Cytokine and C-reactive protein profiles induced by porcine circovirus type 2 experimental infection in 3-week-old piglets.

The purpose of this study was to determine serum profiles of cytokines at a protein level and Creactive protein (CRP) during the development of postweaning multisystemic wasting syndrome (PMWS) in experimentally inoculated pigs. Levels of serum IFN-alpha, IL-6, IL-10, and CRP were examined for a 35-day period in 10 piglets experimentally infected with PCV2 at 3 weeks of age. Four of the infected piglets developed severe PMWS at 14 to 21 days post-infection (d.p.i.) and died prior to termination of the experiment. The remaining six PCV2-infected piglets experienced transient fever, but did not display overt clinical signs of PMWS and were considered as subclinically infected. A bioassay was used to detect IL-6 and ELISAs were used to detect IFN-alpha, IL-10, and CRP. There were no significant differences in cytokine or CRP expression from 0 to 7 d.p.i. between the PMWS-affected and the subclinically infected piglets. Levels of IL-10 and CRP were elevated from 10 and 14 d.p.i. respectively in the PMWS-affected piglets compared to the subclinically infected piglets. There were no significant differences in IFN-alpha and IL-6 expression between the PMWS-affected piglets and the subclinically infected piglets. The present study shows that elevated levels of serum CRP and IL-10 were associated with PCV2-infected piglets that subsequently developed severe PMWS. This may help to provide further insight into the immunoaetiogenesis of this syndrome.

An in vitro immune response model to determine tetanus toxoid antigen (vaccine) specific immunogenicity: Selection of sensitive assay criteria.

Many vaccines employed in childhood vaccination programmes are produced by conventional techniques, resulting in complex biological mixtures for which batch-related quality control requires in vivo potency testing. Monitoring consistency via in vitro tests during the vaccine production has the capacity to replace certain of the in vivo methods. In this respect, determining vaccine antigen immunogenicity through functional immunological tests has high potential. Advances in immunology have made it possible to analyse this biological activity by in vitro means. The present study established such an in vitro test system for tetanus toxoid (TT). This measured vaccine immunogenicity through an antigen-specific secondary (recall) response in vitro, using a porcine model growing in value for its closeness to human immune response characteristics. Discrimination between the specific recall TT antigen and diphtheria toxoid (DT) was possible using both peripheral blood mononuclear cell cultures and monocyte-derived dendritic cells in co-culture with autologous specific lymphocytes. TT-specific activation was detected with highest discrimination capacity using proliferation assays, as well as IFN-gamma and TT-specific antibody ELISPOTS (measuring secreting T and B lymphocytes, respectively). These in vitro systems show a high potential for replacing animal experimentation to evaluate the immunogenicity of complex vaccines.

C-kit positive porcine bone marrow progenitor cells identified and enriched using recombinant stem cell factor.

Porcine haematological studies have been hampered by the lack of monoclonal antibodies against porcine CD34 or CD117 expressed on haematological progenitors. The present report describes the enumeration, phenotyping and isolation of porcine haematopoietic progenitor cells expressing stem cell factor (SCF, c-kit ligand) receptor (c-kit, CD117). Recombinant porcine (rp) SCF and granulocyte-macrophage colony-stimulating factor (GM-CSF) were expressed in the mammalian HEK293 cell-based expression system. Both were biologically active and induced the proliferation of the human erythroleukemic cell line TF-1, as well as of porcine bone marrow haematopoietic cells (BMHC), in a concentration-dependent manner. The effect of rpSCF on BMHC proliferation was synergistic with rpGM-CSF. Furthermore, rpSCF had a synergistic effect on the generation of BMHC-derived dendritic cells (DC) induced by GM-CSF and TNF-alpha. RpSCF was expressed with a 6-histidine epitope, permitting both its purification and immunological detection. Binding studies with BMHC demonstrated ligation of SCF to 4-11% of BMHC. These cells represented the SWC3(low/-)SWC8- BMHC subset, with characteristics of immature proliferative progenitor BMHC. In contrast, no expression was noted on the SWC3+SWC8- monocytic, the SWC3+SWC8+ granulocytic or the SWC3-SWC8+ B cell lineage cells. Using magnetic or fluorescence-activated cell sorting, SCF-ligating BMHC were enriched for pluripotent progenitor cells. In this manner, porcine haematological studies can be pursued in a detailed manner not before possible.

Immunologic features of porcine circovirus type 2 infection.

Clinical expression of porcine circovirus 2 (PCV-2) infection in swine may result in two distinct high mortality disease syndromes. In North America, postweaning multisystemic wasting syndrome (PMWS), while still sporadic in incidence, predominates. In Europe and elsewhere, both PMWS and a second syndrome, porcine dermatitis and nephropathy syndrome (PDNS), occur in endemic and epidemic forms. PMWS but not PDNS has been reproduced in piglets by inoculations with PCV-2 alone or in PCV-2-infected swine co-infected with porcine parvovirus (PPV) or porcine respiratory and reproductive syndrome (PRRS) virus and also if PCV-2-infected piglets are immunostimulated by injections with an immunogen emulsified in an oil-based macrophage-targeted adjuvant. Subclinical but active infection has been achieved by direct inoculation of piglets with cloned PCV-2 DNA and/or progeny virus derived from cloned DNA. Morphologic changes in lymphoid tissues and preliminary functional data suggest that immunosuppression may occur in PMWS-affected swine. This phenomenon appears to be mediated by generalized lymphoid depletion and replacement by infiltrating and proliferating histiocytes and macrophages. Accumulation of virus in both mononuclear phagocytes and follicular dendritic cells is characteristic of PCV-2 infection. Exogenous immunosuppression of PCV-2-infected gnotobiotic piglets with cyclosporine (Cys), but not corticosteroid (St), potentiates PCV-2 replication and promotes productive virus infection of hepatocytes in Cys-treated piglets, a tropism not previously apparent in experimentally induced PMWS in gnotobiotic swine. In the Cys-treated piglets, inflammatory lesions characteristic of PMWS are absent, even though tissues contain high titers of infectious virus, a finding which suggests that the granulomatous inflammatory lesions characteristic of PMWS are immune mediated.