A Fungal Pathogen of Amphibians, Batrachochytrium dendrobatidis, Attenuates in Pathogenicity with In Vitro Passages

Abstract

Laboratory investigations into the amphibian chytrid fungus, Batrachochytrium dendrobatidis (Bd), have accelerated recently, given the pathogen’s role in causing the global decline and extinction of amphibians. Studies in which host animals were exposed to Bd have largely assumed that lab-maintained pathogen cultures retained the infective and pathogenic properties of wild isolates. Attenuated pathogenicity is common in artificially maintained cultures of other pathogenic fungi, but to date, it is unknown whether, and to what degree, Bd might change in culture. We compared zoospore production over time in two samples of a single Bd isolate having different passage histories: one maintained in artificial media for more than six years (JEL427-P39), and one recently thawed from cryopreserved stock (JEL427-P9). In a common garden experiment, we then exposed two different amphibian species, Eleutherodactylus coqui and Atelopus zeteki, to both cultures to test whether Bd attenuates in pathogenicity with in vitro passages. The culture with the shorter passage history, JEL427-P9, had significantly greater zoospore densities over time compared to JEL427-P39. This difference in zoospore production was associated with a difference in pathogenicity for a susceptible amphibian species, indicating that fecundity may be an important virulence factor for Bd. In the 130-day experiment, Atelopus zeteki frogs exposed to the JEL427-P9 culture experienced higher average infection intensity and 100% mortality, compared with 60% mortality for frogs exposed to JEL427-P39. This effect was not observed with Eleutherodactylus coqui, which was able to clear infection. We hypothesize that the differences in phenotypic performance observed with Atelopus zeteki are rooted in changes of the Bd genome. Future investigations enabled by this study will focus on the underlying mechanisms of Bd pathogenicity.

SUSCEPTIBILITY TO THE AMPHIBIAN CHYTRID FUNGUS VARIES WITH ONTOGENY IN THE DIRECT-DEVELOPING FROG,ELEUTHERODACTYLUS COQUI

“…Exposure to a higher Bd dose and prior infection with Bd did not affect survival, and most frogs cleared infections or reduced them to low levels. This finding aligns with other research showing that adult coquis can survive and clear infection when exposed to Bd under normal climate conditions Langhammer et al 2013) and the observation that adult frogs can eliminate natural Bd infections within a few weeks of arriving in the lab (P.F.L. unpubl.…”

“…Clinical signs of chytridiomycosis were not obvious in juveniles, and there were no discernible differences in behavior between exposed and control groups. However, there is little published information on normal behavior of very young juveniles, and adult frogs that die with high Bd loads often show no outward signs of chytridiomycosis (Langhammer et al 2013;P.F.L. unpubl.…”

“…Bd is suspected in the decline and extinction of three frog species from Puerto Rico in recent decades (Burrowes et al 2004), and ongoing declines have been reported for coquis and other Eleutherodacylus species at midelevations in eastern Puerto Rico . Recent laboratory work indicates that adult coquis may have low susceptibility to Bd, at least under normal climatic conditions Langhammer et al 2013), and moribund adults are rarely found in the field (Longo et al 2013). Juvenile coquis show higher prevalence and intensity of Bd infection compared with adults in El Yunque, Puerto Rico , but it is unclear whether juveniles are susceptible to Bd.…”

Evaluating the probability of avoiding disease‐related extinctions of Panamanian amphibians through captive breeding programs

“…Atelopus are charismatic, diurnal toads that normally exist at high densities, so their absences have been particularly noticeable compared to other more cryptic or rare species (Lindquist & Swihart, 1997;La Marca et al, 2005). The hypersensitivity of Atelopus to chytridiomycosis has been demonstrated in both laboratory and field settings that validate this assessment (Woodhams et al, 2006;Bustamante, Livo & Carey, 2010;Richards-Zawacki, 2010;Becker et al, 2012;Langhammer et al, 2013).…”

“…In both cases, efforts should be made to survey known localities of these species and obtain more reliable information on their status, particularly if their congeners are Bd susceptible. Genomic material should be banked for these species including living cell lines, living gametes and cryopreserved tissues (Lermen et al, 2009;Kouba et al, 2013).…”