Taxonomy

In the Class Amphibia, there are three Orders. Order Gymnophiona are a group of limbless worm-like amphibians, the caecilians. Caecilians are not very common in captivity. Most species are fossorial, though the aquatic species are most commonly seen in zoos and the pet trade.

Order Caudata includes the tailed amphibians. There are many fascinating families within three suborders, the Sirenoidea (sirens), the Cryptobranchoidea (hellbenders and Asian giant salamanders), and the largest suborder, the Salamandroidea, which include a diverse collection of salamanders and newts.

By far the most numerous, most widely distributed, and most recognizable order is the Anura, the frogs and toads. Generally speaking, toads are frogs that have specialized adaptations for a more terrestrial lifestyle. Toads tend to have granular skin that protect against dehydration and shorter hind legs for walking rather than hopping. The anuran amphibians are quite common in captivity, are popular with the pet trade and private breeders, and have come to represent amphibians as a whole in popular culture.

Biology

At least a basic understanding of amphibian biology and physiology is essential for the clinician. The most important amphibian adaptation and the most clinically significant feature of their biology are the properties of amphibian skin. The skin is highly permeable and is the primary means of respiration and osmotic balance. Any stress or insult to the skin integrity may result in dehydration, electrolyte imbalance, or sepsis.

Respiration

Respiration occurs through four major routes: cutaneous, buccopharyngeal, pulmonic, and branchial. All amphibians utilize some combination of all four. Most amphibians use the cutaneous route for a significant portion, if not the majority of their respiration. Buccopharyngeal is essentially an extension of the cutaneous route, as it involves gas exchange across the oral membranes. Amphibian lungs are typically thin, simple structured membranes. A large family of salamanders, the plethodontids, has no lungs. Gular pumping is the means of ventilation for both buccopharyngeal and pulmonic respiration. Branchial respiration occurs via gills, which are present in all species during the larval stage and in neotenic species, such as axolotls or sirens, which retain gills through maturity.

Water Homeostasis

The permeable skin is the main route of water loss or absorption. Most species must remain moist for proper osmotic or respiratory exchange across the skin. Availability of fresh water and maintenance of proper humidity in an amphibian enclosure is important to health. Frogs in the genus Phyllomedusa have a waxy lipid secretion and toads have thicker, granular skin for protection in drier environments. In anurans, the ventral surface of the pelvic and inguinal regions, often referred to as the "drink patch", is the most permeable location and is the primary site of water absorption.

Aquatic amphibians have similar requirements to fish. Like fish, aquatic amphibians primarily excrete nitrogenous wastes as ammonia, as opposed to urea in most terrestrial amphibians. Ammonia must quickly be dissipated, filtered, or metabolized by bacteria to avoid toxicity in captive aquatic amphibians.

Calcium Homeostasis

Amphibians possess all the same hormones involved in calcium metabolism as other vertebrates, though the role of each is not known for certain. Likewise, whether ultraviolet light is needed for normal calcium metabolism is also unknown. Specific requirements also likely vary by species. Erring on the side of caution may justify providing access to UV spectrum light in captivity.

Husbandry

Inadequacies of husbandry are probably responsible for a majority of problems in captive amphibians. Species identification and researching an animal's natural history is important for determining optimal environmental parameters and appropriate diet. The animal should feel secure in its environment, with adequate cover and shelter to avoid stress of exposure.

Environmental parameters should be monitored and recorded regularly. This should include at least temperature (high and low), humidity (high and low), and basic water quality parameters such as pH and ammonia.

Water Quality

The ability of amphibian skin to absorb any water it contacts makes water quality an important issue. Any chemicals or contaminants in the water will also be absorbed and can be toxic. Tap water from a municipal water source will contain chlorine or chloramines that will cause skin or gill irritation. Chlorine should be removed from the water prior to use by using dechlorinators, vigorous agitation of the water, or aging the water to allow chlorine to degas naturally. Chloramines must be removed with specific conditioners followed by removal of ammonia that will be released in the process.

Distilled or reverse osmosis water has had all solutes removed, creating hypoosmotic conditions which can be lethal to amphibians over time. Distilled or RO water should be reconstituted by adding essential solutes back prior to use.

Nutrition

Amphibians hunt by sight and require live prey, usually in the form of insects. Large amphibians will also eat rodents, small reptiles, other amphibians, or fish. The most commonly used captive diet is domestic crickets, which are mostly composed of chitin and phosphorous. Attaining proper calcium: phosphorous ratio is the most challenging aspect of the captive diet. Crickets should be gut-loaded with a high calcium supplement (up to 8% calcium DM) at least 24 hours prior to feeding to amphibians. All insects can also be dusted with calcium and vitamin powders immediately prior to feeding. Fine powdered calcium with vitamin D is available commercially.

Various commercial vitamin supplements are also available. One variable to consider is how vitamin A is provided in commercial supplements. Vitamin A deficiency syndromes are reported in several captive species. On the other hand, vitamin A toxicity can easily occur if over-supplemented. Commercial vitamin supplements provide vitamin A in two forms - either as beta carotene, a vitamin A precursor, or as retinol, the active form of vitamin A. The bioavailability of beta carotene to amphibians is not known and is likely to vary among species. Therefore, one must weigh the risks of the possibility of either under-supplementing or over-supplementing vitamin A.

Uneaten prey items should be removed from the enclosure, as the nutritive value of that prey will decrease rapidly over time. Within minutes to hours, dusting will fall off or be groomed off and gut load will be excreted by the insects. Overfeeding and allowing insects to stay in the enclosure will chronically lead to malnutrition. Volume of each feed should be dictated by what the animals can finish within 15 minutes. Overfeeding will also lead to obesity and poor health.

Examination

The same array of diagnostics available in small animal medicine may be used with amphibians. Examination should be performed visually first - observe animals' posture, breathing, mentation, etc. Physical exam should be methodical and should include all major organ systems - skin, eyes, tympanum, oral cavity, limbs and joints, coelomic palpation. Transillumination is a very useful technique for evaluating coelomic contents.

Blood collection is limited by animal size. Fine gauge needles (25 g or smaller) on a 1 ml heparinized syringe work best in animals that are large enough. 0.8 - 1% of body weight is a good rule for maximum volume of blood to draw. Useful blood collection sites include the ventral abdominal vein on the caudal ventral midline, the femoral vein behind the stifle, axillary venous plexus, and ventral tail vein in caudates. Other sites may be possible, such as the brachial vein or lateral abdominal veins in larger species. Transillumination is a useful technique for highlighting the pathways of major vessels.

Radiographs and ultrasound offer all of the same diagnostic uses as in mammals. Radiographic evaluation of skeletal quality is particularly useful and should be included in all new patient exams.

Rigid endoscopy in larger amphibians is valuable for direct visualization and biopsy of coelomic pathologies.

Anesthesia

Tricaine methanesulfonate is a water soluble anesthetic used in fish that is also very reliably used in amphibians. It comes as a white powder that is added to solution to create an anesthetic bath. A good induction dose for most species is a 0.1% solution, or 1 g/L.¹ High quality water safe for amphibians should be used. Bottled spring or distilled water is a convenient source. Tricaine methanesulfonate solution is very acidic, down to pH 3, and must be buffered to pH 7 prior to use. Care should be taken during induction that the nostrils are kept above water so the animal does not drown. Once the animal is induced, it should be transferred to a fresh water bath and a 0.05% solution of tricaine methanesulfonate can be dripped on the skin to maintain anesthesia. Rate of administration of the maintenance solution is altered as necessary to titrate the depth of anesthesia. A new fresh water bath is used to recover the animal.

Other anesthetic agents have been described, though are not typically as safe or effective as tricaine methanesulfonate.

Anesthetic Monitoring

There are three main phases of anesthetic induction. The first phase is excitement and usually results in jumping or escape behavior. Use of a covered padded induction chamber such as an inflated closed bag or a bubble-wrap lined plastic container prevents injury during this phase. The second phase is sedation or light anesthesia, with a loss of righting and corneal reflexes. The final stage is surgical anesthesia, which brings loss of gular respiration and loss of withdrawal reflexes. The heart beat should remain visible on the ventral surface. A decrease in heart rate indicates the animal is becoming too deep and should be rinsed with fresh water and/or decrease the rate of application of the maintenance solution.

A Doppler probe over the heart, or a pulse oximetry device on a foot or limb, are useful for monitoring heart rate.

Diseases

Nutritional Disorders

Metabolic bone disease is common in captive amphibians and is usually due to diets high in phosphorous and low in calcium and vitamin D. Malformed limbs, spine, or jaw are typical hallmarks of the disease. Severely hypocalcemic animals may present in tetany. Animals in tetany should be treated with calcium gluconate at 100 mg/kg topically or by injection. 2.3% calcium gluconate baths for 1 hour a day for at least 30 days, as well as long term oral calcium and vitamin D supplementation will begin to replete calcium stores.¹ Effectiveness of UV-B spectrum lighting is not proven, but is a prudent addition to the treatment protocol.

Obesity is a nutritional disorder and should be addressed by decreasing fat and total calories in the diet. Obese animals may also develop corneal lipidosis, corneal opacity caused by deposition of cholesterol in the corneal stroma. Effective treatment is not reported. If the opacity compromises sight, the animal will be unable to feed itself.

Hypovitaminosis A causes squamous metaplasia of epithelial cells, which results in decreased mucus production, decreased skin integrity, and in severe cases may cause renal failure. Animals may present with swollen conjunctiva, skin lesions, or an inability to capture prey, owing to decreased mucus production and decreased stickiness of the tongue. Treat topically with vitamin A at 2-5 IU/g every 3 days for 2 months.²

"Spindly leg" syndrome is underdevelopment of front limbs in tadpoles morphing to frog-lets. The cause is not known and is likely multifactorial, ranging from nutrition of tadpoles or adults, genetics, water quality, or temperature.² Several treatments have been reported to improve development of future clutches, but vary widely and success is usually only after trial and error. Frog-lets affected with the condition are unable to move well enough to feed and do not survive.

Bacterial Infection

"Red Leg" syndrome is a lay-term for what is more accurately described as bacterial dermato-septicemia. Erythema, ecchymotic hemorrhage, or ulcerative lesions of the skin are dermal manifestations of sepsis. The etiologic agent is most often gram negative bacteria, such as Aeromonas hydrophila, an ubiquitous organism in water environments. The infection is usually secondary to a primary stressor or injury to the skin. Underlying diseases or stressful environmental parameters should be investigated. Treatment should be immediate with an antibiotic with gram-negative spectrum activity. Enrofloxacin (10 mg/kg TO SID x 7-10 d) is a good first-line antibiotic. Injectable enrofloxacin is very caustic and should be diluted at least 1:1 with sterile saline prior to administration.

Mycobacterial infection with environmental Mycobacteria sp. usually manifests as granulomatous or non-healing ulcerative lesions. Granulomas are found in integument or musculoskeletal tissues, but can also be disseminated internally. Intralesional acid-fast bacilli are seen on cytology. Infection implies immunosuppression and primary stressors should be identified. Treatment is typically unsuccessful and infected animals should be humanely euthanized.

Chlamydophila psittaci infection has been seen in Xenopus sp., the African clawed frog. Clinical signs are similar to "red leg" and treatment is with doxycycline (5-10 mg/kg PO SID-BID).¹

Mycotic Infection

Saprolegnia, Basidiobolus, and Mucor spp. are ubiquitous aquatic molds that opportunistically infect local skin defects or contaminate egg masses. Contaminated eggs should be removed and animals can be treated with salt, malachite green, or copper sulfate baths.

The most important mycotic disease in amphibians and the most devastating infectious disease of amphibians worldwide is a chytridiomycete fungus, Batrachochytrium dendrobatidis. The organism is a primary pathogen that invades keratinized epithelial cells of the skin. The skin reacts with hyperplasia of keratinocytes. The only clinical signs that may be noticed are increased shedding and hunched posture to avoid substrate contact with the ventrum. The most common presentation, however, is death. Itraconazole bath (.01% solution 5-10 min/day x 10 d) is an effective treatment if cases are identified prior to death.¹ Antemortem diagnosis may be possible with cytologic examination of skin scrapes or shed skin and infection is confirmed with PCR of skin swabs. Feet and ventral surfaces of the inguinal region and rear legs are infected first and when collecting swabs for PCR, these sites should be included.

Parasites

Fecal examination is by routine methods, though in small samples wet mount is the most useful. Flagellate and ciliate protozoa are commensal organisms and should be seen normally in feces of healthy amphibians. Debilitated amphibians may develop overgrowth of protozoa, especially trichomonad flagellates. Metronidazole (10 mg/kg SID x 5-10 d) can be used to help reduce trichomonad populations to normal levels.¹

Rhabditiform nematodes are common in captive amphibians. Rhabdias sp. lungworms or Strongyloides sp. intestinal worms are typically seen. Both have direct life cycles and parasite burdens will exponentially increase in enclosed environments. Diagnosis is made by identification of nematode larvae or embryonated ova on fecal exam. Larvae have a needle-like tip to the tail, which can be seen microscopically. Fenbendazole (50-100 mg/kg PO, repeat in 10-14 days) is the safest and most effective treatment.¹ Substrate should be replaced with each treatment to remove infective larvae and prevent reinfection.

Edema Syndrome

Edema syndrome is not a disease in itself, but is a common presentation of many different etiologies. Animals present with hydro-coelom, generalized edema, or both. It may be caused by sepsis, osmotic imbalance, renal failure, lymph heart or cardiac dysfunction, chronically retained egg masses, or other systemic disturbances. Treatment should be broad spectrum and supportive until a primary etiology can be determined.

Treatment Notes

When presented with an ill amphibian due to any cause, there are initial supportive treatments that all animals should receive. Most important is fluid therapy to restore or support osmotic balance. A recipe for an amphibian Ringer's stock solution can be made with 6.6 g NaCl, 0.15g KCl, 0.15g CaCl₂, and 0.2g NaHCO₃ in 1 L distilled water.¹ The stock solution is then diluted 1:10 and used as a bath.

Initial treatment of sick amphibians should also include empirical antibiotic therapy. If the animal is not septic at time of presentation, it is safe to assume that it soon will be.

Topical administration of medications is equivalent to a parenteral route in amphibians, as the permeable skin will absorb drugs systemically. The pelvic drink patch is the best location in species with less permeable skin.

A sometimes effective "shotgun" approach to moribund amphibians includes fluid therapy, antibiotics, calcium gluconate (100mg/kg TO or SQ), vitamin B (1-2 drops TO), and cool ambient temperature. Once the animal is stabilized, diagnostics may elucidate need for other treatments, such as metronidazole or itraconazole.

Nutrition of ill patients should not be overlooked. If an animal is not eating on its own within 2-3 days, consider force feeding or tube feeding. Any carnivore based recovery diet is suitable for tube feeding. One product the author has used successfully is Emeraid® Carnivore (Lafeber Co., Cornell, Illinois 61319), which is supplied as a powder that can be mixed with water. The author usually tube feeds small patients every 2-3 days if anorexic and volume can be determined by a rough estimate of the animal's gastric capacity (e.g. 0.1-0.3ml in small dendrobatids).

References

1.  Wright, K.M. and B.R. Whitaker (eds.). 2001. Amphibian Medicine and Captive Husbandry. Krieger Publishing Company, Malabar, Florida.

2.  Wright, K.M. 2006. Overview of amphibian medicine. In: Mader, D.R. (ed.). Reptile Medicine and Surgery, 2^nd ed. Saunders Elsevier, St. Louis, Missouri. Pp. 941-971.