Behavioral Observations for all Amphibian Species
In the observational choice chamber, Greenhouse frogs and pine woods
treefrogs were exposed to Live Bd or Control treatments
(n = 13 or 9 amphibians/treatment, respectively; Table 1, Fig. 1)
as Bd-naïve and Bd-experienced individuals. Southern toads and Cuban
treefrogs were exposed to one of five treatments: Live Bd,
Control, Dead zoospores with metabolites, Dead zoospores alone, andMetabolites alone . The amphibians were randomly assigned to one
of the treatments (n = 20 and 15 amphibians/treatment,
respectively; Table 1).
Observation chambers were created by placing ASW moistened paper towels
on each side of the container leaving a 1 cm gap in the center. For the
Cuban treefrogs, the paper towels and gap covered the bottom of the
container and extended vertically up the wall so that the treefrogs
could not avoid the two treatments. Within each container, the paper
towel on one side of the chamber was inoculated with 1 mL of the
treatment inoculate (see Treatment Inoculate Preparation ; Table
1) and the paper towel on the other side received 1 mL of ASW. The side
inoculated with the treatment was selected randomly and we implemented a
double-blind inoculation process so that the person running the
behavioral trials did not know the treatment location. For each
behavioral trial, the amphibian was placed in the center of the
container. After a 20-minute acclimation period, we recorded the
location of the amphibians in the container (left, center, or right)
every 15 min for 3-4 h depending on the species (southern toads: 4hrs;
all other species: 3hr).