Evaluating Microbial and Molecular Mechanisms that Affect Changes in Antibiotic Resistance during Anaerobic Digestion of Manure

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Livestock production and animal husbandry are considered key contributors responsible for generating and emitting antibiotic-resistant bacteria (ARB) due to the extensive use of veterinary antibiotics. Antibiotic resistance genes (ARG), the genetic units encoding the diverse mechanisms of resistance, play an important role in disseminating antibiotic resistance because they are usually located in mobile genetic elements (MGE) that are easily transferred between microbial species. Therefore, bacteria that are not directly exposed to antibiotics are also susceptible to acquiring antibiotic resistance mechanisms. Since the application of animal manures as soil fertilizers is a common agricultural practice, there is increasing concern about the proliferation of ARB and ARG in agricultural fields and surrounding waterways and ultimately the consequences this will have on public and ecosystem health. One alternative to address this problem is to design manure treatment processes that ensure the reduction of ARB and ARG before they reach the environment. Anaerobic digester installations are readily expanding across Canada (at farm level or larger regional plants) to meet the growing demand in sustainable waste management and renewable energy. We propose using manure anaerobic digestion as a model to better understand microbial and molecular mechanisms involved in the evolution of ARG. Developing effective strategies for ARB and ARG mitigation through manure treatment will provide an additional incentive to build and operate anaerobic digestion systems, which will enhance the long-term sustainability and resilience of Canadian agriculture. The specific objectives of this research program are to: 1) Determine the presence, levels, and prevalence of ARGs in raw and treated manures in dairy farms with different sizes, operations, and manure management systems; 2) Assess the long-term impact of different operational parameters on the resistome (total ARG) and mobilome (total MGE) profiles and microbial population dynamics in anaerobic digesters; 3) Identify specific microbial groups responsible for carrying and spreading ARG, through the use of metagenomic and bioinformatic tools and biostatistics; 4) Validate potential microbial group hosts of ARG by comparing their presence and prevalence (co-occurrence) in other anaerobic digestion applications, such as in municipal wastewater treatment and industrial wastewater treatment; 5) Develop and validate a computational algorithm to estimate the potential risk associated with the presence of ARG, MGE, and microbial communities in raw and treated animal manures that can be used for the systematic comparison between samples and processes. This research program will provide multidisciplinary training in bioprocessing, waste treatment, and metagenomics, and supply highly skilled professionals (2 PhD, 2 MSc, 5 BSc) to Canada's growing bio-economy, where companies reported significant labour and skill shortages.