Nowadays, vaccines are broadly used to prevent porcine circovirus type 2 (PCV2) infection-induced expenditures, but the virus is still spreading among pigs. The current PCV2 vaccines all rely on the immunogenicity of Cap, yet our previous studies found that Cap is also the major component mediating the PCV2 infection-induced immune suppression through its interaction with host gC1qR. Thereby, new vaccines are still necessary for PCV2 prevention and control. In this study, we constructed a new PCV2 DNA vaccine expressing the gC1qR binding site mutant Cap. We introduced the Intron A and WPRE elements into the vector to improve the Cap expression level, and fused the IL-2 secretory signal peptides to the N-terminal of Cap to mediate the secretion of Cap. We also screened and selected chemokines CXCL12, CCL22, and CCL25 to migrate dendritic cells. In addition, we contained the vectors with PEI and then ultrasonic them into nano size to enhance the entrance of the vectors. Finally, the animal experiments showed that the new PCV2 DNA vaccine expressing the gC1qR binding site mutant Cap could induce stronger humoral and cellular immune responses than the PCV2 DNA vaccine expressing the wild-type Cap and the non-ultrasonic treated PCV2 DNA vaccine in mice, and protect the mice from PCV2 infection and lung lesions. The results indicate the new PCV2 DNA vaccine expressing the gC1qR binding site mutant Cap has a certain development value, and provide new insight into the development of novel PCV2 vaccines.

Porcine circovirus 2 (PCV2) has been proved to increase the risk of other pathogens infection via immunosuppression. Although the co-infection of PCV2 and porcine parvovirus (PPV) is commonly observed in worldwide, the relative immune mechanisms promoting PPV infection in PCV2-infected piglets are currently unknown. Herein, we found that PCV2 infection suppressed IFN-β expression and promoted PPV infection in the piglets. Consistent with this finding, we confirmed that PCV2 infection significantly inhibited the induction of IFN-β to promote PPV replication in cell level. Furthermore, PCV2 infection attenuated the K63-linked ubiquitination of STING induced by PPV, blocked the formation of complex of STING, TBK1 and IRF3, and further prevented the phosphorylation of TBK1 and IRF3, resulting in a decreased IFN-β transcription response to PPV infection. Consistently, using cGAMP to direct stimulate STING also appeared a reduced STING-K63 ubiquitination and IFN-β induction in PCV2-infected cells. However, we noted that knockdown of p38-MAPK signaling could markedly attenuate the inhibitory effect of PCV2 on STING-K63 ubiquitination, and improve the induction of IFN-β in PCV2-infected whenever theses cells were challenged with PPV infection or cGAMP stimulation. Meanwhile, we found that PCV2 infection promoted the phosphorylation of USP21 to inhibit the K63 ubiquitination of STING and the transcription of IFN-β via activation of p38-MAPK signaling. Taken together, our results demonstrate that PCV2 infection activates the p38-MAPK signaling pathway-mediated USP21 phosphorylation to inhibit the K63 ubiquitination of STING, which prevents the phosphorylation and transportation to the nucleus of IRF3, leading to an increase risk for PPV infection.

猪自然细胞毒受体(pNCRs)介导的NK细胞异常激活与多种猪病的发生密切相关.本研究通过原核表达获得了pNCR1、pNCR2和pNCR3胞外域可溶性重组目的 蛋白opNCR1、opNCR2和opNCR3,将重组目的 蛋白纯化后分别免疫新西兰白兔,获得抗opNCR1、opNCR2和opNCR3多克隆抗体,ELISA测定效价分别为1∶25600,1∶12800和1∶25600.分离猪血液中的NK细胞,利用制备的兔抗opNCR1、opNCR2和opNCR3多克隆抗体,通过Western-blotting和流式细胞术均能特异性检测到猪NK细胞内或膜上pNCR1、pNCR2和pNCR3的表达.本研究获得的兔抗opNCR1、opNCR2和opNCR3多克隆抗体将为进一步研究pNCRs在病原致病过程中作用机制提供材料.

Effects of different concentrations of Pb2+ (0-15 000 mg · kg-1) on seed germination of four tree species (Pinus tabuli formis,Platycladus orientalis,Koelreuteria paniculata,Robinia pseudoacacia) were investigated.The results showed that low concentration lead stress stimulated the seed germination of P.tabuliformis,P,orientalis and R.pseudoacacia,but the stimulation effect was not significant.With the increase of lead concentration,the promotion effect was gradually changed into the inhibition effect,and the higher the lead concentration,the greater the inhibition effect,then reached a significant level.The seed germination of K.paniculata was inhibited under any lead concentration stress.The seed germination of R.pseudoacacia was completely inhibited under 11 000 mg · kg-1 lead stress.The seed germination inhibition rate to P.tabuliformis and P.orientalis were sequentially reached 100％ under 13 000 mg· kg-1 and 15 000 mg · kg-1 lead stress,and the seed of P.orientalis could still germinate under 15 000 mg · kg-1 lead stress.Comprehensive analysis of the seed germination indexes of four tree species indicated that the effect of lead stress on the seed vigor index was the largest.The foregoing analysis showed that P.orientalis and P.tabuliformis had strong resistance to lead stress,and K.paniculata and R.pseudoacacia were more sensitive to lead stress.So P.orientalis and P.tabuliformis were suitable for the vegetation restoration on lead contaminated soil.

为探讨土壤重金属污染酶指示的可行性,通过对加入外源铅的填土进行控水培养,研究填土中脲酶和碱性磷酸酶活性的变化及其与土壤铅含量、土壤含水量和理化性质之间的关系.结果表明,土壤相对含水量为80％、60％时,土壤中的脲酶和碱性磷酸酶活性均增加;脲酶和碱性磷酸酶活性随铅含量增加而呈下降趋势,当铅含量为2000 mg/kg时,2种酶活性分别仅为对照的43.7％、61.9％.通径分析表明:各因子对脲酶活性的直接作用顺序为:速效钾＞有机质＞铅＞全氮＞速效磷＞pH＞水分,对碱性磷酸酶活性的直接作用顺序为:铅＞有机质＞全氮＞水分＞pH＞速效钾＞速效磷.土壤速效钾对脲酶活性的直接影响较大,铅、有机质是影响碱性磷酸酶活性的主要因素.2种酶活性变化是铅、水分和土壤理化性质共同作用的结果,且碱性磷酸酶活性较脲酶活性更易受影响.

Objective of this paper was to study changes in growth and antioxidant enzyme activity of Platycladus orientalis seedlings under drought and Pb stress. A potting experiment was conducted with four soil water regimes (40%, 60%,80%, 100% of soil relative water content) and five Pb concentrations(0, 300, 500, 1 000, 2 000 mg·kg-1). The results showed that drought stress restrained growth and antioxidase activities of P. orientalis seedlings, and the effects were more significant with the drought aggravated. The distribution pattern of biomass changed under drought stress, manifested as root> stem> leaf. Pb promoted the growth of P. orientalis seedlings within 2 000 mg·kg-1. The drought stress imposed with 60%, 80% water regimes to P. orientalis seedlings was alleviated within 2 000 mg·kg-1 Pb. Growth index and antioxidase activities of P. orientalis seedlings under treatments with 1 000 mg·kg-1 Pb and 80% of soil relative water content were higher than control, however growth index under other treatments were lower than control. With the soil relative water content decreasing, MDA concentrations increased continually, but with Pb concentration increasing, MDA concentrations increased first, then decreased, and reached the maxmum at 2 000 mg·kg-1. When Pb concentration was 1 000 mg·kg-1, regardless of the water content, MDA content was significantly lower than control. In summary, the resistances of P. orientalis to drought stress could be strengthened under proper drought and Pb interaction.

Through simulative methods,the influences of water and lead on the soil urease,glucose,phosphatase activity of soil were studied.A certain extent water stress had a remarkably activated effect on soil urease,glucose,phosphatase activity.Glucose was the most sensitive to the change of soil water content.The effect of lead on glucose showed activation at low concentration and inhibition at high concentration,while the inhibiting effects on urease and phosphatase increased with the aggravation of lead.The inhibition rate achieved 56.31%,53.21% and 38.11% respectively when 2000 mg·kg-1lead concentration.There existed generally mutual action between soil water content and lead.Under mild water stress,the joint of water and lead stress had antagonism to urease;while had synergism at low lead concentration and had antagonism when lead concentration was higher than 1000 mg·kg-1 to glucose;and had synergism to phosphatase as lead concentration was 300 mg·kg-1.Light water stress interacting with lead demonstrated synergism to urease,glucose,antagonism to phosphatase.Under sever water stress,net varying amounts(ΔU) was lower than zero only at 2000 mg·kg-1 of lead concentration.The joint stress of water and lead induced the greatest ΔU of soil enzyme were 88.4%,50.7% and 25.5% of control.

The effects of heavy metal Pb and drought stresses on the antioxidant enzyme activities and malondialdehyde concentration of Robinia pseudoacacia and Amorpha fruticosa in one-year old seedlings were studied with pot experiments for the selection of tree species with strong resistance.The results showed that lead stress at higher concentration(1 000~3 000 mg·kg-1) and at lower(300~500 mg·kg-1) could improve activity,could improve SOD and POD activities,respectively.Both water and lead stresses could improve antioxidant enzyme activities of the leaves.Based on a comprehensive analysis,the resistance of A.fruticosa was better than R.pseudoacacia.Antistress abilities to Pb were in the order of A.fruticosa seedlingA.fruticosa cuttingR.pseudoacacia seedling.

【Objective】 The study was conducted with the purpose of comparing the tolerance of lead to two common drought-enduring trees in northwest China,Robinia pseudoacacia and Amorpha fruticosa,and provided a basis for phytoremediation of soil contaminated by lead in arid and semi-arid regions.【Method】 One-year old seedlings were planted in containers under 6 different lead treatments(soil Pb content 0,300,500,1 000,2 000,3 000 mg/kg,respectively) and 4 water treatments(relative soil water content 100%,80%,60%,40%,respectively).The contents of organic osmotic adjustment substances in the leaves were determined under water,Pb stress and water and Pb interaction stress.【Result】 With the increase of stress level,the changes of Pro,soluble sugar(SS) and soluble protein(SP) content of the two types of trees were complicated.The Pro content in the leaves of R.pseudoacaciawere under water stress was larger than under Pb stress,and when the relative soil water content was 40%,at the same soil Pb content level,the Pro contents in the leaves of R.pseudoacacia were significantly higher than under other relative soil water contents.Under Pb or water stress or interaction,at the same soil Pb content(except for 3 000 mg/kg),the SS contents in the leaves of R.pseudoacacia under different water stress treatments had no significant difference(P0.05).Under water or Pb stress,as the stress level increased,the SP contents in the leaves of R.pseudoacacia did not decrease,so did A.fruticosa.【Conclusion】 The contribution of Pro to raise the resistance to water was stronger than to Pb.R.pseudoacacia's accumulation ability of Pro to raise the resistance to water and Pb stress was stronger than A.fruticosa',vice vercer.A.fruticosa's production of SS and SP to raise the resistance to water and Pb stress was stronger than R.pseudoacacia'.