Blue eye disease (BED) is a swine viral infection that affects the pork industry of Mexico. Porcine orthorubulavirus (PRV) is the etiological agent, and the hemagglutinin-neuraminidase protein (HN) is characterized as the best antigen for serological tests, although other structural proteins, including the nucleoprotein (NP) and the matrix (M) protein, have been investigated during the infection of members of the Paramyxoviridae family, generating promising results. Herein, for the first time, we successfully produced and characterized both the NP and M proteins of PRV by using a recombinant strategy in the E. coli heterologous system. The ORF of the NP and M genes were cloned in-frame with the pET-SUMO expression vector. Recombinant proteins proved to be a sensitive target to detect seroconversion at 7 days until 28 days in vaccinated mice (BALB/c) by indirect ELISAs. Immunoreactivity was also tested using porcine serum samples, in which antibodies were recognized from early stages to a persistence of PRV infection, which is indicative that these proteins contain properties similar to native antigens. The predicted tertiary structure showed that both proteins have a conserved structure that resembles those found in others Paramyxovirus. Our results pave the way for developing biotechnological tools based on these proteins for the control and prevention of BED.

Porcine deltacoronavirus (PDCoV) is an emergent swine coronavirus which infects cells from the small intestine and induces watery diarrhea, vomiting and dehydration, causing mortality in piglets (>40%). The aim of this study was to evaluate the antigenicity and immunogenicity of the recombinant membrane protein (M) of PDCoV (rM-PDCoV), which was developed from a synthetic gene obtained after an in silico analysis with a group of 138 GenBank sequences. A 3D model and phylogenetic analysis confirmed the highly conserved M protein structure. Therefore, the synthetic gene was successfully cloned in a pETSUMO vector and transformed in E. coli BL21 (DE3). The rM-PDCoV was confirmed by SDS-PAGE and Western blot with ~37.7 kDa. The rM-PDCoV immunogenicity was evaluated in immunized (BLAB/c) mice and iELISA. The data showed increased antibodies from 7 days until 28 days (p < 0.001). The rM-PDCoV antigenicity was analyzed using pig sera samples from three states located in “El Bajío” Mexico and positive sera were determined. Our results show that PDCoV has continued circulating on pig farms in Mexico since the first report in 2019; therefore, the impact of PDCoV on the swine industry could be higher than reported in other studies.

Orthorubulavirus suis (LPMV) is the etiologic agent of blue eye disease (BED), which affects pigs of all ages, and it has been endemic in central Mexico since the 1980s. To date, no disease control program has been established. Therefore, there is a need for a serological diagnostic method with high sensitivity and specificity. In this study, the recombinant protein NP of LPMV was produced in the E. coli BL21 system and then purified using affinity chromatography. The purified protein was used to coat plates for an indirect ELISA assay (iELISA). To determine the sensitivity and specificity of the test, a 2 × 2 contingency table was constructed using positive and negative control sera. The specificity and sensitivity levels were 98.1% and 98.7%, respectively. According to our findings, 45% of serum samples (378/839) were positive, with seropositivity percentages in the analyzed states ranging from 72.5% to 6%. To confirm the presence of antibodies, the indirect immunofluorescence technique was applied to iELISA-positive serum samples. In this study, antibodies against the LPMV nucleoprotein were detected, indicating that the virus or defective particles may be circulating in Mexican pigs and highlighting the risk of LPMV spreading to disease-free areas.

Small ruminant lentivirus (SRLV) infects goats and sheep of all breeds and ages worldwide. There are no current records regarding the three-dimensional structure or antigenic capacity of the nucleocapsid protein p14 of FESC-752 Mexican strain. The antigenic structure of p14 protein of a B1 genotype was predicted. cDNA from FESC-752 was used to overexpress the recombinant SRLV-rp14 protein. Then, its antigenicity was verified in vitro by evaluating plasma samples from goats and sheep naturally infected with SRLV. Antigenicity prediction showed a “horseshoe”-type structure shared by different lentiviruses and five epitopes distributed throughout the p14 surface regions where they coincide suggesting conserved epitopes in the zinc-finger structures of the nucleoproteins of the SRLV, Human Immunodeficiency Virus (HIV-1), and Feline Immunodeficiency virus (FIV) retroviruses. Multi-species molecular docking showed a notable structural convergence where caprine, bovine, murine, and human immunoglobulins target a predictive 23 amino acid epitope (residues 41–64) within the core zinc-finger region. Furthermore, CABS docking simulations predicted that p14-derived peptides preferentially bind within the antigen-presenting cleft of both caprine and bovine major histocompatibility complex class I (MHC-I) molecules. The stability of these immunological complexes is mediated by dense networks of hydrophobic interactions and highly conserved aromatic anchoring residues. Antigenicity analysis revealed that 78.7% of samples from naturally infected goats showed immunoreactivity toward SRLV-rp14 and the predictive evidence that p14 can simultaneously stimulate both humoral and cellular pathways makes it a strategic candidate for the design of next-generation vaccines aimed at controlling lentiviruses in small ruminants.