Genomic Analysis And Effect On IPEC-J2 Cells Of Porcine Bocavirus Strain NP08

Porcine bocavirus(PBo V) is a single-stranded DNA virus firstly identified in the lymph node of swine with a background of post-weaning multisystemic wasting syndrome in 2009. PBo V is presented at a high prevalence in either health or diseased swine herds around the world, and may be an important cause for the re-outbreak of piglet diarrheal. So it is significant to conduct the research on PBo V. The initial study of epidemiology, genome and effect on IPEC-J2 of PBo V had been conducted to explore the potential influence caused by PBo V at the level of genes and cells. The results were as follows: 1. The detection of PBo V by PCRThe samples collected from diarrheal pigs during Mar 2013 to Dec 2014 were detected by PCR for PBo V. A total of 134 samples were PBo V positive in 523 samples and the positive rate of PBo V was 25.62%. The positive rate in sera(76%) and feces(49%) were significantly higher than that in tissue(5%). The samples collected from healthy pigs in a slaughterhouse showed a 47.28% positive rate and the virus was detected at a higher rate in feces(68%) than in the mesenteric lymph node(36%) and submandibular lymph node(2%). 2. Full-length genome analysis of PBo V/NP08 strainUsing the DNA template extracted from a PBo V-positive sample and six specific primer pairs, a continuous sequence of approximately 4745 bp of PBo V was obtained and the nearly full-length genome of strain NP08 was submitted to Gen Bank under accession number KR014255. Sequence analysis showed that its nucleotides shared 99%, 99% and 90% homologous to the strain PBo V/Buk-1, PBo V/SX and PBo V/H18, respectively. The genome consists of three open reading frames(ORFs) encoding four virus proteins NS1, NP1, VP1 and VP2. 3. Genetic grouping of PBo V and analysis of putative phospholipase A2(PLA2) domain located at VP1 unique region(VP1u)According to the criteria characterize the bocavirus by International Committee on Taxonomy of Viruses(ICTV), a total of 47 PBo V strains were classified into six Groups(GroupⅠ- GroupⅥ), and the strain PBo V/NP08 belonged to the Group Ⅴ. It was firstly observed that the conserved HDXXY and YXGXF motifs of PLA2 were found only in the VP1 u sequence of PBo V belong to GroupⅠ-Ⅳ, but not in that of PBo V belong to GroupⅤ-Ⅵ. 4. The morphological change and dynamic change of cytokines exhibited by IPEC-J2 inoculated with PBo V/NP08An evident cytopathic effect(CPE) such as swelling, stacking and defulvium could be found in IPEC-J2 at 48 h after inoculated with PBo V/NP08. The expression of IL-1β, IL-6, IL-8, IL-12, TNF-α and IFN-γ were up-regulated during 12-72 h after exposure, especially IL-6 which showed a significantly increase. Meanwhile, the expression of TGF-β1 was also significantly up-regulated during infection.