Background Carbapenemase-producing Enterobacterales pose a serious clinical threat, particularly in high-burden settings of carbapenem-resistant Escherichia coli and Klebsiella pneumoniae (CREK), where rapid detection tools are essential to aid patient management. In this study, we focused on blaNDM, the most frequently reported carbapenemase in the region, and evaluated a combined phenotypic (lateral flow) and genotypic (PCR and WGS) approach for its detection. This research underscores the utility of lateral flow assays as a practical alternative to resource-intensive genotypic methods, offering a scalable solution for settings with limited laboratory capacity. Method One hundred seventy-seven extensively drug-resistant strains were characterized using MALDI-TOF. Isolates were analyzed to detect Carbapenem-resistant Escherichia coli and Klebsiella pneumoniae (CREK) using disk diffusion, MIC test, and PCR targeting blaNDM. Antibiotic susceptibility patterns were analyzed and visualized using single-linkage hierarchical clustering, with results displayed on a permuted heat map. Immunochromatographic assay, RESIST-5 O.K.N.V.I (Coris Bioconcept®) was used for CREK isolates [(n = 17), positive and negative)] and Oxford Nanopore Sequencing was conducted on subsets [(n = 5) blaNDM-positive co-producers of blaNDM and blaOXA, and (n = 2) blaNDM-negative blaOXA producers) to evaluate the reliability of phenotypic and genotypic tests. Result Most of the XDR strains (90%) were CREK, with K. pneumoniae (71.2%) more prevalent than E. coli (28.7%) (p < 0.05). All CREK strains exhibited complete resistance (100%) to multiple antibiotics with 66% showing sensitivity to levofloxacin. Furthermore, K. pneumoniae (57.8%) had higher blaNDM gene prevalence than E. coli (36.9%). Among blaNDM-positive CREK, lateral flow assay revealed approximately half of each bacteria type co-produced blaOXA (E.coli, 52.9%), and (K. pneumoniae, 47%). For blaNDM-negative strains, blaOXA was more prevalent in K. pneumoniae (82.35%) than E. coli (41%) (p < 0.05). Comparing phenotypic to genotypic assays, E. coli showed 100% (CI 80.49 − 100%) sensitivity and specificity with a high Kappa agreement coefficient (0.91) (CI 95% 0.661–1, p < 0.01), whereas K. pneumoniae assays had lower sensitivity and specificity (40%) (CI 5.27 − 85.34%), with a lower Kappa agreement coefficient (0.20) (CI 95% 0.104–0.298, p < 0.01). Conclusion This study demonstrates the value of the RESIST-5 O.K.N.V.I. lateral flow assay as a rapid and reliable diagnostic tool for detecting blaNDM in Escherichia coli, with strong agreement to PCR and WGS. While performance for Klebsiella pneumoniae was lower, the assay offers a practical alternative in resource-limited settings, aiding antimicrobial stewardship and improving diagnostic capacities in high-burden regions.

Exploring New Delhi Metallo Beta Lactamases in Klebsiella pneumoniae and Escherichia coli: genotypic vs. phenotypic insights

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2025年2月8日

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Noor Ul Ain,
Linzy Elton,
Zahra Sadouki,
Timothy D. McHugh,
Saba Riaz

摘要

Background

Carbapenemase-producing Enterobacterales pose a serious clinical threat, particularly in high-burden settings of carbapenem-resistant Escherichia coli and Klebsiella pneumoniae (CREK), where rapid detection tools are essential to aid patient management. In this study, we focused on bla_NDM, the most frequently reported carbapenemase in the region, and evaluated a combined phenotypic (lateral flow) and genotypic (PCR and WGS) approach for its detection. This research underscores the utility of lateral flow assays as a practical alternative to resource-intensive genotypic methods, offering a scalable solution for settings with limited laboratory capacity.

Method

One hundred seventy-seven extensively drug-resistant strains were characterized using MALDI-TOF. Isolates were analyzed to detect Carbapenem-resistant Escherichia coli and Klebsiella pneumoniae (CREK) using disk diffusion, MIC test, and PCR targeting bla_NDM. Antibiotic susceptibility patterns were analyzed and visualized using single-linkage hierarchical clustering, with results displayed on a permuted heat map. Immunochromatographic assay, RESIST-5 O.K.N.V.I (Coris Bioconcept®) was used for CREK isolates [(n = 17), positive and negative)] and Oxford Nanopore Sequencing was conducted on subsets [(n = 5) bla_NDM-positive co-producers of bla_NDM and bla_OXA, and (n = 2) bla_NDM-negative bla_OXA producers) to evaluate the reliability of phenotypic and genotypic tests.

Result

Most of the XDR strains (90%) were CREK, with K. pneumoniae (71.2%) more prevalent than E. coli (28.7%) (p < 0.05). All CREK strains exhibited complete resistance (100%) to multiple antibiotics with 66% showing sensitivity to levofloxacin. Furthermore, K. pneumoniae (57.8%) had higher bla_NDM gene prevalence than E. coli (36.9%). Among bla_NDM-positive CREK, lateral flow assay revealed approximately half of each bacteria type co-produced bla_OXA (E.coli, 52.9%), and (K. pneumoniae, 47%). For bla_NDM-negative strains, bla_OXA was more prevalent in K. pneumoniae (82.35%) than E. coli (41%) (p < 0.05). Comparing phenotypic to genotypic assays, E. coli showed 100% (CI 80.49 − 100%) sensitivity and specificity with a high Kappa agreement coefficient (0.91) (CI 95% 0.661–1, p < 0.01), whereas K. pneumoniae assays had lower sensitivity and specificity (40%) (CI 5.27 − 85.34%), with a lower Kappa agreement coefficient (0.20) (CI 95% 0.104–0.298, p < 0.01).

Conclusion

This study demonstrates the value of the RESIST-5 O.K.N.V.I. lateral flow assay as a rapid and reliable diagnostic tool for detecting bla_NDM in Escherichia coli, with strong agreement to PCR and WGS. While performance for Klebsiella pneumoniae was lower, the assay offers a practical alternative in resource-limited settings, aiding antimicrobial stewardship and improving diagnostic capacities in high-burden regions.