Evaluation of candidate monovalent Bitis arietans antivenom using eggs from hyperimmunized layer chickens in Zaria Kaduna State, Nigeria

Abstract

Aim: This research sought to experiment on a cheaper alternative to snake antivenom production using hyperimmunized chicken eggs. Methods: Twenty-two Swiss albino mice were used to determine the median lethal dose (LD50) and lethality neutralization assay. Twenty-layer chickens, aged 17-weeks-old were divided into two groups of 10 chickens each. Based on body weight, group A were hyperimmunized with 1/5th LD50 of crude Bitis arietans venom in adjuvant (Montanide™ ISA 201) at two-weeks interval over eight-weeks period; while group B were administered normal saline. Using polyethylene glycol 6000, immunoglobulin Y was extracted from 15 pooled egg yolks. Twelve Wistar rats were used for the haemorrhagic and anti-haemorrhagic assays. The haemolytic activity of the venom and anti-haemolytic effect of the candidate antivenom was determined in vitro using caprine erythrocytes. Results: The LD50 of the pooled Bitis arietans venom was 0.9 mg/kg. Agar gel immunodiffusion test confirmed the presence of precipitin antibodies in the candidate antivenom. EchiTab plus-ICP® neutralized lethal effect of all dilutions and the stock (10 mg/ml) venom of Bitis arietans while the candidate antivenom was limited to 3log2 dilution of the stock venom (1.25 mg/ml). In the haemorrhagic assay the venom exhibited haemorrhagic activity at 2.5, 5 and 10 mg/ml and the highest anti-haemorrhagic activity of the candidate antivenom at 10mg/ml. The candidate antivenom exhibited significantly higher (p < 0.05) anti-haemolytic activity against the venom of Bitis arietans at 10 mg/ml. Conclusion: The antibodies have inhibitory properties against the lethal, haemorrhagic and haemolytic properties of Bitis arietans venom.