Comparison of different fluorescent quantitation polymerase chain reaction for genes detection of Streptococcus pneumonia / 中国医师进修杂志

ABSTRACT

Objective To compare the sensitivity and specificity of different fluorescent quantitation polymerase chain reaction(PCR) for genes detection of Streptococcus pneumonia.Methods By designing five sets of fluorescent quantitation PCR primers targeting different genes of Streptococcus pneumonia (lytA,ply,spn9802,psaA,cspA),the sensitivity and specificity of fluorescent quantitation PCR was determined through detecting 37 strains of Streptococcus pneumonia and 28 strains of non-Streptococcus pneumonia,as well as 80 sputum specimens from suspected pneumonia cases.Results These five primers had excellent sensitivity with 90％ amplification efficiency through analysis of the standard curve.The change of Ct value was less than 0.5,indicating that these five primers have good stability.By detecting 37 strains of Streptococcus pneumonia and 28 strains of non-Streptococcus pneumonia,it showed that the specificity of lytA,ply and spn9802 was better than the other two primers (positive rate and negative rate was all 100％).But the detection limit of lytA,ply was one order of magnitude than spn9802 (101 CFU/ml vs.102 CFU/ml).The specificity of psaA and cspA was worse [negative rate of psaA was 89％ (25/28),positive rate of cspA was 97％ (36/37)].By detecting 80 sputum specimens from suspected pneumonia cases,it showed that the specificity of fluorescent quantitation PCR were better than the bacterial culture.The specificity of lytA and ply was the best two [positive rate92.50％(74/80),90.00％(72/80)],and next was spn9802 [86.25％(69/80)],the specificity ofpsaA and cspA was the worst two for detection [76.25％(61/80) and 78.75％(63/80)].Conclusions The sensitivity and specificity of fluorescent quantitation PCR genes used in clinic for detecting Streptococcus pneumonia is different.LytA and ply are the best primers in specificity,next is spn9802,and psaA and cspA are the worst primers for in specificity detection.