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June 3rd, 2025
Version: 2
University of California Riverside
microbiology
biorxiv

Evaluating Best Practices for Isolating Pyrophilous Bacteria and Fungi from Burned Soil

Enright, D. J.Open in Google Scholar•Veerabahu, A.Open in Google Scholar•Quaal, R. J.Open in Google Scholar•Vega, M.Open in Google Scholar•Nguyen, A.Open in Google Scholar•Grindeland, J.Open in Google Scholar•Randolph, J. W. J.Open in Google Scholar•Ordonez, M. E.Open in Google Scholar•Glassman, S. I.Open in Google Scholar

A live microbial culture is invaluable to assess traits and functions, yet culturing immediately from fresh soil is logistically challenging and media selection is not trivial. Isolating pyrophilous bacteria and fungi after wildfires from burned soil, we tested the best: 1) soil storage method to retain culturable viability; 2) media for highest microbial diversity. We tested four soil storage methods (dried, stored at 4{degrees}C, stored at -80{degrees}C alone or in glycerol) and compared to fresh soil obtained 6 months after a severe California shrubland wildfire. We also tested 3 media types: rich (Lysogeny Broth (LB) for bacteria; Malt Yeast Agar (MYA) for fungi), oligotrophic (Reasoner's 2 Agar (R2A)) and media made from pyrogenic organic matter (PyOM). For bacteria, storing soil frozen at -80{degrees}C alone yielded the highest diversity of genera and with glycerol preserved the most species. For fungi, storing soil at -80{degrees}C preserved the most species but preserved equivalent diversity of genera as fresh and refrigerated soil. R2A yielded the highest bacterial and fungal diversity but some species of interest were only captured with PyOM. Using a combination of methods from 2018-2022, we cultured >500 isolates (286 bacteria; 258 fungi) after 7 California wildfires for testing pyrophilous microbial traits.

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