Biochemical characterisation of Cytochrome P450 oxidoreductase from the cattle tick, Rhipicephalus microplus, highlights potential new acaricide target.

Management of the cattle tick, Rhipicephalus microplus, presents a challenge because some populations of this cosmopolitan and economically important ectoparasite are resistant to multiple classes of acaricides. Cytochrome P450 oxidoreductase (CPR) is part of the cytochrome P450 (CYP450) monooxygenases that are involved in metabolic resistance by their ability to detoxify acaricides. Inhibiting CPR, the sole redox partner that transfers electrons to CYP450s, could overcome this type of metabolic resistance. This report represents the biochemical characterisation of a CPR from ticks. Recombinant CPR of R. microplus (RmCPR), minus its N-terminal transmembrane domain, was produced in a bacterial expression system and subjected to biochemical analyses. RmCPR displayed a characteristic dual flavin oxidoreductase spectrum. Incubation with nicotinamide adenine dinucleotide phosphate (NADPH) lead to an increase in absorbance between 500 and 600 nm with a corresponding appearance of a peak absorbance at 340-350 nm indicating functional transfer of electrons between NADPH and the bound flavin cofactors. Using the pseudoredox partner, kinetic parameters for both cytochrome c and NADPH binding were calculated as 26.6 ± 11.4 µM and 7.03 ± 1.8 µM, respectively. The turnover, Kcat, for RmCPR for cytochrome c was calculated as 0.08 s-1 which is significantly lower than the CPR homologues of other species. IC50 (Half maximal Inhibitory Concentration) values obtained for the adenosine analogues 2', 5' ADP, 2'- AMP, NADP+and the reductase inhibitor diphenyliodonium were: 140, 82.2, 24.5, and 75.3 µM, respectively. Biochemically, RmCPR resembles CPRs of hematophagous arthropods more so than mammalian CPRs. These findings highlight the potential of RmCPR as a target for the rational design of safer and potent acaricides against R. microplus.

An immunoinformatics approach for the design of a multi-epitope subunit vaccine for urogenital schistosomiasis.

Discovery of T and B memory cells capable of eliciting long-term immunity against schistosomiasisis is important for people in endemic areas. Changes in schistosomes environment due to developmental cycle, induces up-regulation of Heat Shock Proteins (HSPs) which assist the parasite in coping with the hostile conditions associated with its life cycle. This study therefore focused on exploring the role of HSPs in urogenital schistosomiasis to develop new multi-epitope subunit vaccine against the disease using immunoinformatic approaches. The designed subunit vaccine was subjected to in silico antigenicity, immunogenicity, allergenicity and physicochemical parameters analysis. A 3D structure of the vaccine construct was predicted, followed by disulphide engineering for stability, codon adaptation and in silico cloning for proper expression and molecular protein-protein docking of vaccine construct in the vector against toll-like receptor 4 receptor, respectively. Consequently, a 493 amino acid multi-epitope vaccine construct of antigenicity probability of 0.91 was designed. This was predicted to be stable, non-allergenic in nature and safe for human use.

Development of multiepitope subunit protein vaccines against Toxoplasma gondii using an immunoinformatics approach.

Approximately one-third of the world's human population is estimated to have been exposed to the parasite Toxoplasma gondii. Its prevalence is reportedly high in Ethiopia (74.80%) and Zimbabwe (68.58%), and is 40.40% in Nigeria. The adverse effect of this parasite includes a serious congenital disease in the developing fetus of pregnant women. After several efforts to eliminate the disease, only one licensed vaccine 'Toxovax' has been used to avoid congenital infections in sheep. The vaccine has been adjudged expensive coupled with adverse effects and short shelf life. The potential of vaccine to likely revert to virulent strain is a major reason why it has not been found suitable for human use, hence the need for a vaccine that will induce T and B memory cells capable of eliciting longtime immunity against the infection. This study presents immunoinformatics approaches to design a T. gondii-oriented multiepitope subunit vaccine with focus on micronemal proteins for the vaccine construct. The designed vaccine was subjected to antigenicity, immunogenicity, allergenicity and physicochemical parameter analyses. A 657-amino acid multiepitope vaccine was designed with the antigenicity probability of 0.803. The vaccine construct was classified as stable, non-allergenic, and highly immunogenic, thereby indicating the safety of the vaccine construct for human use.