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Porcine reproductive and respiratory syndrome (PRRS), an economically significant pandemic disease, commonly results in increased impact of bacterial infections, including those by Streptococcus suis (S. suis). In recent years, PRRS virus (PRRSV) NADC30-like strain has emerged in different regions of China, and coinfected with S. suis and PRRSV has also gradually increased in clinical performance. However, the mechanisms involved in host innate responses towards S. suis and their implications of coinfection with NADC30-like strain remain unknown. Therefore, the pathogenicity of NADC30-like strain and S. suis serotype 2 (SS2) coinfection in vivo and in vitro was investigated in this study. The results showed that NADC30-like increased the invasion and proliferation of SS2 in blood and tissues, resulting in more severe pneumonia, myocarditis, and peritonitisas well as higher mortality rate in pigs. In vitro, NADC30-like strain increased the invasion and survival of SS2 in porcine alveolar macrophages (PAM) cells, causing more drastic expression of inflammatory cytokines and activation of NF-ĸB signalling. These results pave the way for understanding the interaction of S. suis with the swine immune system and their modulation in a viral coinfection., (© 2018 Blackwell Verlag GmbH.)

Porcine reproductive and respiratory syndrome is an infectious disease that causes serious economic losses to the swine industry worldwide. To better understand the pathogenesis of the porcine reproductive and respiratory syndrome virus (PRRSV), three PRRSV strains with different molecular markers and virulence were used to infect MARC-145 cells. A total of 1804 proteins were identified, and 233 altered proteins and 72 signaling pathways involved in the proteomic profiling of virus-infected MARC-145 cells increased with the virulence of the PRRSV strain. The three types of viral strains shared a common pathway-the electron transport reaction in mitochondria-in the infected-MARC-145 cells. Moreover, the antisense pathway was the most variable of all significant signaling pathways for the highly virulent SX-1 strain, indicating that this unique pathway may be connected to the high virulence of the SX-1 strain. Our study is the first attempt to provide a proteome profile of MARC-145 cells infected with PRRSV strains with different virulence, and these findings will facilitate a deep understanding of the interactions between this virus and its host.

Epidemiological investigations were conducted on recently emerging porcine reproductive and respiratory syndrome virus (PRRSV) strains in Shandong province in 2014-2015. The proportion of the NADC30 strain identified by ORF7 sequence alignment has been gradually increasing. Three emerging PRRSV strains were successfully isolated, and the complete genomic sequences were determined. Our results indicate the importance of recombinant strains in Shandong province, China. There was a varied degree of recombination of two or three strains (classical, HP-PRRSV and/or NADC30). Moreover, the recombination strains affected the pathogenicity of newly emerged strains.

Porcine reproductive and respiratory syndrome (PRRS), a highly contagious disease, has been constantly causing huge economic losses all over the world. PRRS virus (PRRSV) infection results in immunosuppression and IL-10 up-regulation. The relationship between them is still in dispute. Previous studies demonstrated the protein of PRRSV nucleocapsid (N) protein is able to up-regulate IL-10, yet the underlying molecular mechanisms remain unknown. In this study, the expression kinetics of IL-10 up-regulation induced by PRRSV N protein were analyzed in immortalized porcine alveolar macrophages (PAMs). N protein induced IL-10 expression in a time- and dose-dependent manner. Inhibition experiments of signaling pathways suggested NF-κB and p38 MAPK pathways are both involved in N protein-induced IL-10 up-regulation. Besides, the integrity of N protein is essential for significant IL-10 up-regulation. This research is beneficial for further understanding of the interplay between PRRSV and host immune system., (Copyright © 2017. Published by Elsevier Ltd.)

The nonstructural protein 1α (NSP1α) of porcine reproductive and respiratory syndrome virus (PRRSV) is a nucleo-cytoplasmic protein that suppresses the production of type I interferon (IFN). In this study, we investigated the relationship between the subcellular distribution of NSP1α and its inhibition of type I IFN. NSP1α was found to contain the classical nuclear export signal (NES) and NSP1α nuclear export was CRM-1-mediated. NSP1α was shuttling between the nucleus and cytoplasm. We also showed that the nuclear export of NSP1α was necessary for its ability for type I IFN inhibition. NSP1α was also found to interact with CBP, which implies a possible mechanism of CBP degradation by NSP1α. Taken together, our results describe a novel mechanism of PRRSV NSP1α for type I IFN inhibition and suppression of the host innate antiviral response., (Copyright © 2016. Published by Elsevier Inc.)

PRRS is one of the most important diseases in swine industry. Current PRRS inactivated vaccine provides only a limited protection and cannot induce sufficient cell-mediated immune responses. In this study, we first found that the mRNA and protein levels of Th1-type cytokines (IFN-γ, IL-12) and Th2-type cytokines (IL-6, IL-10) were significantly increased through TRIF/MyD88-NF-κB signaling pathway when porcine peripheral blood monocyte-derived dendritic cells (MoDCs) were treated with poly (I: C) of TLR3 ligand and imiquimod of TLR7 ligand, along with inactivated PRRSV antigen. Meanwhile, the ability of catching PRRSV antigen was also significantly enhanced. In mice experiment, it was found that the PRRSV-specific T lymphocyte proliferation, the percentages of CD4(+), CD8(+) T lymphocytes and PRRSV-specific CD3(+) T cells producing IFN-γ and IL-4, the levels of Th1- and Th2-type cytokines and the titers of neutralization antibody were significantly enhanced in poly (I: C), imiquimod along with inactivated PRRSV group. Taken together, results of our experiments described for the first time that synergy of TLR3 and 7 ligands could significantly enhance the function of DCs to present inactivated PRRSV antigen through TRIF/MyD88-NF-κB signaling pathway and be used as adjuvant candidate for the development of novel PRRS inactivated vaccine.

To improve the efficacy of porcine reproductive and respiratory syndrome virus (PRRSV) subunit vaccine, the immunological enhancement effects of four natural adjuvants were compared, including Astragalus extract, Astragalus/Bacillus production, propolis, and Freund's adjuvant. The results showed that titers of IgG and neutralizing antibody against highly pathogenic PRRSV (HP-PRRSV) glycoprotein 5 (GP5) from the groups of Astragalus/Bacillus adjuvant and Freund's adjuvant co-administered with a recombinant HP-PRRSV GP5 subunit were the highest among all the experimental groups. In cellular immunity, as shown in T lymphocyte proliferation level, Astragalus/Bacillus adjuvant and Freund's adjuvant exhibited the most potent immunological enhancement effects. Exceptionally, pigs inoculated with Freund's adjuvant developed severely delayed humoral immune responses within 28 days post inoculation. The adjuvants of Astragalus extract and propolis also exhibited immunological enhancement effects on humoral immune and cell-mediated immune responses, but which were obviously lower than Astragalus/Bacillus adjuvant and Freund's adjuvant. Following challenge with HP-PRRSV, pigs inoculated with Astragalus/Bacillus adjuvant and Freund's adjuvant co-administered with GP5 subunit showed lower viremia, slighter clinical signs, and less pathological lung lesions compared with the groups inoculated using Astragalus extract and propolis. In conclusion, from immune responses and immunological protection against HP-PRRSV, the Astragalus/Bacillus adjuvant demonstrated the most potent immunological enhancement effect on HP-PRRSV GP5 subunit. Astragalus/Bacillus adjuvant showed promise as a candidate immune adjuvant, to more efficiently prevent and control PRRS.

Highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) had caused catastrophic losses in swine industry in China. The current inactivated vaccine provided only limited protection, and the attenuated live vaccine could protect piglets against the HP-PRRSV but there was a possibility that the attenuated virus returned to high virulence. In this study, the eukaryotic expression vector pVAX1© was modified under the control of rabbit β-globin intron II gene and the modified vector pMVAX1© was constructed. Porcine interleukin-2 (IL-2) and GP3-GP5 fusion protein of HP-PRRSV strain SD-JN were highly expressed by pMVAX1©. Mice inoculated with pMVAX1©-GP35 developed significantly higher PRRSV-specific antibody responses and T cell proliferation than those vaccinated with pVAX1©-GP35. pMVAX1©-GP35 was selected as PRRS DNA vaccine candidate and co-administrated with pVAX1©-IL-2 or pMVAX1©-IL-2 in pigs. pMVAX1©-IL-2+pMVAX1©-GP35 could provide enhanced PRRSV-specific antibody responses, T cell proliferation, Th1-type and Th2-type cytokine responses and CTL responses than pMVAX1©-GP35 and pVAX1©-IL-2+pMVAX1©-GP35. Following homologous challenge with HP-PRRSV strain SD-JN, similar with attenuated PRRS vaccine group, pigs inoculated with pMVAX1©-IL-2+pMVAX1©-GP35 showed no clinical signs, almost no lung lesions and no viremia, as compared to those in pMVAX1©-GP35 and pVAX1©-IL-2+pMVAX1©-GP35 groups. It indicated that pMVAX1©-IL-2 effectively increases humoral and cell mediated immune responses of pMVAX1©-GP35. Co-administration of pMVAX1©-IL-2 and pMVAX1©-GP35 might be attractive candidate vaccines for preventing HP-PRRSV infections.

Porcine reproductive and respiratory syndrome (PRRS) has become one of the most economically important diseases to the global pork industry. Currently, the efficacies of available commercial vaccines remain questionable: the modified live-PRRSV vaccines (MLVs) were generally effective but variable in sufficient protection, and the outcomes of inactivated-PRRSV vaccines (IVs) in the field were not very promising. In the present study, we investigated the effect of swine interleukin 4 (IL-4) on the development of virus-specific immune responses elicited by an MLV. The antibody titer against PRRSV membrane proteins in pigs elicited by MLV plus recombinant plasmid encoding IL-4 (group 3) was significantly higher than those elicited by MLV alone (group 1) and MLV plus empty plasmid (group 2) from 35 days post-inoculation (dpi). Similarly, the neutralizing efficacy of sera from group 3 was markedly enhanced compared with group 1 and group 2. In cellular immunity, the ratio of CD3⁺CD4⁺/CD3⁺CD8⁺ T lymphocyte subpopulations from group 3 monitored by flow cytometry (FCM) was significantly higher than those from group 1 and group 2 from 42 dpi to 21 days post-challenge (dpc). After viral challenge, pigs in group 3 showed significantly lower virus loads in peripheral blood measured by a real-time quantitative PCR (RT-qPCR), as compared with those in group 1 and group 2. Pigs in group 1 and group 2 had a low fever and displayed mild inappetence, lethargy, rough hair coats, and no lung lesions, while those in group 3 showed almost no clinical signs, no lung lesions. The scores of clinical signs of pigs in group 3 were significantly lower than those in both group 1 and group 2. Interestingly, the scores of lung lesions showed no significant differences among the three groups. Our results indicate that swine IL-4 markedly enhanced the protective immune response of pigs and improved the efficacy of the MLV in preventing PRRS disease.

Porcine reproductive and respiratory syndrome virus (PRRSV) has become one of the most economically important diseases to the global pork industry. Current vaccination strategies only provide a limited protective efficacy. In this study, a DNA vaccine, pVAX1(©)-α-γ-GP35, co-expressing GP3 and GP5 of PRRSV with interferon α/γ was constructed, and its immediate and long-lasting protection against highly pathogenic PRRSV (HP-PRRSV) challenge were examined in pigs. For immediate protection, the results showed that pVAX1(©)-α-γ-GP35 could provide partially protective efficacy, which was similar to the pVAX1(©)-α-γ (expressing interferon α/γ). For long-lasting protection, pigs inoculated with pVAX1(©)-α-γ-GP35 developed significantly higher PRRSV-specific antibody response, T cell proliferation, IFN-γ, and IL-4, than those vaccinated with pVAX1(©)-GP35 (expressing GP3 and GP5 of PRRSV). Following homologous challenge with HP-PRRSV strain SD-JN, pigs inoculated with pVAX1(©)-α-γ-GP35 showed almost no clinical signs, no lung lesions, and significantly lower viremia, as compared to those in pVAX1(©)-GP35 group. It indicated that pVAX1(©)-α-γ-GP35 could induce enhanced immune responses and provide both immediate and long-lasting protection against HP-PRRSV challenge in pigs. The DNA vaccine pVAX1(©)-α-γ-GP35 might be an attractive candidate vaccine for the prevention and control of HP-PRRSV infections.

Porcine reproductive and respiratory syndrome (PRRS) has become one of the most economically important diseases to the global pork industry. The etiological agent is the PRRS virus (PRRSV). In the spring of 2006, a highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) appeared in China and caused heavy economic losses. Here we report the complete genomic sequence of a novel PRRSV variant with 174 amino acid deletions in the nonstructural protein 2 (nsp2) gene.

A serologic investigation of porcine reproductive and respiratory syndrome virus (PRRSV) in hybrid wild boar herds was conducted during 2008-2009. PRRSV isolates with novel genetic markers were recovered. Experimental infection of pigs indicated that hybrid wild boars are involved in the epidemiology of PRRSV.

This study was aimed at determining the effect of highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) on Haemophilus parasuis (HPS) in co-infection. A quantitative real-time PCR targeting infB gene, which is conserved among different HPS serotypes, was developed to improve the accuracy and speed of the detection of HPS. A total of 32 four-week-old conventional pigs were distributed randomly into four groups: pigs in group I were intranasally infected with HP-PRRSV first, and were then intraperitoneally inoculated with HPS on 5 days after HP-PRRSV infection; pigs in group II were intranasally inoculated with HP-PRRSV alone; pigs in group III were intraperitoneally inoculated with HPS alone; pigs in group IV were intraperitoneally inoculated with physiological saline. The amount of HPS in serum on 0, 3, 6, 9 and 12 days post-inoculation (dpi) with HPS were detected using the established quantitative real-time PCR. Clinical signs, pathological changes and histopathological lesions were observed. The amount of HPS in serum reached 10(6)copies/μl at 3 dpi with HPS in pigs of group I, while it arrived 10(5.7)copies/μl at 9 dpi with HPS in pigs of group III. The HPS loads in hearts and lungs were much higher than in other tissues. The study showed that HP-PRRSV was able to accelerate HPS infection and loads., (Copyright © 2012 Elsevier B.V. All rights reserved.)

We conducted a serologic investigation of porcine reproductive and respiratory syndrome virus (PRRSV) in hybrid wild boar herds in China during 2008-2009. PRRSV isolates with novel genetic markers were recovered. Experimental infection of pigs indicated that hybrid wild boars are involved in the epidemiology of PRRSV.

To establish a reverse-transcription loop-mediated isothermal amplification (RT-LAMP) method for rapid detection of porcine reproductive and respiratory syndrome virus (PRRSV), four primers specific to six regions of the N gene were designed. After amplification in an isothermal water bath for 1 h, samples containing PRRSV generated the expected ladder-like products while porcine parvovirus, porcine circovirus, classic swine fever virus, pseudorabies virus, and swine testis cells generated no product. The sensitivity and specificity of the RT-LAMP assay were evaluated by comparison with reverse-transcription polymerase chain reaction (RT-PCR) and real-time PCR. Because it is specific and simple, the RT-LAMP assay will be useful for the diagnosis of PRRSV infection.

A highly pathogenic swine disease designated as 'porcine high fever disease (PHFD)' appeared recently in China. Porcine reproductive and respiratory syndrome virus (PRRSV) was identified as an agent associated with PHFD, and two discontiguous sequence deletions were identified as a genetic marker in the Nsp2 region of the viral genome. To examine PHFD in Shandong province, a total of 10 PRRSV isolates were recovered from pig herds that had never been vaccinated for PRRS. Sequence analysis of open reading frame 5 (ORF5) showed that the level of identity among the 10 isolates ranged between 88.2 and 99.2%. For the non-structural protein 2 (Nsp2) gene, three isolates shared high sequence identity with VR-2332, the prototype virus of the North American genotype, while the remaining seven isolates exhibited two discontiguous sequence deletions that were identical to those of PHFD: a one-amino-acid (phenylalanine) deletion at position 482 and a 29-amino-acid deletion at positions 533-561 of Nsp2. Experimental infection of pigs with SD-JN, which was one of the seven isolates containing such deletions, resulted in severe clinical symptoms characterized by red discoloration on the body and hemorrhages in the lungs, kidneys, and inguinal lymph nodes, accompanied by higher mortality and longer duration of viremia. These symptoms were similar to those of PHFD observed in the field. Our results show that VR2332-like PRRSV coexists with PHFD-associated atypical PRRSV in pig herds in the Shandong area, and different PRRSV isolates differ greatly in their pathogenesis and virulence in pigs.