Identification of Paragonimus mexicanus and P. caliensis in freshwater crabs from Costa Rica: evidence of zoonotic lung fluke diversity in Central America.

Paragonimus spp. are foodborne trematodes with complex life cycles involving freshwater snails and decapod crustaceans. In Central America, Paragonimus mexicanus is a known zoonotic agent, whereas the public health significance of P. caliensis remains unclear. Between 2015 and 2016, we surveyed freshwater crabs across 51 sites in 15 river basins spanning three major climatic regions in Costa Rica. A total of 419 crabs (10 species) were examined for metacercariae; male crabs were identified morphologically, and infected females were identified to species by 16S rRNA and COI sequencing. Metacercariae were detected in six crab species, three of which represent new hosts of Paragonimus. The highest prevalence in male crabs occurred in Ptychophallus uncinatus (55.6%, 25/45). Molecular analyses confirmed the presence of two Paragonimus species in crabs: P. mexicanus and P. caliensis. A binomial GLM showed that males had higher odds of infection than females (p = 0.0059). Most infections occurred in areas along the Caribbean coast and in the Northern Region, which drains into Lake Nicaragua and Río San Juan. A binomial GLM revealed that the probability of Paragonimus infection varied significantly across climatic regions. Crabs from the Caribbean slope had the highest odds of infection, followed by the Northern Region. In contrast, crabs from the Pacific slope were significantly less likely to be infected. These patterns support the existence of geographic clusters of transmission within Costa Rica. We provide the first species-level, molecular confirmation of infected female crabs in Costa Rica and identify three freshwater crab species as newly recognized second intermediate hosts. These findings expand the known host range and distribution of Paragonimus in Costa Rica and highlight the need for public health education about the risks of consuming undercooked freshwater crabs.