a single genomic region can lead to global methylome changes across the lineage. We also showed that the distribution of MTases throughout the ST101 lineage was consistent with the presence or absence of mobile genetic elements on which they are encoded. This study shows the diversity of MTases within a single bacterial lineage and shows how strain and lineage-specific methylomes may drive host adaptation. Data SummarySequence data including reads, assemblies and motif summaries have previously been submitted to the National Center for Biotechnology Information (https://www.ncbi.nlm.nih.gov) under the BioProject Accessions: PRJNA580334, PRJNA580336, PRJNA580337, PRJNA580338, PRJNA580339, PRJNA580341 and PRJNA580340 for MS6192, MS6193, MS6194, MS6201, MS6203, MS6204 and MS6207 respectively. All supporting data, code, accessions, and protocols have been provided within the article or through supplementary data files.

genomics↗

bioRxiv2020

Burkholderia pseudomallei clinical isolates are highly susceptible in vitro to cefiderocol, a novel siderophore cephalosporin

Cefiderocol is a novel cephalosporin designed to treat multidrug resistant Gram-negative infections. By forming a chelated complex with ferric iron, cefiderocol is transported into the periplasmic space via bacterial iron transport systems and primarily binds to penicillin-binding protein 3 (PBP3) to inhibit peptidoglycan synthesis. This mode of action results in cefiderocol having greater in vitro activity against many Gram-negative bacilli than currently used carbapenems, {beta}-lactam/{beta}-lactamase inhibitor combinations, and cephalosporins. Thus, we investigated the in vitro activity of cefiderocol (S-649266) against a total of 271 clinical isolates of Burkholderia pseudomallei from Australia. The collection was comprised of primary isolates (92.3%) and subsequent isolates (7.7%). Minimum inhibitory concentrations (MIC) of cefiderocol ranged from [≤]0.03 to 32 mg/L, where the MIC90 was 1 mg/L and 16 mg/L for primary and subsequent isolates, respectively. Based upon non-species specific (Gram-negative bacilli) clinical breakpoints for cefiderocol (MIC [≤] 4 mg/L), twelve isolates (4.4%) would be classified as non-susceptible. Further testing for co-resistance to meropenem, ceftazidime, trimethoprim-sulfamethoxazole, amoxicillin-clavulanate and doxycycline was performed on a subset of isolates with elevated cefiderocol MICs ([≥]2 mg/L, 4.8%) and 84.6% of these isolates exhibited resistance to at least one of these antimicrobials. Cefiderocol was found to be highly active in vitro against B. pseudomallei primary clinical isolates. This novel compound shows great potential for the treatment of melioidosis in endemic countries and should be explored further.

microbiology↗

bioRxiv2020

Dynamics of mcr-1 prevalence and mcr-1-positive Escherichia coli after the cessation of colistin use as a feed additive for animals in China: a prospective cross-sectional and whole genome sequencing based molecular epidemiological study

BackgroundThe global dissemination of colistin resistance encoded by mcr-1 has been attributed to extensive use of colistin in livestock, threatening colistin efficacy in medicine. The emergence of mcr-1 in common pathogens, such as Escherichia coli, is of particular concern. Therefore, China banned the use of colistin in animal feed from May 1ST 2017. We investigated subsequent changes in mcr-1 prevalence, and the genomic epidemiology of mcr-1-positive Escherichia coli (MCRPEC). MethodsSampling was conducted pre- (October-December 2016) and post-colistin ban (October-December, 2017 and 2018, respectively). 3675 non-duplicate pig fecal samples were collected from 14 provinces (66 farms) in China to determine intervention-related changes in mcr-1 prevalence. 15193 samples were collected from pigs, healthy human volunteers, colonized and infected hospital inpatients, food and the environment in Guangzhou, to characterize source-specific mcr-1 prevalence and the wider ecological impact of the ban. From these samples, 688 MCRPEC were analyzed with whole genome sequencing (WGS), plasmid conjugation and S1-PFGE/Southern blots to characterize associated genomic changes. FindingsAfter the ban, mcr-1 prevalence decreased significantly in national pig farms, from 45{middle dot}0% (308/684 samples) in 2016, to 19{middle dot}4% (274/1416) in 2018 (p<0{middle dot}0001). This trend was mirrored in samples from most sources in Guangzhou (overall 19{middle dot}2% [959/5003 samples] in 2016; 5{middle dot}3% [238/4489] in 2018; p<0{middle dot}0001). The population structure of MCRPEC was diverse (23 sequence clusters [SCs]); ST10 clonal complex isolates were predominant (247/688 [36%]). MCRPEC causing infection in hospitalized inpatients were genetically more distinct and appeared less affected by the ban. mcr-1 was predominantly found on plasmids (632/688 [92%]). Common mcr-1 plasmid types included IncX4, IncI2 and IncHI2 (502/656 [76.5%]); significant increases in IncI2-associated mcr-1 and a distinct lineage of mcr-1-associated IncHI2 were observed post-ban. Changes in the frequency of mcr-1-associated flanking sequences (ISApl1-negative MCRPEC), 63 core genome SNPs and 30 accessory genes were also significantly different after the ban, consistent with rapid genetic adaptation in response to changing selection pressures. InterpretationA rapid, ecosystem-wide, decline in mcr-1 was observed after banning the use of colistin in animal feed, with associated genetic changes in MCRPEC. Genomic surveillance is key to assessing and monitoring stewardship interventions. FundingNational Natural Science Foundation of China

microbiology↗

bioRxiv2019

Intensive infection control responses and whole genome sequencing to interrupt and resolve widespread transmission of OXA-181 Escherichia coli in a hospital setting

BackgroundOXA-48-like carbapenemases have become increasingly prevalent in healthcare settings worldwide. Their low-level activity against carbapenems makes them difficult to identify, causing problems for infection control. Here we present an outbreak of Escherichia coli producing OXA-181 (part of the OXA-48 family of carbapenemases) in a Queensland Hospital, and describe how we used whole genome sequencing (WGS) to identify the outbreak strain, determine the extent of transmission within the hospital and support infection control responses. Methods116 isolates were collected and sequenced on an Illumina NextSeq to determine species, sequence type (ST) and presence of resistance genes. Core single nucleotide polymorphisms were used to determine strain relatedness. Three isolates were also sequenced on an Oxford Nanopore MinION to determine the context of the resistance genes. ResultsOf 116 isolates, 85 (84 E. coli and one K. pneumoniae) from 78 patients (and two environmental sources) were related to the ongoing outbreak. The outbreak E. coli strain was found to be ST38 and carried blaOXA-181, blaCTX-M-15 and qnrS1 genes. Long read sequencing revealed blaOXA-181 to be carried on an IncX3 plasmid with qnrS1. blaCTX-M-15 was chromosomally integrated (via ISEcp1 insertion) in close proximity to a second qnrS1 gene. A search of the laboratory database identified an isolate with an identical unusual antibiogram from a patient recently admitted to a hospital in Vietnam, suggesting that the strain was introduced to the hospital. This conclusion was supported by WGS, as comparison of the strain to public data identified a close match to an E. coli recovered from Vietnam in 2011. ConclusionA blaOXA-181-carrying E. coli ST38 strain was introduced to a Brisbane hospital and spread undetected throughout multiple wards over several months. Using WGS, we characterized the outbreak strain and unambiguously detected its presence throughout the hospital. We show how both WGS and infection control measures can be utilized to effectively terminate widespread transmission of an elusory pathogen.

microbiology↗

bioRxiv2019

Genomic characterisation and context of the blaNDM-1 carbapenemase in Escherichia coli ST101.

Carbapenems are last-resort antibiotics; however, the spread of plasmid-encoded carbapenemases such as the New Delhi metallo-{beta}-lactamase 1 (NDM-1) challenges their effectiveness. The rise of NDM-1 has coincided with the emergence of extensively multidrug resistant (MDR) lineages such as Escherichia coli ST101. Here we present a comprehensive genomic analysis of seven E. coli ST101 isolates that carry the blaNDM-1 gene. We determined the complete genomes of two isolates and the draft genomes of five isolates, enabling complete resolution of the plasmid context of blaNDM-1. Comparisons with thirteen previously published ST101 genomes revealed a monophyletic lineage within the B1 phylogroup forming two clades (designated Clade 1 and Clade 2). Most Clade 1 strains are MDR, encoding resistance to at least 9 different antimicrobial classes, including extended spectrum cephalosporins. Additionally, we characterised different pathways for blaNDM-1 carriage and persistence in the ST101 lineage. For IncC plasmids, carriage was associated with recombination and local transposition events within the antibiotic resistance island. In contrast, we revealed recent transfer of a large blaNDM-1 resistance island between F-type plasmids. The complex acquisition pathways characterised here highlight the benefits of long-read Single Molecule Real Time sequencing in revealing evolutionary events that would not be apparent by short-read sequencing alone. These high-quality E. coli ST101 genomes will provide an important reference for further analysis of the role of mobile genetic elements in this emerging multidrug resistant lineage. ImportanceCarbapenem resistant Escherichia coli are urgent priority organisms as they are resistant to our drugs of last resort. E. coli ST101 have been reported as carriers of the New Delhi Metallo-beta-Lactamase 1 gene (blaNDM-1), conferring resistance to carbapenems, however there is limited genomic information available for this lineage. In this study we used long-read genome sequencing to characterise the complete genomes of two E. coli ST101 strains and determine the carriage of blaNDM-1 in a collection of E. coli ST101 strains. We showed that carriage of blaNDM-1 and resistance determinants to eight other antimicrobial classes was confined to a single clade. We also showed two different pathways for the carriage of blaNDM-1, which was dependent on the type of plasmid. Long-read sequencing allowed us to show the full complexities of these resistance regions and highlighted how strains from an emerging E. coli lineage have become resistant to nearly all available antimicrobials.

genomics↗

bioRxiv2019

Genomic investigation reveals contaminated detergent as the source of an ESBL-producing Klebsiella michiganensis outbreak in a neonatal unit

BackgroundKlebsiella species are problematic pathogens in neonatal units and may cause outbreaks, for which sources of transmission can be challenging to elucidate. We describe the use of whole genome sequencing (WGS) to investigate environmental sources of transmission during an outbreak of extended-spectrum-{beta}-lactamase (ESBL)-producing Klebsiella michiganensis colonizing neonates. MethodsCeftriaxone-resistant Klebsiella spp. isolated from neonates (or their mothers) and the hospital environment were included. Short-read (Illumina) and long-read (MinION, Oxford Nanopore Technologies) sequencing was used to confirm species taxonomy, define antimicrobial resistance genes and determine phylogenetic relationships using single nucleotide polymorphism (SNP) profiling. ResultsA total of 21 organisms (10 patient-derived and 11 environmental isolates) were sequenced. Standard laboratory methods identified the outbreak strain as an ESBL-producing Klebsiella oxytoca, but taxonomic assignment from WGS data suggested closer identity to Klebsiella michiganensis. Strains isolated from baby bath drains and multiple detergent dispensing bottles were either identical or closely related by SNP comparison. Detergent bottles contaminated by K. michiganensis had been used for washing milk-expressing equipment. No new cases were identified once the detergent bottles were removed and the baby baths decommissioned. ConclusionsEnvironmental reservoirs may be an important source in outbreaks of multi-drug resistant organisms. WGS, in conjunction with traditional epidemiological investigation, can be instrumental in revealing routes of transmission and guiding infection control responses. Key pointsO_LIK. michiganensis can be misidentified as K. oxytoca and is probably under-recognized as a nosocomial pathogen C_LIO_LIWhole genome sequencing of neonatal and environmental isolates during an outbreak of ESBL-producing K. michiganensis confirmed contaminated detergent and sinks to be the source C_LI

microbiology↗

bioRxiv2017

Dominance of ST131 Escherichia coli carrying blaCTX-M in patients with bloodstream infections caused by cephalosporin-resistant strains in Australia, New Zealand and Singapore: whole genome analysis of isolates from a randomised trial

Synopsis/AbstractO_ST_ABSObjectivesC_ST_ABSTo characterise multi-drug resistant Escherichia coli isolated from patients in Australia, New Zealand and Singapore with bloodstream infection (BSI).\n\nMethodsWe prospectively collected third-generation cephalosporin resistant (3GC-R) E. coli from blood cultures obtained from patients enrolled in a randomised controlled trial. Whole genome sequencing was used to characterise antibiotic resistance genes, sequence types (STs), plasmids and phylogenetic relationships. Antibiotic susceptibility was determined using disk diffusion and Etest.\n\nResultsA total of 70 E. coli were included, of which the majority were ST131 (61.4%). BSI was most frequently from a urinary source (69.6%), community-associated (62.9%) and in older patients (median age 71 years [IQR 64-81]). The median Pitt bacteraemia score at presentation was 1 (IQR 0-2, range 0-3) and ICU admission was infrequent (3.1%). ST131 possessed significantly more acquired resistance genes than non-ST131 (p=0.003). Clade C1/C2 ST131 predominated (30.2% and 53.5% of all ST131 respectively) and these were all resistant to ciprofloxacin. All clade A ST131 were community-associated. The predominant ESBL types were blaCTX-M (78.6% of isolates) and were strongly associated with ST131, with the majority blaCTX-M-15. Clade C1 was associated with blaCTX-M-14 and blaCTX-M-27, whereas blaCTX-M-15 predominated in clade C2. Plasmid-mediated AmpC (p-AmpC) genes (mainly blaCMY-2) were also frequent (17.1%) but were more common with non-ST131 strains (p< 0.001). The majority of plasmid replicon types were IncF.\n\nConclusionsIn a prospective collection of 3GC-R E. coli causing BSI in the Australasian region, community-associated Clade C1/C2 ST131 predominate in association with blaCTX-M ESBLs, although a significant proportion of non-ST131 strains carried blaCMY-2.

microbiology↗

bioRxiv2017

Genomic investigation of an outbreak of carbapenemase-producing Enterobacter cloacae: long-read sequencing reveals the context of blaIMP4 on a widely distributed IncHI2 plasmid

Carbapenem-resistant Enterobacteriaceae (CRE) represent one of the most urgent threats to human health posed by antibiotic resistant bacteria. Enterobacter hormaechei and other members of the Enterobacter cloacae complex are the most commonly encountered Enterobacter spp. within clinical settings, responsible for numerous outbreaks and ultimately poorer patient outcomes. Here we applied three complementary whole genome sequencing (WGS) technologies to characterise a hospital cluster of blaIMP-4 carbapenemase-producing E. hormaechei.\n\nIn response to a suspected CRE outbreak in 2015 within an Intensive Care Unit (ICU)/Burns Unit in a Brisbane tertiary referral hospital we used Illumina sequencing to determine that all outbreak isolates were sequence type (ST)90 and near-identical at the core genome level. Comparison to publicly available data unequivocally linked all 10 isolates to a 2013 isolate from the same ward, confirming the hospital environment as the most likely original source of infection in the 2015 cases. No clonal relationship was found to IMP-4-producing isolates identified from other local hospitals. However, using Pacific Biosciences long-read sequencing we were able to resolve the complete context of the blaIMP-4 gene, which was found to be on a large IncHI2 plasmid carried by all IMP-4-producing isolates. Continued surveillance of the hospital environment was carried out using Oxford Nanopore long-read sequencing, which was able to rapidly resolve the true relationship of subsequent isolates to the initial outbreak. Shotgun metagenomic sequencing of environmental samples also found evidence of ST90 E. hormaechei and the IncHI2 plasmid within the hospital plumbing.\n\nOverall, our strategic application of three WGS technologies provided an in-depth analysis of the outbreak, including the transmission dynamics of a carbapenemase-producing E. hormaechei cluster, identification of possible hospital reservoirs and the full context of blaIMP-4 on a multidrug resistant IncHI2 plasmid that appears to be widely distributed in Australia.

genomics↗