Project description:Porcine reproductive and respiratory syndrome virus (PRRSV) continues to be a major problem to the pork industry worldwide. Increasing data indicate that PRRSV strains differ in virulence in infected pigs and are biologically, antigenically, and genetically heterogeneous. It is evident that the current vaccines, based on a single PRRSV strain, are not effective in protecting against infections with the genetically diverse field strains of PRRSV. The recent outbreaks of atypical or acute PRRS in vaccinated pigs have raised a serious concern about the efficacy of the current vaccines and provided the impetus for developing more effective vaccines. Special attention in this review is given to published work on antigenic, pathogenic and genetic variations of PRRSV and its potential implications for vaccine efficacy and development. Although there are ample data documenting the heterogeneous nature of PRRSV strains, information regarding how the heterogeneity is generated and what clinical impact it may have is very scarce. The observed heterogeneity will likely pose a major obstacle for effective prevention and control of PRRS. There remains an urgent need for fundamental research on this virus to understand the basic biology and the mechanism of heterogeneity and pathogenesis of PRRSV.
Project description:Porcine reproductive and respiratory syndrome virus is the cause of reproductive failure in sows and respiratory disease in young pigs, which has been considered as one of the most costly diseases to the worldwide pig industry for almost 30 years. This study used microarray-based transcriptomic analysis of PBMCs from experimentally infected pigs to explore the patterns of immune dysregulation after infection with two East European PRRSV strains from subtype 2 (BOR and ILI) in comparison to a Danish subtype 1 strain (DAN). Transcriptional profiles were determined at day 7 post infection in three tested groups of pigs and analysed in comparison with the expression profile of control group. Microarray analysis revealed differential regulation (>?1.5-fold change) of 4253 and 7335 genes in groups infected with BOR and ILI strains, respectively, and of 12518 genes in pigs infected with Danish strain. Subtype 2 PRRSV strains showed greater induction of many genes, especially those involved in innate immunity, such as interferon stimulated antiviral genes and inflammatory markers. Functional analysis of the microarray data revealed a significant up-regulation of genes involved in processes such as acute phase response, granulocyte and agranulocyte adhesion and diapedesis, as well as down-regulation of genes enrolled in pathways engaged in protein synthesis, cell division, as well as B and T cell signaling. This study provided an insight into the host response to three different PRRSV strains at a molecular level and demonstrated variability between strains of different pathogenicity level.
Project description:Porcine reproductive and respiratory syndrome virus (PRRSV) infection was diagnosed in 6 Göttingen minipigs (Sus scrofa domestica) with severe interstitial pneumonia. The virus was defined as a North American (NA) subtype virus, which is common in the commercial pig population and might be derived from a widely used attenuated live-virus vaccine in Europe. The ORF5 sequence of the isolated PRRSV was 98% identical to the vaccine virus. The affected pigs were part of a lung transplantation model and received tacrolimus and steroids as well as irradiation or CD8 antibody for immunosuppression. The likely source of the infection was pigs that were shedding the identified PRRSV, which were housed in a separate room of the same building. This case report provides evidence that a virus closely related to an attenuated live vaccine might cause severe pneumonia and death in PRRSVseronegative pigs receiving immunosuppressive treatment. We recommend strict barrier housing for immunocompromised pigs.
Project description:We describe the morphological, biological, and molecular characteristics of Cryptosporidium pig genotype II and propose the species name Cryptosporidium scrofarum n. sp. to reflect its prevalence in adult pigs worldwide. Oocysts of C. scrofarum are morphologically indistinguishable from C. parvum, measuring 4.81-5.96 ?m (mean=5.16)×4.23-5.29 ?m (mean=4.83) with a length to width ratio of 1.07±0.06 (n=400). Oocysts of C. scrofarum obtained from a naturally infected pig were infectious for 8-week-old pigs but not 4-week-old pigs. The prepatent period in 8-week-old Cryptosporidium-naive pigs was 4-6 days and the patent period was longer than 30 days. The infection intensity of C. scrofarum in pigs was generally low, in the range 250-4000 oocysts per gram of feces. Infected pigs showed no clinical signs of cryptosporidiosis and no pathology was detected. Cryptosporidium scrofarum was not infectious for adult SCID mice, adult BALB/c mice, Mongolian gerbils (Meriones unguiculatus), southern multimammate mice (Mastomys coucha), yellow-necked mice (Apodemus flavicollis), or guinea pigs (Cavia porcellus). Phylogenetic analyses based on small subunit rRNA, actin, and heat shock protein 70 gene sequences revealed that C. scrofarum is genetically distinct from all known Cryptosporidium species.
Project description:Porcine reproductive and respiratory syndrome virus (PRRSV), is a highly mutable RNA virus that affects swine worldwide and its control is very challenging due to its formidable heterogeneity in the field. In the present study, DNA vaccines constructed with PRRSV GP5-Mosaic sequences were complexed to cationic liposomes and administered to experimental pigs by intradermal and intramuscular injection, followed by three boosters 14, 28 and 42 days later. The GP5-Mosaic vaccine thus formulated was immunogenic and induced protection from challenge in vaccinated pigs comparable to that induced by a wild type (VR2332) GP5 DNA vaccine (GP5-WT). Periodic sampling of blood and testing of vaccine-induced responses followed. Interferon-? (IFN-?) mRNA expression by virus-stimulated peripheral blood mononuclear cells (PBMCs) of GP5-Mosaic-vaccinated pigs was significantly higher compared to pigs vaccinated with either GP5-WT or empty vector at 21, 35 and 48 days after vaccination. Cross-reactive cellular responses were also demonstrated in GP5-Mosaic vaccinated pigs after stimulation of PBMCs with divergent strains of PRRSV. Thus, significantly higher levels of IFN-? mRNA were detected when PBMCs from GP5-Mosaic-vaccinated pigs were stimulated by four Genotype 2 strains (VR2332, NADC9, NADC30 and SDSU73), which have at least 10% difference in GP5 amino acid sequences, while such responses were recorded only upon VR2332 stimulation in GP5-WT-vaccinated pigs. In addition, the levels of virus-specific neutralizing antibodies were higher in GP5-Mosaic or GP5-WT vaccinated pigs than those in vector-control pigs. The experimental pigs vaccinated with either the GP5-Mosaic vaccine or the GP5-WT vaccine were partially protected from challenge with VR2332, as measured by significantly lower viral loads in sera and tissues and lower lung lesion scores than the vector control group. These data demonstrate that the GP5-Mosaic vaccine can induce cross-reactive cellular responses to diverse strains, neutralizing antibodies, and protection in pigs.
Project description:To determine differences in infection kinetics of two temporally and genetically different type 2 porcine reproductive and respiratory syndrome virus (PRRSV) isolates in vivo with and without concurrent porcine circovirus (PCV) type 2a or 2b infection, 62 pigs were randomly assigned to one of seven groups: negative controls (n=8); pigs coinfected with a 1992 PRRSV strain (VR-2385) and PCV2a (CoI-92-2a; n=9), pigs coinfected with VR-2385 and PCV2b (CoI-92-2b; n=9), pigs coinfected with a 2006 PRRSV strain (NC16845b) and PCV2a (CoI-06-2a; n=9), pigs coinfected with NC16845b and PCV2b (CoI-06-2b; n=9), pigs infected with VR-2385 (n=9), and pigs infected with NC16845b (n=9). Blood samples were collected before inoculation and at day post-inoculation (dpi) 3, 6, 9 and 12 and tested for the presence of PRRSV antibody and RNA, PCV2 antibody and DNA, complete blood counts, and interferon gamma (IFN-?) levels. Regardless of concurrent PCV2 infection, VR-2385 initially replicated at higher levels and reached peak replication levels at dpi 6. Pigs infected with VR-2385 had significantly higher amounts of viral RNA in serum on both dpi 3 and dpi 6, compared to pigs infected with NC16845b. The peak of NC16845b virus replication occurred between dpi 9 and dpi 12 and was associated with a delayed anti-PRRSV antibody response in these pigs. PCV2 coinfection resulted in significantly more severe macroscopic and microscopic lung lesions and a stronger anti-PRRSV IgG response compared to pigs infected with PRRSV alone. This work further emphasizes in vivo replication differences among PRRSV strains and the importance of coinfecting pathogens.
Project description:BACKGROUND: Porcine reproductive and respiratory syndrome virus (PRRSV) is largely responsible for heavy economic losses in the swine industry worldwide because of its high mutation rate and subsequent emergence of virulent strains. However, the immunological and pathological responses of pigs to PRRSV strains with different virulence have not been completely elucidated. METHODS: Twenty-four piglets were divided into 4 groups (n?=?6 each) and inoculated with highly pathogenic PRRSV isolate BB0907 (HP), low pathogenic PRRSV NT0801 (LP), LP derivative strain NT0801-F70 (LP-der), and DMEM medium (control), respectively. The changes in TLR2, 3, 7, and 8 gene expression and TNF-?, IL-1?, IL-6, IFN-?, and IL-10 secretion were evaluated using real-time PCR and ELISA at 6, 9, and 15 days post inoculation (d.p.i.). The cytokine levels were evaluated in the supernatants of porcine alveolar macrophages (PAMs) and peripheral blood mononuclear cells (PBMCs) following stimulation with LTA, poly(I:C), CL097, and PRRSV individually. RESULTS: HP caused more severe clinical signs and pathological lesions in swine than LP and LP-der had almost no virulence compared with LP. The serum levels of IL-1?, IL-6, TNF-?, and IFN-? were increased in HP-infected piglets, which were greater than in those infected with LP or LP-der. The mRNA levels of TLR3, 7, and 8 were significantly up-regulated in PAMs in HP-infected pigs compared to those in groups LP and LP-der. Furthermore, TNF-? and IL-1? secretion in PAMs from group LP was statistically greater than those from the control group after stimulation with either poly(I:C) or CL097. Meanwhile, TNF-?, IL-1?, and IL-6 levels in CL097-stimulated PBMCs from HP-infected pigs were markedly higher than those from the LP- and LP-der-infected groups. CONCLUSIONS: We found that HP was a stronger inducer of TLR 3, 7, and 8 expression and IL-1?, IL-6, TNF-?, and IFN-? production compared to LP and LP-der. HP enhanced production of TNF-?, IL-1?, and IL-6 in PBMCs following CL097-stimulation more than LP and LP-der, whereas LP enhanced the secretion of TNF-? and IL-1? in poly(I:C)- and CL097-stimulated PAMs. Our data regarding cellular reactivity to different isolates should be useful in the development of more efficacious vaccines.
Project description:Porcine reproductive and respiratory syndrome virus (PRRSV) is prevalent in swine farms worldwide and is a major source of economic loss and animal suffering. Rapid genetic variation of PRRSV makes it difficult for current vaccines to confer protection against newly emerging strains. We recently demonstrated that a novel peptide nanofiber hydrogel (H9e) could act as a potent adjuvant for killed H1N1 vaccines. Therefore, the objective of this study was to evaluate H9e as an adjuvant for PRRSV modified live virus (MLV) vaccines. Pigs were vaccinated with Ingelvac PRRSV MLV with or without H9e adjuvant before being challenged with the VR-2332 (parental vaccine strain) or MN184A (genetically diverse strain) PRRSV. Pigs vaccinated with MLV+H9e had higher levels of circulating vaccine virus. More importantly, pigs vaccinated with MLV+H9e had improved protection against challenge by both PRRSV strains, as demonstrated by reduced challenge-induced viremia compared with pigs vaccinated with MLV alone. Pigs vaccinated with MLV+H9e had lower frequency of T-regulatory cells and IL-10 production but higher frequency of Th/memory cells and IFN-? secretion than that in pigs vaccinated with MLV alone. Taken together, our studies suggest that the peptide nanofiber hydrogel H9e, when combined with the PRRSV MLV vaccine, can enhance vaccine efficacy against two different PRRSV strains by modulating both host humoral and cellular immune responses.
Project description:BACKGROUND:Currently, an in vitro immunogenicity screening system for the immunological assessment of potential porcine reproductive and respiratory syndrome virus (PRRSV) vaccine candidates is highly desired. Thus, in the present study, two genetically divergent PRRSVs were characterized in vitro and in vivo to identify an in vitro system and immunological markers that predict the host immune response. Porcine alveolar macrophages (PAMs) and peripheral blood mononuclear cells (PBMCs) collected from PRRSV-negative pigs were used for in vitro immunological evaluation, and the response of these cells to VR2332c or JA142c were compared with those elicited in pigs challenged with the same viruses. RESULTS:Compared with VR2332c or mock infection, JA142c induced increased levels of type I interferons and pro-inflammatory cytokines (TNF-?, IL-1?/?, IL-6, IL-8, and IL-12) in PAMs, and these elevated levels were comparable to the cytokine induction observed in PRRSV-challenged pigs. Furthermore, significantly greater numbers of activated CD4+ T cells, type I helper T cells, cytotoxic T cells and total IFN-?+ cells were observed in JA142c-challenged pigs than in VR2332c- or mock-challenged pigs. CONCLUSIONS:Based on these results, the innate immune response patterns (particularly IFN-?, TNF-? and IL-12) to specific PRRSV strains in PAMs might reflect those elicited by the same viruses in pigs.