MHC genotypes associate with resistance to a frog-killing fungus

Abstract

The emerging amphibian disease chytridiomycosis is caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd). Amphibian populations and species differ in susceptibility to Bd, yet we know surprisingly little about the genetic basis of this natural variation. MHC loci encode peptides that initiate acquired immunity in vertebrates, making them likely candidates for determining disease susceptibility. However, MHC genes have never been characterized in the context of chytridiomycosis. Here, we performed experimental Bd infections in laboratory-reared frogs collected from five populations that show natural variation in Bd susceptibility. We found that alleles of an expressed MHC class IIB locus associate with survival following Bd infection. Across populations, MHC heterozygosity was a significant predictor of survival. Within populations, MHC heterozygotes and individuals bearing MHC allele Q had a significantly reduced risk of death, and we detected a significant signal of positive selection along the evolutionary lineage leading to allele Q. Our findings demonstrate that immunogenetic variation affects chytridiomycosis survival under controlled experimental conditions, confirming that host genetic polymorphisms contribute to chytridiomycosis resistance.

Fig. 1.

Bd infection and mortality vary among populations of L. yavapaiensis in nature and in the laboratory. (A) Field estimates of winter Bd infection prevalence (proportion of frogs testing positive for Bd; filled symbols) and mortality (proportion of Bd-positive frogs that died; open symbols) in five natural populations; 95% Clopper–Pearson binomial confidence intervals are indicated (modified from ref. 37). (B) Survival following Bd infection varies among individuals from the same five populations when reared in a growth chamber. Survival was tracked for 35 DPI with 1 × 10⁵ zoospores of Bd (colored lines) or sham infection (black line). CIC1 and CIC2 denote genetically distinct clutches from population CIC. Numbers of individuals per population (Bd-infected/uninfected) are as follows: AC, 17/11; CIC, 34/22; MR, 5/2; WC, 17/11; SM, 26/15; all, 99/59. (C) Mean Bd infection intensity (zoospore equivalents ± SD) measured at various DPI intervals in populations with no survival (CIC, MR, and WC). (D) Mean Bd infection intensity over time for survivors (open symbols; AC_S and SM_S) and nonsurvivors (filled symbols; AC_D and SM_D) from populations with survivors (AC and SM). Control frogs (black symbols) remained uninfected for the duration of the experiment.

Fig. 2.

(A) Clutch genetic structure inferred by Bayesian assignment of 99 Bd-infected L. yavapaiensis individuals. Thin bars separate individuals, thick bars separate clutches, and asterisks indicate individuals that survived experimental infection. Horizontal bars show the coefficient of membership across genetic demes for each Bd-infected frog, with each color representing a unique genetic deme. (B) ML genealogy of 33 MHC class IIB PBR alleles. Circle size is proportional to frequency, colors correspond to the genetic demes in A, alleles recovered from survivors are shown with open symbols, and alleles recovered from nonsurvivors are shown with filled symbols. Bold denotes terminal branches with significantly elevated nonsynonymous substitution rates (i.e., ω > 1) followed by the codon positions under selection in parentheses. Alleles showing significant associations with survival are enlarged and italicized. Bootstrap values greater than 50% are indicated.

Fig. 3.

MHC class IIB PBR allele and heterozygote frequencies for Bd-infected survivors and nonsurvivors. PBR allele distribution (A) and PBR heterozygote and homozygote frequency (B) among survivors (open bars) and nonsurvivors (filled bars) in AC (red bars) and SM (blue bars), the two genetic demes with survivors. Allele counts and numbers of heterozygous and homozygous individuals are indicated above frequency bars. (C) Survival curves for PBR heterozygotes (solid line) and homozygotes (dotted line) across Bd-infected individuals from all five genetic demes.