Detection of porcine rotavirus by polymerase chain reaction

Rotavirus is an intestinal pathogen that causes gastroenteritis in children and young animals. This pathogen causes significant economic losses in livestock production. Currently, there are no specific drugs for treatment of rotavirus infection, so treatment options for this disease remain limited. Therefore, development of a vaccine against porcine rotavirus infection remains relevant and in demand. A method for detecting and identifying porcine rotavirus based on modern molecular genetic methods, such as RT-PCR, PCR-RFLP, sequencing, and others, is also necessary for genetic control of the vaccine being developed. Pathological material (samples of lungs, spleen and intestines) collected from dead and slaughtered piglets aged 10-12 days from farms in the Republics of Bashkortostan and Tatarstan with high viral disease rates was used for the study. SPEV and RK-15 continuous animal cell lines were used to isolate the rotavirus pathogen. A search for the most specific genetic loci of porcine rotavirus revealed that the VP4, VP6 and VP7 genes are the most specific, and they can be used to detect and identify the pathogen under study. Specific primer combinations with RT-PCR programs were developed for each of these loci. BLASTn analysis confirmed the specificity of developed primers to the porcine rotavirus genome. Using viral isolates adapted to SPEV and RK-15 cells, the performance of developed primer combinations and RT-PCR programs for each identified genetic locus of the porcine rotavirus genome was demonstrated. A positive result was obtained in the analysis of 12 antigen samples (from the lungs, intestines and spleen). An algorithm for the detection and identification of porcine rotavirus using RT-PCR and PCR-RFLP was developed.