Acute kidney injury caused by Crotalus and Bothrops snake venom: a review of epidemiology, clinical manifestations and treatment.

Ophidic accidents are an important public health problem due to their incidence, morbidity and mortality. An increasing number of cases have been registered in Brazil in the last few years. Several studies point to the importance of knowing the clinical complications and adequate approach in these accidents. However, knowledge about the risk factors is not enough and there are an increasing number of deaths due to these accidents in Brazil. In this context, acute kidney injury (AKI) appears as one of the main causes of death and consequences for these victims, which are mainly young males working in rural areas. Snakes of the Bothrops and Crotalus genera are the main responsible for renal involvement in ophidic accidents in South America. The present study is a literature review of AKI caused by Bothrops and Crotalus snake venom regarding diverse characteristics, emphasizing the most appropriate therapeutic approach for these cases. Recent studies have been carried out searching for complementary therapies for the treatment of ophidic accidents, including the use of lipoic acid, simvastatin and allopurinol. Some plants, such as Apocynaceae, Lamiaceae and Rubiaceae seem to have a beneficial role in the treatment of this type of envenomation. Future studies will certainly find new therapeutic measures for ophidic accidents.

INTRODUCTION

Snakes have evoked human curiosity since the ancient Egyptian civilizations and the book of Genesis shows the snake as the personification of Evil. In ancient Brazil, the lethal danger of ophidic accidents scared the Portuguese explorers. With the development of hyperimmune sera, a new phase in the study of ophidic accidents began, which is still of scientific interest to this day.

Ophidic accidents are still an important problem in public health due to their high frequency and severity3. There are approximately three thousand known snakes' species and 20% are venomous34.

There are four clinically relevant venomous snakes' genera in Latin America: Bothrops, Crotalus, Lachesis and Micrurus 34. Snakes of the Bothrops genus are responsible for the majority of accidents, and this is partly due to their vast geographical distribution and aggressive behavior when feeling threatened38.

Due to its high vascularity, the kidney is highly susceptible to toxins44. The snakes of the genus Bothrops and Crotalus can cause severe systemic reactions, and acute kidney injury (AKI) is one of the most severe complications of snakebites2.

The knowledge of these complications by the health care professional is essential for an adequate approach of patients victims of accidents with venomous snakes. The aim of this study is to review the aspects of AKI caused by the venom of the Bothrops and Crotalus genera regarding their diverse characteristics, emphasizing the most appropriate therapeutic approach for these cases.

EPIDEMIOLOGICAL ASPECTS OF OPHIDIC ACCIDENTS

According to the World Health Organization (WHO), the presence of venomous snakes occurs in all the regions of the globe and is a public health problem, especially in tropical areas. Envenomation due to snakebites is considered one of the main neglected tropical diseases, affecting the poor rural populations of Africa, Asia, Latin America and Oceania22.

The WHO estimates the occurrence of 2,500,000 snakebites per year, resulting in 125,000 deaths worldwide and approximately 100,000 survivors with severe consequences16 34. Latin America is the third most affected geographical area, after Africa and Asia5 16.

In Brazil, these accidents have shown a 32.7% increase in the period between 2004 and 2009, according to data from the Ministry of Health (http://portal.saude.gov.br/portal/arquivos/pdf/clipping_30_07_2010.pdf).

The epidemiological pattern of these accidents has remained unchanged for the last 100 years. The most affected individuals are young males (15-49 years), working in rural areas. The accidents occur mainly during the day, preferentially affecting the lower limbs, and most cases are caused by the Bothrops genus6. This pattern has been observed in many regions of Brazil9 25 31.

Regarding seasonality, the occurrence of ophidic accidents is, in general, related to weather factors and to increased human activity in rural areas. There is a higher incidence between the months of September and March in the South, Southeast and Midwest regions of Brazil, and between January and May in the Northeast region3.

The sub-notification of these accidents in some regions of Brazil has been observed in some studies27 38. However, the Northeast region has registered an increase in the incidence of ophidic accidents in the last years, with an incidence similar to the that observed in other regions in 2010 (15.5 accidents/100,000 inhabitants) (http://portal.saude.gov.br/portal/arquivos/pdf/tabela_inc_casos_serpente_01_04_2011.pdf).

PATHOPHYSIOLOGY MECHANISMS OF ACUTE KIDNEY INJURY DUE TO SNAKEBITES

The renal manifestations of ophidic accidents have a broad clinical spectrum. Proteinuria, hematuria and renal failure are among the most common manifestations of snakebites42.

The pathophysiology of AKI due to snakebites has not been completely elucidated. Renal lesions can be caused by the isolated or combined action of different ischemic or nephrotoxic mechanisms, triggered by the venom action in the body3. Experimental studies suggest a multifactorial pathogenesis for this type of AKI, which includes the following mechanisms: renal ischemia resulting from hypovolemia and hypoperfusion, thrombotic microangiopathy due to the deposit of fibrin in the glomerular capillaries and direct cytotoxic action of the venom on the renal tubules23.

Proteinuria varies according to the snake species, and proteinuria levels > 1g/24h are not common24. Hematuria is frequently observed in accidents involving many snakes' species, and can be microscopic or macroscopic. The outcome of hematuria is usually favorable. However it can be associated with acute tubular necrosis, which has a higher severity outcome42.

The occurrence of pigment nephropathy results from the action of the phospholipase A2 enzyme, which induces rhabdomyolysis42. BRAGA et al. 11 demonstrated renal abnormalities after infusion of phospholipase A2 isolated from Bothrops insularis in Wistar rats. Some abnormalities observed in this study were related to increased perfusion pressure, renal vascular resistance, urinary flow and glomerular filtration rate, as well as decreased sodium and chloride transport. The proximal tubular cells showed hydropic alterations, evidenced by brush border discontinuity and desquamation to the tubular lumen, which can reflect the activity of phospholipase A2 and production of derivatives from arachidonic acid.

AKI is one of the main complications of ophidic accidents, which is described in all the types of snakes, being more common in accidents caused by the genus Bothrops and Crotalus in South America, and by the Vipera russelli in Asia, showing relevant morbimortality. AKI is the main cause of death among the victims of snakebites who survive the early effects of envenomation by Bothrops and Crotalus 38. Although bothropic accidents are ten times more frequent than crotalic ones, the incidence of AKI is similar, which suggests a higher nephrotoxicity of the latter13 17 35 40.

PATHOPHYSIOLOGY OF RENAL LESION IN BOTHROPIC ACCIDENTS

The most commonly described anatomopathological lesion in bothropic accidents is acute tubular necrosis (ATN), but interstitial nephritis, cortical necrosis and glomerular changes have also been described34 38.

BURDMANN et al. 13 demonstrated, in an experimental study, the morphological and functional changes induced by these accidents. There was a significant decrease in glomerular filtration rate and renal blood flow, as well as intravascular hemolysis, characterized by a decrease in hematocrit and increase in the levels of LDH and hemoglobin. Optic and electronic microscopy showed massive fibrin deposition in the glomerular capillaries, associated with proximal and distal tubular necrosis. These would be the main mechanisms involved in the genesis of AKI, although a direct effect of the venom on the kidney could not be ruled out.

Experimental studies with Bothrops moojeni venom have demonstrated glomerular injury, tubular degeneration and desquamation, hematuria and decrease in glomerular filtration rate, with increased urinary sodium excretion, no alteration in arterial blood pressure and no fibrin deposit in the glomerular capillaries7. Other studies have demonstrated a direct effect of the bothropic venom on the kidneys, leading to tubular and glomerular injury8 14 30.

A series of variables seem to be involved in the development of AKI after ophidic accidents: patients' age and body surface area, snake's age, amount of inoculated venom, area of the bite and time between the accident and administration of specific anti-venom sera. Associated comorbidities, such as hypertension, diabetes, coronary artery disease and previous nephropathies, can also make the patients more susceptible to the venom effects18.

The prevalence of AKI induced by bothropic accidents varies between 1.4 and 38%, depending on the species involved12 33 38. Mortality varies from 13 to 19%34. Peak AKI incidence varies from a few hours after the accident to 96 h after the snakebite and is commonly diagnosed in the first 24-48 h. Need of dialysis varies from 33 to 75% of cases3 36 45.

PATHOPHYSIOLOGY OF KIDNEY INJURY IN CROTALIC ACCIDENTS

Diverse mechanisms have been proposed to explain the kidney injury caused by crotalic venom, including rhabdomyolysis, hemolysis, shock, intravascular coagulation and a possible direct nephrotoxic effect18.

As the venom is excreted by the kidneys, the mechanisms of concentration and tubular transport favor the occurrence of direct cellular toxicity36. Crotalic venom is a complex mix of enzymes, toxins and peptides, with crotoxin being one of the main responsible for the nephrotoxicity.

MONTEIRO et al. 29 demonstrated that crotoxin causes systemic lesions and skeletal muscle injury. Crotoxin was the main responsible for acute nephrotoxicity in isolated kidneys of rats, being associated with glomerular and tubular changes.

CLINICAL MANIFESTATIONS

Snake venom consists of more than 20 different substances, of which effects have not yet been completely studied. The protein fraction (enzymes, non-enzymatic toxins and non-toxic proteins) comprises 90-95% of its weight38.

The bothropic venom, despite the variability of its composition between species from different regions and within the same species, depending on the snake's age, in general, shows a mechanism characterized by proteolytic, coagulant and hemorrhagic action, leading to local and systemic characteristic manifestations38. The action of proteases, hyaluronidases, phospholipases and inflammatory mediators leads to local tissue injury, with early onset of pain, edema, bleeding and bullous lesions, which can complicate with abscesses and tissue necrosis. Among the systemic manifestations, hemorrhagic events (epistaxis, gingivorrhagia, hematuria, hemoptysis, central nervous system bleeding) are associated to coagulation disturbances secondary to the activation of factor X and to an action similar to that of thrombin, leading to coagulation factor consumption, as it occurs during the process of intravascular coagulation3 22.

Neurotoxicity and myotoxicity are other typically reported consequences, especially in the crotalic accidents. The first is caused by the action of crotoxin, a neurotoxin that comprises 50% of the protein fraction of crotalic venom and acts in the pre-synaptic membrane of the neuromuscular junction, leading to a descending progressive paralysis, which can be fatal when involving the bulbar and respiratory musculature18. In turn, the myotoxic action leads to rhabdomyolysis, which can result in myoglobinemia, hyperkalemia and AKI. Hypotension and shock are other possible complications that have been described23 34.

According to the clinical manifestations, the ophidic accident can be classified as mild, moderate or severe, which is important to guide the therapeutic measures to be adopted. The severity of these accidents is based on local manifestations (presence of pain, edema and ecchymosis), systemic manifestations (hemorrhage, shock, anuria), coagulation time and time to administrate the antiophidic sera. Severe cases are considered when there are intense local manifestations, severe hemorrhage/shock/anuria, abnormal coagulation time and need for a higher dose of anti-ophidic sera3.

The specific treatment consists in the administration of anti-ophidic sera, produced from the sensitization of diverse animals, especially equines. In Brazil, the Butantan Institute (São Paulo), Ezequiel Dias Foundation (Minas Gerais) and Vital Brazil Institute (Rio de Janeiro) are responsible for the production of these immune derivatives for the public health system.

Recently, Brazilian researchers evaluated the neutralizing action of the extract from a native plant against the toxic activity of the bothropic venom in rats. Schizolobium parahyba, popularly known as “guapuruvu” and commonly used in the central region of the country to treat snakebites, was effective to protect against the effects induced by the enzymatic and biological action of the venom of some species from the Bothrops genus, constituting a promising sustainable therapeutic option for these accidents38.

SNAKEBITE-INDUCED ACUTE KIDNEY INJURY APPROACH

Throughout history, the treatment of ophidic accidents varied from surgical excisions to topical therapy46. The recommendations have changed with time and the anti-ophidic sera has been used with increasing frequency. The use of tourniquets was indicated in the past, in order to isolate the injured member in an attempt to decrease venom dissemination in the body, but it is no longer indicated due to its inefficiency.

In kidney injuries caused by animal toxins, AKI secondary to renal ischemia is the most common type43. Multiple factors contribute to this fact, such as hemodynamic changes, inflammatory reactions and direct nephrotoxic effect of the venom (Fig. 1)44. The knowledge of these factors is of huge importance as it provides subsidies to an effective therapy.

Fig. 1

Pathophysiology of kidney injury induced by animal toxins. Adapted from SITPRIJA et al.

The first action to be undertaken when treating a snakebite victim is to soothe the patient, as the hyperdynamic state accelerates venom dissemination1. The patient should be placed in a comfortable position, with the affected limb elevated at the heart's level1.

To better detail the AKI approach, we divided it in support measures (initial approach), classical measures for AKI and specific measures.

The supportive measures includes: hypovolemia correction; broad spectrum antibiotics administration in severe bite cases (for instance, clindamycin and ceftriaxone; ciprofloxacin and clindamycin; ampicillin/sulbactam); tetanus prophylaxis; monitoring of the local edema evolution; careful assessment of the response to specific antivenom therapy; early detection of local and systemic complications; surgical procedures and rehabilitation (in severe cases of local complications)32.

Early treatment of hypovolemia is a fundamental measure for AKI prevention. The use of isotonic solutions (Ringer's lactate or saline) is required to restore circulating volume32. Urinary volume should be measured every hour, mainly in severe cases. Normally, adolescents and adults have a urinary volume higher than 0.5 mL/kg/h (30-40 mL/h) and children higher than 1.0 to 2.0 mL/kg/h.

Antibiotic use is controversial, as there have been few clinical trials and there are different snakes' species with different potentials to cause infection32. However, biological samples should be collected for culture performance in order to provide the identification of any agent causing infection and the correspondent antimicrobial sensitivity pattern32. According to AMARAL et al. 3, the use of antibiotics should be indicated when there is evidence of infection. The bacteria isolated from lesions secondary to snakebites are mainly Morganella morganii, Escherichia coli, Providencia sp and Streptococcus group D.

Monitoring of local edema evolution and bleeding should be performed every hour in the first day and every 6 hours thereafter32.

The therapeutic approach to AKI induced by ophidic accidents is summarized in Table 1.

Table 1

Therapeutic approach to acute kidney injury caused by ophidic accidents

Supportive measures (Initial approach)	Hypovolemia correction, administration of large-spectrum antibiotics in moderate to severe cases, tetanus prophylaxis, surveillance of the response to specific antivenom therapy, early detection of local and systemic complications, surgical procedures and rehabilitation (in severe cases of local complications)
Classic measures for AKI	Correct approach of fluid and electrolyte disturbances, use of diuretics and vasoactive drugs (when indicated)
Specific measures	Antiophidic sera (immunoglobulin, antivenom) is the only specific treatment for envenomation by snakebites.

Attention should also be paid to the systemic signs of severity, including persistent bleeding, neurological symptoms (ptosis, paresthesia, visual abnormalities, vertigo, etc.), cardiovascular abnormalities (hypotension, arrhythmia, shock) and kidney injury (AKI, hemoglobinuria, myoglobinuria, etc.)21. Among the local complications, compartment syndrome is clinically suspected in the presence of tense edema, dysesthesia, proprioception alteration and movement limitation, with or without decreased capillary refill time. Prompt intervention is required in these cases, through the administration of intravenous mannitol or surgery (fasciotomy)32.

The use of intramuscular injections (such as antitetanic vaccine) should be avoided in the first 48 hours after the accident or when there is coagulopathy persistence32.

The classical measures for AKI include fluid correction and electrolyte management, judicious use of diuretics and vasoactive drugs26.

The adequate volemic status is the main measure for AKI prevention and is also indicated to treat AKI. There are no specific guidelines for hemodynamic optimization and renal function preservation; however, one can extrapolate the recommendations from other causes of AKI to that caused by snakebites26. It is important to be careful when administrating fluids to patients with comorbidities, such as heart and liver disease, in order to avoid fluid overload and its complications.

The use of diuretics in AKI treatment is controversial. MACEDO et al. 26 suggest that it is possible to try to increase urinary volume with loop diuretics, but if there is no adequate response, increasing doses should be avoided. Regarding the use of natriuretics, large studies are required to confirm its benefit in AKI treatment26. Vasoactive agents are considered in the cases with organ perfusion impairment and the use of dopamine in order to prevent or treat AKI is not recommended26.

The complications associated to AKI (acute pulmonary edema, electrolyte and acid-base disturbances) that are refractory to clinical treatment should be promptly treated with renal replacement therapy. The need for dialysis in victims of ophidic accidents has been described by SITPRIJA et al. 41, and this treatment also improves myalgia symptoms.

Regarding specific measures, the anti-ophidic sera (immunoglobulin, antivenom) is the only specific treatment for snakebites. The sera are produced by fractioning plasma obtained from domestic animals hyper-immunized against the venoms. When administered to humans, it neutralizes the venom responsible for the accident, and, in some cases, it also neutralizes venoms from related species47. It is very important to have in mind that the early administration of anti-ophidic sera is one of the most effective measures to prevent AKI induced by Bothrops and Crotalus accidents.

The identification of the snake and the classification of accident severity are the first steps for the correct prescription of anti-ophidic sera. Another important step is the anaphylaxis surveillance during its administration, as this is an heterologous immunoglobulin.

In cases of envenomation by Bothrops sp., the administration of antibothropic serum (ABS) should be instituted as early as possible, and when it is not available, the anticrotalic (ACS) or antibothropic- lachesis (ABLS) serum should be administered3.

If the coagulation time remains unchanged 24 hours after serum therapy, an additional dose is indicated, with two ampoules of antivenom sera3.

Some laboratory tests are important in bothropic accidents, including time of coagulation, complete blood count (it can reveal leukocytosis with neutrophilia and thrombocytopenia), urinalysis (to screen for proteinuria, hematuria and leukocyturia) and other tests according to the patient's evolution, with special attention to electrolytes, urea and creatinine to detect AKI3.

In crotalic accidents, the dose of anticrotalic sera varies according to the severity of the case. It is important to note that the same dose is administered to adults and children, and the ACS sera can also be used3, according to what is shown in Table 2.

Table 2

Classification of crotalic accidents regarding severity and recommended specific therapy

Manifestations and treatment	Classification - Initial evaluation
	Mild	Moderate	Severe
Myasthenic fascies and blurred vision	Absent or late	Mild or evident	Evident
Myalgia	Absent or mild	Mild	Intense
Coagulation time (CT)	Normal or altered	Normal or altered	Normal or altered
Red or brown urine	Absent	Poor evident or absent	Present
Oliguria or anuria	Absent	Absent	Present or absent
Serum therapy (# ampoules) ABS/ABCS*	5	10	20
Administration via		Intravenous

ABS = anti-bothropic serum/ABCS = anti-bothropic - crotalic serum.

As the crotalic toxin has myotoxic, neurotoxic and coagulant actions, rhabdomyolysis is an important factor in AKI genesis in these accidents. According to BOSCH et al. 10, preventive and therapeutic measures for rhabdomyolysis-induced AKI includes: (1) monitoring of volemic status, central venous pressure and urinary volume, (2) evaluation of serum levels of creatine kinase (CK), myoglobin, aldolase, lactate-dehydrogenase, alanine aminotransferase, aspartate aminotransferase, (3) monitoring of levels of urea, creatinine, sodium, potassium, total and ionized calcium, phosphorus, magnesium, uric acid, albumin, platelet count, white and red blood cell count, (4) evaluation of urine sediment and dipstick, (5) volemic restoration with isotonic saline solution, at a rate of approximately 400 mL/h, monitoring of central venous pressure, (6) obtaining a urinary output of 3 mL/kg/h, (7) frequent monitoring of potassium levels, (8) correction of hypocalcemia only if it is symptomatic or if there is severe hypercalcemia, (9) if the urine pH is below 6.5, alternate each liter of saline solution with dextrose or 0.45% solution with 100 mmoL bicarbonate, (10) consider treatment with mannitol, discontinuing it if there is improvement in diuresis (> 20 mL/h); if it does not occur, (11) maintain volemic infusion until myoglobinuria decreases and (12) consider renal replacement therapy if there is no response to therapy or if there is persistent hyperkalemia (> 6.5 mEq/L), oliguria (< 0.5 mL/kg/h, in 12 hours), anuria, hypervolemia and refractory metabolic acidosis.

The knowledge of early and late reactions to the administration of anti-ophidic sera and its correct treatment is important to prevent fatal complications. The adverse effects of anti-ophidic sera are: type I acute reactions, due to reactions of circulating immunoglobulin E to horse proteins; anaphylactoid reactions due to direct degranulation of mastocytes and serum sickness (a late allergic reaction caused by immune complexes)1.

The early reactions occur in 4.6 to 87.2% of cases in the first two hours after serum administration. They have low severity and the patient should be under observation for 24 hours due to the risk of onset of more severe late reactions3. Signs and symptoms include urticarial rash, tremors, coughing, nausea, abdominal pain, pruritus and facial rash. More severe manifestations, such as anaphylactic reactions, are rare. In these cases, the patients can also have arrhythmias, hypotension, shock and respiratory failure3.

The late reactions occur five to 24 days after serum administration and can cause fever, arthralgia, lymphadenomegaly, urticarial rash and proteinuria. The incidence of serum sickness is underestimated, as many patients do not return to the health care center. The use of corticosteroids (prednisone) 1 mg/kg/day (maximum 60 mg) for five to seven days3 is indicated for the treatment of this complication.

The use of intravenous corticosteroids, promethazine and adrenaline can be therapeutic for anaphylactic reactions to heterologous sera. As premedication such as H2-antagonists, such as cimetidine (maximum 300 mg) or ranitidine3 can also be used. SILVA et al. 39 demonstrated, in a randomized study with 752 patients with acute reactions to heterologous sera in the first 48 hours, that the use of subcutaneous adrenalin decreased the risk of severe adverse reactions in the first hour; however, neither hydrocortisone nor promethazine showed clear benefits.

The therapeutic measures are of huge importance in the management of victims of ophidic accidents and the early administration is fundamental to prevent AKI occurrence. CASTRO et al. 14, in an experimental study with bothropic venom, showed that the ideal renal protection was obtained when the administration of the venom and the sera were performed at the same time. In a prospective study with 100 victims of crotalic accidents, it was demonstrated that the delay in the administration of antiophidic sera (> 2h) was an independent risk factor for AKI development33.

FUTURE PERSPECTIVES

Some studies have searched for complementary therapies for the treatment of ophidic accidents. BARONE et al. 4, through an experimental study using Bothrops jararaca venom in AKI induction in rats, demonstrated the beneficial effects of lipoic acid and simvastatin on hyperuricemia, renal oxidative stress and serum levels of urea and creatinine, creating new perspectives for the investigation of these drugs as adjuvant agents in bothropic envenomation therapy.

An experimental study with rats with AKI induced by Crotalus durissus terrificus envenomation evidenced the association between hyperuricemia and kidney injury in these animals, as well as the beneficial effect of allopurinol on renal function recovery, preventing death in these animals20.

MOLANDER et al. 28 created a database of plants used in ophidic accidents in the world, selecting countries from different continents (Brazil, Nicaragua, China and Nepal). The most common plants were those from the families Apocynaceae, Lamiaceae and Rubiaceae. It was observed that the use of the same plant in different regions and further studies are required to better investigate the role of these plants in the treatment of snakebite envenomation.

SAUL et al. 37, upon considering that the snakes' toxins first reach the lymph nodes before entering circulation, studied the use of a nitric oxide ointment in 15 patients and obtained satisfactory results regarding the inhibition of lymphatic transport of the toxin.

All these studies can be used as a base for the development of new research in ophidic accidents and its complications.