Amphibian Hymenochirus boettgeri as an experimental model for infection studies with the chytrid fungus Batrachochytrium dendrobatidis

Abstract

Model organisms are crucial in research as they can provide key insights applicable to other species. This study proposes the use of the amphibian species Hymenochirus boettgeri, widely available through the aquarium trade, as a model organism for the study of chytridiomycosis, a disease caused by the fungus Batrachochytrium dendrobatidis (Bd) and linked to amphibian decline and extinction globally. Currently, no model organisms are used in the study of chytridiomycosis, particularly because of the lack of availability and nonstandardized methods. Thus, laboratories around the world use wild local species to conduct Bd infection experiments, which prevents comparisons between studies and reduces reproducibility. Here, we performed a series of Bd infection assays that showed that H. boettgeri has a dose- and genotype-dependent response, can generalize previous findings on virulence estimates in other species, and can generate reproducible results in replicated experimental conditions. We also provided valuable information regarding H. boettgeri husbandry, including care, housing, reproduction, and heat treatment to eliminate previous Bd infections. Together, our results indicate that H. boettgeri is a powerful and low-ecological-impact system for studying Bd pathogenicity and virulence.

Keywords: Batrachochytrium dendrobatidis; Hymenochirus boettgeri; Model organism; amphibians; husbandry; virulence.

Figure 1.

Photographs of housing systems and eggs, tadpoles and adults of H. boettgeri. Tecniplast (1a), standard (1b), and static (1c) systems, female (black arrow; 1d, e) and male (filled white arrow pointing to post-axillary subdermal gland and unfilled white arrow pointing to obiculated tympanum; 1d, e) adult individuals, nursery tank of the Tecniplast system (1f), eggs inside the nursery tank (1g), 4-d-old tadpoles feeding on powdered food (1h), and 12-d-old tadpoles feeding on brine shrimp (1i). Black bars represent 1 cm scale.

Figure 2.

Survival probability curves of JEL423 and CLFT073 Bd genotypes at high (HD) or low (LD) dose. p-value is from log-rank test.

Figure 3.

Survival curves and Bd load boxplot from the model generalizability and validation experiments. Survival curves for H. boettgeri exposed to six Bd genotypes from the model generalizability experiment, built at (a) lineage or (b) genotype level. (c) survival curves and (d) Bd load (log transformed at time of death) boxplots for H. boettgeri exposed to CLFT039 and CLFT150 genotypes from model generalizability (solid line/MG boxes) and validation experiments (dotted line/V boxes). Yellow, blue and green lines/boxes indicate that the Bd genotype belongs to hybrids, Bd-GPL and Bd-Brazil lineages, respectively. p-values are from log-rank test. Letters 'a' and 'b' inside the boxplots represent statistically significant differences from post-hoc multiple comparisons of means test among Bd genotype/experiments.