Different metabolite profiles acrossPenicillium roquefortipopulations associated with ecological niche specialisation and domestication

Abstract

Fungi are known to produce many chemically diversified metabolites, yet their ecological roles are not always fully understood. The blue cheese making fungus Penicillium roqueforti thrives in different ecological niches and is known to produce a wide range of metabolites, including mycotoxins. Three P. roqueforti populations have been domesticated for cheese production and two populations thrive in other anthropized environments, i.e., spoiled food, lumber and silage. Here, we looked for differences in targeted and untargeted metabolite production profiles between populations using HPLC-HR-Q-TOF and UHPLC-Q-TOF-HR-MS/MS. The non-cheese populations produced several fatty acids and different terpenoids, lacking in cheese strains. The Termignon cheese population displayed intermediate metabolite profiles between cheese and non-cheese populations, as previously shown for other traits. The non-Roquefort population, the cheese population with the strongest domestication syndrome, produced the lowest quantities of measured metabolites, including known mycotoxins such as mycophenolic acid (MPA), andrastin A and PR toxin. Its inability to produce MPA was due to a deletion in the mpaC gene, while a premature stop codon in ORF 11 of the PR toxin gene cluster explained its absence and the accumulation of its eremofortin A & B intermediates. In the Roquefort population, we detected no PR toxin nor eremofortins A or B, but found no indel or frameshift mutation, suggesting downregulation. Our results suggest that domesticated cheese populations were selected for lower toxin production while populations from other anthropized environments maintained high metabolite diversity, the bioactivities of these compounds being likely important in these ecological niches.

Generation of diversity in the blue cheese moldPenicillium roquefortiand identification of pleiotropic QTL for key cheese-making phenotypes

“…For three extrolites (MPA, PR toxin and roquefortine C), we identified QTLs with intervals included in their biosynthesis cluster, with non-Roquefort alleles associated with lower production levels. The lower MPA production level associated with the non-Roquefort allele is likely due to the deletion in the mpaC gene that was identified in the non-Roquefort population (Gillot et al 2017; Crequer et al 2024). We also found a QTL in a region including the PR toxin biosynthesis gene cluster, with lower production level of the PR toxin but accumulation of its production intermediates, eremofortins A and B, associated with the non-Roquefort allele at the PR toxin biosynthesis cluster.…”

“…We also found a QTL in a region including the PR toxin biosynthesis gene cluster, with lower production level of the PR toxin but accumulation of its production intermediates, eremofortins A and B, associated with the non-Roquefort allele at the PR toxin biosynthesis cluster. This suggests a disruption in the cluster driving low production levels of the toxin and accumulation of its intermediates in offspring presenting the non-Roquefort allele likely due to a premature STOP codon in the ORF11 gene of the cluster (Crequer et al . 2024).…”

“…Toxin production is a concern in P. roqueforti , in particular the production of PR toxin and roquefortine C. The low toxin concentrations found in cheese and the long history of safe consumption of blue cheeses, combined with the recent discovery that both Roquefort and non-Roquefort populations are unable to produce PR toxin, indicate the innocuity of commercial strains in cheese production (Crequer et al . 2024).…”

“…For extrolite extractions, we used an optimized “high-throughput” extraction method (Lo et al . 2023; Crequer et al . 2024).…”

“…These horizontally transferred regions include in particular two very large regions called Wallaby and CheesyTer , which encompass genes with functions in lactose metabolism or competition against other microorganisms. The non-Roquefort lineage further displays footprints of positive selection in genes involved in volatile compound production (Dumas et al 2020) and has lost the ability to secrete the mycophenolic acid (MPA) mycotoxin, an immunosuppressant used for preventing transplant organ rejection (Matas et al 2013), because of a deletion in a key gene ( mpaC ) of its biosynthesis pathway (Gillot et al 2017; Crequer et al 2024). Penicillium roqueforti can secrete other metabolites, i.e.…”