This thesis embodies experimental studies relating to human and animal envenomation by Australian elapid snakes and the Australian paralysis tick, Ixodes holocyclus.
Epidemiologically, snakebite is a considerable problem worldwide. Australian snakes are among the most dangerous with respect to venom toxicity. Australian venoms have been shown to possess neurotoxic, myotoxic, cardiotoxic and coagulopathic activities.
Venoms have been studied using biochemical techniques for a number of years. In the 1970's new techniques in.radio-immunoassays and enzyme immunoassays were developed. These afforded the tools for toxinological studies of venomous creatures of practical medical importance. In this thesis, immunological tests have been used to study venom injection, degradation, and movement in situations relevant to human snakebite and tick envenomation.
An important consideration in studying snakebite is the quantity of venom delivered when a snake bites. Experimental models were developed to resolve this question. Detailed studies described the mass, range, and delivery patterns of venom delivered during both hunting (aggressive) snakebite, and during defensive bites. The latter are relevant to the situation in human snake bite. For defensive bites, the quantity of venom delivered was: Tiger Snake (Notechis scutatus). 6.8 mg; Taipan (Oxyuranus scutellatus), 30.8 mg; Eastern Brown Snake (Pseudonaja textilis). 3.8 mg; Rough-scaled Snake (Tropidechis carinatus), 19.9 mg. A significant percentage of the total venom delivered remained on the skin at the bite site and is an important source of venom for clinical venom detection.
A new experimental model (a simulated human hand of agar) to study snakebite was developed. Using this technique, venom delivery patterns for medically significant Australian Elapids are described. No significant differences between "defensive" and "aggressive" bites was observed. Venom delivery in a sequence of bites is described. The concept of biting efficiency (ratio of injected skin split venom) was developed and a series of experiments concluded to provide data by species and by bite number in a sequence.
Experiments were performed to test the antigenic stability and physical properties of three Australian snake venoms - Tiger ( Notechis scutatus) , Taipan (Oxyuranus scutellatus) , and Rough-scaled (Tropidechis carinatus) - which have implications for field use, transport , and laboratory procedures relating to immunological characterization of Australian snake venoms. There is significant immunological stability at temperatures of 20°, 37° and 55°C. The venoms are also stable when incubated in muscle, skin and fat homogenates for 24 hours. No loss of detectable venom occurs over the pH range of 5-10. At pH < 4 and > 10, only 30% of venom is detected after 30 minutes incubation. Neither sonication nor freeze-thawing destroys venom detectability. Venom is significantly absorbed onto dacron swabs, with only 13% being detected after 48 hours incubation at 20°C. Venom is fairly robust and easily detected in urine. However serum undiluted appears to interfere with the assaying of venom.
The pressure/immobilization first-aid procedure was studied in two cases of snakebite and found to be effective in retarding the movement of venom. The venom of the Rough-scaled Snake (Tropidechis carinatus) has been studied in detail. The venom has immunological similarity with the Tiger Snake. It was shown by liquid chromatography, high pressure liquid chromatography, sodium dodecylsulphate and discontinuous polyacrylamide gel electrophoresis, and isoelectric focussing-to contain at least five major components. Of these, at least two are neurotoxins both with some rhabdomyolytic activity. One (mw. 10, 000) comprises the major toxic component of the venom and has presynaptic and postsynaptic activity as measured using in vitro electrophysiological techniques. The venom has potent pro-coagulant activity - an incomplete prothrom-bin activator similar to the activity in Tiger Snake (Notechis scutatus) venom.
In vivo, using whole animals and assays specific for individual sub-fractions, it was shown that all components are rapidly cleared from the bloodstream. At least two also affect peripheral nerve sensitivity.
Human cases of Rough-scaled Snake envenomation have been studied in detail. All cases show severe coagulopathy which sometimes masked neurotoxic or myolytic effects.
The enzyme immunoassay technique has been used to study the venom of the paralysis tick (Ixodes holocyclus) . Two assays were developed. The first was a sandwich assay capable of detecting toxins at concentrations of 0.1 wg/ml in the tissues and body fluids of experimentally infested animals. An indirect enzyme immunoassay, capable of detecting antibodies in the serum of immune or exposed animals was also developed. The correlation with in vivo neutralization assays was very good.
This method was studied in conjunction with studies of an artificial vaccine for the paralysis tick, Ixodes holocyclus. This vaccine was found to provide effective immunity against tick paralysis in beagle hounds.
A literature review, to place these studies in perspective is also presented. The implications of the study for the management of snakebite and proposed future experiments are also provided.