Cool Temperatures Reduce Antifungal Activity of Symbiotic Bacteria of Threatened Amphibians – Implications for Disease Management and Patterns of Decline

Chytridiomycosis, caused by the fungus Batrachochytrium dendrobatidis (Bd), is a widespread disease of amphibians responsible for population declines and extinctions. Some bacteria from amphibians' skins produce antimicrobial substances active against Bd. Supplementing populations of these cutaneous antifungal bacteria might help manage chytridiomycosis in wild amphibians. However, the activity of protective bacteria may depend upon environmental conditions. Biocontrol of Bd in nature thus requires knowledge of how environmental conditions affect their anti-Bd activity. For example, Bd-driven amphibian declines have often occurred at temperatures below Bd's optimum range. It is possible these declines occurred due to reduced anti-Bd activity of bacterial symbionts at cool temperatures. Better understanding of the effects of temperature on chytridiomycosis development could also improve risk evaluation for amphibian populations yet to encounter Bd. We characterized, at a range of temperatures approximating natural seasonal variation, the anti-Bd activity of bacterial symbionts from the skins of three species of rainforest tree frogs (Litoria nannotis, Litoria rheocola, and Litoria serrata). All three species declined during chytridiomycosis outbreaks in the late 1980s and early 1990s and have subsequently recovered to differing extents. We collected anti-Bd bacterial symbionts from frogs and cultured the bacteria at constant temperatures from 8 °C to 33 °C. Using a spectrophotometric assay, we monitored Bd growth in cell-free supernatants (CFSs) from each temperature treatment. CFSs from 11 of 24 bacteria showed reduced anti-Bd activity in vitro when they were produced at cool temperatures similar to those encountered by the host species during population declines. Reduced anti-Bd activity of metabolites produced at low temperatures may, therefore, partially explain the association between Bd-driven declines and cool temperatures. We show that to avoid inconsistent antifungal activity, bacteria evaluated for use in chytridiomycosis biocontrol should be tested over a range of environmental temperatures spanning those likely to be encountered in the field.