Brown Snake Antivenom Research Articles

Nephelometry vs turbidimetry in transferrin measurement

Immunoturbidimetry studies the phenomenon of immunoprecipitation of antigens and antibodies in solution, where there is the formation of large, polymeric insoluble immunocomplexes that increase the turbidity of the solution. We used immunoturbidimetry to investigate the interaction between commercial snake antivenoms and snake venoms, as well as cross-reactivity between different snake venoms.Serial dilutions of commercial snake antivenoms (100 μl) in water were placed in the wells of a microtitre plate and 100 μl of a venom solution (50 μg/ml in water) was added. Absorbance readings were taken at 340 nm every minute on a BioTek ELx808 plate reader at 37 °C. Limits imposed were a 30 minute cut-off and 0.004 as the lowest significant maximum increase. Reactions with rabbit antibodies were carried out similarly, except that antibody dilutions were in PBS.Mixing venom and antivenom/antibodies resulted in an immediate increase in turbidity, which either reached a maximum or continued to increase until a 30 minute cut-off. There was a peak in absorbance readings for most Australian snake venoms mixed with the corresponding commercial antivenom, except for Pseudonaja textilis venom and brown snake antivenom. There was cross-reactivity between Naja naja venom from Sri Lanka and tiger snake antivenom indicated by turbidity when they were mixed. Mixing rabbit anti-snake antibodies with snake venoms resulted in increasing turbidity, but there was not a peak suggesting the antibodies were not sufficiently concentrated. The absorbance reading at pre-determined concentrations of rabbit antibodies mixed with different venoms was able to quantify the cross-reactivity between venoms. Indian antivenoms from two manufacturers were tested against four Sri Lankan snake venoms (Daboia russelli, N. naja, Echis carinatus and Bungarus caeruleus) and showed limited formation of immunocomplexes with antivenom from one manufacturer.The turbidity test provides an easy and rapid way to compare and characterise interactions between antivenoms and snake venoms.

Antivenom production in the alpaca (Vicugna pacos): Monovalent and polyvalent antivenom neutralisation of lethal and procoagulant toxins in Australian elapid venoms

Alpaca (Vicugna pacos), like all other members of the camelid family, produce a unique heavy chain only IgG subclass. This form of IgG has been proposed as having superior neutralising properties and improved safety as compared with conventional mammalian IgG-based immunotherapeutics. The objective of this study was to investigate the potential of alpaca antibodies to neutralise the lethal and procoagulant properties of several Australian elapid snake venoms. Various experimental antivenoms were produced by immunisation of alpaca (Vicugna pacos) with combinations of five Australian elapid snake venoms: tiger snake (Notechis scutatus), eastern brown snake (Pseudonaja textilis), Papuan taipan (Oxyuranus scutellatus canni), mulga snake (Pseudechis australis) and common death adder (Acanthophis antarcticus). Venoms were emulsified in Freund’s Incomplete Adjuvant and alpaca repeatedly immunised. Having previously shown that such alpaca produce persistent venom-specific antibody responses with minimal physiological impact on the animal, the current report demonstrates that polyvalent alpaca antivenom neutralises the lethal effects of three elapid venoms (tiger, brown and Papuan taipan venoms). Such lethality neutralisation was, however, of generally lower potency, but of the same order of magnitude, when compared with bioCSL monovalent antivenoms (brown, tiger and taipan antivenoms). By contrast, the monovalent alpaca tiger and brown snake antivenom were found to be superior to the equivalent bioCSL monovalent antivenoms when considering the neutralisation of the procoagulant effects of the respective immunising venoms. The bioCSL polyvalent antivenom also less potent, when compared with the alpaca taipan monovalent antivenom, against Papuan taipan venom procoagulant neutralisation. These findings support the potential of camelid immunoglobulin for the development of novel elapid specific antivenoms.

Incidence of serum sickness after the administration of Australian snake antivenom (ASP-22)

Context: Serum sickness is a delayed immune reaction resulting from the injection of foreign protein or serum. Antivenom is known to cause serum sickness but the incidence and characteristics are poorly defined. Objective: To investigate the incidence and clinical features of serum sickness following the administration of Australian snake antivenoms. Materials and methods: This was a prospective cohort study of patients recruited to the Australian Snakebite Project who received snake antivenom from November 2012 to March 2014. Demographics, clinical information, laboratory tests and antivenom treatment were recorded prospectively. Patients administered antivenom were followed up at 7–10 days and 6 weeks’ post-antivenom. The primary outcome was the proportion with serum sickness, pre-defined as three or more of: fever, erythematous rash/urticaria, myalgia/arthralgia, headache, malaise, nausea/vomiting 5–20 days post-antivenom. Results: During the 16-month period, 138 patients received antivenom. 23 were not followed up (unable to contact, tourist, child, bee sting) and 6 died in hospital. Of 109 patients followed up, the commonest reason for antivenom was venom induced consumption coagulopathy in 77 patients. An acute systemic hypersensitivity reaction occurred post-antivenom in 25 (23%) and 8 (7%) were severe with hypotension. Serum sickness occurred in 32/109 (29%) patients, including 15/37 (41%) given tiger snake, 6/15 (40%) given polyvalent and 4/23 (17%) given brown snake antivenom. There was no association between the volume of antivenom and serum sickness, p = 0.18. The commonest effects were lethargy, headache, muscle/joint aches and fever. Discussion: The incidence of serum sickness after snake antivenom in Australia was higher than earlier investigations which failed to define symptoms or follow-up patients, but similar to more recent studies of antivenoms in the United States. Conclusion: Serum sickness is common with Australian snake antivenom but does not appear to be predictable based on the volume of antivenom administered.

An instructive case of presumed brown snake (Pseudonaja spp.) envenoming

Context. Several species of medically important Australian elapid snakes are frequently involved in human envenoming. The brown snake group (Pseudonaja spp., 9 species) is most commonly responsible for envenoming including life-threatening or fatal cases. Several Pseudonaja spp. can inflict human envenoming that features minor local effects, but may cause serious systemic venom disease including defibrination coagulopathy, thrombocytopenia, micro-angiopathic hemolytic anemia (MAHA) and, rarely, paralysis. Pseudonaja envenoming is typically diagnosed by history, clinical assessment including occasional active clinical bleeding noted on physical examination (e.g. from venipuncture sites, recent cuts, etc.), and laboratory detection of coagulopathy (prolonged activated partial thromboplastin time [APTT]/INR, elevated D-dimer, afibrinogenemia and thrombocytopenia). Lack of verified identity of the envenoming snake species is a common problem in Australia and elsewhere. Identification and confirmation of the envenoming Australian snake taxon is often attempted with enzyme sandwich immunoassay venom detection kits (SVDKs). However, the SVDK has limited utility due to unreliable specificity and sensitivity when used to detect venoms of some Australian elapids. Antivenom (AV) remains the cornerstone of treatment, although there is debate concerning the recommended dose (1 vs. 2 or more vials) necessary to treat serious Pseudonaja envenoming. Envenomed patients receiving timely treatment uncommonly succumb, but a proportion of seriously envenomed patients may exhibit clinical or laboratory evidence of myocardial insult. Case details. An 88-year-old woman presented her dog to a veterinarian after it had sustained a bite by a witnessed snake, reportedly an eastern brown snake (Pseudonaja textilis, Elapidae). The woman became suddenly confused, and lost consciousness at the veterinary office. After transport to hospital, she denied any contact with the snake, but developed large haematomas at intravenous (i.v.) catheter insertion sites; blood tests revealed a severe defibrination coagulopathy, consistent with envenoming by a brown snake. An SVDK-tested urine sample was negative. A non-contrast CT of her head showed a minor subacute infarction of the left corona radiata. A twelve-lead ECG was normal, but her troponins were mildly elevated (39 ng/L). A diagnosis of brown snake envenoming was made and she received 2 vials of brown snake AV i.v., without adverse incident. Thirty min post AV her Glasgow Coma Score (GCS) had improved from 13 to 15 (normal). At 3.5 h post AV all bleeding from i.v. sites ceased, although her troponin T level peaked at 639 ng/L, supporting a diagnosis of non-ST elevated myocardial infarction (NSTEMI). Discussion. Severe brown snake envenoming may occur in the absence of a perceived bite, and AV is temporally associated with improvement in clinical findings and coagulopathy. However, severe envenoming by this species can be complicated by cardiovascular events that in the circumstance of incomplete or absent history may confuse the primary diagnosis and affect patient outcome.

Use of immunoturbidimetry to detect venom–antivenom binding using snake venoms

IntroductionImmunoturbidimetry studies the phenomenon of immunoprecipitation of antigens and antibodies in solution, where there is the formation of large, polymeric insoluble immunocomplexes that increase the turbidity of the solution. We used immunoturbidimetry to investigate the interaction between commercial snake antivenoms and snake venoms, as well as cross-reactivity between different snake venoms. MethodsSerial dilutions of commercial snake antivenoms (100μl) in water were placed in the wells of a microtitre plate and 100μl of a venom solution (50μg/ml in water) was added. Absorbance readings were taken at 340nm every minute on a BioTek ELx808 plate reader at 37°C. Limits imposed were a 30minute cut-off and 0.004 as the lowest significant maximum increase. Reactions with rabbit antibodies were carried out similarly, except that antibody dilutions were in PBS. ResultsMixing venom and antivenom/antibodies resulted in an immediate increase in turbidity, which either reached a maximum or continued to increase until a 30minute cut-off. There was a peak in absorbance readings for most Australian snake venoms mixed with the corresponding commercial antivenom, except for Pseudonaja textilis venom and brown snake antivenom. There was cross-reactivity between Naja naja venom from Sri Lanka and tiger snake antivenom indicated by turbidity when they were mixed. Mixing rabbit anti-snake antibodies with snake venoms resulted in increasing turbidity, but there was not a peak suggesting the antibodies were not sufficiently concentrated. The absorbance reading at pre-determined concentrations of rabbit antibodies mixed with different venoms was able to quantify the cross-reactivity between venoms. Indian antivenoms from two manufacturers were tested against four Sri Lankan snake venoms (Daboia russelli, N. naja, Echis carinatus and Bungarus caeruleus) and showed limited formation of immunocomplexes with antivenom from one manufacturer. DiscussionThe turbidity test provides an easy and rapid way to compare and characterise interactions between antivenoms and snake venoms.

Clinical Effects and Antivenom Dosing in Brown Snake (Pseudonaja spp.) Envenoming — Australian Snakebite Project (ASP-14)

BackgroundSnakebite is a global health issue and treatment with antivenom continues to be problematic. Brown snakes (genus Pseudonaja) are the most medically important group of Australian snakes and there is controversy over the dose of brown snake antivenom. We aimed to investigate the clinical and laboratory features of definite brown snake (Pseudonaja spp.) envenoming, and determine the dose of antivenom required.Methods and FindingThis was a prospective observational study of definite brown snake envenoming from the Australian Snakebite Project (ASP) based on snake identification or specific enzyme immunoassay for Pseudonaja venom. From January 2004 to January 2012 there were 149 definite brown snake bites [median age 42y (2–81y); 100 males]. Systemic envenoming occurred in 136 (88%) cases. All envenomed patients developed venom induced consumption coagulopathy (VICC), with complete VICC in 109 (80%) and partial VICC in 27 (20%). Systemic symptoms occurred in 61 (45%) and mild neurotoxicity in 2 (1%). Myotoxicity did not occur. Severe envenoming occurred in 51 patients (38%) and was characterised by collapse or hypotension (37), thrombotic microangiopathy (15), major haemorrhage (5), cardiac arrest (7) and death (6). The median peak venom concentration in 118 envenomed patients was 1.6 ng/mL (Range: 0.15–210 ng/mL). The median initial antivenom dose was 2 vials (Range: 1–40) in 128 patients receiving antivenom. There was no difference in INR recovery or clinical outcome between patients receiving one or more than one vial of antivenom. Free venom was not detected in 112/115 patients post-antivenom with only low concentrations (0.4 to 0.9 ng/ml) in three patients.ConclusionsEnvenoming by brown snakes causes VICC and over a third of patients had serious complications including major haemorrhage, collapse and microangiopathy. The results of this study support accumulating evidence that giving more than one vial of antivenom is unnecessary in brown snake envenoming.

Clinical effects and treatment of envenoming by Hoplocephalus spp. snakes in Australia: Australian Snakebite Project (ASP-12)

There is limited information on envenoming by snakes of the genus Hoplocephalus from Eastern Australia. We investigated the clinical and laboratory features of patients with definite Hoplocephalus spp. bites including antivenom treatment, recruited to the Australian Snakebite Project. There were 15 definite Hoplocephalus spp. bites based on expert identification including eight by Hoplocephalus stephensi (Stephen’s banded snakes), four by Hoplocephalus bungaroides (broad-headed snake) and three by H. bitorquatus (pale-headed snake). Envenoming occurred in 13 patients and was similar for the three species with venom induced consumption coagulopathy (VICC) in all envenomings. Seven patients had an INR >12 and partial VICC, with only incomplete fibrinogen consumption, occurred in three patients. Systemic symptoms occurred in eight patients. Myotoxicity and neurotoxicity did not occur. H. stephensi venom was detected in all three H. stephensi envenomings (1.1, 44 and 81 ng/mL) for whom pre-antivenom blood samples were available, and not detected in one without envenoming. In two cases with post-antivenom blood samples, venom was not detected after tiger snake antivenom (TSAV) was given. In vitro binding studies demonstrated that TSAV concentrations of 50mU/mL are sufficient to bind the majority of free H. stephensi venom components at concentrations above those detected in envenomed patients (100 ng/mL). Eleven patients received antivenom, median dose 2 vials (Range: 1 to 5 vials), which was TSAV in all but one case, where polyvalent antivenom was used. Immediate hypersensitivity reactions occurred in six cases including one case of anaphylaxis. Envenoming by Hoplocephalus spp. causes VICC and systemic symptoms, making it clinically similar to brown snake (Pseudonaja spp.) envenoming. Based on in vitro studies reported here, patients may be treated with one vial of TSAV, although one vial of brown snake antivenom may also be sufficient.

Extradural haematoma secondary to brown snake (Pseudonaja species) envenomation

A 4-year-old Siberian Husky dog was treated with brown snake antivenom by his regular veterinarian after a witnessed episode of brown snake envenomation. The dog was discharged 5 hours post presentation despite an ongoing coagulopathy. The dog was presented to the emergency centre 2 hours later because the owner believed the dog to be in pain. Initial examination revealed an ambulatory but neurologically normal patient with thoracolumbar pain and laboratory evidence of a coagulopathy. Despite correction of the coagulopathy, the signs progressed to bilateral hind limb paresis after approximately 3 hours of hospitalisation, and continued to deteriorate over the next 56 hours to loss of deep pain perception in the right hind limb. Computed tomography imaging identified the presence of an extradural haematoma which was subsequently removed via a hemilaminectomy. Surgical decompression was successful in treating the spinal compression and the dog recovered with minimal complications. To our knowledge this is the first report of extradural haematoma secondary to coagulopathy induced by brown snake envenomation.

Current use of Australian snake antivenoms and frequency of immediate‐type hypersensitivity reactions and anaphylaxis

To investigate current use of Australian snake antivenoms and the frequency and severity of immediate-type hypersensitivity reactions. Nested prospective cohort study as part of the Australian Snakebite Project. Patients receiving snake antivenom in Australian hospitals between 1 January 2002 and 30 November 2007. The use of CSL Limited antivenom; frequency and severity of hypersensitivity reactions to antivenom; premedication and treatment of these reactions. Snake antivenom was administered to 195 patients, mostly for venom-induced consumption coagulopathy (145 patients, 74%), followed by non-specific systemic effects (12%), neurotoxicity (5%) and myotoxicity (4%). Antivenom was given to nine patients (5%) without evidence of envenoming or who were bitten by a species of snake for which antivenom is not required. The commonest antivenoms used were brown snake (46%), tiger snake (30%) and polyvalent (11%). The median dose was four vials (interquartile range, 2-5 vials), and 24 patients received two different types of antivenom. Immediate-type hypersensitivity reactions occurred in 48 patients (25%); 21 satisfied our definition of anaphylaxis, with 11 moderate and 10 severe cases, including nine in which patients were hypotensive. The remaining 27 reactions were mild (skin only). Adrenaline was used in 26 cases with good effect. The frequency of reactions to tiger snake (41%) and polyvalent (41%) antivenoms was higher than that to brown snake antivenom (10%). Hypersensitivity reactions occurred in 11 of 40 patients receiving any form of premedication (28%) and in 2 of 11 given adrenaline for premedication (18%) versus 20 of 86 not receiving premedication (23%). Antivenom was used appropriately, and most commonly for coagulopathy. Hypersensitivity reactions were common, but most were not severe. The discretionary use of premedication was not associated with any reduction in reactions.

Efficacy of antivenom against the procoagulant effect of Australian brown snake ( Pseudonaja sp.) venom: In vivo and in vitro studies

Snake venom induced consumption coagulopathy (VICC) is a common complication of snake bite due to prothrombin activators or thrombin-like enzymes in the venom. This study aimed to determine the efficacy and dose of antivenom for treating VICC in patients envenomed by brown snakes ( Pseudonaja spp.), including in vitro coagulation studies. In serial blood samples from patients with brown snake envenoming, venom and antivenom concentrations were measured using enzyme immunoassays. In vitro mixtures of brown snake venom and antivenom were used to investigate antivenom binding, neutralisation of prothrombin activity, prevention of venom-mediated clotting and effect on thrombin generation parameters using a thrombinoscope. In 27 envenomed patients the median venom concentration was 20 ng/mL (Interquartile range[IQR]:12–44 ng/mL) prior to antivenom and was not detected after antivenom administration, including 9 patients given one vial. In vitro, 200 μg/mL of antivenom bound all free venom at venom concentrations seen in patients. In vitro prothrombinase activity of the venom (using a chromogenic substrate) was not neutralised by antivenom. However, for venom concentrations seen in humans, 100 μg/mL of antivenom prevented venom clotting activity in human plasma and 479 μg/mL neutralised procoagulant venom activity measured by triggering thrombin generation. One vial of antivenom appears to be sufficient to bind and neutralise all venom in patients with severe brown snake envenoming.

Treatment of snakebite in Australia: The current evidence base and questions requiring collaborative multicentre prospective studies

Despite the wealth of anecdotes and case reports there are fundamental questions of management of snakebite in Australia that remain unresolved or for which the current evidence is limited. The efficacy in the field, potential limitations and possibility of improvements in pressure immobilisation first aid need objective studies in humans. Optimal bandage sizes, stretch and pressure for different sized limbs need further evaluation, as does the use of pressure pads. Better definitions of specific clinical envenoming syndromes attributable to individual snake species are required, including elucidation of within-genus variations, similarities and differences. Venom studies suggest this is especially important for species within the brown snake ( Pseudonaja) and death adder ( Acanthophis) genera. Appropriate antivenom types, doses and dosing intervals for individual snake species should be more formally studied in patients. Especially important are confirmation of the need for higher doses of brown snake antivenom, while possibly limiting unnecessarily high doses, confirmation of the critical importance of early antivenom use to prevent pre-synaptic neurotoxicity in Taipan and tiger snake bites and ascertainment of whether larger doses of antivenom are unhelpful in Taipan bites after specified time delays. Confirmation of clinical efficacy and dosing recommendations for use of tiger snake ( Notechis) antivenom in envenoming from Australian copperhead ( Austrelaps spp.), broad headed ( Hoplocephalus spp.) and rough-scaled snakes ( Tropidechis carinatus) also require formal study in patients. Other examples of clinical relevance of cross-specificity of current and future monospecific antivenoms and whether there are geographical variations in antivenom responses within species will require elucidation. Prospective multicentre collaborative studies with predefined data collection and serial venom level assays are proposed as the way forward in Australia to help resolve therapeutic uncertainties and to establish a firmer evidence base for best-practice treatment guidelines for Australasian elapid snakebite.

Enzyme immunoassays in brown snake ( Pseudonaja spp.) envenoming: Detecting venom, antivenom and venom–antivenom complexes

Although a commercial snake venom detection kit (SVDK) is available to distinguish between the five major snake groups in Australia, there is no assay for quantifying venom or antivenom concentrations in envenomed patients. Serum samples were obtained from patients with brown snake ( Pseudonaja spp.) envenoming before and after the administration of antivenom and patients with suspected brown snake bites but no evidence of envenoming. Enzyme immunoassays (EIAs) were developed for free venom, free antivenom and the venom–antivenom complex. Standard samples measured in duplicate had a coefficient of variation of less than 10%. The EIA for venom was able to detect brown snake venom down to concentrations of 3 ng/mL. A high baseline absorbance was measured in some patients that did not change with the addition of excess antivenom to the samples. In these patients, the baseline absorbance was subtracted from all measurements to calculate the true venom concentration. The EIA for brown snake antivenom had a limit of detection of 20 μg/mL, but 50 μg/mL was used as a cut-off based on assays in patients who had not received antivenom. The EIA for venom–antivenom complexes was unable to detect these at the low venom concentrations that occurred in patients. Quantification of venom and antivenom will help to determine the dose of antivenom required to bind venom and to establish appropriate end points for antivenom treatment.

Toxins not neutralized by brown snake antivenom

The Australian snakes of the genus Pseudonaja (dugite, gwardar and common brown) account for the majority of snake bite related deaths in Australia. Without antivenom treatment, the risk of mortality is significant. There is an accumulating body of evidence to suggest that the efficacy of the antivenom is limited. The current study investigates the protein constituents recognized by the antivenom using 2-DE, immuno-blot techniques and rat tracheal organ bath assays. The 2-DE profiles for all three snake venoms were similar, with major species visualized at 78-132 kDa, 32-45 kDa and 6-15 kDa. Proteins characterized by LC-MS/MS revealed a coagulant toxin ( approximately 42 kDa) and coagulant peptide ( approximately 6 kDa), as well as two PLA(2) ( approximately 14 kDa). Peptides isolated from approximately 78 kDa and 15-32 kDa protein components showed no similarity to known protein sequences. Protein recognition by the antivenom occurred predominantly for the higher molecular weight components with little recognition of 6-32 kDa MW species. The ability of antivenom to neutralize venom activity was also investigated using rat tracheal organ bath assays. The venoms of Pseudonaja affinis affinis and Pseudonaja nuchalis incited a sustained, significant contraction of the trachea. These contractions were attributed to PLA(2) enzymatic activity as pre-treatment with the PLA(2) inhibitor 4-BPB attenuated the venom-induced contractions. The venom of Pseudonaja textilis incited tracheal contractility through a non-PLA(2) enzymatic activity. Neither activity was attenuated by the antivenom treatment. These results represent the first proteomic investigation of the venoms from the snakes of the genus Pseudonaja, revealing a possible limitation of the brown snake antivenom in binding to the low MW protein components.

ANTIVENOM DEVELOPMENT IN AUSTRALIA

Brown snakes, Pseudonaja genus, cause more bites and deaths to animals and humans in Australia than any other terrestrial snake genus. Some aspects of treatment of brown snakebites with antivenom remain poorly managed due to the apparent inability of the antivenom to counter the prothrombin activator in the venom. We present evidence of a new novel antivenom (Antiven Pty Ltd Brown snake antivenom (ABSAV)), which shows high efficacy in reversing the hemostatic abnormality caused by brown snake envenomation. It is also more efficient at reversing overall toxicity. In clotting tests, it is substantially more potent than Commonwealth Serum Laboratories Ltd (CSL) Brown snake antivenom and is twice as effective in reversing overall toxicity than CSL Brown snake antivenom.Antiven Pty. Ltd. Brown snake antivenom is currently being assessed by the National Registration Authority (veterinary regulative body in Australia).

Why do people still die from brown-snake bites?

Brown snakes are the leading cause of snakebites and snakebite fatalities in Australia and the incidence may be rising. Most fatalities are the result of early events after the bite, often a general or cardiac collapse. Lack of appropriate first aid is a common finding. It is probable that an early thrombotic effect of venom procoagulants, prior to defibrination, is responsible for the cardiac collapse in most of these cases. CSL brown-snake antivenom, while not perfect, is usually effective if given in sufficient quantity, but cannot reverse established vital-organ damage. Increased public education about the danger of snakebite and correct first aid may reduce the death toll, and improved professional continuing education on snakebite is advisable but, ultimately, some brown-snake bite deaths are probably inevitable, despite optimum medical care.

Efficacy and Potency of Antivenoms in Neutralizing the Procoagulant Effects of Australian Snake Venoms in Dog and Human Plasma

The efficacy and potency of Commonwealth Serum Laboratories (CSL) snake antivenoms in neutralizing the procoagulant action of Australian snake venoms was studied in vitro. The procoagulant action of venoms from Tiger, Brown and Taipan Snakes were measured in pooled canine or pooled human plasma. Mixtures of the venoms and their appropriate antivenoms, ranging from 0.1 to 15 times the recommended neutralizing dose of antivenom were then added to pooled canine or pooled human plasma and the efficacy and potency of the antivenoms in preventing the procoagulant action was measured. Neutralization was achieved by the addition of the appropriate antivenom in concentrations of 0.5 (Taipan), 10 (Tiger), 10 (Gwardar), 15 (Dugite) and an estimated 20 (Common Brown) times the dose expected. Similar results were obtained in canine and human plasma. The potency of Tiger and especially Brown Snake antivenom is substantially lower (10-20 times) than that predicted by CSL. These findings support an increase in the dose of antivenom to be used for the treatment of Tiger and especially Brown Snake envenomation complicated by coagulopathy. The in vitro canine model may be suitable for studying the procoagulant effects of Australian snake venoms.

Comparative study on the procoagulant from the venom of Australian brown snakes (Elapidae; Pseudonaja spp.)

The procoagulant was isolated from the venom of four clinically significant species of brown snake; Pseudonaja affinis, P. inframacula, P. nuchalis, and P. textilis. HPLC elution profiles and PAGE showed a high degree of homology between the procoagulants from the four species. Antiserum from the CSL Ltd (Brown Snake Antivenom) produced a single band against all four procoagulants. The specific activity of the procoagulant varied between species, while inhibitory studies indicated that the procoagulants were serine proteases with a sialic acid component which also contributes to the coagulant action.

Variation in the composition of the venom from a single specimen of Pseudonaja textilis (common brown snake) over one year

Venom was collected over a 12-month period from a single specimen of Pseudonaja textilis. No trends in the weight of venom extracted or protein content were noted. Amidolytic activity, coagulant and esterolytic activity showed a decrease over the summer months, while a rise in phosphodiesterase and 5′ nucleotidase activity was noted at this time. Precipitin lines developed against the venoms by brown snake antivenom showed variability over the 12-month period. Gel filtration and SDS-polyacrylamide gel electrophoresis profiles showed quantitative and minor qualitative differences; however, the variation noted in the activities of these venoms was not predictable from these profiles.

Ten years of snake bites at Fremantle Hospital

Seventy-six patients (30% children) were admitted to Fremantle Hospital over 10 years with suspected snake bite. Twenty-nine patients were definitely bitten, with 26 bites being witnessed. Of the 13 patients definitely envenomated, 11 had a coagulopathy although seven were asymptomatic; four other patients may have been envenomated. The dugite (Pseudonaja affinis) was probably responsible for most envenomations. Eleven of the 13 envenomated patients received antivenom (six brown snake, four polyvalent and one tiger snake antivenom). The patient envenomated by the tiger snake (Notechis ater occidentalis), a 13-year-old girl, was initially incorrectly treated with brown snake antivenom at a country hospital, and did not receive appropriate antivenom until 50 hours after the bite. She developed profound paralysis, rhabdomyolysis and renal failure, and required prolonged ventilation during her 53-day hospital admission, but survived without disability. Snake bite wounds should not be washed, so that venom can be identified from the wound. Attempts to kill snakes are dangerous, often leading to bites on the fingers. First aid measures of a pressure bandage and immobilisation, used in 13 of the 29 cases (45%), should be more widely publicised.

Variation in venom composition and reactivity in two specimens of yellow-faced whip snake (Demansia psammophis) from the same geographical area

V. Williams and J. White. Variation in venom composition and reactivity in two specimens of yellow-faced whip snake ( Demansia psammophis) from the same geographical area. Toxicon 28, 1351–1354, 1990.—Gel filtration chromatography and SDS-PAGE of venom from two specimens of Demansia psammophis showed little similarity. Amidolytic activity of the venoms, however, was in the same order of reactivity against various chromogenic substrates. The venom from both snakes produced precipitin lines with brown snake antivenom but the venom detection kit (Commonwealth Serum Laboratories) identified one venom as brown snake ( Pseudonaja sp.) and the other as tiger snake ( Notechis sp.). These results raise questions about the phylogeny of this species.