Since penicillin was discovered in 1928, antibiotic usage in human and veterinary medicine and prevalence of antibiotic resistant bacteria (ARB), has been increasing. While antibiotics and antibiotic resistance genes (ARGs) naturally occur in soils, increasing abundances of ARGs correlate with increased antibiotic usage in agricultural settings. When livestock are treated with antibiotics, the antibiotic compounds, ARB, and ARGs can enter soil via manure excreted onto pastures or applied to other fields as fertilizer, thereby spreading antimicrobial resistance (AMR) in the environment. In addition to human health implications, increased AMR has negative impacts on ecosystem services such as carbon and nitrogen cycling. While many studies have researched antibiotic persistence in agricultural systems and their impacts on soil microbial communities, there are still significant knowledge gaps around the long-term effects of antibiotic exposure in soils, how those impacts differ among soils, and how elements of AMR may differentially transport through soil. To address these knowledge gaps, our objectives were to 1) examine the impact of multi-year repeated additions of manure from cattle administered antibiotics on soil microbial communities, 2) determine the interactive effects of soil moisture and type on soil microbial communities exposed to antibiotics and manure, and 3) differentiate between vertical transport of AMR in the form of viable ARB or ARGs in extracellular plasmids.
Our results demonstrate that soil bacterial community structures were consistently altered by 3-year additions of manure from cattle administered antibiotics compared to soil amended with antibiotic-free manure. Furthermore, ARG abundances were higher in soils with manure additions compared to soil without manure, although this was true regardless of whether the cattle were administered antibiotics, suggesting that manure and antibiotic impacts on soil microbial communities can persist over multi-year of repeated manure applications. Additionally, in microcosms, effects of manure from cattle administered antibiotics on ARG abundances, microbial community structures, respiration, and nitrogen pools in soil were seen across multiple soil types and moisture contents, suggesting environmental conditions can alter how manure and antibiotics impact microbial community structure and nutrient cycling. Finally, ARB flowed readily through saturated soil, but were also detectable in the top 5 cm of soil columns. However, ARGs on extracellular plasmids did not flow through soil columns and were not detected in soil, indicating that extracellular DNA does not persist or transport through the soil to any meaningful degree.
Overall, these results indicate a nuanced approach is required to mitigate the environmental spread of AMR. Soil management strategies for addressing the AMR crisis should consider the broader context of manure management, as high ARG abundances can come from application of manure from antibiotic-free cattle, and soil microbial communities in individual environments may have varied responses to manure antibiotic exposure. Furthermore, the transport of AMR through soil is complex and dynamic, as elements of AMR may transport differently through soil and require separate consideration in modeling and management. Future AMR management practices that consider diverse factors that affect persistence and spread of AMR in the environment can help protect livestock productivity and maintain the efficacy of antibiotics to protect human and animal health. / Doctor of Philosophy / Antibiotics are an important tool used to fight infections in humans, pets, and livestock. As antibiotics are used more frequently, the bacteria they target are more likely to develop resistance to the antibiotics, leading to increasing cases of infections that are harder to treat and higher risk. Antibiotic resistance can persist and spread in multiple forms, including the antibiotic compounds themselves, as antibiotic resistant bacteria (ARB), or as the genetic material that encodes for antibiotic resistance genes (ARGs). In agricultural systems, when livestock are treated with antibiotics they can excrete the antibiotics, along with ARB and ARGs, in the manure, which is then applied to land as fertilizer. In addition to the associated health risks, the spread of antibiotic resistance impacts microscopic bacteria and fungi in the soil, which are important for recycling nutrients for plants and maintaining ecosystem health. The overall goal of this dissertation was to gain a better understanding of how manure from cattle given antibiotics impacts these bacteria and fungi when manure is applied to the soil. The specific objectives were to 1) look impacts after long-term (multiple years) of manure addition, 2) examine how bacteria and fungi might respond differently to antibiotics in soils of different type or with different amounts of water, and 3) determine if ARGs that exist as free genetic material outside of living bacteria can be moved through the soil with flowing water in the same way as living bacteria.
Results showed that while the composition of bacterial and fungal communities in the soil vary from year to year, adding manure with and without antibiotics had both caused different and consistent changes on the composition of bacterial communities. There were also higher concentrations of ARGs in soil that had manure added, however antibiotics in the manure did not cause ARGs to increase further, suggesting that even antibiotic-free manure can impact the spread of antibiotic resistance. Experimental work also demonstrated that the soil type and water content of soil can alter how bacteria and fungi respond to antibiotics in manure. The composition of bacterial and fungal communities, their activity rates, and the amount of nitrogen – an important plant nutrient with availability that is strongly affected by microbial activity – all differed with soil type and water content. Thus, while antibiotic resistance antibiotic resistance can cause measurable changes in soil across a range of environmental conditions, it is also likely to persist and spread in different ways in different environments. Finally, when water containing elements of AMR was added to soil, ARB were shown to both move through the soil easily and remain near the top of soil. In contrast, ARGs contained on genetic material outside of living cells did not move through the soil and were broken down within a few days, suggesting that antibiotic resistance likely spreads through living bacteria more than genes outside of cells. Overall, this work highlights the complexity of understanding the role of environmental transmission in the antibiotic resistance crisis and demonstrates the need for nuanced management approaches that take specific environments and conditions into account.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/110497 |
| Date | 08 June 2022 |
| Creators | Shawver, Sarah Elizabeth |
| Contributors | Crop and Soil Environmental Sciences, Badgley, Brian Douglas, Xia, Kang, Stewart, Ryan D., Strickland, Michael, Pruden, Amy |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Language | English |
| Detected Language | English |
| Type | Dissertation |
| Format | ETD, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
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