Dispersion of antibiotic resistance genes from manure to soil

The spreading of antibiotic resistance has raised concerns in recent years. Nonfermenting gram-negative bacteria (NFGNB) harbor many antibiotic-resistant genes and have been studied for their role in antibiotic resistance in clinical setups worldwide. One way antibiotics spread into the environment is the application of antibiotic-treated cattle manure in agricultural soil. Our primary aim was to study the spread of antibiotic-resistant taxa and antibiotic resistance genes into the soil in a manure deposited environment. We simulated the natural manure deposition on the soil using the laboratory microcosm experiment. We used soils from two different bio-farms and fresh manure from a dairy farm different from where the soils were collected. The untreated manure and soils were used as control samples. In the microcosm, the top layer of manure and bottom layer of soil was separated by two sterile plastic nets filled with the same soil as the bottom layer and served as the interlayer in our experiment. We collected samples from the control manure and soil and the interlayer in 2, 14, and 28 days intervals. The samples were cultured on chromagar plates to specifically grow the NFGNB. The colonies were collected, and shotgun metagenomic sequencing was done.