BONCAT-FACS-Seq reveals the active fraction of a biocrust community undergoing a wet-up event.

Ryan V Trexler, Marc Warwick Van Goethem, Danielle Goudeau, Nandita Nath, Rex R Malmstrom, Trent R Northen, Estelle Couradeau

Research output: Contribution to journalArticlepeer-review


Determining which microorganisms are active within soil communities remains a major technical endeavor in microbial ecology research. One promising method to accomplish this is coupling bioorthogonal non-canonical amino acid tagging (BONCAT) with fluorescence activated cell sorting (FACS) which sorts cells based on whether or not they are producing new proteins. Combined with shotgun metagenomic sequencing (Seq), we apply this method to profile the diversity and potential functional capabilities of both active and inactive microorganisms in a biocrust community after being resuscitated by a simulated rain event. We find that BONCAT-FACS-Seq is capable of discerning the pools of active and inactive microorganisms, especially within hours of applying the BONCAT probe. The active and inactive components of the biocrust community differed in species richness and composition at both 4 and 21 h after the wetting event. The active fraction of the biocrust community is marked by taxa commonly observed in other biocrust communities, many of which play important roles in species interactions and nutrient transformations. Among these, 11 families within the Firmicutes are enriched in the active fraction, supporting previous reports indicating that the Firmicutes are key early responders to biocrust wetting. We highlight the apparent inactivity of many Actinobacteria and Proteobacteria through 21 h after wetting, and note that members of the Chitinophagaceae, enriched in the active fraction, may play important ecological roles following wetting. Based on the enrichment of COGs in the active fraction, predation by phage and other bacterial members, as well as scavenging and recycling of labile nutrients, appear to be important ecological processes soon after wetting. To our knowledge, this is the first time BONCAT-FACS-Seq has been applied to biocrust samples, and therefore we discuss the potential advantages and shortcomings of coupling metagenomics to BONCAT to intact soil communities such as biocrust. In all, by pairing BONCAT-FACS and metagenomics, we are capable of highlighting the taxa and potential functions that typifies the microbes actively responding to a rain event.
Original languageEnglish (US)
StatePublished - Jun 26 2023

Bibliographical note

KAUST Repository Item: Exported on 2023-07-18
Acknowledgements: TN, EC, and MV gratefully acknowledge funding from the Office of Science Early Career Research Program, Office of Biological and Environmental Research, of the U. S. Department of Energy under contract number DE-AC02-05CH11231. Research was conducted by the U.S. Department of Energy Joint Genome Institute (, a DOE Office of Science User Facility, was supported by the Office of Science of the U.S. Department of Energy operated under Contract No. DE-AC02-05CH11231. We thank Mary Ann Bruns and José Raúl Román for their insightful discussions and suggestions during data analysis and manuscript preparation.

ASJC Scopus subject areas

  • Microbiology (medical)
  • Microbiology


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