The changes of immune-related molecules within the ileal mucosa of piglets infected with porcine circovirus type 2.

BACKGROUND: Enteritis is one of the most frequently reported symptoms in piglets infected with porcine circovirus type 2 (PCV2), but the immunopathogenesis has not been reported. OBJECTIVES: This study examined the effect of a PCV2 infection on the intestinal mucosal immune function through morphological observations and immune-related molecular detection. METHODS: Morphological changes within the ileum of piglets during a PCV2 infection were observed. The expression of the related-molecules was analyzed using a gene chip. The immunocyte subsets were analyzed by flow cytometry. The secretory immunoglobulin A (SIgA) content was analyzed by enzyme-linked immunosorbent assay. RESULTS: The PCV2 infection caused ileal villus damage, intestinal epithelial cells exfoliation, and an increase in lymphocytes in the lamina propria at 21 days post-infection. Differentially expressed genes occurred in the defense response, inflammatory response, and the complement and coagulation cascade reactions. Most of them were downregulated significantly at the induction site and upregulated at the effector site. The genes associated with SIgA production were downregulated significantly at the induction site. In contrast, the expression of the Toll-like receptor-related genes was upregulated significantly at the effector site. The frequencies of dendritic cells, B cells, and CD8⁺T cells were upregulated at the 2 sites. The SIgA content decreased significantly in the ileal mucosa. CONCLUSIONS: PCV2 infections can cause damage to the ileum that is associated with changes in immune-related gene expression, immune-related cell subsets, and SIgA production. These findings elucidated the molecular changes in the ileum after a PCV2 infection from the perspective of intestinal mucosal immunity, which provides insights into a further study for PCV2-induced enteritis.

Porcine circovirus type 2 upregulates endothelial-derived IL-8 production in porcine iliac artery endothelial cells via the RIG-I/MDA-5/MAVS/JNK signaling pathway.

BACKGROUND: Dysfunction of endothelial cells and vascular system is one of the most important pathological changes of porcine circovirus disease (PCVD) caused by porcine circovirus type 2 (PCV2). PCV2-infected endothelial cells can upregulate the production of endothelial-derived IL-8, which can inhibit the maturation of dendritic cells. Endothelial-derived IL-8 has different structural and biological characteristics compared with monocyte-derived IL-8. However, the mechanism of endothelial-derived IL-8 production is still unclear. RESULTS: Key molecules of RIG-I-like signaling pathway RIG-I, MDA-5, MAVS and a key molecule of JNK signaling pathway c-Jun in PCV2-infected porcine iliac artery endothelial cells (PIECs) were upregulated significantly detected with quantitative PCR, Western blot and fluorescence confocal microscopy, while no significant changes were found in NF-κB signaling pathway. Meanwhile, the expression of endothelial-derived IL-8 was downregulated after RIG-I, MDA-5, or MAVS genes in PIECs were knocked down and PIECs were treated by JNK inhibitor. CONCLUSIONS: PCV2 can activate RIG-I/MDA-5/MAVS/JNK signaling pathway to induce the production of endothelial-derived IL-8 in PIECs, which provides an insight into the further study of endothelial dysfunction and vascular system disorder caused by PCV2.

Complete genome analysis of a novel recombinant isolate of potato virus Y from China.

The complete sequence of GF_YL20, a potato virus Y (PVY) isolate from China, encodes a polyprotein of 3,061 amino acids. Sequence analysis indicates that GF_YL20 has a genomic structure different from previously reported PVY strains. It shares 99 % nucleotide sequence identity with PB209 (PVY(N:O)) except in VPg, but more than 97 % nucleotide sequence identity with the VPg of Mont (PVY(N)), PB312 (PVY(NTN)) and HN2 (SYR-I). Phylogenetic analysis indicates that GF_YL20 is a novel N:O recombinant with three recombination breakpoints.

Sequence variation of P1 gene in Potato virus Y isolated from Fujian Province.

To understand the sequence variation and the putative protein structure of P1 gene in Potato virus Y (PVY) and to identify the sources of the variation, P1 gene in PVY isolated from Fujian Province was amplified by reverse-transcription polymerase chain reaction (RT-PCR) using a pair of degenerate primers designed from the conserved regions of published sequences. Sequence variation and putative protein structure were analyzed, and phylogenetic tree was reconstructed using Bayesian inference method. Expected fragments of 915 bp in size were amplified from 12 samples collected from Fujian Province by RT-PCR. The 12 sequences shared 73%-99% nucleotide identity with the reference sequences from GenBank. A strong recombination signal was identified at position 309 in sequences of isolates QK44, XT02, XT08 and LH05. Among the 12 sequences, 85 amino acid variants were detected, indicating high sequence variation in the P1 protein. However, positions 41-275 in the protein were highly conserved, especially in three active sites (H192, D201 and V235). Phylogenetic analysis grouped the sequences into three clades, each with different Coiled-coil domains and 3D-structures, suggesting divergent phylogenetic relationship among the groups. The above results show P1 gene in PVY is highly variable but contains 3 conserved active sites (H192, D201, V235) and the high genetic variation in the gene is primarily due to mutation and recombination.

[Sequence variation and protein structure of pipo gene in Potato virus Y].

The objectives of this study were to understand the sequence variation and the putative protein structure of pipo gene in the Potato virus Y (PVY) collected from Solanum tuberosum. The pipo gene in PVY was cloned using a pair of degenerate primers designed from its conserved region and its sequences were used to re-construct phylogenetic tree in Potyvirus genera by a Bayesian inference method. An expected fragment of 235 bp was amplified in all 20 samples by RT-PCR and the pipo genes in the 20 samples assayed shared more than 92% nucleotide sequence similarity with the published sequences of PVY strains. Among the 20 pipo gene sequences, 13 polymorphic sites were detected, including 4 parsimony informative sites and 9 singleton variable sites. These results indicate that PVY pipo gene is highly conserved but some sequence variations exist. Further analyses suggest that the pipo gene encodes a hydrophilic protein without signal peptide and transmembrane region. The protein has theoretical isoelectric points (pI) ranging from 11.26 to 11.62 and contains three highly conserved regions, especially between aa 10 and 59. The protein is likely located in the mitochondria and has a-helix secondary structure. Bayesian inference of phylogenetic trees reveals that PVY isolates are clustered in the same branch with high posterior probability, while Sunflower chlorotic mottle virus (SoCMoV) and Pepper severe mosaic virus (PepSMV) are closely related, consisting with the classification of Potyvirus genera using other approaches. Our analyses suggest that the pipo gene can be a new marker for phylogenetic analysis of the genera. The results reported in this paper provide useful insights in the genetic variation and the evolution of PVY and can stimulate further research on structure and function of the PIPO protein.