1887

Abstract

The need for a microbial identification of independent of culture methods has resulted in the introduction of other laboratory principles. The verification of a proper and exclusive gene for the detection of the pneumococcus by nucleic acid-based tests is, however, still unresolved. A previously published -gene-specific real-time PCR method was applied to a panel of bacterial strains to clarify the analytical sensitivity and specificity of a PCR assay targeting this gene. Furthermore, a phylogenetic analysis of published gene sequences was performed to look at gene sequence differences and the theoretical match with the primers and probes. The -gene-specific PCR detected 46/46 isolates. All 49 of the non-pneumococcal isolates tested negative, including 22 isolates from the group streptococci. Phylogenetic analysis of 94 sequences of the gene from different strains of , and showed that 70/87 sequences constituted one cluster and a further six sequences were outside but adjacent to this cluster, all with a complete match with primers and probes. The remaining 11 strains could be placed in a different cluster, which also contained the five and two strains. All strains had no match with primers and probes. The strains in the second cluster were all characterized by being bile-insoluble, an infrequent pneumococcal phenotype. Routine laboratories can utilize the additional observation that pneumococci that were negative by the specific PCR also carried the phenotype of bile insolubility, thereby observing the incidence of false-negative results produced by the PCR assay. The real-time PCR targeting the gene thus constitutes a sensitive and specific assay that distinguishes from its close relatives in the group.

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2012-04-01
2019-12-14
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