Feasibility of Loop-Mediated Isothermal Amplification for Rapid Detection of Methicillin-Susceptible and Methicillin-Resistant Staphylococcus aureus in Tissue Samples

Abstract

Background: To date, few studies have investigated the feasibility of the loop-mediated isothermal amplification (LAMP) assay for identifying pathogens in tissue samples. This study aimed to investigate the feasibility of LAMP for the rapid detection of methicillin-susceptible or methicillin-resistant Staphylococcus aureus (MSSA or MRSA) in tissue samples, using a bead-beating DNA extraction method. Methods: Twenty tissue samples infected with either MSSA (n = 10) or MRSA (n = 10) were obtained from patients who underwent orthopedic surgery for suspected musculoskeletal infection between December 2019 and September 2020. DNA was extract-ed from the infected tissue samples using the bead-beating method. A multiplex LAMP assay was conducted to identify MSSA and MRSA infections. To recognize the Staphylococcus genus, S. aureus, and methicillin resistance, 3 sets of 6 primers for the 16S ribosomal ribonucleic acid (rRNA) and the femA and mecA genes were used, respectively. The limit of detection and sensitivity (de-tection rate) of the LAMP assay for diagnosing MSSA and MRSA infection were analyzed. Results: The LAMP result was positive for samples containing 103 colony-forming unit (CFU)/mL for 16S rRNA, 104 CFU/mL for femA, and 105 CFU/mL for mecA. The limits of detection for 16S rRNA and femA were not different between MSSA and MRSA. For the 10 MSSA-positive samples, the LAMP assay showed 100% positive reactions for 16S rRNA and femA and a 100% negative reaction for mecA. For the 10 MRSA-positive samples, the LAMP assay showed 100% positive reactions for 16S rRNA and mecA but only 90% positive reactions for femA. The sensitivity (detection rate) of the LAMP assay for identifying MSSA and MRSA in infected tissue samples was 100% and 90%, respectively. Conclusions: The results of this study suggest that the LAMP assay performed with tissue DNA samples can be a useful diagnostic method for the rapid detection of musculoskeletal infections caused by MSSA and MRSA.