ABSTRACT
Many different strategies have been used to fight against the Coronavirus disease (COVID-19) pandemic as a therapeutics or prophylaxis approaches. However, not enough attention has been paid to general and specific immune factors and nutritional components found in hyper-immunized dairy products. Hyper-immune bovine colostrum (HBC) has been used against many different respiratory and gastrointestinal tracts infections during past decades. An isolated dairy farm was established, and nine mixed Holstein X Simmental dairy cattle in their 6-7 months of gestation period were chosen for hyper-immunization with inactivated Severe acute respiratory syndrome corona virus-2 (SARS-CoV-2). For this, six cows were inoculated with 2 ml of 109.4/ml (TCID50) of the virus. As a control group, three cows were inoculated with the carrier without virus. Specific IgG level against the SARS-CoV-2 was measured before and after immunization in the sera, and in the colostrum and milk following parturition in hyper-immunized cows using indirect Enzyme-linked immunosorbent assay (ELISA). Neutralizing antibodies in the serum and colostrum was measured by a quantitative ELISA. The safety of the product was determined in40 healthy volunteers aged between 18-65 years old (13 females and 27 males) in the phase 1 clinical trial (https://www.irct.ir/trial/51259). No adverse effects were observed in the experimental cows. A very high level of IgG was observed in the first colostrum that sharply decreased in the following 7 days in the milk. The titer of specific neutralizing antibody in the colostrum samples was 69 times higher than the sera. No adverse effects and clinical complications were reported by the authorized ethics committee, and an official certificate on the safety of the product was issued. Beside other strategies, this approach could be used for large-scale and low-cost production of immune components to be used as a nutritional supplement to confront current SARS-CoV-2 and future pandemics. Clinical Trial Registration: [https://www.irct.ir/trial/51259].
ABSTRACT
Due to its conservation, the extracellular domain of the influenza A M2 protein (M2e) has the potential for being applied as a recombinant vaccine candidate against a wide range of strains, though its immunogenicity may need to be improved. The occurrence of several post-translational modifications within the structure of M2 protein may affect its immunopotency for the induction of humoral immune response. Herein, to construct a recombinant M2e-based vaccine candidate with the appropriate structural conformation and immunogenicity the corresponding nucleotide sequence from an H9N2 influenza strain was fused to the N-terminus of the truncated Mycobacterium tuberculosis HSP70(359-610), as a potent adjuvant, and following its cloning into the pPICZ alpha A plasmid the fusion gene was expressed in Pichia pastoris KM71H yeast. The secreted protein was then easily purified from the culture media, based on the presence of polyhistidine tag and used for the production of rabbit polyclonal antisera. This raised antisera could recognize the native M2e protein on the surface of H9N2 influenza virus-infected MDCK cells at a comparable level with the commercial H2N2-specific anti-M2 antibody, which was evidenced with immunofluorescence and cell-ELISA assays. These results not only re-emphasized on the conservancy of the M2e antigen, but also pointed towards the applicability of the M2e-HSP70(359-610) fusion protein for the induction of specific antibodies capable of binding to the native M2e antigen on the infected cells. Collectively, this study implied that purified M2e-HSP70(359-610) represents a promising vaccine candidate; however, its in vivo potency for the induction of protection remains to be evaluated.
