Intraspecific venom variation in the medically important puff adder (Bitis arietans): comparative venom gland transcriptomics, in vitro venom activity and immunological recognition by antivenom

Abstract

BackgroundVariation in snake venoms is well documented, both between and within species, with intraspecific venom variation often correlated with geographically distinct populations. The puff adder,Bitis arietans, is found widely distributed across sub-Saharan Africa and into the Arabian Peninsula where it is considered a leading cause of the ∼310,000 annual snakebites across the region, with its venom capable of causing substantial morbidity and mortality. Despite its medical importance and wide geographic distribution, there is little known about venom variation between differentB. arietanspopulations and the potential implications of this variation on antivenom efficacy.MethodologyWe applied a range of analyses, including venom gland transcriptomics,in vitroenzymatic assays and reverse phase chromatography to comparatively analyseB. arietansvenoms originating from Nigeria, Tanzania, and South Africa. Immunological assays andin vitroenzymatic neutralisation assays were then applied to investigate the impact of venom variation on the potential efficacy of three antivenom products; SAIMR Polyvalent, EchiTAb-Plus and Fav-Afrique.FindingsThrough the first comparison of venom gland transcriptomes ofB. arietansfrom three geographically distinct regions (Nigeria, Tanzania, and South Africa), we identified substantial variation in toxin expression. Findings of venom variation were further supported by chromatographic venom profiling, and the application of enzymatic assays to quantify the activity of three pathologically relevant toxin families. However, the use of western blotting, ELISA, andin vitroenzymatic inhibition assays revealed that variation withinB. arietansvenom does not appear to substantially impact upon the efficacy of three African polyvalent antivenoms.ConclusionsThe large distribution and medical importance ofB. arietansmakes this species ideal for understanding venom variation and the impact this has on therapeutic efficacy. The findings in this study highlight the likelihood for considerable venom toxin variation across the range ofB. arietans,but that this may not dramatically impact upon the utility of treatment available in the region.Author SummaryThe puff adder (Bitis arietans) is found across sub-Saharan Africa and the Arabian Peninsula and is capable of causing life threatening pathology due to its potent venom. The extensive range ofB. arietansexposes populations to different ecological pressures which may impact upon the composition of venom toxins. In this study, we examined the venom composition ofB. arietansfrom three countries separated by large geographic distance: Nigeria, Tanzanian and South Africa. By integrating venom gland transcriptomes, venom chromatography, andin vitrofunctional assays to profileB. arietansvenom composition, we uncovered extensive variation between the three locales. Given that venom variation can have a significant impact on the efficacy of antivenom treatment, we also investigated the ability of three African antivenoms to recognise and inhibitin vitrovenom activity. Through these analyses, we were able to determine that venom variation did not have a substantial impact on the neutralising effect of selected antivenoms. This study has highlighted the potentially extensive venom variation found across the range ofB. arietansand initiated valuable investigations into the efficacy of African antivenoms to protect human populations vulnerable to snakebite envenoming.