New Immunoinformatics Tools for Swine: Designing Epitope-Driven Vaccines, Predicting Vaccine Efficacy, and Making Vaccines on Demand.

Novel computational tools for swine vaccine development can expand the range of immunization approaches available to prevent economically devastating swine diseases and spillover events between pigs and humans. PigMatrix and EpiCC are two new tools for swine T cell epitope identification and vaccine efficacy analysis that have been integrated into an existing computational vaccine design platform named iVAX. The iVAX platform is already in use for the development of human vaccines, thus integration of these tools into iVAX improves and expands the utility of the platform overall by making previously validated immunoinformatics tools, developed for humans, available for use in the design and analysis of swine vaccines. PigMatrix predicts T cell epitopes for a broad array of class I and class II swine leukocyte antigen (SLA) using matrices that enable the scoring of sequences for likelihood of binding to SLA. PigMatrix facilitates the prospective selection of T cell epitopes from the sequences of swine pathogens for vaccines and permits the comparison of those predicted epitopes with "self" (the swine proteome) and with sequences from other strains. Use of PigMatrix with additional tools in the iVAX toolkit also enables the computational design of vaccines in silico, for testing in vivo. EpiCC uses PigMatrix to analyze existing or proposed vaccines for their potential to protect, based on a comparison between T cell epitopes in the vaccine and circulating strains of the same pathogen. Performing an analysis of T cell epitope relatedness analysis using EpiCC may facilitate vaccine selection when a novel strain emerges in a herd and also permits analysis of evolutionary drift as a means of immune escape. This review of novel computational immunology tools for swine describes the application of PigMatrix and EpiCC in case studies, such as the design of cross-conserved T cell epitopes for swine influenza vaccine or for African Swine Fever. We also describe the application of EpiCC for determination of the best vaccine strains to use against circulating viral variants of swine influenza, swine rotavirus, and porcine circovirus type 2. The availability of these computational tools accelerates infectious disease research for swine and enable swine vaccine developers to strategically advance their vaccines to market.

Screening of PRRSV- and ASFV-encoded proteins involved in the inflammatory response using a porcine iGLuc reporter.

Inflammasome plays a major role in innate immune responses by activating caspase-1, resulting in secretion of interleukin-1ß (IL-1ß) and inflammatory pathologic responses. IL-1ß release is widely used as an indirect readout to study inflammasome activation. Here we report an iGLuc reporter (pro-IL-1ß-Gluc) of pig origin to monitor cytosolic pro-IL-1ß cleavage and mature IL-1ß release. Based on the iGLuc reporter, we reconstructed the inflammasome system in vitro and screened PRRSV- and ASFV-encoded proteins involved in regulating inflammasome activation. We found that three non-structural proteins (nsps) of PRRSV, nsp1ß, nsp2 and nsp5, activate the NLRP3 inflammasome, and four nsps of PRRSV, nsp1ɑ, nsp7, nsp10 and nsp11, inhibit NLRP3 inflammasome activation, of which nsp10 and nsp11 have a highly significant inhibitory effect. In addition, we also found that four ASFV-encoded proteins, S183L, E199L, O61R and I7L activate the inflammatory response and four ASFV-encoded proteins, I226L, A151R, NP419L and QP383R, inhibit the inflammatory response. Our results provide a highly sensitive and high-throughput tool to screen for proteins that regulate inflammasome activation in vitro.

Recombinant antigen targets for serodiagnosis of African swine fever.

African swine fever (ASF) is an infectious and economically important disease of domestic pigs. There is no vaccine, and so reliable diagnosis is essential for control strategies. The performance of four recombinant ASF virus (ASFV) protein (pK205R, pB602L, p104R, and p54)-based enzyme-linked immunosorbent assays (ELISAs) was evaluated with European porcine field sera that had been established by Office International des Epizooties (OIE)-approved tests to be ASFV negative (n = 119) and ASFV positive (n = 80). The kappa values showed that there was almost perfect agreement between the results of the "gold standard" test (immunoblotting) and the results obtained by the p54-specific ELISA (kappa = 0.95; 95% confidence interval [CI], 0.90 to 0.99) and the pK205R-specific ELISA or the pB602L-specific ELISA (kappa = 0.92; 95% CI, 0.86 to 0.97). For the pA104R-specific ELISA, there was substantial to almost perfect agreement (kappa = 0.81; 95% CI, 0.72 to 0.89). Similar results were observed by the OIE-approved ELISA (kappa = 0.89; 95% CI, 0.82 to 0.95). Importantly, antibodies against these proteins were detectable early after infection of domestic pigs. Preliminary testing of 9 positive and 17 negative serum samples from pigs from West Africa showed identical results by the recombinant protein-based ELISA and the OIE-approved tests. In contrast, there was a high degree of specificity but a surprisingly a low level of sensitivity with 7 positive and 342 negative serum samples from pigs from East Africa. With poorly preserved sera, only the p104R-specific ELISA showed a significant reduction in sensitivity compared to that of the OIE-approved ELISA. Finally, these recombinant proteins also detected antibodies in the sera of the majority of infected warthogs. Thus, recombinant ASFV proteins p54, pB602L, and pK205R provide sensitive and specific targets for the detection of antibodies in European and West African domestic pigs and warthogs.

African swine fever: morphopathology of a viral haemorrhagic disease.

Twelve miniature pigs were infected experimentally with the moderately-virulent Spanish isolate of African swine fever virus E75. The disease was reproduced acutely and subacutely, and six days after infection the animals showed severe clinical and pathological signs characterised by diffuse haemorrhages in lymphoid and non-lymphoid organs. An immunohistopathological study, using two different methods, and a monoclonal antibody (mAb) (18BG3) were used to analyse the distribution of the virus protein 73 (VP73) and IgM and IgG. VP73 was observed primarily in cells of the mononuclear phagocyte system, and in many megakaryocytes, some tubular epithelial cells of the kidneys, tonsillar epithelium, some hepatocytes, and in a few endothelial cells and neutrophils in the later stages of the infection. The histological findings and the distribution of VP73 and IgM and IgG indicated that the virus caused haemorrhagic lesions through its effect on various haemostatic mechanisms.