One pair of primers was designed based on the 3D gene sequences of porcine kobuvirus,and the two-step RT-PCR assay was established in this study.The positive templates of CSFV，PRRSV，JEV，SIV，PEDV，TGEV，GARV and recombinant plasmids (pMD18T-PEV3D,pMD18T-FMDV3D) which contain either FMDV’s or PEV’s 3D gene were used to test the specificity of the porcine kobuvirus RT-PCR assay we established and the assay showed excellent specificity.Sensitivity test showed that the minimal concentration of positive template the assay can detect is 180 fg/mL.The epidemiology of porcine kobuvirus in Hubei Province was then investigated by applying this assay.Among 165 samples of pigs with diarrhea,118 samples were kobuvirus positive.To investigate the distribution of porcine kobuvirus in different age groups in pig farm with diarrhea,the samples of different age groups from 4 different pig farms were collected.The results clearly indicated that porcine kobuvirus were centralized distributed in the pig group with diarrhea,which suggested that positive relationship exists between diarrhea disease outbreak and porcine kobuvirus.