Novel adjuvants derived from attenuated lipopolysaccharides and lipid As of purple non-sulfur photosynthetic bacteria

Adjuvants are substances that enhance adaptive immune response to antigen. Development of a safe and effective immunostimulant adjuvant is essential for the efficacy of a vaccine to protect against infectious pathogens. Purple non-sulfur photosynthetic bacteria exhibited nontoxic natural lipid A variants that are distinct in their chemical structures from that of the Escherichia coli-type lipid A. In this study, the adjuvant efficacy of attenuated lipid A variants and their corresponding lipopolysaccharides (LPSs), derived from purple photosynthetic bacteria (Rhodocyclus tenuis and Rhodobacter sphaeroides) were evaluated. LPS was extracted using modified phenol-chloroform-petroleum ether method and lipid A was separated by mild acid hydrolysis. Trinitrophenol (TNP) was conjugated to hen egg albumin (TNP-HEA) and used as haptenic antigen. The LPS and lipid A adjuvant candidates were formulated in oil-in-water emulsion (OIWE) and evaluated to elicit anti-TNP IgG against TNP-HEA conjugate in BALB/c female mice. The anti-TNP IgG titers were measured using ELISA. The intact LPS-based adjuvants present in OIWE formulation showed significantly higher efficacy to elicit anti-TNP IgG titers against TNP-HEA conjugate compared to their corresponding lipid A-based adjuvants. As expected, the OIWE formulations of all LPS- and lipid A-based adjuvant candidates showed higher activities compared to the aqueous formulations. Slow reduction in the levels of anti-TNP IgG antibodies in the serum was observed over 4 months after immunization using the LPS- and lipid A-based adjuvant candidates which may provide a long protection against pathogens. The attenuated LPSs and lipid A’s from the photosynthetic bacteria showed promising results to develop novel safe and effective adjuvants that can evoke the immune response. The most promising adjuvant candidate was the LPS-based adjuvant from R. tenuis.     

A simple and rapid assay to evaluate purity of foot-and-mouth disease vaccine before animal experimentation

Currently, foot-and-mouth disease (FMD) vaccine purity is tested in cattle to detect antibodies against the non-structural protein (NSP) after repeated immunization with the final vaccine product. In case of vaccine failure, the manufacturing company would suffer significant economic loss. To prevent such unfortunate losses with the final vaccine product, in vitro testing is required to quantitate an NSP antigen during the manufacturing process prior to animal experiments. A novel lateral-flow assay device was developed using a monoclonal antibody (MAb) against the 3B NSP. To determine the minimal amount of NSP required to elicit antibodies in livestock, goats were immunized several times with various concentrations of either the recombinant 3AB (rec.3AB) protein or FMD virus culture supernatant. Antibodies against 3AB were elicited after a second immunization with 10.6 ng to 42.5 ng of rec.3AB and a third immunization with a 10-fold diluted FMD virus culture supernatant in goats. The lateral-flow assay device detected the minimal amount of rec.3AB and native NSP in FMD virus culture supernatant required to induce NSP antibodies in goats. The in vitro assay device is simple and economical, provides rapid results, and should be useful for FMD vaccine-manufacturing companies prior to conducting animal experiments to test the vaccine purity.