Rapid and direct real-time detection of blakpc and blandm from urveillance samples

Shawn Vasoo, Scott A. Cunningham, Peggy C. Kohner, Jayawant Mandrekar, Karen Lolans, Mary K. Hayden, Robin Patel

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

We assessed the performance of a duplex real-time PCR assay for blaKPC and blaNDM performed directly (D-PCR) on perianal and perirectal swabs and stool. Spiked specimens and 126 clinical surveillance swabs (comprising a sensitivity panel of 46 perirectal double swabs previously determined to be culture positive for blaKPC-PCR-positive Enterobacteriaceae and a specificity panel of 80 perianal swabs from patients at risk of carbapenemase-producing Enterobacteriaceae [CPE] colonization) were studied. For the surveillance swabs, D-PCR was compared to PCR after broth enrichment (BE-PCR) and two culture-based methods: the HardyCHROM ESBL agar (HC-A) and the CDC screening (CDC-A) methods. PCR was performed on morphologically distinct colonies that were isolated by culture. All of the initial PCR testing was done without extraction using a simple lysis procedure. The analytical sensitivities of D-PCR for blaKPC were 9 CFU/l (for swabs) and 90 CFU/l (for stool), and for blaNDM, it was 1.9 CFU/l (for both swabs and stool). In the clinical sensitivity panel, D-PCR and BE-PCR were initially positive for blaKPC in 41/46 (89.1%) and 43/46 (93.5%) swabs, respectively. The swabs that were initially negative by D-PCR (n5) and BE-PCR (n3) were visibly stool soiled; all swabs were blaKPC positive upon repeat testing after lysate extraction. The CDC-A and HC-A yielded blaKPC-positive Enterobacteriaceae from 36/46 (78.3%) and 35/46 (76.1%) swabs, respectively (sensitivities of D-PCR/BEPCR postextraction of soiled specimens versus HC-A, P-0.0009, and versus CDC-A, P=0.0016). All swabs in the specificity panel were negative for CPE by all four methods. D-PCR allows for the timely detection of blaKPC and blaNDM carriage with excellent sensitivity when specimens visibly soiled with stool undergo preparatory extraction.

Original languageEnglish (US)
Pages (from-to)3609-3615
Number of pages7
JournalJournal of Clinical Microbiology
Volume51
Issue number11
DOIs
StatePublished - Nov 2013

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Polymerase Chain Reaction
Enterobacteriaceae
Centers for Disease Control and Prevention (U.S.)
Agar
Real-Time Polymerase Chain Reaction

ASJC Scopus subject areas

  • Microbiology (medical)

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Rapid and direct real-time detection of blakpc and blandm from urveillance samples. / Vasoo, Shawn; Cunningham, Scott A.; Kohner, Peggy C.; Mandrekar, Jayawant; Lolans, Karen; Hayden, Mary K.; Patel, Robin.

In: Journal of Clinical Microbiology, Vol. 51, No. 11, 11.2013, p. 3609-3615.

Research output: Contribution to journalArticle

Vasoo, Shawn ; Cunningham, Scott A. ; Kohner, Peggy C. ; Mandrekar, Jayawant ; Lolans, Karen ; Hayden, Mary K. ; Patel, Robin. / Rapid and direct real-time detection of blakpc and blandm from urveillance samples. In: Journal of Clinical Microbiology. 2013 ; Vol. 51, No. 11. pp. 3609-3615.
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N2 - We assessed the performance of a duplex real-time PCR assay for blaKPC and blaNDM performed directly (D-PCR) on perianal and perirectal swabs and stool. Spiked specimens and 126 clinical surveillance swabs (comprising a sensitivity panel of 46 perirectal double swabs previously determined to be culture positive for blaKPC-PCR-positive Enterobacteriaceae and a specificity panel of 80 perianal swabs from patients at risk of carbapenemase-producing Enterobacteriaceae [CPE] colonization) were studied. For the surveillance swabs, D-PCR was compared to PCR after broth enrichment (BE-PCR) and two culture-based methods: the HardyCHROM ESBL agar (HC-A) and the CDC screening (CDC-A) methods. PCR was performed on morphologically distinct colonies that were isolated by culture. All of the initial PCR testing was done without extraction using a simple lysis procedure. The analytical sensitivities of D-PCR for blaKPC were 9 CFU/l (for swabs) and 90 CFU/l (for stool), and for blaNDM, it was 1.9 CFU/l (for both swabs and stool). In the clinical sensitivity panel, D-PCR and BE-PCR were initially positive for blaKPC in 41/46 (89.1%) and 43/46 (93.5%) swabs, respectively. The swabs that were initially negative by D-PCR (n5) and BE-PCR (n3) were visibly stool soiled; all swabs were blaKPC positive upon repeat testing after lysate extraction. The CDC-A and HC-A yielded blaKPC-positive Enterobacteriaceae from 36/46 (78.3%) and 35/46 (76.1%) swabs, respectively (sensitivities of D-PCR/BEPCR postextraction of soiled specimens versus HC-A, P-0.0009, and versus CDC-A, P=0.0016). All swabs in the specificity panel were negative for CPE by all four methods. D-PCR allows for the timely detection of blaKPC and blaNDM carriage with excellent sensitivity when specimens visibly soiled with stool undergo preparatory extraction.

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