Abstract

How microorganisms survive and persist in subsurface environments is a fundamental question to the C-DEBI community. The main driver of this research is to investigate the potential for microorganisms to utilize extracellular electron transfer as a source for acquiring electrons/reducing power from minerals—a strategy that has been implicated as important for microbes in subsurface ecosystems, but that has been challenging to study due to the dearth of known mechanisms and organism identified with this capability. With funding from this research grant we are identifying candidate biomarkers for mineral oxidation processes in marine sediment microbes, previously isolated using electro-cultivation techniques as part of a C-DEBI funded postdoctoral fellowship. To date we have completed the genomes of ten organisms from eight different genera. We have also performed a pilot TN-seq study and are amid analyzing a statistically robust dataset from the same organism, Thioclava electrotropha ElOx9. Through the work of several undergraduate researchers, we have developed conjugation techniques for introducing a plasmid into organism and have been successful at removing genes in T. electrotrophica via homologous recombination. We are also in the process of developing CRISPR CAS-9 based techniques for genome editing in this strain. In our future work, application of these techniques will be applied to confirm activity of the gene(s) involved in EET that are being identified using high throughput genetic screens. The results of this work are presently being synthesized into two publications, and the results stemming from this work are the foundation of a pending NSF-Career Proposal.