Challenges in care of snake envenomation in rural Pakistan: a narrative review

Abstract Snake envenomation remains a neglected tropical disease in Pakistan per the World Health Organization, with approximately 40,000 bites and 8,200 fatalities annually. To understand the factors leading to increased mortality, we performed a narrative review of the medical literature pertaining to snake envenomation in Pakistan and surrounding countries. Several factors have led to increased mortality in rural communities. First, geographical variations in snake venom composition between countries lead to a decreased efficacy of polyvalent antivenom in Pakistan that is produced using snake species from different areas. Furthermore, healthcare providers in rural communities are inadequately trained on antivenom dosing regimens or adverse reactions. Long transport times and the use of traditional healers lead to delays in administration of antivenom. Additionally, many types of antivenom must be stored at 2-8 degrees Celsius. Lack of refrigeration to store antivenom limits access in rural locations. Research advances are being developed to create new generations of antivenom that may be better suited for rural communities. Lastly, increased educational efforts may further reduce mortality.


Introduction
Snake envenomation remains a neglected tropical disease per the World Health Organization (WHO).South Asia constitutes approximately 70% of the global snake bite mortality [1].Approximately 40,000 bites are reported annually in Pakistan with approximately 8,200 fatalities [2].A total of 72 species of snakes are distributed throughout Pakistan.Of those 72 species, 40 are venomous [3].However, there is only one available polyvalent antivenom produced by several manufacturers targeted against the "Big Four" (the common krait, the saw-scaled viper, Russell's viper, and the Indian cobra); the most common causes of snakebite fatalities in the region [4].
Most snake bite fatalities in Pakistan occur in the Sindh province, particularly in the Thar desert [5].Rural communities in the Thar desert have an increased incidence of snake bites due to both environmental and occupational exposures [6].A recent study looking at the Tharparkar district of the Sindh province demonstrated that increased agricultural practices and low socioeconomic status lead to an increased snake-human interaction [6].Snake bites occur most commonly during warm and rainy months when snakes are more active [7].Importantly, snake bites are often under-reported in rural populations, thus the true number of snake bite fatalities is unknown [2,6].
Currently, there are limited data regarding snake envenomation mortality, particularly in rural areas of Pakistan.This literature search was conducted to better understand the multitude of factors that lead to an increased mortality rate in these rural areas, and to increase awareness of the current research deficiency.

Methods
We performed a narrative review of the scientific databases PubMed, ScienceDirect, and Web of Science.Search terms included relevant snake species with the keywords (envenomation, antivenom, Sindh) and Pakistan.These search terms, along with any additional relevant articles found, were used to identify articles meeting the inclusion criteria.Articles were excluded if they were not geographically relevant, not relevant to the topic, full text was not available, or the article was not available in English.

Results and data summary
A total of 2,718 articles were obtained in the initial search.After removing duplicate articles, 2,020 articles were screened.Twenty-six articles met inclusion criteria (Figure 1).A summary of articles discussing the Big Four snakes is provided in Table 1.A summary of Pakistan-specific articles in provided in Table 2.

Venomous snakes in Pakistan
The main venomous snakes in South Asia are the common krait (Bungarus caeruleus), Indian cobra (Naja naja), Russell's viper (Daboi russelii), and the saw-scaled viper (Echis carinatus), also known as the "Big Four" [6].Each snake venom is composed of a variety of proteins and peptides that target specific enzyme pathways, receptors, and ion channels [5].Due to the complexity of  adverse reactions to snake antivenom, and their prevention and treatment de silva 2015 [8] not applicable adverse antivenom reactions range from acute anaphylaxis to pyrogenic reactions, to delayed serum sickness.The high incidence of adverse events can be an indicator of poor-quality control and manufacturing of antivenom.
Rediscovery of severe saw-scaled viper (Echis sochureki) envenoming in the Thar desert region of Rajasthan, india Kochar 2007 [9] Rajasthan, india The southern india antivenom used patients envenomated by saw-scaled vipers was not effective in reversing the hematotoxicity.
In vivo neutralization of dendrotoxin-mediated neurotoxicity of black mamba venom by oligoclonal human igg antibodies laustsen 2018 [10] african savannah showed the potential use of fully human monoclonal iggs against animal toxins and the first use of oligoclonal human igg mixtures against experimental snakebite envenoming.
Beyond the 'big four': Venom profiling of the medically important yet neglected indian snakes reveals disturbing antivenom deficiencies laxme 2019 [11] india There are several differences in the venom profiles of the neglected venomous snakes of india compared to the Big Four current antivenoms are minimally effective against these species.
severe neurotoxic Envenoming and cardiac complications after the Bite of a 'sind Krait' (Bungarus cf.sindanus) in maharashtra, india Pillai 2012 [12] india case report of a patient who developed paralysis, respiratory failure, and autonomic dysfunction resulting in multiple episodes of pulseless ventricular tachycardia after being bit by a sind krait (Bungarus sindanus) Envenoming bites by kraits: the biological basis of treatment-resistant neuromuscular paralysis Prasarnpun 2005 [13] southeast asia Rats inoculated with Beta-bungarotoxin were found to have denervation that occurred through several preceding co-occurring mechanisms 12 h after exposure and lasted for approximately 3-5 days.
Pharmacoinformatic approach to Explore the antidote Potential of Phytochemicals on Bungarotoxin from indian Krait, Bungarus caeruleus Rajendran 2018 [14] india Bioactive phytochemical compounds from medicinal plants were found to have potential as a bungarotoxin antidote based on simulation of toxin-ligand interaction and molecular dynamics.
The timing is right to end snakebite deaths in south asia Ralph 2019 [1] south asia There is a high burden of morbidity and mortality of snakebites in south asia, with current treatment practices and gaps leading to poor outcomes.
neuromuscular Effects of common Krait (Bungarus caeruleus) Envenoming in sri lanka silva 2016 [15] sri lanka seventeen patients admitted following snake bite by the common krait (Bungarus caeruleus) developed severe progressive descending neurotoxicity that was not improved with administration of antivenom.
a Wolf in another Wolf's clothing: Post-genomic Regulation Dictates Venom Profiles of medically-important cryptic Kraits in india sunagar 2021 [16] south asia Performed phylogenetic and comparative venomics investigations of kraits against commercial antivenoms and found there to be poor antivenom function against subspecies.
Poisoning by bites of the saw-scaled or carpet viper (Echis carinatus) in nigeria Warrell 1977 [17] nigeria Treatment included specific antivenom therapy, blood transfusions, and early surgical debridement of necrotic tissue for patients poisoned by saw-scaled or carpet Viper (Echis carinatus) bites studied in nigeria.

Venomous and Poisonous animals, Expedition medicine
Warrell 2014 [18] not applicable a review of snake identification, the geographical distribution of snake species, the epidemiology of snake bites, and venom composition.
Venom, antivenom production and the medically important snakes of india Whitaker 2012 [19] india Barriers to snake bite treatment in india include the intra-regional variations that exist between the Big Four.The current polyvalent antivenom does not cover for many subspecies.
Ending the drought: new strategies for improving the flow of affordable, effective antivenoms in asia and africa Williams 2011 [20] africa and asia Review of the problems leading to the shortage of safe, effective, and affordable antivenom in asia and africa.Proposed the global snakebite initiative be the leaders of an international collaboration develops new regional antivenoms and evaluates existing antivenom.Karachi, Pakistan There is an increased morbidity in snake bite victims that have delayed presentation to tertiary care centers.
a contextual approach to managing snake bite in Pakistan: snake bite treatment with particular reference to neurotoxieity and the ideal hospital snake bite kit.
Quraishi 2008 [20] Pakistan Provided guidelines to aid in the treatment of snake envenomation patients.
comparative cost and efficacy trial of Pakistani versus indian anti snake venom Qureshi 2013 [19] Pakistan and india Pakistani antivenom had statistically significant increased improvement in coagulopathy after 1 dose and 2.5 times lower cost when compared to indian antivenom.
a study of the current knowledge base in treating snake bite amongst doctors in the high-risk countries of india and Pakistan: does snake bite treatment training reflect local requirements?
simpson 2008 [25] india and Pakistan current textbooks and medical education do not adequately prepare doctors in india and Pakistan to treat snake bites.
Haemotoxic snake venoms: their functional activity, impact on snakebite victims and pharmaceutical promise slagboom 2017 [14] Pakistan and india Reviews the functional activities of hemotoxic venom proteins, the pathologies they cause in snakebite victims and how their exquisite selectivity and potency make them amenable for use as therapeutic and diagnostic tools.
snake bite in the Thar Desert suleman 1998 [6] Thar Desert, Pakistan studied the incidence of snake bite, its mortality and the management in public health facilities located in the Thar Desert Province of sindh, Pakistan.
Venom and Purified Toxins of the spectacled cobra (naja naja) from Pakistan: insights into Toxicity and antivenom neutralization Wong 2016 [13] Pakistan investigated the toxicity and neutralization profile of the venom and toxins from Pakistani spectacled cobra, naja naja, using Vins polyvalent antivenom (VPaV, india), naja kaouthia monovalent antivenom (nKmaV, Thailand), and neuro bivalent antivenom (nBaV, Taiwan).Results suggest that optimal neutralization for Pakistani n. naja venom may be achieved by improving the formulation of antivenom production to enhance antivenom immunoreactivity.
snake venom and wide variation of snake species, there are several ways snake envenomation can clinically present [6,21].
Common krait (Bungarus caeruleus) venom consists of two neurotoxins, the alpha and beta bungarotoxins [15].Beta bungarotoxins cause neurotoxicity through presynaptic phospholipase A 2 activity [15].Alpha bungarotoxins bind to postsynaptic neuromuscular receptors and block the action of acetylcholine at the postsynaptic membrane [14].These neurotoxins cause paralysis leading to respiratory depression [13].Importantly, there are regional variations in species of the "Big Four." The Sind krait (Bungarus sindanus), a subspecies of the common krait, is found in the Thar desert [12].The Sind krait venom contains the same neurotoxins as the common krait, albeit the venom is six to eleven times more potent than that of the common krait [16].
Indian cobra (Naja naja) venom contains a similar neurotoxin to that of the common krait that acts on the postsynaptic neuromuscular receptors to block the action of acetylcholine and leads to muscle paralysis and ultimately respiratory depression [22].
Saw-scaled vipers (Echis carinatus) venom acts through a similar hematotoxin that causes a consumptive coagulopathy, leading to disseminated intravascular coagulation and life-threatening spontaneous hemorrhage [17].Both Russell's vipers and saw-scaled vipers are commonly found in the rural Sindh province.

Current antivenom treatment
The treatment for snake envenomation includes supportive care and can include polyvalent antivenom.Polyvalent antivenom is produced by extracting venom from the Big Four species, injecting the venom into equine subjects, and then harvesting antibodies [19].Antivenom neutralizes venom by binding to components of the snake venom and either reversing or inhibiting the effects of the snake bite [5].The Indian polyvalent antivenom, which is imported to Pakistan for use, is produced in two forms: the liquid and the lyophilized form [1].While the imported Indian polyvalent antivenom is the predominant antivenom used in Pakistan, Pakistan also produces antivenom at one facility in the liquid form only [24].Liquid polyvalent antivenom is stored at 2-8 degrees Celsius, whereas the lyophilized antivenom can be stored at room temperature [1].Rural communities with poor access to electricity and refrigeration have difficulty storing the liquid form [1].However, the lyophilized form requires up to one hour reconstituting with distilled water [25].
There are many complications of polyvalent antivenom [8].Severe reactions may include systemic anaphylaxis, hypotension, and loss of consciousness [8].Acute reactions are thought to be due to type I hypersensitivity [8].Main treatments for acute allergic reactions include intramuscular epinephrine, fluid resuscitation, airway management, and discontinuing antivenom administration [8].
Pyrogenic reactions can also occur due to contamination during manufacture and presents as fever, chills, rigors, and occasionally hypotension [8].This reaction usually occurs within one hour of antivenom use and is treated by cooling methods and intravenous fluids [8].Serum sickness may also occur between 5 to 14 days after administration, with symptoms including fever and lymphadenopathy [8].Serum sickness is routinely treated with corticosteroids [8].

Limitations to treatment
Importantly, there are geographic and regional variations in snake venom, even within the same species [5].Therefore, antivenom produced in India is not as effective against snakes in Pakistan [24].In Pakistan, there exists only one authorized site of antivenom production and also targets the Big Four [24].However, there is low domestic supply of the product [24].As a result, Pakistan depends upon polyvalent antivenom imported primarily from India, and secondarily Saudi Arabia [5].There are no current data on where imported versus domestically produced antivenoms are distributed throughout Pakistan.
One study focused on the Bharat polyvalent antivenom (BPAV) produced in southern India showed that the BPAV does not have complete neutralization effects against the Big Four snakes in Pakistan [5].Likely due to both different ecologies and diet, the Big Four and other subspecies vary greatly in their venom profiling from different countries [11].While BPAV does have some neutralization effects against the Big Four snakes in Pakistan, the response is variable [5].
Furthermore, the polyvalent antivenom is not as effective against species variations outside the Big Four.In particular, the polyvalent antivenom was poor at neutralizing the venom of the Sind Krait (Bungarus sindanus) commonly located in the Thar desert [11].Additionally, the Indian polyvalent antivenom was created against the species of saw-scaled viper (Echis carinatus), not the Echis sochureki subspecies found in the Thar desert, the Pakistani antivenom [9].The Indian polyvalent antivenom is not as effective against Echis sochureki in preventing hematotoxicity [9].One study found that Indian polyvalent antivenom restored coagulation in patients in India with saw-scaled viper bites with an average of 15.4 vials, compared to over 25 vials required by Sindh victims [25].
A recent study demonstrated that the domestically produced Pakistani antivenom is both more effective and cheaper than the Indian antivenom [24].In this study, the Pakistani antivenom produced blood coagulation at a rate of 60.5% compared to 36.11% with the Indian antivenom after the first dose [24].The Pakistani antivenom was also noted to be 2.5 times cheaper and required less doses [24].Therefore, not only is the Pakistani antivenom more effective, but also economically advantageous.
In both Pakistan and India, there are currently no protocols to guide antivenom production [2,19].This can lead to variable antivenom effectiveness [1].While the Central Drugs Laboratory in India screens antivenom for potency, it only screens four of the major six manufacturers [1].Prior to the mid-1950s, India required antivenom to be much more potent to combat the effects of both Russell's viper (Daboi russelii) and Indian cobra (Naja naja) [20].As a result of more relaxed protocols, many victims require massive doses of antivenom to combat venom effects leading to increased cost and adverse side effects [20].
There is also a significant time delay to treatment from those in rural communities.The causes for delay include first obtaining treatment through nontraditional and local practices, transportation issues, and longer distance to travel [26].The delay to treatment leads to severe complications, including bleeding, renal failure, tissue necrosis, and wound infections that if left untreated can lead to sepsis [26].
In rural communities, poor literacy and decreased awareness lead to lack of implementation of preventative measures against snake bites [1].In a study interviewing the rural population of Sindh, 44.5% of interviewees did not know that snake antivenom existed [3].Additionally, poverty is common and results in inability to afford medical treatment [4].In a recent study looking at snake bites in Karachi, five out of the eighty included patients who died from snake envenomation all had a delayed arrival to the hospital, ranging from nine to twenty-seven hours [4].
There is limited education of proper snake bite management among healthcare providers [27].A recent study assessing knowledge of doctors from the Sindh province showed that even with the use of major Western references, doctors failed to identify more than 30% of the correct answers [27].The lack of current guidelines dedicated to local infrastructure and available resources leads to additional barriers in treatment [27].
Despite these limitations, there is promising new research on the horizon utilizing human oligoclonal IgG antibodies [10].A recent study found the human IgG antibodies were both safer and cost effective compared to polyvalent antivenom [10].Oligoclonal human IgG antibodies may produce lower levels of serum sickness and allergic reactions, however further research is needed.

Discussion
There remain several barriers to treating snake envenomation in Pakistan.Our review shows the limitations of using the current imported polyvalent antivenom, including poor efficacy against regional snake variations in Pakistan.This highlights the need for either domestically produced polyvalent antivenom that targets a larger range of species, monovalent antivenom, or novel antivenoms to create species-specific antivenom.
A large limitation to determining the need for snake antivenom in rural communities in Pakistan stems from under-reporting.The creation of a streamlined process for reporting snake bites may result in a more accurate number of reported snake bites.Furthermore, efforts should be placed into educating rural communities regarding proper care of snake bites and developing region-specific snake bite management protocols.
Increasing the number of sites for domestic production or increasing production capacity at a single site may improve care, given the increased efficacy of domestically produced antivenom.Additionally, more affordable antivenom would be beneficial to rural communities that cannot otherwise afford antivenom.Further research should be conducted to determine the distribution of domestically produced antivenom versus imported polyvalent antivenom to determine which populations are receiving imported antivenom, and whether or not this may play a role in increased mortality in rural communities.More stringent protocols for antivenom production are also necessary.

Conclusion
Few studies describe snake bite incidence, antivenom use, and outcomes.The paucity of data is greatest for snake bites in the Thar Desert.Creating more affordable antivenom, increasing production of antivenom, researching specific antivenom to the regional species of Pakistan, and a standardized educational curriculum for healthcare providers are all key to mitigating this neglected tropical disease.Despite the barriers to eradicating snake envenomation mortality and morbidity, with the support of government organizations, hospitals, and public health agencies, several of these barriers can be leveraged to prevent the death of so many from a disease for which a cure is so close within reach.

Disclosure statement
No potential conflict of interest was reported by the author(s).

Table 1 .
summary of included big four snake articles.

Table 2 .
summary of included Pakistan-specific articles.