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Diseases of Xenopus tropicalis


PREMETAMORPHS
Bacterial Infection of Embryos
Symptoms: Embryos disintegrate due to a bacterial infection.
Treatment: Maintaining embryos in gentamicin (100 µg/mL) from fertilization to stage 41, when they are free-swimming, is effective at killing bacterial pathogens. Also, it is essential to keep the embryos at a low density, because a high density breeds infection, and to quickly remove any unfertilized eggs or dead embryos. In fact, at high density, embryos are susceptible to bacterial infection and death despite gentamicin. In a 90 mm Petri dish, we recommend raising only 100 embryos. We have raised higher numbers than this (200 or so), but at much higher numbers, you greatly increase the risk of bacterial infection. The embryos should look very sparse in the dish.


Protozoal Infestation of Tadpoles

Symptoms: Tadpoles that should, developmentally, be free swimming appear lethargic and sessile. The ciliated protozoan can be viewed at high magnification under a stereomicroscope (Fig. 1). Heavy infestation of the protozoans will lead to tadpole death.

Treatment: Although there is currently no effective means of ridding the pathogens from the water, the effects of the protozoa on the tadpoles can be diminished by releasing the tadpoles into a circulating flow of water at stage 41. Stage 41 correlates with day 2-3 of life if tadpoles are raised at 28ºC. If tadpoles are raised at 22ºC, the infestation becomes obvious at a stage when the tads are too young to be effectively transferred to a flow through system. Therefore, we recommend 28ºC as an incubation temperature.

Ciliated Protozoa

POSTMETAMORPHS
Mycobacteriosis – Caused by an acid-fast bacilli of the genus Mycobacterium. Many mycobacterial species have been found to cause infection in amphibians, including M. marinum, M. chelonae, and M. xenopi. The causative agent in our X. tropicalis colony has been proven, through molecular and phenotypic characterization, to be M. ulcerans-like and named M. liflandii. M. ulcerans is a water-borne pathogen endemic to tropical regions of Africa, Australia, Central America, and Eastern Asia that causes dermal-epidermal ulcerations in humans. The organism identified in our X. tropicalis colony, M. liflandii strain KT1, is most closely related, although not identical, to M. ulcerans by molecular analysis and has the growth characteristics of M. ulcerans. However, since the bacteria is related to M. ulcerans, a known human pathogen, it is recommended that personnel wear protective eyewear and gloves when handling the frogs.

Mycobacteriosis is a serious threat to the health of the colony. It is currently our leading cause of death, and the prevention of disease should always be a high priority.

Symptoms: The symptoms of amphibian mycobacteriosis are characterized by cutaneous and visceral milliary granulomas (white millet-like bumps) and ulcerative lesions on the skin. (Fig. 2a,c). The M. ulcerans-like pathogen present in our X. tropicalis colony, however, also causes coelomitis (infection of the coelom/abdomen) and subsequent general or local edema (bloating) (Fig. 2b). Cutaneous lesions and bloating both can occur independently. Once these signs develop, the disease is consistently fatal and affected frogs are usually humanely or obligatorily euthanized. An early sign can be loss of the diving reflex. X. tropicalis often float motionless at the surface of the water, but when startled, will dive to the bottom of the tank. Unhealthy frogs often lose this reflex, which can be used to easily screen a large tank of frogs.

A simple assay for the presence of any species of Mycobacterium is the acid-fast stain; both Ziehl Neelson and Auromine-Rhodamine Fluorescent stain work well (Fig. 2d,e). Both aspirated coelomic pus or heart blood and impression smears of infected organs are suitable samples for staining and confirmation of mycobacteriosis. In addition, we have developed assays for distinguishing M. liflandii from M. ulcerans.

Treatment: Currently, there is no cure for mycobacteriosis in frogs. Some species of Mycobacterium are susceptible to antibiotics in higher vertebrates, but there has been little success in curing mycobacteriosis in amphibians. M. ulcerans, specifically, is resistant to the majority of anti-mycobacterial drugs. Therefore, as of now, the only treatment is prevention. If a frog is suspected of having mycobacteriosis, it should immediately be removed from the colony, humanely euthanized and assayed for the presence of mycobacteria. The mycobacterial species affecting amphibians are water-borne, and therefore, a frog with a fulminant infection is highly likely to act as a source of infection for other frogs. Frogs from tanks with known exposure to mycobacteria should not be placed in the same container as non-exposed frogs. We have had deaths of frogs due to mycobacteriosis whose only contact with an exposed frog was through a natural mating overnight. Therefore, if a colony or group of frogs is identified as potential carriers of mycobacteria, they should remain physically isolated from other frogs at all times. We are currently attempting to develop a pathogen-free line of frogs and investigating the onset of mycobacteriosis in X. tropicalis.




Chytridiomycosis (Batrachochytrium dendrobatidis) – Chytrid pathogen of amphibian skin.

Symptoms: X. tropicalis infected with a chytrid show hyperpigmentation, dysecdysis (excessive sloughing of skin), loss of slime layer, excessive buoyancy, and death (Fig 3,4).

Treatment: Infected frogs should be treated with a solution of formalin and malachite green for 24 hours, with the concentration of formalin at 25ppm and 0.10 ml/L of malachite green, and repeated every other day for a total of four treatments, switching the frogs into fresh water on the off days (Parker et al). Note: X. laevis are known to harbor chytrids and be asymptomatic. Therefore X. laevis can act as a source of chytrid infection for X. tropicalis. We recommend strict isolation of X. tropicalis from X. laevis for this reason. Additionally warm temperatures inhibit chytrid growth so we keep our trops in rooms with ambient temperatures of 28-30ºC (slightly lower water temperatures).

*Parker, John M.; Mikaelian, Igor; Hahn, Nina; Diggs, Helen E. Clinical diagnosis and treatment of epidermal chytridiomycosis in African clawed frogs (Xenopus tropicalis). Comparative Medicine. June, 2002. 52 (3): 265-268.

Incidental Findings – Intestinal parasites found upon gross examination and not believed to be the causative agent of death or illness (Fig. 5,6).

Nematode -

Intestinal Ciliated Protozoan

contributed by Kristin Trott and John Parker