Detection of Streptococcus pyogenes using rapid visual molecular assay

• Published in: FEMS microbiology letters 2015-09, Vol.362 (18), p.fnv148-fnv148
• Main Authors: Zhao, Xiangna, He, Xiaoming, Li, Huan, Zhao, Jiangtao, Huang, Simo, Liu, Wei, Wei, Xiao, Ding, Yiwei, Wang, Zhaoyan, Zou, Dayang, Wang, Xuesong, Dong, Derong, Yang, Zhan, Yan, Xiabei, Huang, Liuyu, Du, Shuangkui, Yuan, Jing
• Format: Article
• Language: English
• Subjects: Assaying; Calcein; Deoxyribonucleic acid; DNA; DNA Primers; DNA, Bacterial - analysis; Fluoresceins; Humans; Limit of Detection; Microbiology; Nucleic Acid Amplification Techniques - methods; Pathogens; Primers; Sensitivity and Specificity; Streptococcal Infections - diagnosis; Streptococcus infections; Streptococcus pyogenes; Streptococcus pyogenes - genetics; Streptococcus pyogenes - isolation & purification; Target detection; Target recognition; Turbidity
• ISSN: 0378-1097; 1574-6968; 1574-6968
• DOI: 10.1093/femsle/fnv148

Abstract Streptococcus pyogenes is an increasingly important pathogen in many parts of the world. Rapid and accurate detection of S. pyogenes aids in the control of the infection. In this study, a loop-mediated isothermal amplification (LAMP) assay was developed and validated for the specific detection of S. pyogenes. The assay incorporates two methods: a chromogenic analysis using a calcein/Mn2+ complex and real-time turbidity monitoring to assess the reaction. Both methods detected the target DNA within 60 min under 64°C isothermal conditions. The assay used specifically designed primers to target spy1258, and correctly identified 111 strains of S. pyogenes and 32 non-S. pyogenes strains, including other species of the genus Streptococcus. Tests using reference strains showed that the LAMP assay was highly specific. The sensitivity of the assay, with a detection limit of 1.49 pg DNA, was 10-fold greater than that of PCR. The LAMP assay established in this study is simple, fast and sensitive, and does not rely upon any special equipment; thus, it could be employed in clinical diagnosis. A rapid and visual loop-mediated isothermal amplification assay with good specificity and sensitivity was developed and validated for the specific detection of Streptococcus pyogenes. Graphical Abstract Figure. A rapid and visual loop-mediated isothermal amplification assay with good specificity and sensitivity was developed and validated for the specific detection of Streptococcus pyogenes.