UNCOVERING THE MOLECULAR AND MICROECOLOGICAL BASIS FOR THE BIOTRANSFORMATION OF ANTIMICROBIALS BY RHIZOBACTERIA AND ENDOPHYTE

<b>Project Methods</b><br> Efforts.The prevalence and persistency of antibiotics and antibiotic resistance represent a biggest challenge for soil health and agricultural sustainability. Unfortunately, no feasible approaches are available to conjugate the control and mitigation of both issues of antibiotics and antibiotic resistance. Taking advantages of their unique ecological niches, diverse catabolic activities, and mutualistic relationship with the crop plants, rhizobacteria and endotypes are well suited for their irreplaceable yet uncharacterized roles in the agricultural ecosystem to sustainably contributing to biological attenuation of antibiotics and thus shunting the dissemination of antibiotic resistance as the selection pressure of antibiotics is reduced. In this project, we will combine conventional isolation (Task 1) and high-throughput omics and single-cell analysis (Task 3). Both culturable and unculturable microbes will be identified on the basis of their involvement of antibiotic decomposition. A novel technology named emulsion, paired isolation, and concatenation PCR (epicPCR) will be employed to uncover the transport and distribution of these antibiotic degraders along the manure-soil-plant passage without the need of isolation. Further, the molecular foundations of antibiotic biotransformation will be comprehensively characterized in bacterial isolates (Task 2). Genome analysis and metatranscriptomic assays are employed to screen putative genes/enzymes in association with the biotransformation pathways predicted from the detection of metabolites by advanced mass spectrometry approach. The catalytic roles of putative enzymes will be further unequivocally evaluated when they are expressed in a foreign host that lacks the ability of transforming the target antibiotics. Finally, the feasibility of using these isolates to remove antibiotics in the contaminated agricultural soils will be assessed and optimized using microcosm assays (Task 4). The potential to eliminate or outcompete with indigenous microbes carrying the antibiotic resistance will be investigated to discern the effectiveness of controlling the antibiotic resistance (as an ancillary goal of this proposal) via the proposed antibiotic biotransformation mechanism. Evaluation.This is a four-year project. The first two year will be spent obtaining and identifying novel isolates with unique biodegradation potentials on SAs (Task 1). Their genomic and molecular basis will be further characterized (Task 2). Distribution of SAs and microbial communities in targeted Maize farms will be surveyed seasonally for up to 2 years (Task 3.1 and 3.2). Based upon the genetic characterization and environmental metagenomics, targeted SAs degradation genes and associated phylogenies will be uncovered using epicPCR (Task 3.3) and quantified using biomarker assays (Task 3.4). Microcosm assays with agricultural soil samples to assess the feasibility of bioremediation will be mainly conducted in the fourth year after the biomarker design and genetic validation (Task 4). All proposed experimental and analytical techniques are fully operational and routinely used by our labs. An interim report and a final project summary will be provided at the midway and the end of the project, respectively.