Xenopus Care, Health & Disease: A Brief Overview
Masha Rand and Jennifer Kalishman,
The Kelley lab maintains colonies
of several species of the pipid Xenopus:
Xenopus laevis, Xenopus tropicalis, and a Gabonese Xenopus species. The adult Xenopus laevis colony is derived from
commercial breeders (Nasco, http://www.enasco.com/prod/Home
and Xenopus One, http://www.xenopusone.com).
The Xenopus (Silurana) tropicalis are derived from a Nigerian stock from the
Grainger laboratory at The University of Virginia (http://faculty.virginia.edu/xtropicalis). In addition, we are developing a colony of
animals that Drs. Kelley and Tobias brought back from their 2001 field trip to
Following is a broad description of each colony, maintenance, mating, and rearing.
These colonies have provided a rich case study of both common and rare diseases and infections affecting the different Xenopus species. Here we describe the diseases encountered in our colonies, including symptoms, diagnosis, and treatment.
The Xenopus laevis colony ordinarily consists of 10-50 animals. These are housed at 18.3oC on a 12/12, light/dark cycle.
We use carbon-filtered water (mixed bed deionizer, US Filter), to which we add Novaqua (0.5ml/gallon) and NaCl (13.3g/gallon). The water is stored in large Nalgene carboys. After the tank is filled, the water is allowed to stand for one day before use. Water in the animals' tank is changed completely twice per week via a static renewal system.
The frogs are kept in clear,
polycarbonate tanks measuring 1'6"l x 9"w x 8"h. The tank covers are made of stainless steel
rims to which, neoprene mesh with 1/4" openings is attached. The lids are secured to the tanks with wire
and inspected frequently for holes. Frogs dessicate rapidly out of water. A maximum of five adult males or three adult
females are housed per tank in 4L of water.
The frogs are fed every other day with Nasco Frog Brittle (http://www.enasco.com/prod/Static?page=xen_brittle&seqid=4). They are fed approximately 3-5 pellets per frog or until they are satisfied (as judged by a cessation of feeding).
Of all the colonies, the X. laevis seem the most susceptible to disease. This is probably due to the fact that, with the high turnover rate, this colony most frequently contains recently shipped animals. Shipping animals causes stress, which leads to significant immunosuppression and flora or fauna that are present ordinarily may become pathogenic as a result (Tinsley, 1996). In addition, new animals could introduce novel pathogens into the colony.
Health & Disease:
Adult male X. laevis with snout lesions caused by Aeromonas infection.
For several months we witnessed the widespread occurrence of dermal ulcers. Snout lesions, such as the one above, were accompanied by similar lesions on the digits of the forelimbs and less frequently of the hind limbs; the skin appeared to be eaten away, as did the upper phalanges. In a few severe cases, the nasal septum was visible on the snout. The symptoms were either present in a nascent form upon arrival from the breeder, or else developed soon thereafter. Cultures of snout lesion tissue showed heavy growth of Aeromonas hydrophila/caviae. Necropsy of several symptomatic animals showed no obvious internal sign of infection. However, histological evidence indicated considerable liver damage, also due to Aeromonas infection.
Antibiotic treatment was attempted, administering 4 courses of gentamicin, subcutaneously, at a dose of 2mg/kg. However, the treated animals fared no better than a control group of equally symptomatic animals from the same frog shipment. Significantly, all animals lived - the lesions healed, leaving a faint scar. We did not try other antibiotics; possible alternatives are tetracycline or doxycycline.
Aeromonas hydrophila makes up a normal part of the fauna encountered by Xenopus in their environment and only under immunosuppressive conditions does it become pathogenic. Aeromonas, along with a host of other gram-negative bacteria, is responsible for Red Leg, a commonly encountered disease plaguing stressed Xenopus colonies. It is characterized by reddening on the abdomen and inner thighs, caused by vasodilation, and by internal hemorrhaging. Red Leg has a very high mortality rate if left untreated.
Finally, complete isolation of infected animals was successful in controlling the spread of infection. New animals were quarantined for a minimum of two weeks: they were housed in separate tanks and kept only on the bottom shelves to prevent water from one tank splashing into tanks with uninfected animals. When animals were transferred from shipping containers, none of the water or plant material in which they were shipped was transferred with them. Animals already in the colony are removed to quarantine tanks immediately upon detection of infection.
sign of infection appeared immediately following a change of water: a female
a tank of two began expelling blood from her mouth. Upon closer inspection, we noticed that the
extension of the dorso-lateral surface due to the
underlying, inflated lung, was missing from the right
side and the frog was tilted towards that side.
Necropsy revealed a fully deflated, right lung filled with
black/pustulous fluid. The bottom 3/4 of
the left lung was normally air-filled, but it was similarly necrotic at the
apex. The only other sign of infection
was a copious amount of recently clotted blood between the surface skin
and the muscle layers. Histological
examination of the lungs revealed a chromomycosis infection. Chromomycosis is an opportunistic fungal
infection caused by a variety of pigmented fungi.
The first symptoms of lymphosarcoma appeared as unusual antero-lateral swellings. These swellings increased in size. After approximately a month swellings were also present caudally in the dorsum as discrete lumps. All swellings were symmetrical, as is typical of lymphosarcoma (the lymph vessels are symmetrically arranged). Histological evidence supported the anatomical evidence, showing tissue from the lymphatic bumps, as well as from most of the internal organs (ovaries, fat bodies, liver, etc.) to be overrun by lymphocytes of a uniform size and shape.
Above: dorsal view of lymphatic tumors. Below: ventral view of same.
In addition, tumorous growths were evident on the hind limbs, both dorsally and ventrally. The entire lymphatic system was affected, in essence creating a perfect anatomical map of the otherwise difficult to visualize system.
Lymphosarcoma occurs very rarely in Xenopus laevis. There is some suggestion that it is related to mycobacterial infectious granuloma (Asfari, 1988). However, it is not clear whether the presence of Mycobacterium marinum in such cases is causative or a secondary infection due to weakening of the immune system (Clothier and Balls, 1973). In our case, no Mycobacterium marinum grew in culture. However, a negative result is not conclusive since mycobacteria are notoriously hard to culture.
The X. tropicalis colony is housed in the same 1'5"x9"x8" polycarbonate tanks and mesh covers as the X. laevis, in a 26oC warm room with a 12/12 hr light/dark cycle. Each tank contains a maximum of 12 animals. They are fed Zeigler's Salmon Starter Pellets, size #3, twice a week (http://www.zeiglerfeed.com/finfish.asp). Their water is made up of carbon-filtered water with Novaqua (0.5ml Novaqua/1L water) allowed to stand for 24 hours to release gases and to to warm to warm-room temperature. The water is changed twice a week, completely, several hours after feeding to allow frogs time to digest.
The colony consists of 40-70
animals, originally from the Grainger lab
as well as approximately 50 lab bred adults and juveniles. So far, the minimum
period from fertilization to sexual maturity has been three months.
This colony was founded with frogs
collected by Drs. Kelley and Tobias in Cap Esterias,
X. tropicalis cap esterias housing at the Kelley lab.
The colony currently contains approximately fifty wild-collected animals and approximately 300 lab-bred (both in vitro (link to in vitro protocol) and natural) tadpoles and juveniles. The animals are housed at 26oC on a 12/12hr, light/dark cycle in 5L or 7L Rubber MaidÔ containers with perforated snap-shut lids; maximum 4 animals per container.
The X. tropicalis cap esterias are cared for
almost exactly like the X. tropicalis
Health & Disease:
have encountered a dermal infection, occurring only in the X. tropicalis cap esterias colony. The symptoms are rust colored, diffuse
patches occurring dorsally on the torso and legs. White, fuzzy tufts localized
to the discolored areas can be distinguished in profile. Histological evidence
indicates a parasitic infection. Slides
of naturally shed skin with areas of discoloration show lengthy, winding
tunnels, occurring in localized areas and containing numerous brown eggs
inside. This matches Tinsley’s
description of the nematode Pseudocapillaroides xenopodis of which he says they “create tunnels in the
epidermis within which embryonated eggs and all
developmental stages occur” (Tinsley, 1996).
Oval shaped eggs in tunnel (indicated by arrow) surrounded by epithelial cells.
Slide made from skin shed by X. tropicalis cap esterias.
Treatment with a 5% Potassium Permanganate (KMnO4) solution and with MarOxy, a commercially available freshwater fungal and bacterial medication, were not successful in eliminating the skin lesions. The infection does not to affect the general well being of the animal: level of activity and food intake remain normal in affected frogs. Finally, the symptoms remit and reappear spontaneously.
Able, D.J. 1988.
An economical, balanced diet for Xenopus.
Brown, L.E. and R.R. Rosati. 1997. Effects of three different diets on survival and growth of larvae of the African clawed frog Xenopus laevis. Progressive Fish-Culturist 59(1): 54-58.
Davys, J.S. 1986.
The breeding of Xenopus laevis
on a large scale in the laboratory.
Animal Technology: Journal of the
Dawson, D., T.W. Schultz and E.C. Shroeder. 1992. Laboratory care and breeding of the African clawed frog. Lab Animal 21(4): 31-36.
Deuchar, Elizabeth Marion. 1975. Xenopus: The South African Clawed
Etheridge, Albert L. and Stephen M.A.
Richter. 1978. Xenopus
laevis: Rearing and Breeding the African Clawed Frog. NASCO, Publishing Agencies,
sHilken, G., F. Iglauer, H.-P. Richter, Kathleen M. Cromwell, J. Dimigen and H. Kahler.
Xenopus laevis als Labortier: Biologie, Haltung, Zucht und experimentelle Nutzung (The Clawed Frog Xenopus laevis as a Laboratory Animal). F. Enke, Publishing
Hilken, G., J. Dimigen and F. Iglauer. 1995. Growth of Xenopus laevis under different laboratory rearing conditions. Laboratory Animals 29: 152-162.
Hoogstraten-Miller, S. and D. Dunham. 1997. Practical Identification Methods for African Clawed Frogs (Xenopus laevis). Lab Animal 26(7): 36-38.
Kaplan, M.L. 1993. An Enriched Environment for the African Clawed Frog (Xenopus laevis). Lab Animal 22(5): 25-27.
Major, N. and R.J. Wassersug. 1998. Survey of current techniques in the care and maintenance of the African clawed frog (Xenopus laevis). Contemporary Topics in Laboratory Animal Science 37(5): 57-60.
1978. South African Clawed Frog, Xenopus laevis: Rearing & Breeding
Mrozek, M., R. Fischer, M. Trendelenburg and U. Zillman. 1995. Microchip Implant System Used for Animal Identification in Laboratory Rabbits, Guineapigs, Woodchucks and in Amphibians. Laboratory Animals 29(3): 339-344
1979. The Care and Induced
Breeding of Xenopus laevis Laboratory
Rizzo, A.M., R. Gornati, F. Rossi, G. Bernardini and B. Berra. 1999. Effect of Maternal Diet on the Distribution of Phospholipids and their Fatty Acid Composition in Xenopus laevis Embryos. The Journal of Nutritional Biochemistry 10(1): 44-48.
Sackin, N. and Sackin, H. 1991. A new method for feeding captive frogs (Xenopus laevis). Lab Animal 20(8): 44-46.
Smith, J.M. and K.C. Stump. 2000. Isoflurane anesthesia in the African clawed frog (Xenopus laevis). Contemporary Topics in Animal Science 39(6): 39-42.
Asfari, Maryam. 1988. Mycobacterium-induced infectious Granuloma in Xenopus: histopathology and transmissibility. Cancer Research 48: 958-963.
Ber, Artur and Zieleniewski. Schozenia Zab Xenopus Laevis Daudin Obserwowane W Hodowli Zakladu Endokrynologii Am W Lodzi (Diseases of the Frogs Xenopus laevis daudin Observed in the
Colony at Endocrinological Department of the
Bercovier, H. & V. Vincent. 2001. Mycobacterial infections in domestic and wild animals. Revue scientifique et techinique (International Office of Epizootics) 20(1): 265-90.
Clothier, R.H. & M. Balls. 1973. Mycobacteria and lymphoreticular tumors in Xenopus laevis, the South African clawed toad. Oncology 28: 458-480.
Elsner, H.A., H.H. Honck, F. William, H.J. Kreienkamp & F. Iglauer. 2000. Poor quality of oocytes for Xenopus laevis used in laboratory experiments: prevention by use of antiseptic surgical techniques and antibiotic supplementation. Comparative medicine 50(2): 206-211.
Green, S.L., D.M. Bouley, R.J. Tolwani, K.S. Waggie, B.D. Lifland, G.M. Otto & J.E. Ferrell, Jr. 1999. Identification and management of an outbreak of Flavobacterium and meningosepticum infection in a colony of South African clawed frogs. Journal of the American Veterinary Medical Association 214(12): 1833-1838.
Howeth, E.W. 1984. Pathology of naturally occurring chlamydiosis in African clawed frogs (Xenopus laevis) [Chlamydia]. Veterinary pathology 21(1): 28-32.
Hubbard, G.B. 1981. Aeromonas hydrophila infection in Xenopus laevis water borne bacillus. Laboratory animal science 31(3): 297-300.
Iglauer, F., F. Willmann, G. Hilken, E. Huisinga & J. Dimigen. 1997. Anthelmintic treatment to eradicate cutaneous capillariasis in a colony of South African clawed frogs (Xenopus laevis). Laboratory animal science 47(5): 477-482.
Miller, J.C. & R. Landesman. 1977. Magnesium deficiency in embryos of Xenopus laevis. J. Embryol. Exp. Morpholo. 39, 97-113.
Pearson, M. 1996. Studies on the Role of Aeromonas spp. in bacterial septicaemias of cultured frogs. Aquaculture News 21(24).
Tinsley, R.C. 1996. Parasites in Xenopus. Pp. 233-261 in
R.C. Tinsely and H.R. Kobel,
(eds.), The Biology
IACUC Learning Module, includes section on health and disease http://www.ahsc.arizona.edu/uac/iacuc/xenopus/xenopus.shtml
Xenopus laevis Frog Colony Care, includes section on Health & Disease http://www.xlaevis.com
Xenopus Express husbandry and disease sites
Bibliography of amphibian diseases http://www.jcu.edu.au/school/phtm/PHTM/frogs/bibliog.htm
African Clawed Frog links