Usefulness of reverse transcriptase-polymerase chain reaction for detection of rabies RNA in archival samples

The examination of archival samples by molecular techniques has been found to be a valuable tool in providing retrospective and epidemiological data. In contrast to DNA, there are fewer comprehensive studies quantitatively addressing the feasibility of assessing RNA from archival samples. In tropical countries like India, such studies are even more rare. One problem is that the warm climate and the fluctuations in the ambient temperature during transport and storage of tissues lead to early decomposition of the tissues. The present study was carried out to evaluate the usefulness of reverse transcriptase-polymerase chain reaction (RT-PCR) in unfixed archival samples to assess whether a retrospective diagnosis of human rabies could be made from archival brain samples from patients suspected to have died of rabies. A nested RT-PCR was performed on 6 rabies confirmed and 8 rabies negative brain tissue samples. Rabies viral RNA could be detected in all 6 positive brain tissue samples and was absent in all mouse inoculation test negative samples. None of the control samples revealed cross-reaction with rabies primers, indicating its specificity. Our results demonstrate the importance of RT-PCR in the detection of rabies virus RNA in 5- to 6-year-old preserved samples without substantial loss. The study proves the role of retrospective diagnosis by RT-PCR in archival autopsy material and decomposed tissues and paves the path for further molecular epidemiologic and phylogenetic studies.