作者机构：
[Wang, Naidong; Wang, Dongliang; Mai, Jinhui; Yang, Yi] Hunan Agr Univ, Hunan Prov Key Lab Prot Engn Anim Vaccines, Lab Funct Prote LFP, Res Ctr Reverse Vaccinol RCRV,Coll Vet Med, Changsha 410128, Peoples R China.
通讯机构：
[Wang, Naidong] H;Hunan Agr Univ, Hunan Prov Key Lab Prot Engn Anim Vaccines, Lab Funct Prote LFP, Res Ctr Reverse Vaccinol RCRV,Coll Vet Med, Changsha 410128, Peoples R China.
关键词：
PPV7;Cap;B cell and T cell epitopes;vaccine;evolution
摘要：
Porcine parvovirus 7 (PPV7) belonging to the genus Chapparvovirus in the family Parvoviridae, has been identified in the USA, Sweden, Poland, China, South Korea and Brazil. Our objective was to determine the phylogeny, estimate the time of origin and evolutionary dynamics of PPV7, and use computer-based immune-informatics to assess potential epitopes of its Cap, the main antigenic viral protein, for vaccines or serology. Regarding evolutionary dynamics, PPV7 had 2 major clades, both of which possibly had a common ancestor in 2004. Furthermore, PPV7 strains from China were the most likely ancestral strains. The nucleotide substitution rates of NS1 and Cap genes were 8.01 × 10−4 and 2.19 × 10−3 per site per year, respectively, which were higher than those reported for PPV1-4. The antigenic profiles of PPV7 Cap were revealed and there were indications that PPV7 used antigenic shift to escape from the host’s immune surveillance. Linear B cell epitopes and CD8 T cell epitopes of Cap with good antigenic potential were identified in silico; these conserved B cell epitopes may be candidates for the PPV7 vaccine or for the development of serological diagnostic methods.
作者机构：
[Zhan, Yang; Wang, Dongliang; Zhou, Wenfeng; Wan, Naidong; Yu, Wanting; Mai, Jinhui; Yang, Yi] Hunan Agr Univ, Coll Vet Med, Res Ctr Reverse Vaccinol RCRV, Hunan Prov Key Lab Prot Engn Anim Vaccines,Lab Fu, Changsha 410128, Peoples R China.;[Yu, Wanting] Peking Univ, Biodynam Opt Imaging Ctr BIOPIC, Sch Life Sci, State Key Lab Membrane Biol, Beijing 100871, Peoples R China.;[Epstein, Neal D.] NHLBI, Cell & Dev Biol Ctr, NIH, Bldg 10, Bethesda, MD 20892 USA.
通讯机构：
[Yang, Yi] H;[Epstein, Neal D.] N;Hunan Agr Univ, Coll Vet Med, Res Ctr Reverse Vaccinol RCRV, Hunan Prov Key Lab Prot Engn Anim Vaccines,Lab Fu, Changsha 410128, Peoples R China.;NHLBI, Cell & Dev Biol Ctr, NIH, Bldg 10, Bethesda, MD 20892 USA.
关键词：
SARS-CoV-2;S protein;B-cell and T-cell epitopes;vaccine
摘要：
Currently, there is limited knowledge about the immunological profiles of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). We used computer-based immunoinformatic analysis and the newly resolved 3-dimensional (3D) structures of the SARS-CoV-2 S trimeric protein, together with analyses of the immunogenic profiles of SARS-CoV, to anticipate potential B-cell and T-cell epitopes of the SARS-CoV-2 S protein for vaccine design, particularly for peptide-driven vaccine design and serological diagnosis. Nine conserved linear B-cell epitopes and multiple discontinuous B-cell epitopes composed of 69 residues on the surface of the SARS-CoV-2 trimeric S protein were predicted to be highly antigenic. We found that the SARS-CoV-2 S protein has a different antigenic profile than that of the SARS-CoV S protein due to the variations in their primary and 3D structures. Importantly, SARS-CoV-2 may exploit an immune evasion mechanism through two point mutations in the critical and conserved linear neutralization epitope (overlap with fusion peptide) around a sparsely glycosylated area. These mutations lead to a significant decrease in the antigenicity of this epitope in the SARS-CoV-2 S protein. In addition, 62 T-cell epitopes in the SARS-CoV-2 S protein were predicted in our study. The structure-based immunoinformatic analysis for the SARS-CoV-2 S protein in this study may improve vaccine design, diagnosis, and immunotherapy against the pandemic of COVID-19.
关键词：
Antiviral activity;PK15 cells;Porcine circovirus 2;Tyrosine kinase inhibitor STI-571 and PP2;Virus like particles
摘要：
Porcine circovirus type 2 (PCV2) causes huge economic losses in the global swine industry and has a complex and poorly understood virus-host interaction mechanism. We reported that the C-terminal of the capsid protein of all PCV2 isolates shared a strictly conserved PXXP motif that may interact with SH3 domain-containing tyrosine kinases; however, its roles in PCV2 cell entry and replication remain unknown. In this study, we determined that mRNA levels of two SH3 domain-containing tyrosine kinases family (Abl and Src) had distinct profiles (wild-type and PXXP-mutated) during PCV2 infections of PK15 cells. Therefore, we hypothesized that activities of tyrosine kinases (Abl and Fyn) in PK15 cells may be hijacked by PCV2 via its PXXP motif of the Cap, to favor virus replication. Specific inhibitors PP2 of Lck/Fyn and STI-571 of Abl family kinases decreased viral production through suppression of DNA and Cap synthesis at the replication stage. However, based on indirect immunofluorescence assay (IFA), entry of PCV2 virus-like particles (VLPs) into PK15 cells was not altered. Elucidating mechanisms of PCV2-host interactions should provide new insights for development of new compounds to prevent or reduce PCV2 infections.
摘要：
Porcine circovirus type 3 (PCV3) has recently been isolated from diseased pigs within the USA. The objective was to detect the presence of PCV3 in dogs. Nested polymerase chain reactions (PCR) with PCV3-specific primers for the capsid gene were used to detect PCV3 genomic DNA in serum samples from dogs (n = 44) in China. There was PCV3 DNA detected in 4 of 44 dogs [all were negative for PCV2 and canine circovirus (CanineCV)]. Based on sequence analysis, positive sequences were grouped into PCV3 genotypes. However, these isolates had close evolutionary relationships with FoxCV (KP941114) and CanineCV (JQ821392). Further investigations of the epidemiology, evolutionary biology, and pathobiology of PCV3 to dogs are warranted.
摘要：
Targeted integration of exogenous genes into so-called safe harbors/friend sites, offers the advantages of expressing normal levels of target genes and preventing potentially adverse effects on endogenous genes. However, the ideal genomic loci for this purpose remain limited. Additionally, due to the inherent and unresolved issues with the current genome editing tools, traditional embryonic stem (ES) cell-based targeted transgenesis technology is still preferred in practical applications. Here, we report that a high and repeatable homologous recombination (HR) frequency (>95%) is achieved when an approximate 6kb DNA sequence flanking the MYH9 gene exon 2 site is used to create the homology arms for the knockout/knock-in of diverse nonmuscle myosin II (NM II) isoforms in mouse ES cells. The easily obtained ES clones greatly facilitated the generation of multiple NM II genetic replacement mouse models, as characterized previously. Further investigation demonstrated that though the targeted integration site for exogenous genes is shifted to MYH9 intron 2 (about 500bp downstream exon 2), the high HR efficiency and the endogenous MYH9 gene integrity are not only preserved, but the expected expression of the inserted gene(s) is observed in a pre-designed set of experiments conducted in mouse ES cells. Importantly, we confirmed that the expression and normal function of the endogenous MYH9 gene is not affected by the insertion of the exogenous gene in these cases. Therefore, these findings suggest that like the commonly used ROSA26 site, the MYH9 gene locus may be considered a new safe harbor for high-efficiency targeted transgenesis and for biomedical applications.
摘要：
The ability of liver to respond to changes in nutrient availability is essential for the maintenance of metabolic homeostasis. Autophagy encompasses mechanisms of cell survival, including capturing, degrading, and recycling of intracellular proteins and organelles in lysosomes. During negative nutrient status, autophagy provides substrates to sustain cellular metabolism and hence, tissue function. Severe negative energy balance in dairy cows is associated with fatty liver. The aim of this study was to investigate the hepatic autophagy status in dairy cows with severe fatty liver and to deter- mine associations with biomarkers of liver function and inflammation. Liver and blood samples were collected from multiparous cows diagnosed as clinically healthy (n = 15) or with severe fatty liver (n = 15) at 3 to 9 d in milk. Liver tissue was biopsied by needle puncture, and serum samples were collected on 3 consecutive days via jugular venipuncture. Concentrations of free fatty acids and beta-hydroxybutyrate were greater in cows with severe fatty liver. Milk production, dry matter intake, and concentration of glucose were all lower in cows with severe fatty liver. Activities of serum aspartate aminotransferase, alanine aminotransferase, glutamate dehydrogenase, and gamma-glutamyl transferase were all greater in cows with severe fatty liver. Serum concentrations of haptoglobin and serum amyloid A were also markedly greater in cows with severe fatty liver. The mRNA expression of autophagosome formation-related gene ULK1 was lower in the liver of dairy cows with severe fatty liver. However, the expression of other autophagosome formation-related genes, beclin 1 (BECN1), phosphatidylinositol 3-kinase catalytic subunit type 3 (P1K3C3), autophagy-related gene (ATG) 3, ATG5, and ATG12, did not differ. More important, ubiquitinated proteins, protein expression of sequestosome-1 (SQSTM1, also called p62), and microtubule-associated protein 1 light chain 3 (MAP1LC3, also called LC3)-II was greater in cows with severe fatty liver. Transmission electron microscopy revealed an increased number of autophagosomes in the liver of dairy cows with severe fatty liver. Taken together, these results indicate that excessive lipid infiltration of the liver impairs autophagic activity that may lead to cellular damage and inflammation.
摘要：
A newly emerging porcine circovirus, designated PCV3, has been reported in various countries (USA, Poland, South Korea and China) since 2017. Its presence may be associated with porcine dermatitis and nephropathy syndrome (PDNS), reproductive failure, and multisystem inflammation. In this study, we report identification of PCV3 in cases of reproductive failure in various regions in Hunan, China. From January 2015 to December 2016, sera were collected from 190 sows from seven farms with reproductive problems. Specifically, 85 samples were from sows with a history of reproductive failure, whereas the remaining 105 were from healthy sows. The PCV3-positive rate was significantly higher in sows with reproductive failure (45.9%) than in healthy sows (21.9%), based on quantitative PCR (qPCR) assays. Although phylogenetic analysis based on the cap gene suggested that these PCV3 isolates belonged to the clade PCV3a, amino acid sequence variations in the Cap protein still occurred among these isolates, and these might have contributed to antigenic alterations of the Cap protein, based on the Jameson-Wolf antigenic index. Finally, we concluded that PCV3 was circulating in sows in Hunan province, China. However, the association of PCV3 with reproductive failure in sows and its potential for vertical transmission need to be studied further.
通讯机构：
[Yang, Yi] H;[Jiang, Ping] N;Hunan Agr Univ, LFP, Coll Vet Med, Changsha 410128, Hunan, Peoples R China.;Hunan Agr Univ, RCRV, Coll Vet Med, Changsha 410128, Hunan, Peoples R China.;Nanjing Agr Univ, Key Lab Anim Dis Diagnost & Immunol, Minist Agr, Coll Vet Med, Nanjing 210095, Jiangsu, Peoples R China.
关键词：
Porcine circovirus type 2 (PCV2);Virus-like particles (VLPs);Osmolytes;Stability
摘要：
Although porcine circovirus type 2 (PCV2) virus-like particles (VLPs) have been successfully harvested from various protein expression systems, conditions to promote their stability and integrity during long-term storage have not been well defined since only the intact VLPs, instead of the monomeric capsid protein (Cap), can induce neutralizing antibodies in pigs in previous studies. In this study, freshly prepared PCV2 VLPs were stored in several media (various concentrations of NaCl, sorbitol, sucrose and trehalose) at three temperatures (4 °C, −20 °C and −80 °C) and their stability and integration were evaluated after 7 month. Addition of 15% trehalose in storage buffer promoted long-term preservation of PCV2 VLPs. In contrast, storage buffer with 5% osmolytes (sucrose, trehalose and sorbitol) did not confer stabilization for long-term storage. These refined storage conditions for stabilization of PCV2 VLPs should enhance their use in vaccines.
期刊：
Archives of Virology,2017年162(7):2015-2020 ISSN：0304-8608
通讯作者：
Wang, Aibing
作者机构：
[Hu, Yi; Liu, Tanbin; Liu, Wei; Wang, Aibing; Lei, Hongyu] Hunan Agr Univ, Coll Vet Med, Res Ctr Reverse Vaccinol RCRV, Lab Anim Models & Funct Genom LAMFG, Changsha 410128, Hunan, Peoples R China.;[Zhan, Yang; Xie, Xiaohong; Wang, Naidong; Wang, Dongliang; Deng, Zhibang; Yang, Yi] Hunan Agr Univ, Coll Vet Med, Res Ctr Reverse Vaccinol RCRV, Lab Funct Prote LFP, Changsha 410128, Hunan, Peoples R China.
通讯机构：
[Wang, Aibing] H;Hunan Agr Univ, Coll Vet Med, Res Ctr Reverse Vaccinol RCRV, Lab Anim Models & Funct Genom LAMFG, Changsha 410128, Hunan, Peoples R China.
关键词：
PCV2 Strain;Major Open Reading Frame;PCV2 Genome;PCV2 Genotype;Porcine Circoviruses
摘要：
Porcine circovirus type 2 (PCV2) is the causative pathogen of porcine circovirus-associated diseases (PCVAD). This virus evolves mostly through point mutations and genome recombination between different PCV2 genotypes (e.g. PCV2a and PCV2b), as has been confirmed in swine herds. In the current work, the complete PCV2 genome sequences of 69 clones derived from various tissues (lymph node, spleen and lung,) of an infected individual, were subjected to phylogenetic and alignment analyses. The results not only demonstrate co-infection with distinct PCV2b subtypes (e.g. 1B and 1C) in the same animal, but also highlight another mechanism of evolution - diverse point mutations acquired during immune evasion by this virus.
