Ergosterol’s Double Game: How a Fungal Membrane Molecule Shapes Termite Immunity

Deep in the soil, subterranean termites wage a constant chemical war against one of nature’s most lethal insect killers: the fungus Metarhizium. These social insects, whose thin cuticles offer little physical protection, have evolved a remarkable collective defense in which workers detect fungal spores clinging to their nestmates and groom them away before germination can begin. A new study published in The Science of Nature by Jacob H. Rosenbloom, Bhawana Luitel and Mark S. Bulmer of Towson University now dissects the molecular logic behind this hygiene system, and the findings reveal an unexpected evolutionary trade-off centered on a single fungal molecule: ergosterol, the cholesterol-like sterol that stabilizes fungal cell membranes.

When Reticulitermes flavipes workers encounter spores of Metarhizium adhering to a nestmate’s cuticle, they respond with brief bouts of rapid, whole-body shaking, a behavior the researchers call longitudinal oscillatory movement. This alarm display functions as a call for help, soliciting allogrooming from nearby colony members. Workers use their mouthparts to strip conidia from each other’s bodies, and the ingested spores are effectively destroyed, aided by salivary antimicrobial compounds including beta-1,3-glucanases and a small, cysteine-rich antifungal peptide called termicin. The critical open question has been which molecular signature, or pathogen-associated molecular pattern, actually triggers this rapid hygienic response. Ergosterol, an essential component of fungal membranes, was a leading candidate, particularly after earlier work showed that applying ergosterol to the cuticle of the Formosan subterranean termite Coptotermes formosanus elicited increased allogrooming.

To test this hypothesis rigorously, the team collected thirteen clonally propagated Metarhizium isolates from soils near active termite foraging sites across five US states, representing three species: Metarhizium robertsii, M. brunneum and M. guizhouense. Species and strains were confirmed by sequencing the translation elongation factor 1-alpha gene along with ITS and intergenic spacer regions. The researchers then quantified ergosterol in each isolate’s conidia using a saponification and spectrophotometric method originally developed for measuring ergosterol in Candida, calculating sterol content as a percentage of wet spore weight and correcting for the late pathway intermediate 24(28)-dehydroergosterol, which absorbs at a different wavelength.

The behavioral experiments produced a striking and initially counterintuitive result. In preference assays, termites given a choice between filter paper halves treated with ergosterol, cholesterol or solvent alone aggregated on the sterol-treated sides, but only ergosterol significantly elevated alarm shaking and allogrooming. Filter papers treated with methanol extracts of ergosterol from conidia also triggered alarm responses that correlated positively with the ergosterol content of the source isolates. Yet when termites were challenged with intact, living conidia, the intensity of the alarm response was not positively correlated with ergosterol content at all. Instead, the relationship trended in the opposite direction, suggesting that while ergosterol can serve as a recognition cue on its own, it is not the initial signal that alerts termites to dangerous contamination on a nestmate’s body.

Even more revealing was the correlation between ergosterol and pathogenicity. Across the thirteen isolates, spore ergosterol content was significantly and positively associated with the hazard ratio of death in Cox regression survival analyses, meaning that isolates richer in membrane ergosterol were more lethal to termites, not less. Adhesion assays ruled out the possibility that the most alarming isolate simply deposited more spores on the cuticle, and thin-layer chromatography showed no peroxidation or amino acid acylation of the sterol that might have altered its detectability. The picture that emerged was of a fungus in which high ergosterol somehow confers a survival advantage during the critical early hours after contact with a termite host.

The key to that advantage appears to lie in termicin, the termite’s principal antifungal weapon. When the researchers incubated conidia from each isolate in a five micromolar solution of recombinant R. flavipes termicin, the peptide’s fungicidal activity varied dramatically across isolates and correlated strongly with both virulence and ergosterol content. Termicin was most effective against the most lethal isolates, those with the highest ergosterol levels, and least effective against the least alarming, low-ergosterol strains. In vivo confirmation followed: pretreating conidia of the high-ergosterol isolate MD 002 with termicin significantly improved termite survival over fourteen days, while the same treatment offered no protection against the low-ergosterol isolate MD 353. Because termicin is an amphipathic peptide believed to disrupt fungal membranes, membrane ergosterol content plausibly determines how vulnerable a spore is to this assault.

Together, these findings sketch a mechanistic model of coupled, counterbalancing traits. Fungal cell wall integrity is known to be influenced by membrane ergosterol, and studies of antifungal drugs that block ergosterol synthesis show that reducing membrane sterol is compensated by reinforcing the cell wall. The Towson team proposes that Metarhizium isolates with lower membrane ergosterol develop cell walls richer in whatever unidentified pathogen-associated molecular pattern termites actually detect, making those spores easier to recognize and triggering vigorous allogrooming. Conversely, isolates with high ergosterol are harder for termicin to kill but present a weaker initial alarm signal. The fungus thus faces a genuine trade-off between a shield that protects against the peptide weapon and a signal that betrays it to the colony’s hygiene patrol, and the termites appear to have adaptively exploited exactly this vulnerability.

The evolutionary dimension of the story is equally compelling. Previous work documented a recent selective sweep for an advantageous termicin variant in sympatric populations of R. flavipes and R. virginicus in Maryland, and the two species respond differently to M. brunneum and M. robertsii, hinting at co-adaptation to local fungal lineages. In the new study, genetic distances between Metarhizium isolates correlated significantly with differences in the alarm responses their conidia elicited, even after controlling for collection latitude, indicating that the true recognition cue has diverged along with fungal phylogeny. Ergosterol content, by contrast, showed no correlation with genetic distance, suggesting it is a more plastic trait shaped by environmental conditions. Consistent with that idea, prior research on Argentine ants found that serial laboratory exposure to small ant groups drove Metarhizium to reduce its ergosterol and evade social immunity, a shift that would simultaneously increase the wall-based signal termites detect.

Ergosterol may also play a subtler secondary role in the hygiene cascade. The authors speculate that when termicin disrupts the membranes of high-ergosterol spores, released sterol could then act as a secondary elicitor, amplifying aggregation and allogrooming precisely when the initial wall-based alarm is weak. Residual ergosterol has been detected on the cuticles of C. formosanus workers an hour after Metarhizium exposure, lending plausibility to this compensation mechanism. Candidate molecules for the primary cue include cell wall beta-glucans, although inhibiting termite beta-1,3-glucanases did not reduce allogrooming in earlier experiments, leaving the identity of the true trigger unresolved. What is clear is that termite survival rests on a finely tuned balance between a behavioral response that is energetically costly and a peptide weapon whose effectiveness varies with the biochemistry of each fungal strain it meets.

Beyond its elegance as an evolutionary case study, the work carries practical implications. Metarhizium species are widely deployed as biological control agents against termites and other soil pests, and understanding why some isolates fail, namely because they trigger strong social immunity while resisting termicin, could guide the selection or engineering of strains that slip past termite defenses. Conversely, the finding that ergosterol content can shift plastically under selection pressure suggests that pathogens can rapidly adapt to dense, patchily distributed social insect populations. For now, the termites hold their ground through a two-layered strategy: an unidentified cell wall signal that sounds the alarm, and a membrane-targeting peptide that strikes hardest at the strains most adept at staying quiet. The arms race beneath the forest floor, it turns out, is negotiated one sterol molecule at a time.

Subject of Research: The role of ergosterol in termite social immunity and fungal pathogen detection against Metarhizium infections

Article Title: The influence of ergosterol on termites’ defenses against Metarhizium infections

Article References: Rosenbloom, J. H., Luitel, B., & Bulmer, M. S. (2026). The influence of ergosterol on termites’ defenses against Metarhizium infections. The Science of Nature, 113(6), Article 130. https://doi.org/10.1007/s00114-026-02183-3

Image Credits: AI Generated

DOI: 10.1007/s00114-026-02183-3

Keywords: ergosterol, termites, Metarhizium, social immunity, allogrooming, termicin, antifungal peptide, pathogen-associated molecular pattern, Reticulitermes flavipes, entomopathogenic fungus, fungal cell membrane, host-pathogen coevolution