Genomic Science Program
U.S. Department of Energy | Office of Science | Biological and Environmental Research Program

2024 Abstracts

Towards a Mechanistic Understanding of Rhodanobacter Dominance in the Contaminated Subsurface


H. K. Carlson1* ([email protected]), A.M. Deutschbauer1,2, J. V. Kuehl1, B. W. Biggs1, Y. Fan4, H. P. Lesea1, X. Tao4, J. P. Michael4, M. N. Price1, A. E. Kazakov1, M. Chen1, S. Priya1, V. Bhanot1, A. Zhou4, L. M. Lui1, V. V. Trotter1, T. N. Nielsen1, R. Chakraborty1, A. Mukhopadhyay1, M. W. Fields3, P. J. Walian1, J-Z. Zhou4, A. P. Arkin1,2, P. D. Adams1,2


1Lawrence Berkeley National Laboratory; 2University of California–Berkeley, CA; 3Montana State University–Bozeman; 4University of Oklahoma–Norman



The goal of ENIGMA (Ecosystems and Networks Integrated with Genes and Molecular Assemblies) is to develop theoretical, technological, and scientific approaches to gain a predictive and mechanistic understanding of the biotic and abiotic factors that constrain microbial communities’ assembly and activity in dynamic environments. To link genetic, ecological, and environmental factors to the structure and function of microbial communities, ENIGMA uses a systems biology approach to integrate and develop laboratory, field, and computational methods.


Over decades of measurements, Rhodanobacter species are consistently dominant in the most contaminated groundwater at the Oak Ridge Field Research Center (ORFRC). Scientists know that Rhodanobacter tend to be low pH and high metal tolerant relative to other bacteria but know less of the precise genetic and physiological mechanisms that enable them to survive and persist in the contaminated subsurface. Here, the team highlights work that exemplifies the ENIGMA Environmental Atlas experimental strategy with the aim of understanding the mechanisms behind Rhodanobacter survival at the ORFRC. In particular, high-throughput culturing and biofilm assays reveal phenotypic variability in metal stress resistance within the Rhodanobacter genus and between Rhodanobacter and other bacteria from the field site. Researchers have also generated random-barcode transposon sequencing (RB-TNSeq) mutant libraries in multiple ORFRC Rhodanobacter strains. Using these high-throughput genetics resources, the team has identified genes important for resistance to the key selective inorganic ion stressors at the ORFRC, including 33 efflux genes important for tolerance to 22 different inorganic ions. In addition, researchers have also implemented the DOE Joint Genome Institute DNA affinity purification sequencing (DAP-seq) approach to examine transcriptionally acting response regulators of signaling systems for two different Rhodanobacter strains. Additionally, the project has also developed CRISPR-based tools for precision genetics in ORFRC Rhodanobacter strains. Despite these advances, cultivation and analysis of the most highly abundant Rhodanobacter strain present in groundwater has remained a challenge. The project has made advances on this front by leveraging long-read metagenomics to identify key traits such as its unusual genomic capacity for carbon fixation.

The team has also identified an unusually high number of toxin-antitoxin systems in this genome, which may suggest rampant phage infection in the contaminated groundwater. As the project gains more insights into functional genomics and physiology of cultured Rhodanobacter, researchers will improve the ability to predict traits in uncultured strains.

Funding Information

This material by ENIGMA, a Science Focus Area at Lawrence Berkeley National Laboratory, is based upon work supported by the U.S. DOE, Office of Science, BER program under contract number DE-AC02-05CH11231.