Direct detection of methicillin-resistant in Staphylococcus spp. in positive blood culture by isothermal recombinase pol
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(2020) 36:162
ORIGINAL PAPER
Direct detection of methicillin-resistant in Staphylococcus spp. in positive blood culture by isothermal recombinase polymerase amplification combined with lateral flow dipstick assay Arpasiri Srisrattakarn1 · Patcharaporn Tippayawat1 · Aroonwadee Chanawong1 · Ratree Tavichakorntrakool1 · Jureerut Daduang1 · Lumyai Wonglakorn2 · Pirom Sooksongsoontorn2 · Aroonlug Lulitanond1 Received: 24 March 2020 / Accepted: 17 September 2020 © Springer Nature B.V. 2020
Abstract Methicillin-resistant staphylococci (MRS) are important antimicrobial-resistant pathogens in sepsis. Conventional blood cultures take 24–72 h. The polymerase chain reaction (PCR)-based methods give faster results (2–3 h) but need expensive thermal cyclers. We therefore developed an isothermal recombinase polymerase amplification (RPA) combined with lateral flow dipstick (LFD) assay for rapid detection of MRS in spiked blood culture samples. Fifty-six clinical isolates including 38 mecA-carrying staphylococci and 18 non-mecA-carrying organisms as confirmed by PCR methods were studied. RPA primer set and probe specific for mecA gene (encoding penicillin-binding protein 2a) were designed. RPA reaction was carried out under isothermal condition (45 °C) within 20 min and read by LFD in 5 min. The RPA-LFD provided 92.1% (35/38) sensitivity for identifying MRS in positive blood culture samples, and no cross-amplification was found (100% specificity). This test failed to detect three mecA-carrying S.sciuri isolates. The detection limits of RPA-LFD method for identifying MRS were equal to those of PCR method. The RPA-LFD is simple, fast, and user-friendly. This method could detect the mecA gene directly from the positive blood culture samples without requirement for special equipment. This method would be useful for appropriate antibiotic therapy and infection control, particularly in a low-resource setting. Keywords Lateral flow dipstick · Methicillin-resistant staphylococci · mecA gene · Recombinase polymerase amplification · Sepsis
Introduction Electronic supplementary material The online version of this article (https://doi.org/10.1007/s11274-020-02938-8) contains supplementary material, which is available to authorized users.
Bloodstream infection or sepsis is a major cause of morbidity and mortality among hospitalized patients in intensive care units (ICU) as described by Mayr et al. (2014).
* Aroonlug Lulitanond [email protected]
Lumyai Wonglakorn [email protected]
Arpasiri Srisrattakarn [email protected]
Pirom Sooksongsoontorn [email protected]
Patcharaporn Tippayawat [email protected]
1
Centre for Research and Development of Medical Diagnostic Laboratories, Faculty of Associated Medical Sciences, Khon Kaen University, 40002 Khon Kaen, Thailand
2
Clinical Microbiology Unit, Srinagarind Hospital, Khon Kaen University, 40002 Khon Kaen, Thailand
Aroonwadee Chanawong [email protected] Ratree Tavichakorntrakool [email protected] Jureerut Daduang [email protected]
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