Development of a semi-nested reverse transcription polymerase chain reaction assay for the retrospective diagnosis of canine distemper virus infection.

Formalin-fixed, paraffin-embedded tissue blocks from 14 dogs were used to test the utility of a newly developed semi-nested reverse transcription polymerase chain reaction (RT-PCR) assay for canine distemper virus (CDV). The results from this new test were compared with those of histopathologic examination, fluorescent antibody detection (FA), and immunohistochemistry (IHC). The semi-nested RT-PCR protocol was used to detect CDV RNA in 9 of the 10 cases that were positive by at least 1 of the immunologic (FA and IHC) methods. Sequence data indicated that the amplified strains of CDV are more closely related to a naturally occurring strain than to a vaccine strain. Thus, the semi-nested RT-PCR assay is a useful diagnostic method applicable to the retrospective diagnosis of CDV infection. Sequence determination may yield molecular epidemiologic information regarding vaccine efficacy.

Retrospective differentiation of canine distemper virus and phocine distemper virus in phocids.

Formalin-fixed paraffin-embedded tissues from one Caspian seal (Phoca caspica), one harp seal (Phoca groenlandica), one hooded seal (Cystophora cristata), and one harbor seal (Phoca vitulina vitulina) were used to compare the utility of immunohistochemistry (IHC) versus that of a novel seminested reverse transcriptase polymerase chain reaction (RT-PCR) to detect and differentiate canine distemper virus (CDV) and phocine distemper virus (PDV). Four antibodies made against PDV were able to detect both viruses. Two antibodies made against cetacean morbillivirus (CMV) did not label antigens from either CDV or PDV. A third anti-CMV antibody inconsistently stained CDV antigens but did not label PDV antigens. The seminested RT-PCR was able to detect RNA of the phosphoprotein gene in all positive cases. Nucleotide sequence analyses of seminested RT-PCR products were used to differentiate CDV RNA from PDV RNA. From these data, it was determined that IHC using antibodies generated against PDV provided a rapid means of detection for both CDV and PDV antigens; however, differentiation between CDV and PDV was achieved only with the RT-PCR assay.