Real-time reverse transcriptase-polymerase chain reaction assay with modified probe for the diagnosis of rabies viruses and other lyssaviruses

摘要

A real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay that utilizes multiplex primers and probes with degenerate nucleotides to detect divergent species of lyssavirus is described. The probes used in the RT-PCR assay target a highly conserved region at the 5′ end of the lyssavirus genome and are modified with either a minor groove binder (MGB) or locked nucleic acid (LNA) nucleotides to increase their melting temperature. The described assay detects all known lyssavirus species with a sensitivity and specificity superior to traditional hemi-nested PCR and the direct fluorescent antibody (DFA) test.