Sex-dependent resistance to the pathogenic fungus Cryptococcus neoformans

Abstract

Sex differences occur in most species and affect a variety of biological traits including morphology, behavior, and life history. The nematode Caenorhabditis elegans exists as a population of self-fertile hermaphrodites with occasional males, which differ anatomically and behaviorally from hermaphrodites. Here we show that male C. elegans also differ from hermaphrodites in their susceptibility to a fungal pathogen, Cryptococcus neoformans. Wild-type males show greater resistance than hermaphrodite animals to killing by this pathogen and this resistance can be induced in hermaphrodite animals by inappropriate activation of the male sex-determination pathway. Resistance is molecularly determined, rather than resulting from behavioral changes or reproductive differences, and requires the activity of the stress-response transcription factor DAF-16. Finally, we demonstrate that resistance to C. neoformans correlates broadly with longevity within the Caenorhabditis genus. Our results hint at an overlap between the pathways controlling immunity and longevity and raise the possibility that differential regulation of these pathways may contribute to sex-dependent and species-dependent variation.

Figure 1.

Male and hermaphrodite animals at the L4 stage were singled onto wells preseeded with C. neoformans JEC21 and monitored for survival over 8 days. While hermaphrodite animals show almost 100% lethality over this period, male survival is significantly higher (P < 0.0001), with >50% of animals surviving >8 days. This increase in survival rate is not due simply to the absence of “matricidal killing” in males, since sterile hermaphrodite animals (BA1 and SS104) do not suffer from matricidal killing but remain more sensitive than males (P < 0.001) to Cryptococcus-mediated killing.

Figure 2.

Phenotypically male (JK987, CB2823, CB2590) and hermaphrodite (NL4317, CB5475) L4 animals were singled onto wells preseeded with C. neoformans JEC21 and monitored for survival over 8 days. Note that sexually transformed tra-2 animals are significantly more resistant than either wild-type or sexually transformed hermaphrodite animals (P < 0.0001) to Cryptococcus-mediated killing. Sexually transformed males carrying mutations in tra-1, however, are fully susceptible to Cryptococcus-mediated killing.

Figure 3.

Male and hermaphrodite L4 animals were exposed to JEC21 and monitored for survival over 8 days. Wild-type and daf-16-deficient hermaphrodite animals show identical susceptibility to Cryptococcus-mediated killing. daf-16 males, however, are far more susceptible than wild-type males (P < 0.0001) to the pathogen, but remain significantly more resistant to killing than either wild-type or daf-16 hermaphrodites (P < 0.01). Thus DAF-16 is largely, but not exclusively, required for male-dependent resistance to Cryptococcus-mediated killing.

Figure 4.

(A) L4-stage hermaphrodite or female animals were singled onto C. neoformans JEC21 and monitored for survival. Female C. remanei and C. sp CB5161 animals were significantly more resistant to C. neoformans-mediated killing than wild-type C. elegans hermaphrodites (P < 0.0001 for both) or C. japonica females (P < 0.0001 for both). In contrast, females/hermaphrodites of the other species tested showed remarkably similar levels of susceptibility (P > 0.05 for each pairwise comparison). (B) L4-stage male and hermaphrodite/female animals were singled onto C. neoformans JEC21 and monitored for survival. Hermaphrodite/female animals from each of the four species tested showed similar levels of susceptibility to killing (P > 0.05), while both C. elegans (CB4856) males and C. japonica (DF5078) males were significantly more resistant to killing than hermaphrodites/females of the same species (P < 0.0001 and P < 0.001, respectively). Interestingly, however, male C. briggsae were either equally (AF16) or more (VT847) susceptible to killing by Cryptococcus than were hermaphrodite animals. Thus male-dependent resistance is not a ubiquitous feature of the Caenorhabditis genus.

Figure 4.

(A) L4-stage hermaphrodite or female animals were singled onto C. neoformans JEC21 and monitored for survival. Female C. remanei and C. sp CB5161 animals were significantly more resistant to C. neoformans-mediated killing than wild-type C. elegans hermaphrodites (P < 0.0001 for both) or C. japonica females (P < 0.0001 for both). In contrast, females/hermaphrodites of the other species tested showed remarkably similar levels of susceptibility (P > 0.05 for each pairwise comparison). (B) L4-stage male and hermaphrodite/female animals were singled onto C. neoformans JEC21 and monitored for survival. Hermaphrodite/female animals from each of the four species tested showed similar levels of susceptibility to killing (P > 0.05), while both C. elegans (CB4856) males and C. japonica (DF5078) males were significantly more resistant to killing than hermaphrodites/females of the same species (P < 0.0001 and P < 0.001, respectively). Interestingly, however, male C. briggsae were either equally (AF16) or more (VT847) susceptible to killing by Cryptococcus than were hermaphrodite animals. Thus male-dependent resistance is not a ubiquitous feature of the Caenorhabditis genus.