Evaluation of multiplex tandem PCR (MT-PCR) assays for the detection of bacterial resistance genes among Enterobacteriaceae in clinical urines

Schmidt, K., Stanley, K. K., Hale, R., Smith, L., Wain, J., O'Grady, J. and Livermore, D. M. ORCID: https://orcid.org/0000-0002-9856-3703 (2019) Evaluation of multiplex tandem PCR (MT-PCR) assays for the detection of bacterial resistance genes among Enterobacteriaceae in clinical urines. Journal of Antimicrobial Chemotherapy, 74 (2). 349–356. ISSN 0305-7453

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Abstract

Background: Increasing resistance drives empirical use of less potent and previously reserved antibiotics, including for urinary tract infections (UTIs). Molecular profiling, without culture, might better guide early therapy.  Objectives: To explore the potential of AusDiagnostics multiplex tandem (MT) PCR UTI assays.  Methods: Two MT-PCR assays were developed successively, seeking 8 or 16 resistance genes. Amplification was tracked in real time, with melting temperatures used to confirm product identity. Assays were variously performed on: (i) extracted DNA; (ii) cultured bacteria; (iii) urine spiked with reference strains; and (iv) bacteria harvested from clinical urines. Results were compared with those from sequencing, real-time SybrGreen PCR or phenotypic susceptibility.  Results: Performance was similar irrespective of whether DNA, cultures or urines were used, with >90% sensitivity and specificity with respect to common β-lactamases, dfr genes and aminoglycoside resistance determinants except aadA1/A2/A3, for which carriage correlated poorly with streptomycin resistance. Fluoroquinolone-susceptible and -resistant Escherichia coli (but not other species) were distinguished by the melting temperatures of their gyrA PCR products. The time from urine to results was <3 h.  Conclusions: The MT-PCR assays rapidly identified resistance genes from Gram-negative bacteria in urines as well as from cultivated bacteria. Used directly on urines, this assay has the potential to guide early therapy.

Item Type: Article
Faculty \ School: Faculty of Medicine and Health Sciences > Norwich Medical School
UEA Research Groups: Faculty of Medicine and Health Sciences > Research Groups > Public Health and Health Services Research
Faculty of Medicine and Health Sciences > Research Groups > Epidemiology and Public Health
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Depositing User: LivePure Connector
Date Deposited: 11 Dec 2018 16:30
Last Modified: 04 Aug 2023 14:30
URI: https://ueaeprints.uea.ac.uk/id/eprint/69282
DOI: 10.1093/jac/dky419

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