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Background <p>Antimicrobial resistance (AMR) is a major global challenge that disproportionately affects low- and middle-income countries (LMICs). Environmental contamination by resistant bacteria from animal production facilities is a major driver of the spread of AMR through the food chain, requiring a robust one-health control approach. Traditional culture-based AMR surveillance is time-consuming and less sensitive, and fails to fully capture the spectrum of AMR, evolutionary trends, and epidemiological patterns of AMR spread. Whole-genome sequencing (WGS) has revolutionized AMR surveillance capabilities. Rapid WGS captures the full AMR spectrum with minimum samples, aids source attribution, and provides insights into trends in AMR spread. The portable Oxford Nanopore® Technology (ONT) platform, coupled with open-source software such as Galaxy and Konstanz Information Miner (KNIME), enables the establishment of a potentially portable, transferable workflow for low-resource settings. This study aimed to assess the AMR burden on four dairy farms in Kandy, Sri Lanka, via a resource-limited LMIC using a low-cost high-throughput screening assay and rapid WGS via ONT with Galaxy and KNIME processing to obtain full antibiotic resistomes.</p> Results <p>The four isolates exhibiting the highest minimum inhibitory concentrations for amoxicillin were identified as <i>Enterobacter cloacae</i> and <i>E. hormaechei</i> by WGS. Chromosomes (4.8 to 4.9&#xa0;Mb) carry the strain-specific resistance genes <i>blaCMH-1</i>, <i>blaACT-25, fosA_7,</i> and <i>ramA</i>, which are associated with diverse antibiotic classes. Plasmids, including IncFIB (pECLA), IncFII (pECLA), and IncX3, carry multiple resistance genes, including <i>AAC(3)-IIe</i>, <i>AAC(6’)-Ib-cr</i>, <i>APH(3″)-Ib</i>, <i>APH(6)-Id</i>, <i>blaCTX-M-15</i>, <i>blaNDM</i>, <i>blaOXA-1</i>, <i>blaTEM-1</i>, <i>dfrA14</i>, <i>QnrB17</i>, <i>catII</i>, <i>determinant-of-bleomycin-resistance</i>, and <i>sul2</i>. Novel arrangements of insertion sequences were observed in <i>E. hormaechei</i> plasmids. The phenotypic resistance of all the isolates matched the genotypic MDR profiles, including resistance to chloramphenicol, gentamicin, tetracycline, and cotrimoxazole.</p> Conclusions <p>ONT WGS with Galaxy and KNIME processing may be a feasible option for AMR surveillance in resource-limited LMICs. To the best of our knowledge, this is the first in-house whole-genome analysis workflow in the country tailored for AMR surveillance. The presence of potentially pathogenic high-MIC, MDR <i>Enterobacter</i> spp. with wide resistomes, including the <i>blaNDM</i> gene, emphasizes the urgent need to address AMR in animal production facilities within a one-health framework.</p>

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Rapid whole genome sequencing for AMR surveillance in low- and middle-income countries: Oxford Nanopore Technology reveals multidrug-resistant Enterobacter cloacae complex from dairy farms in Sri Lanka

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  • 发表日期：

    2025年5月17日

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  • Lakmini S. Kumari,
  • Dinushika M. Siriwardhana,
  • Veranja Liyanapathirana,
  • Rasika Jinadasa,
  • Priyanga Wijesinghe

摘要

Background

Antimicrobial resistance (AMR) is a major global challenge that disproportionately affects low- and middle-income countries (LMICs). Environmental contamination by resistant bacteria from animal production facilities is a major driver of the spread of AMR through the food chain, requiring a robust one-health control approach. Traditional culture-based AMR surveillance is time-consuming and less sensitive, and fails to fully capture the spectrum of AMR, evolutionary trends, and epidemiological patterns of AMR spread. Whole-genome sequencing (WGS) has revolutionized AMR surveillance capabilities. Rapid WGS captures the full AMR spectrum with minimum samples, aids source attribution, and provides insights into trends in AMR spread. The portable Oxford Nanopore® Technology (ONT) platform, coupled with open-source software such as Galaxy and Konstanz Information Miner (KNIME), enables the establishment of a potentially portable, transferable workflow for low-resource settings. This study aimed to assess the AMR burden on four dairy farms in Kandy, Sri Lanka, via a resource-limited LMIC using a low-cost high-throughput screening assay and rapid WGS via ONT with Galaxy and KNIME processing to obtain full antibiotic resistomes.

Results

The four isolates exhibiting the highest minimum inhibitory concentrations for amoxicillin were identified as Enterobacter cloacae and E. hormaechei by WGS. Chromosomes (4.8 to 4.9 Mb) carry the strain-specific resistance genes blaCMH-1, blaACT-25, fosA_7, and ramA, which are associated with diverse antibiotic classes. Plasmids, including IncFIB (pECLA), IncFII (pECLA), and IncX3, carry multiple resistance genes, including AAC(3)-IIe, AAC(6’)-Ib-cr, APH(3″)-Ib, APH(6)-Id, blaCTX-M-15, blaNDM, blaOXA-1, blaTEM-1, dfrA14, QnrB17, catII, determinant-of-bleomycin-resistance, and sul2. Novel arrangements of insertion sequences were observed in E. hormaechei plasmids. The phenotypic resistance of all the isolates matched the genotypic MDR profiles, including resistance to chloramphenicol, gentamicin, tetracycline, and cotrimoxazole.

Conclusions

ONT WGS with Galaxy and KNIME processing may be a feasible option for AMR surveillance in resource-limited LMICs. To the best of our knowledge, this is the first in-house whole-genome analysis workflow in the country tailored for AMR surveillance. The presence of potentially pathogenic high-MIC, MDR Enterobacter spp. with wide resistomes, including the blaNDM gene, emphasizes the urgent need to address AMR in animal production facilities within a one-health framework.