A burning question: How to best culture fire-adapted microbes?

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Marely Vega, Jorge Ponce, Dylan Enright and Sydney I. Glassman, Microbiology and Plant Pathology, University of California, Riverside, Riverside, CA

Background/Question/Methods
The impact of wildfires on Earth system processes is growing rapidly as fires surge in frequency, size, and severity across the globe. Fires reduce bacterial and fungal biomass and richness and can select for distinct groups of pyrophilous microbes that are rare or undetectable pre-fire but have high dominance post-fire. In order to understand the traits that allow pyrophilous microbes to survive or even thrive after fires, it is necessary to have them in culture, so that genomics and biophysical assays can be performed. Yet, obtaining a culture collection of pyrophilous microbes is not trivial. Fires happen rapid-fire in southern California, so it is not possible to always culture from fresh soils, and it remains unclear which media is best to obtain the widest range of pyrophilous microbes. Here, we ask 1) what is the effect of different soil storage methods (fresh versus dried, frozen, or frozen in glycerol) 2) and different media types (nutrient rich LB or MYA versus nutrient poor R2A or fire simulating pyrogenic organic matter (PyOM) media) on colony forming unit (CFU) counts and diversity of pyrophilous microbes? Finally, we present a growing culture collection of 100+ pyrophilous bacteria and fungi isolated from Southern California wildfires.
Results/Conclusions
In total, we cultured, sequenced, and permanently stored a diverse group of fungi (1 phylum, 12 families, 16 genera) and bacteria (3 phyla, 17 families, 22 genera). When compared to the fresh soils, dried soil had the highest bacterial and fungal diversity and yielded CFU counts closest to fresh soil (reduction of 13%, compared to 29% for frozen and 98% for frozen in glycerol). For bacteria, PyOM media had the highest CFU counts, but all media yielded similar numbers of unique species (~7 each), with LB yielding four times more unique genera. Fungal diversity was greater when grown in the PyOM media while fungal growth was greater in the MYA media. Additionally, our methods allowed us to culture previously identified pyrophilous microbes such as bacterial Actinobacteria Arthrobacter and fungal Ascomycota Penicillium species. Based on our findings we conclude that 1) storing soils dried is the best method for preserving culture-viable bacteria and fungi, and 2) a combination of rich media (LB for bacteria and MYA for fungi) and media made from PyOM garners the widest diversity of pyrophilous microbes. Future experiments include exploring additional media types and dilution to extinction culturing to further widen the diversity of microbes obtained.