Swabbing Often Fails to Detect Amphibian Chytridiomycosis under Conditions of Low Infection Load

Abstract

The pathogenic chytrid fungus, Batrachochytrium dendrobatidis (denoted Bd), causes large-scale epizootics in naïve amphibian populations. Intervention strategies to rapidly respond to Bd incursions require sensitive and accurate diagnostic methods. Chytridiomycosis usually is assessed by quantitative polymerase chain reaction (qPCR) amplification of amphibian skin swabs. Results based on this method, however, sometimes yield inconsistent results on infection status and inaccurate scores of infection intensity. In Asia and other regions where amphibians typically bear low Bd loads, swab results are least reliable. We developed a Bd-sampling method that collects zoospores released by infected subjects into an aquatic medium. Bd DNA is extracted by filters and amplified by nested PCR. Using laboratory colonies and field populations of Bombina orientalis, we compare results with those obtained on the same subjects by qPCR of DNA extracted from swabs. Many subjects, despite being diagnosed as Bd-negative by conventional methods, released Bd zoospores into collection containers and thus must be considered infected. Infection loads determined from filtered water were at least 1000 times higher than those estimated from swabs. Subjects significantly varied in infection load, as they intermittently released zoospores, over a 5-day period. Thus, the method might be used to compare the infectivity of individuals and study the periodicity of zoospore release. Sampling methods based on water filtration can dramatically increase the capacity to accurately diagnose chytridiomycosis and contribute to a better understanding of the interactions between Bd and its hosts.

Host contact and shedding patterns clarify variation in pathogen exposure and transmission in threatened tortoiseGopherus agassizii: implications for disease modelling and management

“…These solitary positives could indicate low infection levels that avoid detection (Shin et al . ), but we suspect such isolated positives in our study are not indicative of an established infection. False‐positives can occur in single PCR tests, but are less likely when repeated sampling methods are used to diagnose infection (Cuenca‐Estrella et al .…”

American Bullfrogs (Lithobates catesbeianus) Resist Infection by Multiple Isolates of Batrachochytrium dendrobatidis, Including One Implicated in Wild Mass Mortality

“…Furthermore, the only previous indication of American bullfrog susceptibility to chytridiomycosis in a laboratory setting (Gervasi et al 2013b), which used Bd strain JEL 274 isolated from Colorado, is atypical in that it attributed mortality to disease at zoospore loads (<10 ZE) far below those commonly thought necessary for severe chytridiomycosis that leads to death (*10,000 ZE) (Carey et al 2006;Voyles et al 2009;Vredenburg et al 2010;Kinney et al 2011). We note that detection of Bd via swabbing can be challenging, especially at low infection intensities (Shin et al 2014), and were evaluated using a quantitative PCR assay. Data from Finley Lake Bd-exposed animals are shown as squares connected by dotted lines, Carter Meadow Bd-exposed animals as circles connected by dashed lines, and Section Line Bd-exposed animals as triangles connected by solid lines.…”

Microbiome Variation Across Amphibian Skin Regions: Implications for Chytridiomycosis Mitigation Efforts

“…Thus, the bacterial community structure on the ventral skin should be more affected than other body regions by Bd infection. Also, adults with sublethal infection and those tolerant of-or resistant to-the disease may have localized Bd infections [29,30]. Then, changes in bacterial community structure may be observable only in small areas of infection.…”