Open AccessHemoculture and Direct Sputum Detection of mecA‐Mediated Methicillin‐Resistant Staphylococcus aureus by Loop‐Mediated Isothermal Amplification in Combination With a Lateral‐Flow Dipstick
Author(s) -
Nawattanapaiboon Kawin,
Prombun Photchanathorn,
Santanirand Pitak,
Vongsakulya Apirom,
Srikhirin Toemsak,
Sutapun Boonsong,
Kiatpathomchai Wansika
Publication year - 2016
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.21935
Subject(s) - loop mediated isothermal amplification , sputum , dipstick , microbiology and biotechnology , staphylococcus aureus , medicine , biology , bacteria , dna , tuberculosis , pathology , urine , genetics , endocrinology
This study reports loop‐mediated isothermal amplification (LAMP) for rapid detection of methicillin‐resistant Staphylococcus aureus from direct clinical specimens. Four primers including outer and inner primers were specifically designed on the two target sequences— fem B to identify S. aureus and mec A to identify antibiotic‐resistant gene. Reference strains including various species of gram‐positive/gram‐negative isolates were used to evaluate and optimize LAMP assays. The optimum LAMP condition was found at 63°C within 70 min assay time (include hybridization with FITC probe for 5 min and further 5 min for reading the results on the lateral flow dipstick). The detection limits of LAMP for mec A was 10 pg of total DNA or 100 CFU/ml. The LAMP assays were applied to a total of 155 samples of direct DNA extraction from sputum and hemoculture bottles. The sensitivity of LAMP for mec A detection in sputum and hemoculture bottles was 93.3% (28/30) and 100% (52/52), respectively. In conclusion, LAMP assay is an alternative technique for rapid detection of MRSA infection with a technical simplicity and cost‐effective method in a routine diagnostic laboratory.