This study is the first to highlight the clinical sequelae and laboratory findings recorded during the clinical care and management of dogs envenomed by H.hypnale, one of the commonest venomous snakes in Sri Lanka. Consistent with studies of snakebite incidence from other venomous snakes this study observed a higher proportion of males suffered HNPV bites. It is hypothesised this reflects the more prominent territorial behaviour of male dogs [8, 22, 23]. This observation parallels the incidence of snakebite in human males in farming communities due to social norms governing occupational roles [4, 24].
The majority of HNPV bite victims were young dogs. It is speculated that younger dogs have a greater tendency than old dogs to sniff peculiar moving objects, although the reason is poorly understood but may be related to the development of recognition and aversion to threats. A study in 2016 drew a connection between this exploratory behaviour and the facilitation of allowing snakes to enter households and consequently heighten risks to humans . Another study indicates dogs may play a significant role as sentinels leading to avoidance of human snakebite . Correspondingly, the economically active age groups are the most susceptible to become HNPV bite victims in man [2, 4, 24]. Therefore, it is important to note that to identify hotspots of snakebite risk to both human and domestic animals a new concept called “citizen science and action ecology” has emerged based on digital and social innovations [26, 27]. Such exploration will help herpetologists to design and implement policies on prevention and treatment of snakebite in humans and animals.
In Sri Lanka H. hypnale is largely nocturnal  and the majority of bites recorded in this study occurred during darkness or the crepuscular hours. H. hypnale is most commonly found under stones, logs and leaf litter in and around human settlements. Therefore, the pattern of behaviour of both dog and HNPV increases the possibility of envenoming in dogs at dusk and dawn . By contrast the majority of HNPV bites in human are reported in day time in their home gardens . The preference for anthropogenic habitats by HNPV reflects the relationship of bite time and active hours of the day of both man and dog.
Fang marks among the majority of dog victims of N. naja and Dabia russelii [21, 22] are seen in the head and neck region and a similar bite distribution pattern was seen in the majority of HNPV bitten dogs. Sharp canine teeth have evolved as the major weapons for protection in dogs and they fight facing ahead. Dogs have a highly sensitive of olfaction; the detection and recognition of odorants plays an essential role in many adaptive dog behaviours, such as escape from danger, so they tend to sniff strange objects . Therefore the observation of fang marks around head and neck in the majority of snakebite victims is not unexpected. By comparison the vast majority of fang marks in humans are observed on either the feet or hands [2, 4, 30]. Failure to use protective measures when entering environments with poor visibility where snakes may dwell is instrumental in acquiring snakebite in man . The education of owners to restrict dogs’ activities, particularly young dogs, during the hours of darkness or twilight should significantly lessen the possibility of becoming a HNPV bite victim.
Our work describes, for the first time, the detection of local clinical signs of mild swelling, extensive swelling, hemorrhagic blistering and hemorrhagic bullae due to the venom of H.hypnale in all dogs. Zinc-dependent metalloproteases are almost as abundant as phospholipase A2 in H. hypnale venom and belong to ‘Pel protease’ subclass, and are another major toxin class of concern for clinic-pathological correlation . The venom metalloprotiases of H. hypnale venom have been recognized as capable of inducing local hemorrhage, activate prothrombin and cause fibrin (ogen)olysis [31–34]. PeI protease contributes to local tissue symptoms in H. hypnale envenoming in 90% of cases of man . The clinical manifestations are inflammation (swelling and pain), wound hemorrhage, blistering, and irreversible tissue destruction i.e. necrosis, and involve disruption of capillary vessel integrity, matrix degradation, dermal-epidermal junction separating and secretion of proinflammatory mediators ie leading to tissue ischemia, disrupted tissue repair, and a varied degree of tissue damages (e.g. dermonecrosis, myonecrosis) [1, 15, 31, 35]. Mild swelling and extensive swelling observed in this study are consistent with the observation of envenoming in dogs due to N. naja in Sri Lanka . Further similarity of varying degrees of swelling, haemorrhagic blisters, local tissue necrosis and regional lymphadenopathy are reported in different studies in human victims of HNPV bite envenoming [1–4, 15]. Mild and extensive swelling at the site of the bite resolved spontaneously without any adverse outcome as observed in previous studies in both man and animals [21, 36]. However, to address snakebite-induced local tissue damage and implement wound care, surgical management, including limb amputation on rare situations, had to be employed for HNPV envenomed dogs, as has been necessary for N.naja envenomed dogs . As well as palliative treatment measures, wound care, surgical intervention, skin grafting and occasional limb amputation, are practiced for HNPV envenomed human patients . There is thus, some evidence that early surgical intervention for such dogs is beneficial to minimize tissue necrosis, cost of treatment and to improve quality of life for dogs. However, in addition to the risk of mortality, a venomous bite by a member of Viperidae or Elapidae families also causes a severe local tissue trauma which frequently leads to amputation of the bitten limb to save the life of the victim . Therefore, surviving victims suffer a substantially-reduced quality of life due to the multiple effects of permanent physical and psychological disabilities. Hence, exploring the potential utility of small molecules as community-based therapeutics to prevent local tissue damage following snakebite will help to prevent and minimize the extent of local tissue damage.
The development of systemic clinical manifestations were fewer compared to local clinical signs in patients in this study. Severe systemic envenoming of neurotoxicity, coagulopathy and AKI were present in only approximately 20% of victims. Positive correlations between LH and extent of local tissue injury (rs = 0.78, p < 0 .0001); length of hospitalization and CT (rs = 1.0, p < 0.0001); PT and aPTT (rs = 0.47, p < 0 .00) were observed in dogs with coagulopathy. Therefore, care must be taken when managing patients with coagulopathy. The effects of HNPV venom on coagulation factors and renal tissues in this study contrasts with the pronounced effects of elevated PT, aPTT ,CT, BUN, creatinine and decreased concertation of fibrinogen. Therefore, it is crucial that all the HNPV bitten dogs are observed and have investigations done to assess the presence of coagulopathy or AKI. Accordingly, it is important to note that the more subtle effects of HNPV venom on the coagulation pathway and renal tissues may yet be found by more detailed biochemical analysis since the mechanism of AKI in HNPV envenoming remains unidentified .
In humans, systemic effects: nephrotoxicity, coagulopathy, thrombocytopenia and spontaneous haemorrhage, as well as less common effects (nausea, vomiting) are less common [6, 13, 38]. However, despite being rare the unpredictability of sporadic and potentially fatal systemic effects of HNPV envenoming confirms the significance of this species . Similar to the current study, systemic clinical signs of coagulopathy and AKI have been reported, although not frequently, in HNPV bite envenoming in humans [30, 39 ].
Although exact mechanism has yet to be identified, all the HNPV envenomed dogs showed Leukocytosis; which is consistent with N. naja bite envenoming ad D.russelii bite envenoming in dogs and humans. [21, 22, 40]. However, the cause of leukocytosis following such snakebite envenoming is yet to be explored [21, 22] and elucidation of this parameter would have the potential to open avenues to estimate the severity of envenoming .
Currently available AVS in Sri Lanka is imported from India and uses venom from the Indian species of Naja. naja, Bangarus caeruleus, D. russelii and Echis carinatus. HNPV, which is common in Sri Lanka and South East Asia is not included in the immunization mixtures used in the production of this AVS . Scarcity of AVS has resulted in quantitative measurement of venom concentration within 24 hours of snakebite . When AVS becomes available for HNPV, these assessments will still be beneficial in clinical decision making. Moreover, in depth analysis of venom cytotoxins will be beneficial in exploring refinements to therapeutic intervention for snake envenoming.