Multi-Analytic Based Determination of Substrate Fate From in situ Stable Isotope Labeled Exposures of Natural Microbial Mats

Abstract:

Microbial communities play impactful roles in almost every aspect of our society including the environment, climate, agriculture and human health, expanding the functional capacity of life on earth. The recent emergence of a suite of omics driven technologies offers powerful tools for investigating functionality of this community. However, these tools provide only a static snapshot of the community in space and time. The temporal nature of stable isotope probing (SIP) experiments expands the depth at which microbial communities can be investigated and understood. While selectively targeting only metabolically active organisms in a community, the labeled substrate can be tracked spatially, temporally and phylo-genetically and linked to active functions, organism interactions and exchanges.

Single SIP technologies are limited in their ability to describe the biological system as a whole. However, integration of multiple SIP based analytics offers a more comprehensive description of substrate fate. The phototroph based microbial mat community resident in Hot Lake, a hypersaline lake located in Washington State, offers a tractable system for testing the multi analytic approach. We exposed the mat to three different ¹³C-labeled substrates (HCO₃^-, glucose and acetate) in situ at midday, and subsequently analyzed the mat 24 hours after incubation.

The approach revealed different metabolic fates and organism specific uptake. When compared to acetate, glucose and HCO₃^- showed a greater incorporation into extracellular material, while acetate had a greater conversion to intracellular fatty acids, suggesting that HCO₃^- and glucose could be more readily shared as a community currency than acetate. All substrates were converted to amino acids and proteins, but while glucose and HCO₃^- demonstrated considerable incorporation into heterotrophic proteins, the conversion of acetate to these proteins was minimal, potentially implying that acetate derived intermediates are not a currency of metabolic exchange in the community. Functional analyses revealed the predominance of synthesis of photosynthetic proteins in the HCO₃^- incubations and transporters in the glucose and acetate incubations potentially indicating the repression of photosynthetic activity in the presence of an alternative carbon source.