DAO 80:63-67 (2008)  -  DOI: https://doi.org/10.3354/dao01919

ABSTRACT: Batrachochytrium dendrobatidis, an aquatic amphibian fungus, has been implicated in many amphibian declines and extinctions. A real-time polymerase chain reaction (PCR) TaqMan® assay is now used to detect and quantify B. dendrobatidis on amphibians and other substrates via tissue samples, swabbing and filtration. The extreme sensitivity of this diagnostic test makes it necessary to rigorously avoid cross-contamination of samples, which can produce false positives. One technique used to eliminate contamination is to destroy the contaminating DNA by chemical means. We tested 3 concentrations of sodium hypochlorite (NaOCl) (1, 6 and12%) over 4 time periods (1, 6, 15 and 24 h) to determine if NaOCl denatures B. dendrobatidis DNA sufficiently to prevent its recognition and amplification in PCR tests for the fungus. Soaking in 12% NaOCl denatured 100% of DNA within 1 h. Six percent NaOCl was on average 99.999% effective across all exposure periods, with only very low numbers of zoospores detected following treatment. One percent NaOCl was ineffective across all treatment periods. Under ideal, clean conditions treatment with 6% NaOCl may be sufficient to destroy DNA and prevent cross-contamination of samples; however, we recommend treatment with 12% NaOCl for 1 h to be confident all B. dendrobatidis DNA is destroyed.

KEY WORDS: Chytridiomycosis · Denaturation · Field hygiene · PCR · Diagnostic test