DEVELOPMENT OF A POLYMERASE CHAIN-REACTION ASSAY TO DETECT THE PRESENCE OF STREPTOCOCCUS-PNEUMONIAE DNA

In this study, we have developed a chemically sensitive and specific p olymerase chain reaction (PCR) assay to detect the presence of Strepto coccus pneumoniae genomic DNA. The target DNA sequence was a 322-base pair segment of the S. pneumoniae DNA polymerase I gene (pol I). PCR p roducts of pure cultures of a set of pneumococcal serotypes commonly a ssociated with human infection could be amplified in water and in bloo d cultures of clinical isolates containing S. pneumoniae. We were able to detect 2 fg of purified S. pneumoniae DNA. There were no false-pos itive reactions when the assay was performed on samples containing the following clinically encountered bacteria: Haemophilus influenzae typ e B, Neisseria meningitidis, Escherichia coli, Klebsiella pneumoniae, Pseudomonas spp. nontypeable H. influenzae, Staphylococcus aureus, coa gulase-negative staphylococci, and Streptococcus pyogenes. The additio n of EDTA and citrate-anticoagulated whole blood to the PCR reaction m ixture inhibited the PCR assay, whereas the addition of lithium hepari n, sodium heparin, and sodium polyanetholesulfonate-anticoagulated who le blood to PCR reaction mixture did not interfere with the ability to detect the presence of S. pneumoniae DNA.