Naja annulifera Snake: New insights into the venom components and pathogenesis of envenomation

Background Naja annulifera is a medically important venomous snake occurring in some of the countries in Sub-Saharan Africa. Accidental bites result in severe coagulation disturbances, systemic inflammation and heart damage, as reported in dogs, and death, by respiratory arrest, in humans. Despite the medical importance of N. annulifera, little is known about its venom composition and the pathogenesis of envenomation. In this paper, the toxic, inflammatory and immunogenic properties of N. annulifera venom were analyzed. Methodology/Principal findings Venom proteomic analysis identified 79 different proteins, including Three Finger Toxins, Cysteine Rich Secretory Proteins, Metalloproteinases, Phospholipases A(2) (PLA(2)), Hyaluronidase, L-amino-acid oxidase, Cobra Venom Factor and Serine Proteinase. The presence of PLA(2), hyaluronidase, fibrinogenolytic and anticoagulant activities was detected using functional assays. The venom was cytotoxic to human keratinocytes. In an experimental murine model of envenomation, it was found that the venom induced local changes, such as swelling, which was controlled by anti-inflammatory drugs. Moreover, the venom caused death, which was preceded by systemic inflammation and pulmonary hemorrhage. The venom was shown to be immunogenic, inducing a strong humoral immune response, with the production of antibodies able to recognize venom components with high molecular weight and to neutralize its lethal activity. Conclusions/Significance The results obtained in this study demonstrate that N. annulifera venom contains toxins able to induce local and systemic inflammation, which can contribute to lung damage and death. Moreover, the venom is immunogenic, an important feature that must be considered during the production of a therapeutic anti-N. annulifera antivenom. Author summary N. annulifera is a dangerous snake that belongs to the Elapidae family. It is found in some of the countries in Sub-Saharan Africa and has caused accidents in humans and dogs. In this study, we characterized some of the biochemical, toxic and immunogenic properties of N. annulifera venom. We showed that the venom is composed of several proteins, some of which display enzymatic activities, such as phospholipase A2, hyaluronidase, metalloproteinases and serine proteinases. The venom promoted disturbances in the human coagulation system and was cytotoxic to human epidermal cells. Using a mouse model, we showed that the venom promotes local reactions that were reduced with anti-inflammatory drugs. The venom caused systemic inflammation, lung hemorrhage and death. Further, the venom stimulated production of high antibody titers when injected into mice and the antiserum produced was able to inhibit venom-induced death. This study demonstrated that N. annulifera venom contains toxins that trigger inflammatory process, which may contribute to the envenomation pathology. Moreover, the venom is immunogenic, an important aspect for the production of an efficient N. annulifera antivenom. (AU)