Lu Xia

Porcine epidemic diarrhea virus (PEDV) is a highly virulent enterovirus that is responsible for severe diarrhea in neonatal piglets. For no effective vacci

Background/Objectives: Streptococcus suis (SS), an important zoonotic pathogen, has caused significant economic losses to the global pig industry. Existing commercial vaccines for SS mainly provide effective protection against a single serotype. Due to the existence of many serotypes and their robust immune evasion capabilities, the development of multi-component subunit vaccines or multi-epitope vaccines that provide effective cross-protection against different strains of SS is a key focus of current research. Methods: We applied two-dimensional electrophoresis (2-DE) and immunoblotting to screen for candidate immunogens among the immunogenic cell wall proteins of SS. BALB/c mice were immunized intradermally with a multi-component, multi-epitope vaccine. The vaccine’s safety and immunogenicity were assessed via clinical monitoring, antibody titer detection, cytokine assays, and survival curve analyses. Results: In this study, eight immunogenic cell wall proteins (GH25, Pk, PdhA, Ldh, ExoA, Pgk, MalX, and Dnak) were successfully identified using MALDI-TOF-MS, all of which could induce high IgG antibody titers. Based on the conservation and immunoprotection demonstrated by these eight protective antigenic proteins, PdhA, Ldh, and MalX were screened to construct a multi-component subunit vaccine as a candidate vaccine for providing cross-protection against SS isolates of multiple serotypes. Challenge studies showed that mice immunized with the multi-component subunit vaccine (PdhA, Ldh, and MalX) were protected against challenges with the SS2 virulent strain ZY05719 (62.5% protection) and the SSChz virulent strain CZ130302 (75% protection). Subsequently, we utilized immunoinformatics techniques to design a novel multi-epitope vaccine (MVPLM) derived from the immunogenic proteins PdhA, Ldh, and MalX. However, challenge tests revealed that the MVPLM offered limited protection against SS. Conclusions: These data demonstrate that a multi-component subunit vaccine composed of PdhA, Ldh, and MalX proteins shows promise as a candidate universal vaccine against multiple SS serotypes.

Porcine parvovirus (PPV) is one of the most important pathogens that cause reproductive failure in pigs. However, the pathogenesis of PPV infection remains unclear. Proteomics is a powerful tool to understand the interaction between virus and host cells. In the present study, we analyzed the proteomics of PPV-infected PK-15 cells. A total of 32 and 345 proteins were differentially expressed at the early and replication stages, respectively. Subsequent gene ontology annotation and Kyoto Encyclopedia of Genes and Genomes enrichment analysis showed these differentially expressed proteins were significantly enriched in pathways including toll-like receptor signaling pathway, tumor necrosis factor signaling pathway, and viral carcinogenesis. The expression of poly (rC) binding protein 1 (PCBP1) was observed to decrease after PPV infection. Overexpressed or silenced PCBP1 expression inhibited or promoted PPV infection. Our studies established a foundation for further exploration of the multiplication mechanism of PPV. IMPORTANCE: Porcine parvovirus (PPV) is a cause of reproductive failure in the swine industry. Our knowledge of PPV remains limited, and there is no effective treatment for PPV infection. Proteomics of PPV-infected PK-15 cells was conducted to identify differentially expressed proteins at 6 hours post-infection (hpi) and 36 hpi. Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis showed that various pathways participate in PPV infection. Poly (rC) binding protein 1 was confirmed to inhibit PPV replication, which provided potential targets for anti-PPV infection. Our findings improve the understanding of PPV infection and pave the way for future research in this area.

Porcine sapelovirus (PSV) is an emerging swine enteric virus that can cause various disorders including acute diarrhea, respiratory distress, reproductive failure, and polioencephalomyelitis in pigs. In this study, we isolated a PSV strain HNHB-01 from a clinical porcine deltacoronavirus- (PDCoV-) positive intestinal content of a diarrheic piglet. PSV was first identified using the small RNA deep sequencing and assembly, and further identified by the electron microscopic observation and the immunofluorescence assay. Subsequently, this virus was serially passaged in swine testis (ST) cells, and the complete genomics of PSV HNHB-01 passage 5 (P5), P30, P60, and P100 were sequenced and analyzed. 9 nucleotide mutations and 7 amino acid changes occurred in the PSV HNHB-01 P100 strain when compared with the PSV HNHB-01 P5. Pathogenicity investigation showed that orally inoculation of PSV HNHB-01 P30 could cause obvious clinical symptoms and had broad tissue tropism in 5-day-old piglets. Epidemiological investigation revealed that PSV infections and the coinfections of diarrhea coronaviruses were highly prevalent in swine herds. The complete genomes of 8 representative PSV epidemic strains were sequenced and analyzed. Phylogenetic analysis revealed that the PSV epidemic strains were closely related to other PSV reference strains that located in the Chinese clade. Furthermore, recombination analysis revealed that the recombination events were occurred in downstream of the 2C region in our sequenced PSV HNNY-02/CHN/2018 strain. Our results provided theoretical basis for future research studies of the pathogenic mechanism, evolutionary characteristics, and the development of vaccines against PSV.

Porcine parvovirus 1 (PPV1) mainly induces severe reproductive failure in pregnant swine, and causes huge economic losses to the swine industry. Cell apoptosis induced by PPV1 infection has been identified the major cause of reproductive failure. However, the molecular mechanism was not fully elucidated. In this study, the potential mechanism of PPV1 induced apoptosis in PK-15 cells was investigated. Our results showed that PPV1 induced apoptosis in PK-15 cells. Further studies revealed toll-like receptor 2 (TLR2) was involved in the PPV1-mediated apoptosis. TLR2 siRNA significantly decreased the apoptosis. Finally, our study showed NF-κB was activated by TLR2 during PPV1-induced apoptosis. The activation of NF-κB signaling was demonstrated by the phosphorylation of p65, p65 nuclear translocation and degradation of inhibitor of kappa B α (IκBα). Together, these results provided evidence that the recognition between PPV1 and PK-15 cells was mainly through TLR2, and then induction of the NF-κB signaling pathway activation, which further induces apoptosis. Our study could provide information to understand the molecular mechanisms of PPV1 infection.

Porcine deltacoronavirus (PDCoV) mainly causes severe diarrhea and intestinal pathological damage in piglets and poses a serious threat to pig farms. Currently, no effective reagents or vaccines are available to control PDCoV infection. Single-chain fragment variable (scFv) antibodies can effectively inhibit virus infection and may be a potential therapeutic reagent for PDCoV treatment. In this study, a porcine phage display antibody library from the peripheral blood lymphocytes of piglets infected with PDCoV was constructed and used to select PDCoV-specific scFv. The library was screened with four rounds of biopanning using the PDCoV N protein, and the colony with the highest affinity to the PDCoV N protein was obtained (namely, N53). Then, the N53-scFv gene fragment was cloned into plasmid pFUSE-hIgG-Fc2 and expressed in HEK-293T cells. The scFv-Fc antibody N53 (namely, scFv N53) was purified using Protein A-sepharose. The reactive activity of the purified antibody with the PDCoV N protein was confirmed by indirect enzyme-linked immunosorbent assay (ELISA), western blot and indirect immunofluorescence assay (IFA). Finally, the antigenic epitopes that the scFv N53 recognized were identified by a series of truncated PDCoV N proteins. The amino acid residues ⁸²GELPPNDTPATTRVT⁹⁶ of the PDCoV N protein were verified as the minimal epitope that can be recognized by the scFv-Fc antibody N53. In addition, the interaction between the scFv-Fc antibody N53 and the PDCoV N protein was further analyzed by molecule docking. In conclusion, our research provides some references for the treatment and prevention of PDCoV.

Porcine parvovirus (PPV) mainly induces severe reproductive failure in pregnant swine, and causes huge economic losses to the swine industry. Cell apoptosis induced by PPV infection has been identified the major cause of reproductive failure. However, the molecular mechanism was not fully elucidated. In this study, the potential mechanism of PPV induced apoptosis in PK-15 cells was investigated. Our results showed that PPV induced apoptosis in PK-15 cells. Further studies revealed that PPV induces apoptosis by activating the nuclear factor-kappa B (NF-κB) signaling. The activation of NF-κB signaling was demonstrated by the phosphorylation of p65, p65 nuclear translocation and degradation of inhibitor of kappa B α (IκBα). Finally, our studies provided evidence that toll-like receptor 2 (TLR2) was involved in the PPV-mediated apoptosis and activation of NF-κB signaling. The expression of TLR2 were increased after PPV infection. TLR2 siRNA significantly decreased the apoptosis and activation of NF-κB after PPV infection. Together, these results provided evidence that the recognition between PPV and PK-15 cells was mainly through TLR2, and then induction of the NF-κB signaling pathway activation, which further induces apoptosis. Our study could provide information to understand the molecular mechanisms of PPV infection.

Zhang Y, et al. J Vet Sci. 2019 Oct;21:e12. https://doi.org/10.4142/jvs.2020.21.e12

Transmissible gastroenteritis virus (TGEV) is a coronavirus that causes severe diarrhea in suckling piglets. TGEV primarily targets and infects porcine intestinal epithelial cells, which play an important role in nutrient absorption. However, the effects of TGEV infection on nutrient absorption in swine have not yet been investigated. In this study, we evaluated the impact of TGEV infection on arginine uptake using the porcine small intestinal epithelial cell line IPEC-J2 as a model system. High performance liquid chromatography (HPLC) analyses showed that TGEV infection leads to reduced arginine uptake at 48 hours post-infection (hpi). Expression of cationic amino acid transporter 1 (CAT-1) was attenuated as well. TGEV infection induced activation of phospho-protein kinase C α (p-PKC α), phospho-epidermal growth factor receptor (p-EGFR), and enhanced the expression of caveolin-1, all of which appear to be involved in down-regulating arginine uptake and CAT-1 expression. These results illuminate the relationship between TGEV infection and nutrient absorption, and further our understanding of the mechanisms of TGEV infection.