Design- and model-based recommendations for detecting and quantifying an amphibian pathogen in environmental samples.

Area of Science:

• Environmental microbiology
• Wildlife disease ecology
• Amphibian pathology

Background:

• Accurate pathogen detection is crucial for managing emerging infectious diseases in wildlife.
• Sampling free-living pathogens via environmental DNA (eDNA) offers advantages but faces challenges.
• Batrachochytrium dendrobatidis (Bd) is a significant threat to amphibian populations globally.

Purpose of the Study:

• To evaluate the impact of pathogen concentration, water type, and qPCR inhibitors on Bd detection and quantification using water filtration.
• To compare results before and after inhibitor removal and assess inferential differences between single and multiple sample collections.
• To determine the influence of these factors on Bd occurrence and abundance estimates in environmental samples.

Main Methods:

• Laboratory experiment using water filtration to detect and quantify Batrachochytrium dendrobatidis (Bd).
• Comparison of quantitative polymerase chain reaction (qPCR) results pre- and post-inhibitor removal.
• Assessment of inferential biases resulting from single versus multiple spatial or temporal sample collections.

Main Results:

• qPCR inhibition significantly biased Bd detection and quantification in natural water samples, affecting inferences on occurrence and abundance.
• Collecting multiple samples in space or time mitigated occurrence biases but did not resolve quantification biases.
• Varying Bd concentrations influenced detection probability, suggesting occupancy modeling for abundance heterogeneity.

Conclusions:

• Environmental DNA (eDNA) detection of Bd is susceptible to qPCR inhibition in natural waters.
• Study design, particularly sample replication, is critical for accurate inference of amphibian pathogen dynamics.
• Findings inform eDNA-based wildlife health monitoring and species distribution studies.