A novel approach for tracing mycophagous small mammals and documenting their fungal diets

We developed a method combining passive baiting (animals that are not trapped) with DNA metabarcoding of the feces acquired, to study fungi in the diet of small mammals. Mammal and fungal species were identified using genomic DNA of 596 fecal samples collected in five regions of the eastern Canadian boreal forest. For identification of the small mammal species, the cytochrome b region was used. A total of eight species of small mammals displayed hypogeous fungi consumption, with northern flying squirrels (Glaucomys sabrinus) and red- backed voles (Myodes gapperi) as the top consumers. For identification of their fungal diets, the ribosomal internal transcribed spacer (ITS) region was used. We recovered 722 taxa of Ascomycota, 429 Basidiomycota, 81 Zygomycota, 4 Chytridiomycota, 1 Glomeromycota, and 44 unidentified fungal taxa. Of these, 28 were hypogeous sequestrate fungi (underground fructification), which presumably are dug out by small mammals for consumption. Otherwise, for the remaining fungi [epigeous (above ground fructification) or microscopic fungal species], it is unclear which ones are selected by the animal as a dietary source or result from incidental contamination. Our paper presents a promising approach for tracing mycophagy in small mammals, and our results suggest that fungal diversity is important for the diet of some small mammals.