Outbreak of parasite-induced limb malformations in a declining amphibian species in Colorado

The detection of severe limb malformations in metamorphosing northern leopard frogs (Rana pipiens) from a Colorado pond in August 2022 prompted questions about the cause(s) and concern over the implications. Northern leopard frogs, which are a Tier 1 Species of Greatest Conservation Need in Colorado, have declined over much of their range in the state, particularly along the Front Range. Although malformations in amphibians have been reported in other parts of the USA, they are rare in Colorado, and the current case represents the most severe hotspot reported in the state for over 70 years. Across three survey events in late summer and early fall of 2022, approximately 68% of captured leopard frogs (late-stage larvae and metamorphic frogs) exhibited one or more malformations. Malformations exclusively affected the hind limbs and were dominated by skin webbings (51.7% of the total), bony triangles (32.2%), and extra limbs or digits (11%). Many animals had multiple malformations that limited the movement of one or both limbs (average of 2.3 malformations per malformed frog). Dissection of a subset of animals coupled with 28S rDNA genetic sequencing revealed the occurrence of the trematode Ribeiroia ondatrae at an average of 75.2 trematode cysts (metacercariae) per frog. The parasite was also detected in 2.6% of dissected snails (Helisoma trivolvis), which function as the trematode's first intermediate host. The relatively high loads of infection detected here - coupled with the similarity of observed malformations to those previously linked to R. ondatrae in experimental studies and from other malformation hotspots in the USA - offer compelling evidence that the current case is the result of parasite infection. Unresolved questions include why malformation prevalence was so high in 2022 and the degree to which such abnormalities will affect population persistence for local leopard frogs, particularly if malformations continue.