Development of a Loop-mediated Isothermal Amplification Assay for Rapid Detection of Streptococcus pneumoniae Isolates in Clinical Sputum Samples

Abstract

Streptococcus pneumoniae is one of the leading causes of invasive infections. Traditional culture and identification methods require a long time period and are liable to be complicated by normal flora. In this study, a new method was developed based on loop-mediated isothermal amplification. A series of primers for autolysin (LytA) from S. pneumoniae were designed with a bioinformatics technique, and a loop-mediated isothermal amplification system for the detection of S. pneumoniae was set up. Comparative analysis of specificity and sensitivity from the culture and polymerase chain reaction methods in 50 samples collected from patients with respiratory infections was performed. The data demonstrated that the selected primers and reaction system in the loop-mediated isothermal amplification were respondent to Streptococcus pneumoniae and did not cross-react with other bacteria. The detection of sensitivity to the loop-mediated isothermal amplification method was 102 CFU/ml. From all of the clinical specimens tested, 9 cases (18.0 %) were confirmed to be positive for Streptococcus pneumoniae using the loop-mediated isothermal amplification method, whereas only 4 cases (8.0 %) were diagnosed using the culture method. Loop-mediated isothermal amplification technology is a specific, simple and accurate method for the efficient differentiation of S. pneumoniae in clinical specimens.