Evaluation of the humoral and mucosal immune response of a multiepitope vaccine against COVID-19 in pigs.

Introduction: This study evaluated the immune response to a multiepitope recombinant chimeric protein (CHIVAX) containing B- and T-cell epitopes of the SARS-CoV-2 spike's receptor binding domain (RBD) in a translational porcine model for pre-clinical studies. Methods: We generated a multiepitope recombinant protein engineered to include six coding conserved epitopes from the RBD domain of the SARS-CoV-2 S protein. Pigs were divided into groups and immunized with different doses of the protein, with serum samples collected over time to determine antibody responses by indirect ELISA and antibody titration. Peptide recognition was also analyzed by Western blotting. A surrogate neutralization assay with recombinant ACE2 and RBDs was performed. Intranasal doses of the immunogen were also prepared and tested on Vietnamese minipigs. Results: When the immunogen was administered subcutaneously, it induced specific IgG antibodies in pigs, and higher doses correlated with higher antibody levels. Antibodies from immunized pigs recognized individual peptides in the multiepitope vaccine and inhibited RBD-ACE2 binding for five variants of concern (VOC). Comparative antigen delivery methods showed that both, subcutaneous and combined subcutaneous/intranasal approaches, induced specific IgG and IgA antibodies, with the subcutaneous approach having superior neutralizing activity. CHIVAX elicited systemic immunity, evidenced by specific IgG antibodies in the serum, and local mucosal immunity, indicated by IgA antibodies in saliva, nasal, and bronchoalveolar lavage secretions. Importantly, these antibodies demonstrated neutralizing activity against SARS-CoV-2 in vitro. Discussion: The elicited antibodies recognized individual epitopes on the chimeric protein and demonstrated the capacity to block RBD-ACE2 binding of the ancestral SARS-CoV-2 strain and four VOCs. The findings provide proof of concept for using multiepitope recombinant antigens and a combined immunization protocol to induce a neutralizing immune response against SARS-CoV-2 in the pig translational model for preclinical studies.

Immunogenicity of a recombinant hemagglutinin neuraminidase-Porcine rubulavirus produced by Escherichia coli of Porcine rubulavirus gives protective immunity of litter after challenge.

Porcine rubulavirus (PRV) is a contagious virus that affects the Mexican swine industry. This work aimed to evaluate the immunogenicity of an recombinant hemagglutinin neuraminidase-Porcine rubulavirus (rHN-PorPV) candidate vaccine on pregnant sows, and the protective efficacy afforded to their 7-day-old suckling piglets against PRV lethal challenge. Three sows were immunized with rHN-PorPV formulated with immune-stimulating complex (ISCOMs) and two sows with rHN-PorPV protein alone as well as a mock-immunized pregnant sow (negative control). Quantitative ELISA detected a high concentration of anti-rHN-PorPV Immunoglobulin G (IgG) antibodies in sow sera after the second dose of vaccine administered on day 14 until farrowing, showing viral-neutralizing and cross-neutralization activity against different variants of PRV. Sera samples from piglets of immunized sows (with or without adjuvant), showed high concentrations of IgG antibodies. As expected, piglets from the negative control sow (n=5), exhibited severe signs of disease and 100% of mortality after PRV challenge study. Conversely, 75% and 87.5% of the piglets born from the rHN-PorPV and the rHN-PorPV-ISCOMs-immunized sows (n=8), survived, respectively, showing milder PRV clinical signs. Our data indicate that rHN-PorPV candidate vaccine produced in Escherichia coli induces efficient humoral response in pregnant sows and that the maternally derived immunity provides high protection to suckling piglets against PRV lethal challenge.

CD71+ Erythroid Cells in Human Neonates Exhibit Immunosuppressive Properties and Compromise Immune Response Against Systemic Infection in Neonatal Mice.

Newborns are highly susceptible to infectious diseases. The underlying mechanism of neonatal infection susceptibility has generally been related to their under-developed immune system. Nevertheless, this notion has recently been challenged by the discovery of the physiological abundance of immunosuppressive erythroid precursors CD71+erythroid cells (CECs) in newborn mice and human cord blood. Here, as proof of concept, we show that these cells are also abundant in the peripheral blood of human newborns. Although their frequency appears to be more variable compared to their counterparts in mice, they rapidly decline by 4 weeks of age. However, their proportion remains significantly higher in infants up to six months of age compared to older infants. We found CD45 expressing CECs, as erythroid progenitors, were the prominent source of reactive oxygen species (ROS) production in both humans and mice. Interestingly, a higher proportion of CD45+CECs was observed in the spleen versus bone marrow of neonatal mice, which was associated with a higher ROS production by splenic CECs compared to their siblings in the bone marrow. CECs from human newborns suppressed cytokine production by CD14 monocytes and T cells, which was partially abrogated by apocynin in vitro. Moreover, the depletion of CECs in neonatal mice increased the number of activated effector immune cells in their spleen and liver, which rendered them more resistant to Listeria monocytogenes infection. This was evident by a significant reduction in the bacteria load in the spleen, liver and brain of treated-mice compared to the control group, which enhanced their survival rate. Our finding highlights the immunoregulatory processes mediated by CECs in newborns. Thus, such tightly regulated immune system in newborns/infants may explain one potential mechanism for the asymptomatic or mild COVID-19 infection in this population.

Mucosal and systemic immune responses to Aujeszky's disease virus (ADV) in early vaccinated piglets.

Newborn humans and animals are highly susceptible to viral infections. The Aujeszky´s disease virus (ADV) is a porcine herpes virus 1 which infects the respiratory tract and is lethal during the first weeks of life. Current intramuscular vaccines, applied at weaning, induce poor mucosal immunity and frequently fail to prevent and control the disease. Additionally, early vaccination has not been studied thoroughly. Therefore, we studied a systemic/mucosal route of immunization using an inactivated ADV vaccine in two-and fourteen-day-old groups of unweaned SPF miniature Vietnamese pigs, measuring the anti ADV antibody (ELISA) and cytokine (qPCR) responses in systemic and mucosal samples. The results showed that the serum ADV-specific IgG response was higher in the 14-day groups. However, the nasal IgA responses were similar in immunized groups, although the response in saliva was higher in the 2-day old group. Moreover, in vitro ADV stimulated peripheral blood mononuclear cells and lung cells from immunized pigs showed higher IFN-γ mRNA production in the 14-day old group than in younger animals and similar levels of IL-4 and IL-10 transcripts. Our data suggest that early mucosal immunization induce humoral and cellular systemic and mucosal immune responses against ADV in young pigs and younger animals may have compensatory mechanisms to overcome early immaturity and maternal-driven immune interference. Therefore, early protection in susceptible animals could be induced using this immunization protocol, opening the possibility for its application against other viral pathogens of pigs and for traslational studies in humans.

Exercise modulates the expression of IL-1ß and IL-10 in the articular cartilage of normal and osteoarthritis-induced rats.

After a joint lesion, high-impact exercise is a risk factor for the development of osteoarthritis (OA). The degradation of articular cartilage in OA has been associated with the activation of inflammatory cytokine signaling pathways. However, differences in cytokine expression in healthy and injured cartilage after exercise have not yet been analyzed. We used immunofluorescence and Western blot to study the expression of IL-1ß and IL-10 in the articular cartilage of normal (N), sham-operated (S), and menisectomized (OA) rats subjected or not to high-impact exercise (E) for 3, 6, and 10 days (N, NE, S, SE, and OA groups). Cartilage integrity and proteoglycan content were only affected in the OA groups. Exercise increased the amount of IL-1ß and IL-10 positive chondrocytes in NE and SE groups compared with non-exercised groups (N and S). The expression of IL-1ß was up-regulated over time in the NE and OA groups, although in the late stages the increase was higher in the OA groups. In contrast, the expression of anti-inflammatory IL-10 was low in the OA group, whereas in the NE groups expression levels were higher at each time point analyzed. These results suggest that anti- and pro-inflammatory molecules in the cartilage might be tightly regulated to maintain the integrity of the tissue and that when this equilibrium is broken (when the meniscus is removed), the pro-inflammatory cytokines take over and OA develops.

Perfil inmunológico de los cerdos durante las primeras diez semanas de edad / Immune profile in pigs during the first ten weeks of age

Se muestrearon semanalmente 14 animales para determinar el perfil inmunológico de los cerdos durante las primeras diez semanas de vida. Durante la lactancia, de la primera a la cuarta semana de edad, fue evidente una baja concentración de leucocitos, linfocitos T totales (TE), T de alta afinidad (Taa) y B con receptor Fc (Bfc), determinados por rosetas con eritrocitos de borrego. Asimismo, se incrementó la respuesta inradérmica a la fitohemaglutinina (PHA). En la cuarta semana los animales fueron destetados: entre la quinta y sexta semana de edad continuó el incremento de leucocitos y linfocitos TE, Taa y Bfc y disminuyeron los linfocitos Null. Sin embargo, hubo una disminución marcada de la respuesta intradérmica a la PHA. A partir de la octava semana de edad, los valores que se encontraron en las subpoblaciones celulares y la respuesta intradérmica fueron semejantes a los que se informan en adultos. Se concluye que el sistema inmune de los cerdos madura durante las primeras ocho semanas de vida, aunque durante el destete hubo cambios drásticos en las concentraciones de las subpoblaciones celulares y se hicieron anérgicos a la prueba intradérmica, lo que sugiere una inmunosupresión transitoria