Rapid detection of Enterococcus and vancomycin resistance using recombinase polymerase amplification

Pimchanok Panpru; Arpasiri Srisrattakarn; Nuttanun Panthasri; Patcharaporn Tippayawat; Aroonwadee Chanawong; Ratree Tavichakorntrakool; Jureerut Daduang; Lumyai Wonglakorn; Aroonlug Lulitanond

doi:10.7717/peerj.12561

Rapid detection of Enterococcus and vancomycin resistance using recombinase polymerase amplification

PeerJ. 2021 Dec 7:9:e12561. doi: 10.7717/peerj.12561. eCollection 2021.

Authors

Pimchanok Panpru¹, Arpasiri Srisrattakarn¹, Nuttanun Panthasri², Patcharaporn Tippayawat¹, Aroonwadee Chanawong¹, Ratree Tavichakorntrakool¹, Jureerut Daduang¹, Lumyai Wonglakorn³, Aroonlug Lulitanond¹

Affiliations

¹ Centre for Research and Development of Medical Diagnostic Laboratories, Faculty of Associated Medical Sciences, Khon Kaen University, Khon Kaen, Thailand.
² The 8th Regional Medical Science Center, Udon Thani, Thailand.
³ Clinical Microbiology Unit, Srinagarind Hospital, Khon Kaen University, Khon Kaen, Thailand.

Abstract

Vancomycin-resistant enterococci (VRE), especially Enterococcus faecium, have been a global concern, often causing serious healthcare-associated infections. We established a rapid approach for detecting E. faecium and vancomycin-resistance genes (vanA and vanB) in clinical samples using isothermal recombinase polymerase amplification (RPA) combined with a lateral-flow (LF) strip. Specific RPA primer sets and probes for ddl (to identify the presence of E. faecium) vanA and vanB genes were designed. The RPA reaction was performed under isothermal condition at 37 °C within 20 min and read using the LF strip within a further 5 min. A total of 141 positive blood-cultures and 136 stool/rectal swab samples were tested using RPA-LF method compared to the conventional PCR method. The RPA-LF method exhibited 100% sensitivity in both blood-culture (60 E. faecium; 35 vanA type and two vanB type) and stool/rectal-swab samples (63 E. faecium and 36 vanA type) without cross-reaction (100% specificity). The lower detection limit of the RPA-LF was approximately 10 times better than that of the conventional PCR method. The RPA-LF method is an alternative rapid method with excellent sensitivity and specificity for detecting E. faecium, vanA, and vanB, and it has the potential to be used as a point-of-care device for VRE therapy and prevention.

Keywords: Enterococcus faecium; Lateral flow strips; Recombinase polymerase amplification; VanA; VanB; Vancomycin-resistant enterococci.

Grants and funding

This study was supported by the KKU Research Fund (Project no. I62-00-19-03). Pimchanok Panpru was awarded by Graduate School, Khon Kaen University Research Grant (Grant no. 611JH107). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.