Coinfection with chytrid genotypes drives divergent infection dynamics reflecting regional distribution patterns

Abstract

By altering the abundance, diversity, and distribution of species-and their pathogens-globalization may inadvertently select for more virulent pathogens. In Brazil's Atlantic Forest, a hotspot of amphibian biodiversity, the global amphibian trade has facilitated the co-occurrence of previously isolated enzootic and panzootic lineages of the pathogenic amphibian-chytrid (Batrachochytrium dendrobatidis, 'Bd') and generated new virulent recombinant genotypes ('hybrids'). Epidemiological data indicate that amphibian declines are most severe in hybrid zones, suggesting that coinfections are causing more severe infections or selecting for higher virulence. We investigated how coinfections involving these genotypes shapes virulence and transmission. Overall, coinfection favored the more virulent and competitively superior panzootic genotype, despite dampening its transmission potential and overall virulence. However, for the least virulent and least competitive genotype, coinfection increased both overall virulence and transmission. Thus, by integrating experimental and epidemiological data, our results provide mechanistic insight into how globalization can select for, and propel, the emergence of introduced hypervirulent lineages, such as the globally distributed panzootic lineage of Bd.

Fig. 1. Phylogenetic relationships among Batrachochytrium dendrobatidis (Bd) genotypes, geographical distribution of their lineages, and experimental design for single- and mixed-genotype infections.

A Phylogenetic tree of Bd genotypes used in the challenge assay. B Spatial distribution of Bd lineages or hybrid genotypes identified from multiple amphibian species across the Brazilian Atlantic Forest (highlighted area in map) (Supplementary Data 2). On the map, symbols represent the location where we isolated the pathogen genotypes used in this experiment. White triangle: P1, black triangle: P2, diamond: E2, star: E1 and H. C Infection assay procedure. In order to prepare the Bd solutions, we flooded Bd cultures with distilled water, quantified Bd zoospore density using a hemocytometer, and adjusted all solutions to the same concentration (see “Methods” for additional experimental details). C was created with BioRender.com.

Fig. 2. Competitiveness and transmission potential of Batrachochytrium dendrobatidis (Bd) in both single- and mixed-genotype infections.

A–C The competitiveness of each genotype is represented as mean Bd load (± standard deviation [SD]) during the first time point of the assay (day 21 post inoculation). D–F Transmission potential is represented as the average Bd load (± SD) after death or on day 76 at the end of the experiment. Competitive suppression could either help reduce the transmission potential of more virulent genotypes (P1; E) or increase the transmission potential of less virulent genotypes (P2; F). Gray shades in figures highlight single-genotype infections. Infection loads are represented by Bd concentration in pg/μl.

Fig. 3. Variation in virulence across infection treatments.

Virulence, measured as host life span (A–C) and survival probability (D–F), varied across hosts infected with single- and mixed-genotypes of Batrachochytrium dendrobatidis (Bd). The presence of multiple genotypes reduced the overall virulence (i.e., increased host life span and survival) in those treatments involving the most virulent genotype (P1; B, E). Coinfections with the least virulent genotype (P2), however, did not influence host life span (C), though they had an effect on survival or median survival time (F). For survival curves, P values are from log-rank tests. Gray shades in figures highlight single-genotype infections. Points represent the mean host life span and bars the standard deviation.

Fig. 4. Competitive outcomes in mixed-genotype infections of Batrachochytrium dendrobatidis (Bd).

A–C Competitive outcomes for the mixed-genotype infections were based on the proportion of the total pathogen load (in pg/μl) that was either the most and least virulent genotype (P1 and P2, respectively).