See and Sequence: Integrating Whole-Genome Sequencing Within the National Antimicrobial Resistance Surveillance Program in the Philippines

AbstractDrug-resistant bacterial infections constitute a growing threat to public health globally 1. National networks of laboratory-based surveillance of antimicrobial resistance (AMR) monitor the emergence and spread of resistance and are central to the dissemination of these data to AMR stakeholders 2. Whole-genome sequencing (WGS) can support these efforts by pinpointing resistance mechanisms and uncovering transmission patterns 3, 4. However, genomic surveillance is rare in low- and middle-income countries (LMICs), which are predicted to be the most affected by AMR 5. We implemented WGS within the established Antimicrobial Resistance Surveillance Program (ARSP) of the Philippines via ongoing technology transfer, capacity building in and binational collaboration. In parallel, we conducted an initial large-scale retrospective sequencing survey to characterize bacterial populations and dissect resistance phenotypes of key bug-drug combinations, which is the focus of this article. Starting in 2010, the ARSP phenotypic data indicated increasing carbapenem resistance rates for Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae and Escherichia coli. We first identified that this coincided with a marked expansion of specific resistance phenotypes. By then linking the resistance phenotypes to genomic data, we revealed the diversity of genetic lineages (strains), AMR mechanisms, and AMR vehicles underlying this expansion. We discovered a previously unreported plasmid-driven hospital outbreak of carbapenem-resistant K. pneumoniae, uncovered the interplay of carbapenem resistance genes and plasmids in the geographic circulation of epidemic K. pneumoniae ST147, and found that carbapenem-resistant E. coli ST410 consisted of diverse lineages of global circulation that carried both international and local plasmids, resulting in a combination of carbapenemase genes variants previously unreported for this organism. Thus, the WGS data provided an enhanced understanding of the interplay between strains, genes and vehicles driving the dissemination of carbapenem resistance in the Philippines. In addition, our retrospective survey served both as the genetic background to contextualize local prospective surveillance, and as a comprehensive dataset for training in bioinformatics and genomic epidemiology. Continued prospective sequencing, capacity building and collaboration will strengthen genomic surveillance of AMR in the Philippines and the translation of genomic data into public-health action. We generated a blueprint for the integration of WGS and genomic epidemiology into an established national system of laboratory-based surveillance of AMR through international collaboration that can be adapted and utilized within other locations to tackle the global challenge of AMR.

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Integrating whole-genome sequencing within the National Antimicrobial Resistance Surveillance Program in the Philippines

Nature Communications ◽

10.1038/s41467-020-16322-5 ◽

2020 ◽

Vol 11 (1) ◽

Author(s):

Silvia Argimón ◽

Melissa A. L. Masim ◽

June M. Gayeta ◽

Marietta L. Lagrada ◽

Polle K. V. Macaranas ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

The Philippines ◽

Surveillance Program ◽

Whole Genome ◽

Antimicrobial Resistance Surveillance

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Whole-genome sequencing as part of national and international surveillance programmes for antimicrobial resistance: a roadmap

BMJ Global Health ◽

10.1136/bmjgh-2019-002244 ◽

2020 ◽

Vol 5 (11) ◽

pp. e002244

Author(s):

Keyword(s):

Public Health ◽

Antimicrobial Resistance ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Low Income ◽

The Philippines ◽

Surveillance Systems ◽

Whole Genome ◽

Global Public Health ◽

Bacterial Strains

The global spread of antimicrobial resistance (AMR) and lack of novel alternative treatments have been declared a global public health emergency by WHO. The greatest impact of AMR is experienced in resource-poor settings, because of lack of access to alternative antibiotics and because the prevalence of multidrug-resistant bacterial strains may be higher in low-income and middle-income countries (LMICs). Intelligent surveillance of AMR infections is key to informed policy decisions and public health interventions to counter AMR. Molecular surveillance using whole-genome sequencing (WGS) can be a valuable addition to phenotypic surveillance of AMR. WGS provides insights into the genetic basis of resistance mechanisms, as well as pathogen evolution and population dynamics at different spatial and temporal scales. Due to its high cost and complexity, WGS is currently mainly carried out in high-income countries. However, given its potential to inform national and international action plans against AMR, establishing WGS as a surveillance tool in LMICs will be important in order to produce a truly global picture. Here, we describe a roadmap for incorporating WGS into existing AMR surveillance frameworks, including WHO Global Antimicrobial Resistance Surveillance System, informed by our ongoing, practical experiences developing WGS surveillance systems in national reference laboratories in Colombia, India, Nigeria and the Philippines. Challenges and barriers to WGS in LMICs will be discussed together with a roadmap to possible solutions.

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Genomic surveillance of Neisseria gonorrhoeae in the Philippines, 2013–2014

Western Pacific Surveillance and Response ◽

10.5365/wpsar.2020.11.1.005 ◽

2021 ◽

Vol 12 (1) ◽

pp. 17-25

Author(s):

Alfred Villamin ◽

Silvia Argimon ◽

Karis Bohme ◽

Melissa Masim ◽

June Gayeta ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Neisseria Gonorrhoeae ◽

Sequence Data ◽

The Philippines ◽

Sequence Type ◽

Surveillance Program ◽

Gyra Gene ◽

Genotypic Resistance ◽

Genomic Epidemiology ◽

Epidemiology Studies

Antimicrobial-resistant Neisseria gonorrhoeae is a major threat to public health and is of particular concern in the Western Pacific Region, where the incidence of gonorrhoea is high. The Antimicrobial Resistance Surveillance Program (ARSP) has been capturing information on resistant gonorrhoea since 1996, but genomic epidemiology studies on this pathogen are lacking in the Philippines. We sequenced the whole genomes of 21 N. gonorrhoeae isolates collected in 2013–2014 by ARSP. The multilocus sequence type, multiantigen sequence type, presence of determinants of antimicrobial resistance and relatedness among the isolates were all derived from the sequence data. The concordance between phenotypic and genotypic resistance was also determined. Ten of 21 isolates were resistant to penicillin, ciprofloxacin and tetracycline, due mainly to the presence of the blaTEM gene, the S91F mutation in the gyrA gene and the tetM gene, respectively. None of the isolates was resistant to ceftriaxone or cefixime. The concordance between phenotypic and genotypic resistance was 92.38% overall for five antibiotics in four classes. Despite the small number of isolates studied, they were genetically diverse, as shown by the sequence types, the N. gonorrhoeae multiantigen sequence typing types and the tree. Comparison with global genomes placed the Philippine genomes within global lineage A and led to the identification of an international transmission route.

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Whole-Genome Sequencing for Molecular Characterization of Carbapenem-Resistant Enterobacteriaceae Causing Lower Urinary Tract Infection among Pediatric Patients

Antibiotics ◽

10.3390/antibiotics10080972 ◽

2021 ◽

Vol 10 (8) ◽

pp. 972

Author(s):

Hassan Al Mana ◽

Sathyavathi Sundararaju ◽

Clement K. M. Tsui ◽

Andres Perez-Lopez ◽

Hadi Yassine ◽

...

Keyword(s):

Urinary Tract ◽

Antimicrobial Resistance ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Bacterial Species ◽

Whole Genome ◽

E Coli ◽

Carbapenem Resistant ◽

Tract Infections ◽

Carbapenem Resistant Enterobacteriaceae

Antibiotic resistance is a growing public health problem globally, incurring health and cost burdens. The occurrence of antibiotic-resistant bacterial infections has increased significantly over the years. Gram-negative bacteria display the broadest resistance range, with bacterial species expressing extended-spectrum β-lactamases (ESBLs), AmpC, and carbapenemases. All carbapenem-resistant Enterobacteriaceae (CRE) isolates from pediatric urinary tract infections (UTIs) between October 2015 and November 2019 (n = 30). All isolates underwent antimicrobial resistance phenotypic testing using the Phoenix NMIC/ID-5 panel, and carbapenemase production was confirmed using the NG-Test CARBA 5 assay. Whole-genome sequencing was performed on the CREs. The sequence type was identified using the Achtman multi-locus sequence typing scheme, and antimicrobial resistance markers were identified using ResFinder and the CARD database. The most common pathogens causing CRE UTIs were E. coli (63.3%) and K. pneumoniae (30%). The most common carbapenemases produced were OXA-48-like enzymes (46.6%) and NDM enzymes (40%). Additionally, one E. coli harbored IMP-26, and two K. pneumoniae possessed mutations in ompK37 and/or ompK36. Lastly, one E. coli had a mutation in the marA porin and efflux pump regulator. The findings highlight the difference in CRE epidemiology in the pediatric population compared to Qatar’s adult population, where NDM carbapenemases are more common.

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The Role of Whole Genome Sequencing in the Surveillance of Antimicrobial Resistant Enterococcus spp.: A Scoping Review

Frontiers in Public Health ◽

10.3389/fpubh.2021.599285 ◽

2021 ◽

Vol 9 ◽

Author(s):

Lindsay A. Rogers ◽

Kayla Strong ◽

Susan C. Cork ◽

Tim A. McAllister ◽

Karen Liljebjelke ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

Scoping Review ◽

Companion Animals ◽

Surveillance Program ◽

Whole Genome ◽

Antimicrobial Resistance Genes ◽

Wide Range ◽

Scoping Reviews

Enterococcus spp. have arisen as important nosocomial pathogens and are ubiquitous in the gastrointestinal tracts of animals and the environment. They carry many intrinsic and acquired antimicrobial resistance genes. Because of this, surveillance of Enterococcus spp. has become important with whole genome sequencing emerging as the preferred method for the characterization of enterococci. A scoping review was designed to determine how the use of whole genome sequencing in the surveillance of Enterococcus spp. adds to our knowledge of antimicrobial resistance in Enterococcus spp. Scoping review design was guided by the PRISMA extension and checklist and JBI Reviewer's Guide for scoping reviews. A total of 72 articles were included in the review. Of the 72 articles included, 48.6% did not state an association with a surveillance program and 87.5% of articles identified Enterococcus faecium. The majority of articles included isolates from human clinical or screening samples. Significant findings from the articles included novel sequence types, the increasing prevalence of vancomycin-resistant enterococci in hospitals, and the importance of surveillance or screening for enterococci. The ability of enterococci to adapt and persist within a wide range of environments was also a key finding. These studies emphasize the importance of ongoing surveillance of enterococci from a One Health perspective. More studies are needed to compare the whole genome sequences of human enterococcal isolates to those from food animals, food products, the environment, and companion animals.

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Genomic Analysis of a Pan-Resistant Isolate ofKlebsiella pneumoniae, United States 2016

mBio ◽

10.1128/mbio.00440-18 ◽

2018 ◽

Vol 9 (2) ◽

Cited By ~ 16

Author(s):

Tom J. B. de Man ◽

Joseph D. Lutgring ◽

David R. Lonsway ◽

Karen F. Anderson ◽

Julia A. Kiehlbauch ◽

...

Keyword(s):

Public Health ◽

United States ◽

Antimicrobial Resistance ◽

Klebsiella Pneumoniae ◽

Whole Genome Sequencing ◽

Genome Sequencing ◽

The United States ◽

Whole Genome ◽

Beta Lactams ◽

Content Type

ABSTRACTAntimicrobial resistance is a threat to public health globally and leads to an estimated 23,000 deaths annually in the United States alone. Here, we report the genomic characterization of an unusualKlebsiella pneumoniae, nonsusceptible to all 26 antibiotics tested, that was isolated from a U.S. patient. The isolate harbored four known beta-lactamase genes, including plasmid-mediatedblaNDM-1andblaCMY-6, as well as chromosomalblaCTX-M-15andblaSHV-28, which accounted for resistance to all beta-lactams tested. In addition, sequence analysis identified mechanisms that could explain all other reported nonsusceptibility results, including nonsusceptibility to colistin, tigecycline, and chloramphenicol. Two plasmids, IncA/C2 and IncFIB, were closely related to mobile elements described previously and isolated from Gram-negative bacteria from China, Nepal, India, the United States, and Kenya, suggesting possible origins of the isolate and plasmids. This is one of the firstK. pneumoniaeisolates in the United States to have been reported to the Centers for Disease Control and Prevention (CDC) as nonsusceptible to all drugs tested, including all beta-lactams, colistin, and tigecycline.IMPORTANCEAntimicrobial resistance is a major public health threat worldwide. Bacteria that are nonsusceptible or resistant to all antimicrobials available are of major concern to patients and the public because of lack of treatment options and potential for spread. AKlebsiella pneumoniaestrain that was nonsusceptible to all tested antibiotics was isolated from a U.S. patient. Mechanisms that could explain all observed phenotypic antimicrobial resistance phenotypes, including resistance to colistin and beta-lactams, were identified through whole-genome sequencing. The large variety of resistance determinants identified demonstrates the usefulness of whole-genome sequencing for detecting these genes in an outbreak response. Sequencing of isolates with rare and unusual phenotypes can provide information on how these extremely resistant isolates develop, including whether resistance is acquired on mobile elements or accumulated through chromosomal mutations. Moreover, this provides further insight into not only detecting these highly resistant organisms but also preventing their spread.

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Genomic Surveillance of Neisseria gonorrhoeae in the Philippines from 2013-2014

10.1101/2020.03.19.998435 ◽

2020 ◽

Author(s):

Alfred S. Villamin ◽

Silvia Argimón ◽

Manuel C. Jamoralin ◽

Karis D. Boehme ◽

Melissa L. Masim ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Neisseria Gonorrhoeae ◽

Sequence Data ◽

The Philippines ◽

Sequence Type ◽

Western Pacific ◽

Surveillance Program ◽

Genotypic Resistance ◽

Genomic Epidemiology ◽

The Western Pacific

AbstractAntimicrobial-resistant Neisseria gonorrhoeae is a major threat to public health, and of particular concern in the Western Pacific Region, where the incidence of gonorrhoea is high. The Antimicrobial Resistance Surveillance Program (ARSP) has been capturing information on resistant gonorrhoea since 1996, but studies of the genomic epidemiology of gonorrhoea in the Philippines are lacking.We sequenced the whole genomes of 21 N. gonorrhoeae isolates collected in 2013-2014 by the ARSP. The multi-locus sequence type, multi-antigen sequence type, presence of antimicrobial resistance (AMR) determinants, and relatedness between the isolates were all derived from the sequence data. The concordance between phenotypic and genotypic resistance was also determined.Ten out of 21 isolates were resistant to penicillin, ciprofloxacin and tetracycline, mostly linked to the presence of blaTEM gene, the S91F mutation in the gyrA gene, and the tetM gene, respectively. None of the isolates were resistant to azithromycin, ceftriaxone or cefixime, although we identified the A24-deletion in the mtrR promoter in one isolate. The concordance between phenotypic and genotypic resistance was 92.38% overall for 5 antibiotics in 4 classes. Despite the small number of isolates studied, they were genetically diverse, as shown by the sequence types, the NG-MAST types and the tree. Comparison with global genomes placed the Philippine genomes within global Lineage A and led to the identification of an international transmission route.This first genomic survey of N. gonorrhoeae isolates collected by ARSP will be used to contextualize ongoing prospective surveillance, and it highlights the importance of genomic surveillance in the Western Pacific and other endemic regions to understand the spread of drug-resistant gonorrhoea worldwide.

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Molecular characterization of rectal isolates of carbapenemase-negative carbapenem-resistant enterobacterales obtained from ICU patients in Kuwait by whole-genome sequencing

Journal of Medical Microbiology ◽

10.1099/jmm.0.001409 ◽

2021 ◽

Vol 70 (9) ◽

Author(s):

Amani Al Fadhli ◽

Wafaa Jamal ◽

Vincent O. Rotimi

Keyword(s):

Whole Genome Sequencing ◽

Genome Sequencing ◽

Carbapenem Resistance ◽

Efflux Pumps ◽

Multidrug Resistant ◽

Whole Genome ◽

Genetic Characteristics ◽

Carbapenem Resistant ◽

Genes Encoding ◽

Amoxicillin Clavulanic Acid

Introduction. Carbapenem-resistant enterobacterales (CRE) are listed among the most urgent antibiotic resistance threats. Hypothesis. Previous studies on the mechanisms of CRE in Kuwait have focused on carbapenemases. There have been no studies on non-carbapenemase-producing CRE in Kuwait. Aim/Gap Statement. The aim of this study was to investigate the genetic characteristics of non-carbapenemase-producing carbapenem-resistant enterobacterales (NCPE) isolates using whole-genome sequencing (WGS). Methodology. Fourteen confirmed NCPE isolates that were negative for genes encoding carbapenemase production by polymerase chain reaction (PCR) assays using rectal swabs from intensive care unit patients were characterized using phenotypic, PCR and WGS methods. Susceptibility testing was performed via Etest and clonality via multi-locus sequence typing (MLST). Results. All of the isolates were resistant to ertapenem; 78.6 % were resistant to imipenem, meropenem and trimethoprim–sulfamethoxazole. Resistance to the other antibiotics was variable, ranging from 28.5 (colistin) through 50 (tigecycline) and 64.3 (amikacin) up to 85.7 % against both amoxicillin–clavulanic acid and ciprofloxacin. WGS detected several resistance genes mediating the production of β-lactamases, genes encoding an outer-membrane porin permeability mutation resulting in reduced susceptibility to β-lactams, including carbapenems, and genes for multidrug-resistant (MDR) efflux pumps. The isolates also possessed global activator protein MarA, which mediated reduced permeability to β-lactams. The existence of β-lactamase genes, overexpression of MDR efflux pumps and reduced permeability mediated by the porin genes were responsible for carbapenem resistance. Conclusions. This finding reflects the superior detection capabilities offered by WGS analysis, which can be used to complement traditional methods and overcome their limited resolution in clinical settings.

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2461. Community-Acquired in Name Only: A Cluster of Carbapenem-Resistant Acinetobacter baumannii in a Burn Intensive Care Unit

Open Forum Infectious Diseases ◽

10.1093/ofid/ofz360.2139 ◽

2019 ◽

Vol 6 (Supplement_2) ◽

pp. S852-S852

Author(s):

Erica S Shenoy ◽

Virginia M Pierce ◽

Mohamad Sater ◽

Febriana Pangestu ◽

Ian Herriott ◽

...

Keyword(s):

Intensive Care Unit ◽

Intensive Care ◽

Whole Genome Sequencing ◽

Acinetobacter Baumannii ◽

Genome Sequencing ◽

Carbapenem Resistance ◽

Environmental Sampling ◽

Whole Genome ◽

Single Source ◽

Carbapenem Resistant

Abstract Background Detection of nosocomial outbreaks often relies on epidemiological definitions of community and nosocomial acquisition. We report a cluster of three carbapenem-resistant Acinetobacter baumannii (CRAB) infections linked to a single source patient with infections occurring within 2 days of admission to a burn intensive care unit (ICU). The epidemiological investigation was supplemented by whole-genome sequencing (WGS) of clinical and environmental isolates. Methods Study participants included burn ICU patients identified with infections caused by CRAB. A detailed review of patient demographic and clinical data was conducted. Clinical A. baumannii isolates were assessed by antimicrobial susceptibility testing and WGS. Review of infection control practices on the affected unit was followed by environmental sampling. A. baumannii isolates obtained through environmental sampling were assessed for carbapenem resistance and then underwent WGS for comparison to the clinical isolates. Results Three cases of CRAB infection in the affected unit spanning a period of 3 months were linked to a preceding source patient, with CRAB isolates from the four patients differing by 5–7 single nucleotide variations. All case patients had been admitted to the same room within 2 days before development of CRAB infection. Environmental sampling performed while the third case patient occupied the room identified highly contaminated areas, and environmental CRAB isolates linked the patient isolates. The contaminated areas were subsequently re-sampled after enhanced terminal cleaning of the room. No additional CRAB was isolated, but other pathogenic organisms were recovered. Conclusion We report a cluster of three infections caused by highly resistant A. baumannii that occurred in a burn intensive care unit over a period of 3 months, linked to a single source patient. Three case patients developed infections classified as community-acquired using standard epidemiological definitions, however, whole-genome sequencing revealed clonality. An extensive investigation identified the role of environmental reservoirs. Burn patients may be particularly vulnerable to early-onset nosocomial infection from environmental contamination. Disclosures All authors: No reported disclosures.

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Characterization of an Enterobacter cloacae Strain Producing both KPC and NDM Carbapenemases by Whole-Genome Sequencing

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01275-15 ◽

2015 ◽

Vol 59 (10) ◽

pp. 6625-6628 ◽

Cited By ~ 27

Author(s):

Wenjing Wu ◽

Yu Feng ◽

Alessandra Carattoli ◽

Zhiyong Zong

Keyword(s):

Whole Genome Sequencing ◽

Bloodstream Infection ◽

Genome Sequencing ◽

Carbapenem Resistance ◽

Enterobacter Cloacae ◽

Whole Genome ◽

Content Type ◽

Carbapenem Resistant

ABSTRACTA carbapenem-resistantEnterobacter cloacaestrain, WCHECl-14653, causing a fatal bloodstream infection, was characterized by genome sequencing and conjugation experiments. The strain carried two carbapenemase genes,blaNDM-1andblaKPC-2, on separate IncF plasmids. The coexistence ofblaNDM-1andblaKPC-2conferred slightly higher-level carbapenem resistance compared with that ofblaNDM-1orblaKPC-2alone, and the coexistence of two IncF plasmids may generate new platforms for spreading carbapenemase genes.

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