Trachymyrmex septentrionalisants promote fungus garden hygiene usingTrichoderma-derived metabolite cues

Abstract

AbstractFungus-growing ants depend on a fungal mutualist that can fall prey to fungal pathogens. This mutualist is cultivated by these ants in structures called fungus gardens. Ants exhibit weeding behaviors that keep their fungus gardens healthy by physically removing compromised pieces. However, how ants detect diseases of their fungus gardens is unknown. Here, we applied the logic of Koch’s postulates using environmental fungal community gene sequencing, fungal isolation, and laboratory infection experiments to establishTrichodermaspp. as previously unrecognized pathogens ofTrachymyrmex septentrionalisfungus gardens. Our environmental data showed thatTrichodermaare the most abundant non-cultivar fungi in wildT. septentrionalisfungus gardens. We further determined that metabolites produced byTrichodermainduce an ant weeding response that mirrors their response to liveTrichoderma. Combining ant behavioral experiments with bioactivity-guided fractionation and statistical prioritization of metabolites inTrichodermaextracts demonstrated thatT. septentrionalisants weed in response to peptaibols, a specific class of secondary metabolites known to be produced byTrichodermafungi. Similar assays conducted using purified peptaibols, including the two new peptaibols trichokindins VIII and IX, suggested that weeding is likely induced by peptaibols as a class rather than by a single peptaibol metabolite. In addition to their presence in laboratory experiments, we detected peptaibols in wild fungus gardens. Our combination of environmental data and laboratory infection experiments strongly support that peptaibols act as chemical cues ofTrichodermapathogenesis inT. septentrionalisfungus gardens.Significance StatementAn extended defense response may exist in any relationship where one partner benefits from defending a mutualistic partner. Such a response is observed in the fungus-growing ant symbiosis, where ants must identify and remove pathogens of their symbiotic fungus gardens. Here we describe the fungal pathogenTrichodermaand its associated metabolites, which induceTrachymyrmex septentrionalisant weeding behavior. Ants removed fungus garden pieces inoculated withTrichodermaspores or peptaibol-richTrichodermaextracts, and peptaibols as a class cued ant defensive behavior, allowingT. septentrionalisto differentiate healthy from diseased fungus gardens. Extended defense responses mediated by chemical cues may be underappreciated mechanisms that structure symbiotic interactions.