Otimização e avaliação da técnica da RT -PCR para o diagnóstico Ante e Post Mortem da infecção pelo vírus da cinomose canina

A reverse transcription-polymerase chain reaction (RT-PCR) assay was used to detect the nucleoprotein gene of canine distemper virus (CDV) ante and post mortem in dogs. In optimization of the technique we compared four RNA extraction methods and two (specific and random hexamer primer) protocols in reverse transcription for CDV detection in body fluids and tissues. The best results were obtained with silica / guanidine isothiocyanate for RNA extraction and performing reverse transcription with specific sense primer. CDV RNA was amplified from dogs with signs of distemper in leukocytes (18/18) and plasma (15/18). RT-PCR was positive in cerebrospinal fluid (CSF) from others 12 (65%) dogs with distemper encephalopathies (n=20). We also amplified CDV RNA from urinary bladder, brain, cerebellum, and lungs of dogs that died of distemper. CDV RNA was not obtained in seven control dogs. The RT-PCR standardized here is simple and specific to diagnose distemper ante and post mortem in dogs. Leukocytes, and to a lesser degree plasma, are the preferred samples to diagnose dogs during the epithelial or systemic phase of the disease, whereas CSF appears to be the best sample to ante mortem diagnose distemper encephalitis.