Biology subjects

Aanensen, D. M.

Publications and source records attributed to Aanensen, D. M..

At least 19 recordsLinked to original sources

bioRxiv2026

Mobile element-mediated carbapenem resistance in Enterobacter hormaechei in a Nigerian intensive care unit

Carbapenem-resistant Gram-negative bacteria pose a critical public health threat. The role of mobile genetic elements in driving their transmission and persistence remains poorly defined. In 2022, we investigated a suspected outbreak of carbapenem-resistant Acinetobacter baumannii (CRAB) in a Nigerian adult intensive care unit (ICU), using short-read whole genome sequencing (WGS) of carbapenem-resistant clinical and environmental isolates during the cluster period. Mobile element dynamics were then inferred from hybrid assemblies of Illumina and Oxford Nanopore reads. The suspected CRAB outbreak was ruled out by WGS but a carbapenem-resistant Enterobacter hormaechei ST114 bloodstream isolate was found to be indistinguishable from two environmental isolates, all recovered during the Acinetobacter surge. Hybrid assemblies revealed a strikingly conserved [~]19 Kb resistance island shared across all ST114 genomes. The island contained a blaNDM-5 cassette alongside many other antimicrobial resistance genes, within class 1 integronns and flanked by insertions sequences, located on a 46,176 bp plasmid. Using the ST114 plasmids hybrid assembly as scaffold, the same plasmid was identified in the genome of a Klebsiella pneumoniae ST15 isolate from the ICU environment during the same period. Additionally, re-interrogation of genomic surveillance data uncovered four clonal 2020 ST109 Enterobacter bloodstream isolates from the same facility that carried the resistance genes in the same context on a large 267,242 bp plasmid. Carbapenem resistance in hospital Enterobacterales is driven by both clonal expansion and horizontal spread of mobile resistance elements. These findings underscore the need to track mobile elements alongside bacterial lineages to inform evidence-based infection control, especially in low-resource settings. Impact StatementCarbapenem resistance among Enterobacterales remains a major public health threat, yet how mobile genetic elements contribute to their persistence and spread in hospital settings is still poorly understood. In this study, we investigated a suspected outbreak of carbapenem-resistant Acinetobacter baumannii in an adult intensive care unit in Nigeria. Although the outbreak was eventually ruled out, genomic analysis has shown the importance of careful interpretation of suspected outbreak cases in hospital settings. Our findings highlight the importance of close monitoring of ICU environments, the implementation of blood culture-based diagnostics, and the value of genomic support in outbreak investigations. These findings demonstrate that carbapenem resistance in hospital Enterobacterales is driven not only by clonal expansion but also by the horizontal dissemination of a highly stable blaNDM-5-associated MDR island capable of integrating into diverse plasmid backbones. This study emphasizes the need for genomic surveillance that tracks both mobile elements and bacterial lineages to strengthen outbreak investigations, especially in low-resource settings. It further underscores the links between clinical and environmental AMR reservoirs and reinforces the value of a One Health approach to controlling carbapenem resistance. Data summaryFASTQ sequences were deposited in the NCBI BioSample database under accession numbers SAMN55915584 - SAMN55915597.

microbiology↗

bioRxiv2025

Ciprofloxacin resistance in Klebsiella pneumoniae: phenotype prediction from genotype and global distribution of resistance determinants

BACKGROUNDCiprofloxacin resistant Klebsiella pneumoniae is common or emerging in many geographies, and knowledge of local resistance rates is important for empirical therapy. Whilst there are known K. pneumoniae ciprofloxacin resistance determinants, there is a lack of systematic data on the effect of determinants, alone and in combination, and there are no publicly accessible tools for predicting resistance from whole genome sequence data. METHODSThe KlebNET-GSP AMR Genotype-Phenotype Group aggregated a matched genotype-phenotype dataset of n=12,167 K. pneumoniae species complex (KpSC) isolates from 27 countries between 2001-2021. We developed a rules-based classifier to predict ciprofloxacin resistance by categorizing the number of quinolone resistance determining regions mutations in gyrA and parC, the number of plasmid-mediated quinolone resistance genes, and the presence/absence of aac(6')-Ib-cr (which can acetylate ciprofloxacin). Predictive performance was assessed using the discovery dataset, for which we re-phenotyped discrepant isolates; and validated using externally contributed datasets (n=7,030 KpSC isolates). RESULTSThe rules-based classifier predicted R vs S/I with categorical agreement, sensitivity, and specificity >96%, and major/very major error rates <4%. Performance was similar across diverse KpSC sources (human, animal, other), species, and intra-species lineages. External validation of the classifier yielded overall 93.12% categorical agreement [95% confidence interval (CI), 92.50-93.74%], 8.65% major errors [95% CI, 7.34-9.97%], and 6.20% very major errors [95% CI, 5.51-6.90%]. We implemented the classifier in Kleborate, a command-line tool that is integrated into the Pathogenwatch web platform. Using this to assess the global distribution of ciprofloxacin resistance determinants in KpSC genomes available in Pathogenwatch (n=31,319, from 109 countries between years 2000-2023), we observed a significant positive association between national quinolone consumption rates and predicted ciprofloxacin resistance (R2=0.20, p=0.004). CONCLUSIONS Ciprofloxacin resistance phenotypes can be reasonably predicted from genotypes, which is sufficient for informing surveillance. However, unexplained resistance remains and accuracy is insufficient for clinical applications. We demonstrate the value of aggregating genotype-phenotype data to explore resistance mechanisms and develop predictors, but highlight complexities in combining phenotype data from different assays and standards.

genomics↗

bioRxiv2025

Monitoring of vaccine targets and interventions using global genome data: vaccines.watch

The expansion of pathogen genome sequencing into routine disease surveillance programmes is set to bring rapidly-growing volumes of increasingly structured data on a global scale. This has the potential to deliver exciting opportunities for accelerating vaccine development and monitoring. Here we present an interactive platform, vaccines.watch (https://vaccines.watch), which aims to support decision-making around vaccine formulations and roll-out by enabling interrogation of vaccine target diversity from global genome data. We have initially focused on targets included in existing or prospective multivalent polysaccharide- based vaccines for Streptococcus pneumoniae, Klebsiella pneumoniae (and related species) and Acinetobacter baumannii. The platform currently displays data for >100k high-quality genomes with geotemporal sampling information (post-2010), with new genomes assembled, analysed and incorporated on an ongoing basis (every 4 hours) as public data are newly deposited. Crucially, users can view vaccine target information in the broader context of genotypic variants (e.g. sequence types) and antimicrobial resistance markers. The platform also enables users to review the composite serotypes of pneumococcal vaccine formulations among the available genomes. For example, using data in vaccines.watch from 3 June 2025, we observed that serotypes included in the PCV13 and PCV21 formulations accounted for 36.2% (11,907/32,918) and 87.4% (28,764/32,918) of global public genomes, respectively. The platform also enables continuous review of the global genomic landscape of the included pathogens, enabling identification of gaps (e.g. in geographic coverage) that should be targeted with increased genomic surveillance. Indeed we demonstrate that substantial geographic gaps remain in the coverage of available genomes, with over half of countries contributing no genomes for each of the three pathogens. However, while caution in interpretation is important, as global representativeness of genome data grows, vaccines.watch is positioned to support different stages of the vaccine pipeline, from selection of target antigens to post-rollout monitoring of population changes.

genomics↗

bioRxiv2025

amr.watch - monitoring antimicrobial resistance trends from global genomics data

BackgroundWhole genome sequencing (WGS) is increasingly supporting routine pathogen surveillance at local and national levels, providing comparable data that can inform on the emergence and spread of antimicrobial resistance (AMR) globally. However, the potential for shared WGS data to guide interventions around AMR remains under-exploited, in part due to challenges in collating and transforming the growing volumes of data into timely insights. We present an interactive platform, amr.watch (https://amr.watch), that enables interrogation of AMR trends from public WGS data on an ongoing basis to support research and policy. MethodsThe amr.watch platform incorporates, analyses and visualises high-quality WGS data from WHO-defined priority bacterial pathogens. Analytics are performed using community-standard methods with bespoke species-specific curation of AMR mechanisms. FindingsBy 31 March 2025, amr.watch included data from 620,700 pathogen genomes with geotemporal information, with highly variable representation of different species and geographic regions. By integrating WGS data with sampling information, amr.watch enables users to assess geotemporal trends among genotypic variants (e.g. sequence types) and AMR mechanisms, with implications for interventions including antimicrobial prescribing and drug and vaccine development. Interpretationamr.watch is an information platform for scientists and policy-makers delivering ongoing situational awareness of AMR trends from genomic data. As broad adoption of WGS continues, amr.watch is positioned to monitor both pathogen populations and our global efforts in genomic surveillance, guiding control strategies tailored to each pathogens characteristics. FundingUK National Institute for Health Research & Gates Foundation. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSWhole genome sequencing (WGS) approaches enable us to track the spread of bacterial pathogens and antimicrobial resistance (AMR) with high resolution at local, national and global levels. To date, genomic studies assessing AMR dynamics have largely used retrospective data collected for specific research agendas. However, the growing volumes of publicly-shared WGS data, generated increasingly from routine surveillance, provide improved power to detect novel trends and guide interventions. Efforts to collate public bacterial genome data exist, such as AllTheBacteria, although these are aimed at the research community and do not facilitate data usage and interpretation by non-genomics experts, particularly those in public health. To our knowledge, no platforms exist for readily interrogating AMR dynamics from continuously updated public genome data across the array of different WHO priority bacterial pathogens. Of note, however, TyphiNet provides an interactive online dashboard for examining AMR trends exclusively from Salmonella Typhi using a periodically-updated data set from the Pathogenwatch platform. These findings are based on searching PubMed without language restrictions from Jan 1 2000 to December 31 2024, using terms related to "genomic surveillance" and "antimicrobial resistance". Added value of this studyWe have developed amr.watch which, to our knowledge, is the first platform that enables ongoing analysis and visualisation of AMR trends from public genome data across the spectrum of WHO priority bacterial pathogens via an accessible interface. Crucially, the platform incorporates processed genome data via a live always-on stream, enabling insights that are delayed only by the time to data deposition. We reviewed public genomes with available geotemporal information up until 31 March 2025, providing a contemporary global landscape of pathogen genome sequencing. While the number of genomes available annually grew over five-fold globally between 2010 and 2018, we also revealed the extent of differences in geographic representation, with 89.6% of genomes originating from high-income countries and 89 countries contributing no genome data from the priority pathogens. Implications of all the available evidenceRapid generation and sharing of global genome data enables us to more precisely track the spread and define the characteristics of contemporary circulating resistant pathogens. The amr.watch platform provides a solution for retrieving, curating and translating shared genomic data into relevant insights that are accessible and actionable by diverse stakeholders. It thereby forms a basis for monitoring progress in surveillance efforts, alerting on ongoing population changes and guiding enhanced precision for surveillance and interventional development. Additionally, our review of available bacterial genome data highlights the need for additional efforts to increase and sustain the implementation of genomic surveillance of AMR globally, and improve the timely sharing of WGS data and its associated metadata.

genomics↗

bioRxiv2024

Whole genome Sequencing Reveals Enterobacter hormaechei as a key Bloodstream Pathogen in six tertiary care hospitals in southwestern Nigeria.

Enterobacter spp. are an important cause of healthcare-associated bloodstream infections uncommonly reported in Africa. This study used whole genome sequencing (WGS) to characterise Enterobacter spp. from hospitals in Nigerias antimicrobial resistance (AMR) surveillance system. Blood-culture isolates of Enterobacter from six such tertiary-care hospitals recovered between 2014 and 2020 were re-identified and antimicrobial susceptibility-tested using VITEK2. Illumina technology provided whole genome sequences for genome nomenclature, antimicrobial resistance gene prediction, Single Nucleotide Polymorphism (SNP) phylogeny, and multi-locus sequence typing via publicly available bioinformatics pipelines. Initial biochemical delineation often misclassified Enterobacter, necessitating whole-genome sequencing for accurate classification. Among 98 Enterobacter received, Enterobacter hormaechei subspecies xiangfangensis predominated (43), followed by other E. hormachei subspecies (18), E. cloacae (26), E. roggenkampii (4), E. bugandensis (3), E. kobei (2), E. asburiae (1) and E. cancerogenous (1). Cephalosporins, aminoglycoside, chloramphenicol, macrolide, and carbapenem resistance in E. hormaechei was attributed to known resistance genes. They belonged to clusters III, IV, and VIII based on hsp60 typing and clades A, B, C, and D according to Sutton and Cos nomenclature. These isolates and other Enterobacter species recently reported from Nigeria reveal extensive E. hormaechei diversity, as well as clusters representing potential outbreaks. Enterobacter hormaechei, often misidentified and rarely reported from Nigeria, is this studys most common blood culture isolated Enterobacter spp. Uncovering underappreciated species as important bloodstream pathogens and retrospective detection of likely outbreaks emphasise the value of genomic surveillance in resource-limited settings. DATA SUMMARYAll sequence reads were submitted to the European Nucleotide Archive (ENA) under the project ID PRJEB29739 (https://www.ebi.ac.uk/ena/browser/view/PRJEB29739). Accessions can be found in Table S1. IMPACT STATEMENTAccurate identification of Enterobacter is essential in healthcare settings as misidentification can lead to selecting antimicrobials to the genus is intrinsically resistant resistant before susceptibility testing results are available. Also, misidentification can compromise microbiology support for infection prevention and control. We show that E. hormaechei, which is never reported from clinical laboratories in Nigeria, is frequently misidentified using conventional methods like tube- or strip biochemical testing and VITEK systems. Whole genome sequence data demonstrates that E. hormaechei and E. cloacae are the most common Enterobacter isolated from bloodstream infections in Nigeria. Enhanced identification methods for surveillance play pivotal roles in improving patient care, optimising antibiotic stewardship, and combating the evolving challenges posed by this pathogen. Overall, this study reveals the effectiveness of WGS in correctly identifying this important pathogen.

microbiology↗

bioRxiv2024

Bacterial strain nomenclature in the genomic era: Life Identification Numbers using a gene-by-gene approach

Unified strain taxonomies are needed for the epidemiological surveillance of bacterial pathogens and international communication in microbiological research. Core genome multilocus sequence typing (cgMLST) holds great promise for standardized high-resolution strain genotyping. However, this approach faces challenges including classification instability and disconnection of new nomenclature from widely adopted classical MLST identifiers. This essay discusses the cgMLST-based Life Identification Number (LIN) method, recently proposed as a stable multilevel strain taxonomy system applicable to most bacterial pathogens. We describe how LIN codes are implemented and used in practice for precise strain definitions and epidemiological tracking. Glossary Multilocus sequence typing (MLST)A genotyping method applied mostly to microbial strains to study population structure and epidemiology, based on comparing the nucleotide sequences of a small number (typically seven) of housekeeping protein-coding genes. In MLST, allele numbers are assigned to each sequence variant (allele) of a given gene. The MLST genotype of a bacterial strain is defined by the combination of the allele numbers observed at the genes that are included in the genotyping scheme. A sequence type (ST) is assigned to each unique combination of alleles, called an MLST profile. MLST was invented in 1998 and became a de-facto standard taxonomy of bacterial strains, albeit at low resolution. Core genome MLSTAn extension of MLST that analyzes sequence variation across hundreds to thousands of conserved (core) genes, shared by all strains of a species, providing higher resolution typing for genomic epidemiology and evolutionary studies. cgMLST schemes typically comprise 2000 to 4000 genes, depending on the genome size and genetic variation (in terms of presence/absence of genes) within bacterial species. A core genome sequence type (cgST) can be assigned to unique cgMLST profiles, i.e., a unique combination of cgMLST allelic numbers. Whole Genome Sequencing (WGS)A method that determines the complete DNA sequence of an organisms genome in a single process, providing comprehensive information for comparative genetic analyses based on cgMLST or other analytic methods. Single Nucleotide Polymorphisms (SNPs)Variations at a single base position in the DNA sequence among individuals isolates, strains or species, used as genetic markers for studying for example, evolutionary relationships or strain identity. Average nucleotide identity (ANI)A measure of genomic similarity between two organisms, calculated as the average percentage of identical nucleotides in orthologous genomic regions; commonly used to assess species-level relatedness in prokaryotes. TaxonomyHere, we apply the word taxonomy to bacterial strains as a system of classifying, naming and identifying strains based on shared genetic characteristics as defined by e.g., cgMLST.

microbiology↗

bioRxiv2023

High genetic diversity and blaNDM-1 prevalence among Acinetobacter baumannii in Nigerian hospitals

BackgroundAcinetobacter baumannii cause difficult-to-treat infections mostly among immunocompromised patients. Clinically relevant A. baumannii lineages and their carbapenem resistance mechanisms are sparsely described in Nigeria. ObjectiveThis study aimed to characterise the diversity and genetic mechanisms of carbapenem resistance among A. baumannii strains isolated from hospitals in southwestern Nigeria. MethodsWe sequenced the genomes of all A. baumannii isolates submitted to Nigerias antimicrobial resistance surveillance reference laboratory between 2016 - 2020 on an Illumina platform and performed in silico genomic characterisation. Selected strains were sequenced using the Oxford Nanopore technology to characterise the genetic context of carbapenem resistance genes. ResultsThe 86 A. baumannii isolates were phylogenetically diverse and belonged to 35 distinct sequence types (STs), 16 of which were novel. Thirty-eight (44.2%) isolates belonged to none of the known international clones (ICs). Over 50% of the isolates were phenotypically resistant to 10 of 12 tested antimicrobials. Majority (n=54) of the isolates were carbapenem-resistant, particularly the IC7 (100%) and IC9 (>91.7%) strains. blaOXA-23 (34.9%) and blaNDM-1 (27.9%) were the most common carbapenem resistance genes detected. All blaOXA-23 genes were carried on Tn2006 or Tn2006-like transposons. Our findings suggest that the mobilisation of a 10kb Tn125 composite transposon is the primary means of blaNDM-1 dissemination. ConclusionOur findings highlight an increase in blaNDM-1 prevalence and the widespread transposon-facilitated dissemination of carbapenemase genes in diverse A. baumannii lineages in southwestern Nigeria. We make the case for improving surveillance of these pathogens in Nigeria and other understudied settings. ImportanceAcinetobacter baumannii are increasingly clinically relevant due to their propensity to harbour genes conferring resistance to multiple antimicrobials, as well as their ability to persist and disseminate in hospital environments and cause difficult-to-treat nosocomial infections. Little is known about the molecular epidemiology and antimicrobial resistance profiles of these organisms in Nigeria, largely due to limited capacity for their isolation, identification, and antimicrobial susceptibility testing. Our study characterised the diversity and antimicrobial resistance profiles of clinical A. baumannii in southwestern Nigeria using whole-genome sequencing. We also identified the key genetic elements facilitating the dissemination of carbapenem resistance genes within this species. This study provides key insights into the clinical burden and population dynamics of A. baumannii in hospitals in Nigeria and highlights the importance of routine whole-genome sequencing-based surveillance of this and other previously understudied pathogens in Nigeria and other similar settings.

genomics↗

bioRxiv2022

Nanopore-only assemblies for genomic surveillance of the global priority drug-resistant pathogen, Klebsiella pneumoniae

BackgroundOxford Nanopore Technologies (ONT) sequencing has rich potential for genomic epidemiology and public health investigations of bacterial pathogens, particularly in low-resource settings and at the point of care, due to its portability and affordability. However, low base-call accuracy has limited the reliability of ONT data for critical tasks such as antimicrobial resistance (AMR) and virulence gene detection and typing, serotype prediction and cluster identification. Thus, Illumina sequencing remains the standard for genomic surveillance despite higher capital and running costs. MethodsWe tested the accuracy of ONT-only assemblies for common applied bacterial genomics tasks (genotyping and cluster detection, implemented via Kleborate, Kaptive and Pathogenwatch), using data from 54 unique Klebsiella pneumoniae isolates. ONT reads generated via MinION with R9.4 flowcells were basecalled using three alternative models (Fast, High-accuracy (HAC) and Super-accuracy (SUP), available within ONTs Guppy software), assembled with Flye and polished using Medaka. Accuracy of typing using ONT-only assemblies was compared with that of Illumina-only and hybrid ONT+Illumina assemblies, constructed from the same isolates as reference standards. ResultsThe most resource-intensive ONT-assembly approach (SUP basecalling, with or without Medaka polishing) performed best, yielding reliable capsule (K) type calls for all strains (100% exact or best matching locus), reliable multi-locus sequence type (MLST) assignment (98.3% exact match or single-locus variants), and good detection of acquired AMR genes and mutations (88% - 100% correct identification across the various drug classes). Distance-based trees generated from SUP+Medaka assemblies accurately reflected overall genetic relationships between isolates; however, the definition of outbreak clusters from ONT-only assemblies was problematic. HAC basecalling + Medaka polishing performed similarly to SUP basecalling without polishing, and polishing introduced errors into HAC- or Fast-basecalled assemblies. Therefore, we recommend investing compute resources into basecalling (SUP model) over polishing, where compute resources and/or time are limiting. ConclusionsOverall, our results show that MLST, K type and AMR determinants can be reliably identified with ONT-only data. However, cluster detection remains challenging with this technology.

genomics↗

bioRxiv2022

Genomic Surveillance of Salmonella spp. in the Philippines, 2013-2014

Increasing antimicrobial resistance (AMR) in Salmonella has been observed in the Philippines. This study aims to utilize whole genome sequencing (WGS) to characterize the population and AMR mechanisms of Salmonella captured by the Philippine Antimicrobial Resistance Surveillance Program (ARSP) and contrast to traditional laboratory methods. We sequenced the whole genomes of 148 Salmonella Typhi (S. Typhi) and 65 non-typhoidal Salmonella (NTS) collected in the Philippines in 2013-2014. From the genome sequences, we determined the serotype, multilocus sequence type, presence of determinants of antimicrobial resistance and relatedness between isolates. We also compared the genotypic predictions of serotype and AMR to the phenotypic data. AMR rates in S. Typhi were low, with sparse acquisition of mutations associated with reduced susceptibility to fluoroquinolones or extended-spectrum beta-lactamases (ESBL) genes. In contrast, three quarters of NTS isolates were insusceptible to at least one antimicrobial, with more than half carrying mutations and/or genes linked to resistance to fluoroquinolones. ESBL genes were detected in five genomes that also carried other AMR determinants. The population of S. Typhi was dominated by the likely endemic genotype 3.0, which also caused of a putative local outbreak susceptible to antibiotics. The main NTS clades were global epidemic S. Enteritidis ST11 and the monophasic variant of S. Typhimurium (I 4,[5],12:i:-) ST34, which had frequently been serotyped as S. Typhimurium in the laboratory. This was the first time that Salmonella isolated from the Philippines were characterized by WGS and we provide evidence of its utility for ongoing surveillance at the ARSP.

genomics↗

bioRxiv2022

Recurrent emergence of carbapenem resistance in Klebsiella pneumoniae mediated by an inhibitory ompK36 mRNA secondary structure

Outer membrane porins in Gram-negative bacteria facilitate antibiotic influx. In Klebsiella pneumoniae (KP), modifications in the porin OmpK36 are implicated in increasing resistance to carbapenems. Analysis of large KP genome collections, encompassing major healthcare-associated clones, revealed the recurrent emergence of a synonymous cytosine to thymine transition at position 25 (25c>t) in ompK36. We show that the 25c>t transition increases carbapenem resistance through depletion of OmpK36 from the outer membrane. The mutation attenuates KP in a murine pneumonia model, which accounts for its limited clonal expansion observed by phylogenetic analysis. However, in the context of carbapenem treatment, the 25c>t transition tips the balance towards treatment failure, thus accounting for its recurrent emergence. Mechanistically, the 25c>t transition mediates an intramolecular mRNA interaction between a uracil encoded by 25t and the first adenine within the Shine-Dalgarno sequence. This specific interaction leads to the formation of an RNA stem structure, which obscures the ribosomal binding site thus disrupting translation. While mutations reducing OmpK36 expression via transcriptional silencing are known, we uniquely demonstrate the repeated selection of a synonymous ompK36 mutation mediating translational suppression in response to antibiotic pressure.

microbiology↗

bioRxiv2021

Circulation of third-generation cephalosporin resistant Salmonella Typhi in Mumbai, India

We report the persistent circulation of third-generation cephalosporin resistant Salmonella Typhi in Mumbai, linked to the acquisition and maintenance of a previously characterized IncX3 plasmid carrying the ESBL gene blaSHV-12 and the fluoroquinolone resistance gene qnrB7 in the genetic context of a triple mutant also associated with fluoroquinolone resistance.

genomics↗

bioRxiv2021

Complexity of genomic epidemiology of carbapenem-resistant Klebsiella pneumoniae isolates in Colombia urges the reinforcement of whole genome sequencing based surveillance programs

BackgroundCarbapenem-resistant Klebsiella pneumoniae (CRKP) is an emerging public health problem. This study explores the specifics of CRKP epidemiology in Colombia based on whole genome sequencing (WGS) of the National Reference Laboratory at Instituto Nacional de Salud (INS)s 2013-2017 sample collection. MethodsA total of 425 CRKP isolates from 21 departments were analyzed by HiSeq-X10(R)Illumina high-throughput sequencing. Bioinformatic analysis was performed, primarily using the pipelines developed collaboratively by the National Institute for Health Research Global Health Research Unit (GHRU) on Genomic Surveillance of AMR, and AGROSAVIA. ResultsOf the 425 CRKP isolates, 91.5% were carbapenemase-producing strains. The data support a recent expansion and the endemicity of CRKP in Colombia with the circulation of 7 high-risk clones, the most frequent being CG258 (48.39% of isolates). We identified genes encoding carbapenemases blaKPC-3, blaKPC-2, blaNDM-1, blaNDM-9, blaVIM-2, blaVIM-4, and blaVIM-24, and various mobile genetic elements (MGE). The virulence of CRKP isolates was low, but colibactin (clb3) was present in 25.2% of isolates, and a hypervirulent CRKP clone (CG380) was reported for the first time in Colombia. ST258, ST512, and ST4851 were characterized by low levels of diversity in the core genome (ANI> 99.9%). ConclusionsThe study outlines complex CRKP epidemiology in Colombia. CG258 expanded clonally and carries specific carbapenemases in specific MGEs, while the other high-risk clones (CG147, CG307, and CG152) present a more diverse complement of carbapenemases. The specifics of the Colombian situation stress the importance of WGS-based surveillance to monitor evolutionary trends of STs, MGE, and resistance and virulence genes. summaryIn Colombia, the dissemination of carbapenemases in carbapenem-resistant Klebsiella pneumoniae is attributed to horizontal gene transfer and successful circulation of CG258, and, to a lesser extent, other clones such as ST307, ST147, and ST152.

genomics↗

bioRxiv2021

Clones and Clusters of Antimicrobial-Resistant Klebsiella from Southwestern Nigeria

IntroductionKlebsiella pneumoniae is a World Health Organization high-priority antibiotic-resistant pathogen. However, little is known about the population structure and evolution of Klebsiella circulating in Nigeria. MethodsWe performed whole genome sequencing (WGS) of 141 Klebsiella isolated between 2016 and 2018 from clinical specimens at 3 antimicrobial-resistance (AMR) sentinel surveillance tertiary hospitals in southwestern Nigeria. We conducted in silico multilocus sequence typing, AMR gene, virulence gene, plasmid, and K and O loci profiling, as well as phylogenetic analyses, using publicly available tools and Nextflow pipelines. ResultsPhylogenetic analysis revealed that the majority of the 134 K. pneumoniae and 5 K. quasipneumoniae isolates from Nigeria characterized are closely related to globally disseminated multidrug-resistant clones. Of the 39 K. pneumoniae sequence types (STs) identified, the most common were ST307 (15%), ST5241 (12%), ST15 (~9%), and ST25 (~6%). ST5241, one of 10 novel STs detected, is a single locus variant of ST636 carrying dfrA14, tetD, qnrS, and oqxAB resistance genes. The extended-spectrum {beta} lactamase (ESBL) gene blaCTX_M-15 was seen in 72 % of K. pneumoniae genomes, while 8% encoded a carbapenemase. Four likely outbreak clusters from one facility, within STs 17, 25, 307, and 5241, were ESBL but not carbapenemase-bearing clones. ConclusionThis study uncovered known and novel K. pneumoniae lineages circulating in Nigeria that include multidrug-resistant ESBL producers. Carbapenemase-producing isolates remain uncommon. WGS retrospectively identified outbreak clusters, pointing to the value of genomic approaches in AMR surveillance for improving infection prevention and control in Nigerian hospitals. summaryWe performed whole genome sequencing (WGS) of 141 Klebsiella isolated in 2016-2018 at 3 antimicrobial-resistance (AMR) sentinel surveillance tertiary hospitals in southwestern Nigeria. This study uncovered known and novel K. pneumoniae lineages circulating in Nigeria that include multidrug-resistant ESBL producers. FUNDINGThis work was supported by Official Development Assistance (ODA) funding from the National Institute of Health Research [16/136/111: NIHR Global Health Research Unit on Genomic Surveillance of Antimicrobial Resistance]. This research was commissioned by the National Institute of Health Research using Official Development Assistance (ODA) funding. INO is an African Research Leader supported by the UK Medical Research Council (MRC) and the UK Department for International Development (DFID) under the MRC/DFID Concordat agreement that is also part of the EDCTP2 program supported by the European Union. The funders had no role in the content, crafting or submission of this paper. The views expressed in this publication are those of the authors and not necessarily those of the funders or their affiliates. CONFLICT OF INTERESTThe authors: No reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest.

genomics↗

bioRxiv2021

Rapid Genomic Characterization and Global Surveillance of Klebsiella Using Pathogenwatch

BackgroundKlebsiella species, including the notable pathogen K. pneumoniae, are increasingly associated with antimicrobial resistance (AMR). Genome-based surveillance can inform interventions aimed at controlling AMR. However, its widespread implementation requires tools to streamline bioinformatic analyses and public health reporting. MethodsWe developed the web application Pathogenwatch, which implements analytics tailored to Klebsiella species for integration and visualization of genomic and epidemiological data. We populated Pathogenwatch with 16,537 public Klebsiella genomes to enable contextualization of user genomes. We demonstrated its features with 1,636 genomes from four low- and middle-income countries (LMICs) participating in the NIHR Global Health Research Unit (GHRU) on AMR. ResultsUsing Pathogenwatch, we found that GHRU genomes were dominated by a small number of epidemic drug-resistant clones of K. pneumoniae. However, differences in their distribution were observed (e.g. ST258/512 dominated in Colombia, ST231 in India, ST307 in Nigeria, ST147 in the Philippines). Phylogenetic analyses including public genomes for contextualization enabled retrospective monitoring of their spread. In particular, we identified hospital outbreaks, detected introductions from abroad, and uncovered clonal expansions associated with resistance and virulence genes. Assessment of loci encoding O-antigens and capsule in K. pneumoniae, which represent possible vaccine candidates, showed that three O-types (O1-O3) represented 88.9% of all genomes, whereas capsule types were much more diverse. ConclusionsPathogenwatch provides a free, accessible platform for real-time analysis of Klebsiella genomes to aid surveillance at local, national and global levels. We have improved representation of genomes from GHRU participant countries, further facilitating ongoing surveillance. summaryPathogenwatch is a free web-application for analysis of Klebsiella genomes to aid surveillance at local, national and global levels. We improved the representation of genomes from middle-income countries through the Global Health Research Unit on AMR, further facilitating ongoing surveillance.

genomics↗

bioRxiv2021

Genome Sequencing Identifies Previously Unrecognized Klebsiella pneumoniae Outbreaks in Neonatal Intensive Care Units in the Philippines

BackgroundKlebsiella pneumoniae is a critically important pathogen in the Philippines. Isolates are commonly resistant to at least two classes of antibiotics, yet mechanisms and spread of its resistance are not well studied. MethodsA retrospective sequencing survey was performed on carbapenem-, extended spectrum beta-lactam- and cephalosporin-resistant Klebsiella pneumoniae isolated at 20 antimicrobial resistance (AMR) surveillance sentinel sites from 2015-2017. We characterized 259 isolates using biochemical methods, antimicrobial susceptibility testing, and whole genome sequencing (WGS). Known AMR mechanisms were identified. Potential outbreaks were investigated by detecting clusters from epidemiologic, phenotypic and genome-derived data. ResultsPrevalent AMR mechanisms detected include blaCTX-M-15 (76.8%) and blaNDM-1 (37.5%). An epidemic IncFII(Yp) plasmid carrying blaNDM-1 was also detected in 46 isolates from 6 sentinel sites and 14 different sequence types (ST). This plasmid was also identified as the main vehicle of carbapenem resistance in 2 previously unrecognized local outbreaks of ST348 and ST283 at 2 different sentinel sites. A third local outbreak of ST397 was also identified but without the IncFII(Yp) plasmid. Isolates in each outbreak site showed identical STs, K- and O-loci, and similar resistance profiles and AMR genes. All outbreak isolates were collected from blood of children aged <1. ConclusionWGS provided an in-depth understanding of the epidemiology of AMR in the Philippines, which was not possible with only phenotypic and epidemiologic data. The identification of three previously unrecognized Klebsiella outbreaks highlights the utility of WGS in outbreak detection, as well as its importance in public health and in implementing infection control programs. summaryWhole genome sequencing identified three distinct previously unrecognized local outbreaks in a retrospective study in the Philippines, along with an epidemic plasmid carrying antimicrobial resistance genes, highlighting its importance in antimicrobial resistance surveillance, outbreak detection and infection control.

genomics↗

bioRxiv2021

Genomic Surveillance of Acinetobacter baumannii in the Philippines, 2013-2014

Acinetobacter baumannii is an opportunistic nosocomial pathogen that has increasingly become resistant to carbapenems worldwide. In the Philippines, carbapenem resistance and multi-drug resistance (MDR) rates are above 50%. We undertook a genomic study of carbapenem resistant A. baumannii in the Philippines to characterize the population diversity and antimicrobial resistance (AMR) mechanisms. We sequenced the whole genomes of 117 A. baumannii isolates recovered by 16 hospitals in the Philippines between 2013 and 2014. We determined the multi-locus sequence type (MLST), presence of acquired AMR determinants and relatedness between isolates from the genome sequences. We also compared the phenotypic and genotypic resistance results. Carbapenem resistance was mainly explained by the acquisition of class-D beta-lactamase gene blaOXA-23. The concordance between phenotypic and genotypic resistance to imipenem was 98.15% and 94.97% overall for the seven antibiotics analysed. Twenty-two different sequence types (ST) were identified, including 7 novel STs. The population was dominated by high-risk international clone 2 (i.e., clonal complex 92), in particular by ST195 and ST208 and their single locus variants. With WGS we identified local clusters representing potential undetected nosocomial outbreaks, as well as multi-hospital clusters indicating inter-hospital transmission. Comparison with global genomes suggested that the establishment of carbapenem-resistant IC2 clones in the Philippines is likely the result of clonal expansion and geographical dissemination and at least partly explained by inadequate hospital infection control and prevention. This study is the first extensive genomic study of carbapenem-resistant A. baumannii in the Philippines and underscores the importance of hospital infection control and prevention to contain high-risk clones.

genomics↗

bioRxiv2020

MAJORA: Continuous integration supporting decentralised sequencing for SARS-CoV-2 genomic surveillance

Genomic epidemiology has become an increasingly common tool for epidemic response. Recent technological advances have made it possible to sequence genomes rapidly enough to inform outbreak response, and cheaply enough to justify dense sampling of even large epidemics. With increased availability of sequencing it is possible for agile networks of sequencing facilities to collaborate on the sequencing and analysis of epidemic genomic data. In response to the ongoing SARS-CoV-2 pandemic in the United Kingdom, the COVID-19 Genomics UK (COG-UK) consortium was formed with the aim of rapidly sequencing SARS-CoV-2 genomes as part of a national-scale genomic surveillance strategy. The network consists of universities, academic institutes, regional sequencing centres and the four UK Public Health Agencies. We describe the development and deployment of Majora, an encompassing digital infrastructure to address the challenge of collecting and integrating both genomic sequencing data and sample-associated metadata produced across the COG-UK network. The system was designed and implemented pragmatically to stand up capacity rapidly in a pandemic caused by a novel virus. This approach has underpinned the success of COG-UK, which has rapidly become the leading contributor of SARS-CoV-2 genomes to international databases and has generated over 60,000 sequences to date.

bioinformatics↗

bioRxiv2020

A global resource for genomic predictions of antimicrobial resistance and surveillance of Salmonella Typhi at Pathogenwatch

BackgroundMicrobial whole-genome sequencing (WGS) is now increasingly used to inform public health investigations of infectious disease. This approach has transformed our understanding of the global population structure of Salmonella enterica serovar Typhi (S. Typhi), the causative agent of typhoid fever. WGS has been particularly informative for understanding the global spread of multi-drug resistant (MDR) typhoid. As WGS capacity becomes more decentralised, there is a growing opportunity for collaboration and sharing of surveillance data within and between countries to inform disease control policies. This requires freely available, community driven tools that reduce the barriers to access genomic data for public health surveillance and that deliver genomic data on a global scale. MethodsHere we present the Pathogenwatch (https://pathogen.watch/styphi) scheme for S. Typhi, a web application enabling the rapid identification of genomic markers of antimicrobial resistance (AMR) and contextualization with public genomic data to identify high-risk clones at a population level. Data are delivered in single genome reports or in collections of genomes combined with geographic and other data using trees, maps and tables. ResultsWe show that the clustering of S. Typhi genomes in Pathogenwatch is comparable to established bioinformatics methods, and that genomic predictions of AMR are largely concordant with phenotypic drug susceptibility data. We demonstrate the public health utility of Pathogenwatch with examples selected from over 4,300 public genomes available in the application. ConclusionsPathogenwatch democratises genomic epidemiology of S. Typhi by providing an intuitive entry point for the analysis of WGS and linked epidemiological data, enabling international public health monitoring of the emergence and spread of high risk clones.

genomics↗