K.L.Ravikumar
KIMS Hospital and Research Centre, India
Title: New, rapid PCRSeqTyping strategy to identify and serotype S.pneumoniae from culture and culture negative serum samples
Biography
Biography: K.L.Ravikumar
Abstract
Identification and continued monitoring of pneumococcal serotypes is needed to access vaccine efficacy, serotype replacement and to identify emerging new types. Presently available serological methods are expensive, labor intensive and prone to misidentification, while current molecular methods have limited serotype coverage, multistep and time consuming. In PCRSeqTyping method two step amplification and sequencing of cpsB and adjoining genes within the pneumococcal capsular locus is implemented. The assay was compared with conventional blood culture, qmPCR and Quellung reaction for its specificity and sensitivity. ply, lytA, pspA and spn9802 primers were used in qmPCR. 1504 blood and serum samples from children clinically suspected of Invasive Pneumococcal Disease with raised CBC, CRP, PCT and abnormal chest X-ray were subjected to testing. Conventional blood culture and qmPCR were positive in 7% and 30% samples respectively. PCRSeqTyping assay identified 456 serum samples positive for S.pneumoniae showing 100% correlation with qmPCR. The assay detected serotype of 456 qmPCR positive serum samples which included 108 culture positives and 348 culture negatives. The results showed 100% correlation with quellung test. Our study reveals the usefulness of PCRSeqTyping for direct detection and subsequent serotyping of S.pneumoniae from culture negative serum samples. An added advantage of this assay was the precise identification of the homologous types and detection of 91 specific serotypes. This simple, efficient PCR sequence based method with total serotype coverage will improve the accuracy of pneumococcal surveillance, identify non typeable and vaccine escape strains.