Extreme low-flow conditions in a dual-chamber denitrification bioreactor contribute to pollution swapping with low landscape-scale impact.

Denitrification bioreactors are an effective edge-of-field conservation practice for nitrate (NO3) reduction from subsurface drainage. However, these systems may produce other pollutants and greenhouse gases during NO3 removal. Here a dual-chamber woodchip bioreactor system experiencing extreme low-flow conditions was monitored for its spatiotemporal NO3 and total organic carbon dynamics in the drainage water. Near complete removal of NO3 was observed in both bioreactor chambers in the first two years of monitoring (2019-2020) and in the third year of monitoring in chamber A, with significant (p < 0.01) reduction of the NO3-N each year in both chambers with 8.6-11.4 mg NO3-N L-1 removed on average. Based on the NO3 removal observed, spatial monitoring of sulfate (SO4), dissolved methane (CH4), and dissolved nitrous oxide (N2O) gases was added in the third year of monitoring (2021). In 2021, chambers A and B had median hydraulic residence times (HRTs) of 64 h and 39 h, respectively, due to varying elevations of the chambers, with drought conditions making the differences more pronounced. In 2021, significant production of dissolved CH4 was observed at rates of 0.54 g CH4-C m-3 d-1 and 0.07 g CH4-C m-3 d-1 in chambers A and B, respectively. In chamber A, significant removal (p < 0.01) of SO4 (0.23 g SO4 m-3 d-1) and dissolved N2O (0.21 mg N2O-N m-2 d-1) were observed, whereas chamber B produced N2O (0.36 mg N2O-N m-2 d-1). Considering the carbon dioxide equivalents (CO2e) on an annual basis, chamber A had loads (~12,000 kg CO2e ha-1 y-1) greater than comparable poorly drained agricultural soils; however, the landscape-scale impact was small (<1 % change in CO2e) when expressed over the drainage area treated by the bioreactor. Under low-flow conditions, pollution swapping in woodchip bioreactors can be reduced at HRTs <50 h and NO3 concentrations >2 mg N L-1.

Amending woodchip bioreactors with corncobs reduces nitrogen removal cost.

Woodchip denitrification bioreactors are an effective agricultural practice to reduce nitrogen (N) export from subsurface drainage via the conversion of nitrate (NO3-) to nitrogen gas (N2), but there are challenges associated with limited woodchip supplies and increasing prices. Previous lab studies indicate that corncobs could be a promising woodchip alternative from the perspectives of N removal rate and cost. This field study aimed to provide early performance and cost assessment of denitrification bioreactors with two woodchip-corncob treatments. The objectives were to i) compare N removal rates of bioreactors with different carbon and hydraulic retention time (HRT) treatments, ii) compare bioreactor N removal costs, and iii) conduct sensitivity analysis on full-scale bioreactors (FBR) N removal costs with varying corncob lifespans and prices. Nine replicated field pilot-scale bioreactors (PBRs) using three carbon treatments and three HRTs were assessed for N removal efficiency. The carbon treatments were woodchip-only (WC100), 25% (by vol.) corncobs + 75% woodchips media (CC25) in series, and 75% corncobs + 25% woodchips (CC75) in series set at HRTs of 2, 8, and 16 h. N concentrations were monitored at each PBR inlet and outlet, and the PBR N removal efficiencies were used to estimate FBR N removal rates and costs. At respective HRTs, the estimated N removal rates of CC75 were 1.6- to 10.1-fold higher than WC100, but CC25 exhibited 0.9-fold lower (at 8-hr HRT) to 2.8-fold higher than WC100. A 15-yr cost assessment indicated CC75 ($10.56 to $13.89 kg-1 N) was the most cost-efficient treatment, followed by WC100 ($13.30 to $88.11 kg-1 N) and CC25 ($22.41 to $60.13 kg-1 N). This assessment showed CC75 as a promising alternative to WC100 in terms of N removal rate and cost, but CC25 did not provide sufficient N removal rate increase for it to be a cost-efficient option. Nonetheless, using corncobs as a bioreactor medium is a relatively new approach, and we encourage more field studies to explore the long-term opportunities and challenges.

Allelic variation of Escherichia coli outer membrane protein A: Impact on cell surface properties, stress tolerance and allele distribution.

Outer membrane protein A (OmpA) is one of the most abundant outer membrane proteins of Gram-negative bacteria and is known to have patterns of sequence variations at certain amino acids-allelic variation-in Escherichia coli. Here we subjected seven exemplar OmpA alleles expressed in a K-12 (MG1655) ΔompA background to further characterization. These alleles were observed to significantly impact cell surface charge (zeta potential), cell surface hydrophobicity, biofilm formation, sensitivity to killing by neutrophil elastase, and specific growth rate at 42°C and in the presence of acetate, demonstrating that OmpA is an attractive target for engineering cell surface properties and industrial phenotypes. It was also observed that cell surface charge and biofilm formation both significantly correlate with cell surface hydrophobicity, a cell property that is increasingly intriguing for bioproduction. While there was poor alignment between the observed experimental values relative to the known sequence variation, differences in hydrophobicity and biofilm formation did correspond to the identity of residue 203 (N vs T), located within the proposed dimerization domain. The relative abundance of the (I, δ) allele was increased in extraintestinal pathogenic E. coli (ExPEC) isolates relative to environmental isolates, with a corresponding decrease in (I, α) alleles in ExPEC relative to environmental isolates. The (I, α) and (I, δ) alleles differ at positions 203 and 251. Variations in distribution were also observed among ExPEC types and phylotypes. Thus, OmpA allelic variation and its influence on OmpA function warrant further investigation.

The Human Health Implications of Antibiotic Resistance in Environmental Isolates from Two Nebraska Watersheds.

One Health field-based approaches are needed to connect the occurrence of antibiotics present in the environment with the presence of antibiotic resistance genes (ARGs) in Gram-negative bacteria that confer resistance to antibiotics important in for both veterinary and human health. Water samples from two Nebraska watersheds influenced by wastewater effluent and agricultural runoff were tested for the presence of antibiotics used in veterinary and human medicine. The water samples were also cultured to identify the bacteria present. Of those bacteria isolated, the Gram-negative rods capable of causing human infections had antimicrobial susceptibility testing and whole-genome sequencing (WGS) performed to identify ARGs present. Of the 211 bacterial isolates identified, 37 belonged to pathogenic genera known to cause human infections. Genes conferring resistance to beta-lactams, aminoglycosides, fosfomycins, and quinolones were the most frequently detected ARGs associated with horizontal gene transfer (HGT) in the watersheds. WGS also suggest recent HGT events involving ARGs transferred between watershed isolates and bacteria of human and animal origins. The results of this study demonstrate the linkage of antibiotics and bacterial ARGs present in the environment with potential human and/or veterinary health impacts. IMPORTANCE One health is a transdisciplinary approach to achieve optimal health for humans, animals, plants and their shared environment, recognizing the interconnected nature of health in these domains. Field based research is needed to connect the occurrence of antibiotics used in veterinary medicine and human health with the presence of antibiotic resistance genes (ARGs). In this study, the presence of antibiotics, bacteria and ARGs was determined in two watersheds in Nebraska, one with agricultural inputs and the other with both agricultural and wastewater inputs. The results presented in this study provide evidence of transfer of highly mobile ARG between environment, clinical, and animal-associated bacteria.

Diversity of Antibiotic Resistance genes and Transfer Elements-Quantitative Monitoring (DARTE-QM): a method for detection of antimicrobial resistance in environmental samples.

Effective monitoring of antibiotic resistance genes and their dissemination in environmental ecosystems has been hindered by the cost and efficiency of methods available for the task. We developed the Diversity of Antibiotic Resistance genes and Transfer Elements-Quantitative Monitoring (DARTE-QM), a method implementing TruSeq high-throughput sequencing to simultaneously sequence thousands of antibiotic resistant gene targets representing a full-spectrum of antibiotic resistance classes common to environmental systems. In this study, we demonstrated DARTE-QM by screening 662 antibiotic resistance genes within complex environmental samples originated from manure, soil, and livestock feces, in addition to a mock-community reference to assess sensitivity and specificity. DARTE-QM offers a new approach to studying antibiotic resistance in environmental microbiomes, showing advantages in efficiency and the ability to scale for many samples. This method provides a means of data acquisition that will alleviate some of the obstacles that many researchers in this area currently face.

Hydrophobic laser-induced graphene potentiometric ion-selective electrodes for nitrate sensing.

Current solid-contact ion-selective electrodes (ISEs) suffer from signal-to-noise drift and short lifespans partly due to water uptake and the development of an aqueous layer between the transducer and ion-selective membrane. To address these challenges, we report on a nitrate ISE based on hydrophobic laser-induced graphene (LIG) coated with a poly(vinyl) chloride-based nitrate selective membrane. The hydrophobic LIG was created using a polyimide substrate and a double lasing process under ambient conditions (air at 23.0 ± 1.0 °C) that resulted in a static water contact angle of 135.5 ± 0.7° (mean ± standard deviation) in wettability testing. The LIG-ISE displayed a Nernstian response of - 58.17 ± 4.21 mV dec-1 and a limit-of-detection (LOD) of 6.01 ± 1.44 µM. Constant current chronopotentiometry and a water layer test were used to evaluate the potential (emf) signal stability with similar performance to previously published work with graphene-based ISEs. Using a portable potentiostat, the sensor displayed comparable (p > 0.05) results to a US Environmental Protection Agency (EPA)-accepted analytical method when analyzing water samples collected from two lakes in Ames, IA. The sensors were stored in surface water samples for 5 weeks and displayed nonsignificant difference in performance (LOD and sensitivity). These results, combined with a rapid and low-cost fabrication technique, make the development of hydrophobic LIG-ISEs appealing for a wide range of long-term in situ surface water quality applications.

Prairie strips' effect on transport of antimicrobial resistance indicators in poultry litter.

Poultry litter is a valuable nutrient resource for agricultural production but is also a potential source for introducing antibiotic resistance genes (ARGs) and litter-associated bacteria (LAB) to the environment. Prairie strips have been demonstrated as an effective conservation practice to improve environmental quality in agroecosystems. This research aims to assess prairie strips' potential for reducing the transport of LAB and ARGs in runoff after litter application. Plot-scale rainfall simulations were performed using a replicated block design, with soil and surface runoff samples taken during the rainfall event. Microbial taxa and ARGs were characterized in the litter, soil, and water samples. In plots with litter application, LAB and ARGs were mainly detected in runoff, with very low detection in soils. Detection of ARGs in runoff, irrespective of strip installations, is consistent with previous observations of litter as a source of antimicrobial resistance (AMR) risks. The effectiveness of prairie strips to remove LAB and ARGs varied. In two of the three prairie strip plots, fewer AMR indicators were detected relative to control plots, suggesting that the prairie strips can potentially reduce these risks. In one plot, which was also associated with increased flow rate, we observed increased AMR indicators despite the installation of a prairie strip. Our observations highlight the need to prioritize understanding of soil properties even within the same site. Although we show that prairie strips can potentially reduce AMR risks, further research is needed to better understand the influence of rainfall timing, soil, and litter characteristics.

Comparison of microbial communities in replicated woodchip bioreactors.

Denitrification in woodchip bioreactors is a microbial process, but the effects of variations in bioreactors operation on microbial community structure are not well understood. Here, our goals were to understand hydraulic retention time (HRT) as a factor that influences woodchip bioreactor microbial community variation and structure in replicated field bioreactors and to evaluate relationships between microbial community membership and marker genes for denitrification. We used a combination of quantitative polymerase chain reaction of nirS, nirK, nosZI, and nosZII and 16S rRNA amplicon sequencing to characterize the microbial communities of nine pilot-scale woodchip bioreactors located at Iowa State University. Our results showed dynamic microbial communities but with persistent taxa between two sampling years and three HRTs. Similarities between functional gene copy numbers across sampling year and HRT indicate that the potential for denitrification is conserved despite differences in the microbial communities. These results are evidence that there are specific and persistent taxa within replicated bioreactors. Woodchip bioreactor microbial community membership is recommended to be the focus of future studies to better understand the relationship between microbial community functions and bioreactor management.

Stacked conservation practices reduce nitrogen loss: A paired watershed study.

Combinations of best management practices (BMPs) are needed to achieve nutrient reduction goals in the Mississippi/Atchafalaya River Basin (MARB), but field results are crucial to encourage stacked adoption of BMPs. A paired catchment-scale study (2015-18) was done to assess the impact of (i) BMPs, (ii) precipitation patterns, and (iii) seasonality on nitrogen (N) export. Flow-weighted samples were collected and analyzed for total ammonia nitrogen (TAN), nitrate (NO3-N), and total nitrogen (TN). Catchments Low-BMP 11 and High-BMP 12 had 27.6% and 87.6% areal coverage of BMPs, respectively. No significant difference (p > 0.05) in TAN concentrations was found between Low-BMP 11 (0.023 mg L-1) and High-BMP 12 (0.020 mg L-1). However, NO3-N and TN concentrations were significantly higher (p < 0.05) at Low-BMP 11 (NO3-N: 26.0 mg L-1, TN: 28.7 mg L-1) than at High-BMP 12 (NO3-N: 8.8 mg L-1, TN: 9.2 mg L-1). Two precipitation factors that affected N export patterns were observed. First, N flushing could continue for several years after a drought as elevated NO3-N concentrations were observed in 2015 (i.e., two years after the 2011-2013 drought). Second, higher annual N export was observed when more precipitation occurred during the pre-planting or early-growing season versus later periods. For both catchments, the highest 50% of flows were responsible for majority of the NO3-N export. We estimated that 33-37%, 61-62%, and 82-85% of the NO3-N loads occurred in the 90th, 75th, and 50th flow percentiles, respectively. As demonstrated in High-BMP 12, stacked BMP application effectively lowered NO3-N and TN loads by 60.3% and 59.1%, respectively, relative to Low-BMP 11. Although 27.6% BMP coverage area in Low-BMP 11 was considered low for this study, this coverage area is higher than many other parts of the MARB. This research highlights the importance of joint efforts between landowners in a watershed to meet downstream water quality goals.

Denitrifying bioreactor microbiome: Understanding pollution swapping and potential for improved performance.

Denitrifying woodchip bioreactors are a best management practice to reduce nitrate-nitrogen (NO3 -N) loading to surface waters from agricultural subsurface drainage. Their effectiveness has been proven in many studies, although variable results with respect to performance indicators have been observed. This paper serves the purpose of synthesizing the current state of the science in terms of the microbial community, its impact on the consistency of bioreactor performance, and its role in the production of potential harmful by-products including greenhouse gases, sulfate reduction, and methylmercury. Microbial processes other than denitrification have been observed in these bioreactor systems, including dissimilatory nitrate reduction to ammonia (DNRA) and anaerobic ammonium oxidation (anammox). Specific gene targets for denitrification, DNRA, anammox, and the production of harmful by-products are identified from bioreactor studies and other environmentally relevant systems for application in bioreactor studies. Lastly, cellulose depletion has been observed over time via increasing ligno-cellulose indices, therefore, the microbial metabolism of cellulose is an important function for bioreactor performance and management. Future work should draw from the knowledge of soil and wetland ecology to inform the study of bioreactor microbiomes.

Impact of Swine and Cattle Manure Treatment on the Microbial Composition and Resistome of Soil and Drainage Water.

Evaluating potential environmental and clinical impacts of industrial antibiotic use is critical in mitigating the spread of antimicrobial resistance. Using soil columns to simulate field application of swine or cattle manure and subsequent rain events, and a targeted qPCR-based approach, we tracked resistance genes from source manures and identified important differences in antimicrobial resistance gene transport and enrichment over time in the soil and water of artificially drained cropland. The source manures had distinct microbial community and resistance gene profiles, and these differences were also reflected in the soil columns after manure application. Antibiotic resistance genes (ARGs) were only significantly enriched in effluent samples following the first rain event (day 11) for both soil types compared to the control columns, illustrating the high background level of resistance present in the control soils chosen. For swine, the genes tetQ, tet(36), tet44, tetM, sul2 and ant(6)-ib persisted in the soil columns, whereas tetO, strB and sul1 persisted in effluent samples. Conversely, for cattle manure sul2 and strB persisted in both soil and effluent. The distinct temporal dynamics of ARG distribution between soil and effluent water for each manure type can be used to inform potential mitigation strategies in the future.

Antimicrobial resistance in integrated agroecosystems: State of the science and future opportunities.

As the Journal of Environmental Quality (JEQ) celebrates 50 years of publication, the division of environmental microbiology is one of the newest additions to the journal. During this time, significant advances in understanding of the interconnected microbial community and impact of the microbiome on natural and designed environmental systems have occurred. In this review, we highlight the intractable challenge of antimicrobial resistance (AMR) on humans, animals, and the environment, with particular emphasis on the role of integrated agroecosystems and by highlighting contributions published in JEQ. From early studies of phenotypic resistance of indicator organisms in waters systems to current calls for integrating AMR assessment across "One Health," publications in JEQ have advanced our understanding of AMR. As we reflect on the state of the science, we emphasize future opportunities. First, integration of phenotypic and molecular tools for assessing environmental spread of AMR and human health risk continues to be an urgent research need for a one health approach to AMR. Second, monitoring AMR levels in manure is recommended to understand inputs and potential spread through agroecosystems. Third, baseline knowledge of AMR levels is important to realize the impact of manure inputs on water quality and public health risk; this can be achieved through background monitoring or identifying the source-related genes or organisms. And finally, conservation practices designed to meet nutrient reduction goals should be explored for AMR reduction potential.

Antibiotic resistance gene dissipation in soil microcosms amended with antibiotics and swine manure.

The use of antibiotics in animal agriculture has exacerbated the presence of both antibiotic resistance genes (ARGs) and residual antibiotics excreted in animal manure. Field application of this manure is a common practice because its nutrient rich material can benefit crop growth. However, this practice can also introduce antibiotics and ARGs into nonagricultural settings. The integration of prairie buffer strips within and at the edge of crop fields is a potential management solution to reduce concentrations of ARGs commonly transported via water runoff and infiltration. An incubation experiment was conducted to investigate the fate of ARGs in directly manured crop field soils and the surrounding affected prairie strip soils. Row crop and prairie strip soils sampled from three sites received either an antibiotic spike and swine manure addition or a control water addition. The concentrations of select ARGs were then monitored over a 72-d period. Although soil vegetation and site location were not observed to influence ARG dissipation, the select genes did display different half-lives from one another. For example, tetM demonstrated the fastest dissipation of the genes quantified (average half-life, 5.18 d). Conversely, sul1 did not conform to the first-order linear regression kinetics used to describe the other investigated genes and was highly abundant in control prairie strip soils. The quantified half-lives of these select ARGs are comparable to previous studies and can inform monitoring and mitigative efforts aimed at reducing the spread of ARGs in the environment.

Midwestern cropping system effects on drainage water quality and crop yields.

Grain producers are challenged to maximize crop production while utilizing nutrients efficiently and minimizing negative impacts on water quality. There is a particular concern about nutrient export to the Gulf of Mexico via loss from subsurface drainage systems. The objective of this study was to investigate the effects of crop rotation, tillage, crop residue removal, swine manure applications, and cereal rye (Secale cereale L.) cover crops on nitrate-N (NO3 -N) and total reactive phosphorus (TRP) loss via subsurface drainage. The study was evaluated from 2008 through 2015 using 36 0.4-ha plots outfitted with a subsurface drainage water quality monitoring system. Results showed that when swine manure was applied before both corn (Zea mays L.) and soybean [Glycine max (L.) Merr.], drainage water had significantly higher 8-yr-average flow-weighted NO3 -N concentrations compared with swine manure applied before corn only in a corn-soybean rotation. The lowest NO3 -N loss was 15.2 kg N ha-1  yr-1 from a no-till corn-soybean treatment with rye cover crop and spring application of urea-ammonium nitrate (UAN) to corn. The highest NO3 -N loss was 29.5 kg N ha-1  yr-1 from swine manure applied to both corn and soybean. A rye cover crop reduced NO3 -N loss, whereas tillage and residue management had little impact on NO3 -N loss. Losses of TRP averaged <32 g P ha-1  yr-1 from all treatments. Corn yield was negatively affected by both no-till management and cereal rye cover crops. Results showed that cropping management affected N leaching but impacts on P leaching were minimal.

Seasonal variations in export of antibiotic resistance genes and bacteria in runoff from an agricultural watershed in Iowa.

Seasonal variations of antimicrobial resistance (AMR) indicators in runoff water can help improve our understanding of AMR sources and transport within an agricultural watershed. This study aimed to monitor multiple areas throughout the Black Hawk Lake (BHL) watershed (5324 ha) in central Iowa during 2017 and 2018 that consists of both swine and cattle feeding operations as well as known areas with manure application. The measured indicators included plate counts for fecal indicator bacteria (FIB) E. coli, Enterococcus, antibiotic resistant fecal indicator bacteria (ARBs) tylosin resistant Enterococcus, tetracycline resistant Enterococcus, and antibiotic resistance genes (ARGs): ermB, ermF (macrolide), tetA, tetM, tetO, tetW (tetracycline), sul1, sul2 (sulfonamide), aadA2 (aminoglycoside), vgaA, and vgaB (pleuromutilin). Both the plate count and the ARG analyses showed seasonal trends. Plate counts were significantly greater during the growing season, while the ARGs were greater in the pre-planting and post-harvest seasons (Wilcoxon Rank-Sum Test p < 0.05). The ermB gene concentration was significantly correlated (p < 0.05) with E. coli and Enterococcus concentrations in 2017, suggesting a potential use of this ARG as an indicator of environmental AMR and human health risk. Flow rate was not a significant contributor to annual variations in bacteria and AMR indicators. Based on observed seasonal patterns, we concluded that manure application was the likely contributor to elevated ARG indicators observed in the BHL watershed, while the driver of elevated ARB indictors in the growing season can only be speculated. Understanding AMR export patterns in agricultural watersheds provides public health officials knowledge of seasonal periods of higher AMR load to recreational waters.

Spatial and temporal distribution of E. coli contamination on three inland lake and recreational beach systems in the upper Midwestern United States.

Swimming advisories are commonly posted at public beaches across the United States every year. In Iowa, weekly monitoring of public swimming areas at state and county beaches have resulted in the impairment of numerous lakes for fecal indicator bacteria (FIB) contamination, as detected by E. coli. An extensive study was established to assess the relationships between E. coli contamination of nearshore beach water environments, open lake conditions and beach sands in three recreational beach/lake systems currently impaired for FIB contamination across Iowa. A transect/grab sample based sampling design was implemented across the systems with collections spanning from April through October of 2015 and 2016. Collections of E. coli along water transects identified strong near to far shore gradients of decreasing concentrations in all systems. Results indicate that concentrations of E. coli observed in swimming waters consistently disassociate with concentrations in the broader lake environment. Swimming water E. coli concentrations correlated with elevated beach sand E. coli, samples collected from beach sands uncovered concentrations up to 86,500 times higher than adjacent swimming waters. Results from this study indicate that foreshore beach sands and other beach proximate FIB sources serve as the major contributing source for swimming zone advisories. The current methodology used by state and federal officials includes impairing entire lake waterbodies for FIB contamination of the swimming area. These impairment listings do not accurately reflect the condition(s) of the larger lake environment outside the swimming area and fail to account for beach proximate conditions in the assessment process. Further, this approach provides potentially misleading information to the public and may undermine implementation strategies deployed by resource managers aimed at addressing FIB contamination at recreational swimming areas. Views expressed in this paper are those of the authors and do not necessarily reflect the views or policies of the Iowa Department of Natural Resources.

Catchment-scale export of antibiotic resistance genes and bacteria from an agricultural watershed in central Iowa.

Antibiotics are administered to livestock in animal feeding operations (AFOs) for the control, prevention, and treatment of disease. Manure from antibiotic treated livestock contains unmetabolized antibiotics that provide selective pressure on bacteria, facilitating the expression of anti-microbial resistance (AMR). Manure application on row crops is an agronomic practice used by growers to meet crop nutrient needs; however, it can be a source of AMR to the soil and water environment. This study in central Iowa aims to directly compare AMR indicators in outlet runoff from two adjacent (221 to 229 ha) manured and non-manured catchments (manure comparison), and among three catchments (600 to 804 ha) with manure influence, no known manure application (control), and urban influences (mixed land use comparison). Monitored AMR indicators included antibiotic resistance genes (ARGs) ermB, ermF (macrolide), tetA, tetM, tetO, tetW (tetracycline), sul1, sul2 (sulfonamide), aadA2 (aminoglycoside), vgaA, and vgaB (pleuromutilin), and tylosin and tetracycline resistant enterococci bacteria. Results of the manure comparison showed significantly higher (p<0.05) tetracycline and tylosin resistant bacteria from the catchment with manure application in 2017, but no differences in 2018, possibly due to changes in antibiotic use resulting from the Veterinary Feed Directive. Moreover, the ARG analysis indicated a larger diversity of ARGs at the manure amended catchment. The mixed land use comparison showed the manure amended catchment had significantly higher (p<0.05) tetracycline resistant bacteria in 2017 and significantly higher tylosin resistant bacteria in 2017 and 2018 than the urban influenced catchment. The urban influenced catchment had significantly higher ermB concentrations in both sampling years, however the manure applied catchment runoff consisted of higher relative abundance of total ARGs. Additionally, both catchments showed higher AMR indicators compared to the control catchment. This study identifies four ARGs that might be specific to AMR as a result of agricultural sources (tetM, tetW, sul1, sul2) and optimal for use in watershed scale monitoring studies for tracking resistance in the environment.

Impact of stacked conservation practices on phosphorus and sediment export at the catchment scale.

Best management practices (BMPs) are effective in reducing nutrient and sediment export, but further understanding of the benefits of the stacked BMPs is needed. This catchment-scale study was established to evaluate the impact of hydrology and BMPs on phosphorus (P) and sediment losses. Two adjacent catchments, one with a lower level of BMP adoption (Low-BMP #11) and one with a higher level (High-BMP #12), were compared for total P (TP) and total suspended solids (TSS) export. The BMPs include nutrient management plans, reduced tillage, grassed waterways, terraces, and perennial vegetation. The TP event-flow-weighted (EFW) concentration was significantly higher at Low-BMP #11 (0.293 mg L-1) than at High-BMP #12 (0.069 mg L-1). There was no significant difference in TP base-flow-weighted (BFW) concentrations between Low-BMP #11 (0.035 mg L-1) and High-BMP #12 (0.037 mg L-1). The TSS-EFW (148.0 vs. 18.6 mg L-1) and TSS-BFW (13.3 vs. 6.9 mg L-1) concentrations were also higher at Low-BMP #11 than at High-BMP #12. High-BMP #12 had lower TP (0.36 vs. 0.59 kg ha-1 yr-1) and TSS (253 vs. 1,961 kg ha-1 yr-1) loading than Low-BMP #11. The lower TP export at High-BMP #12 was likely attributed to the effectiveness of stacked erosion control BMPs and nutrient management plans. Overall, lower P and sediment loading was observed when a greater areal extent of stacked practices was implemented at the catchment level. This finding provides vital information to encourage wider BMP adoption at the watershed scale.

Long-term impact of poultry manure on crop yield, soil and water quality, and crop revenue.

A long-term poultry manure fertilizer study was initiated in 1998 and continued until 2009 under corn-soybean (CS) rotation. To match changing landscape trends, the plots were switched to continuous corn (CC) from 2010 to 2017. In both CS and CC phases, poultry manure (PM) was applied at the crop rotation recommended agronomic N rate and either half (CC phase) or double (CS phase) the recommended rate. Urea-ammonium nitrate (UAN) was applied to comparison plots at the crop recommended application rate (168 kg N ha-1 and 224 kg N ha-1 for the CS and CC phases, respectively) throughout the study. The objectives of this study include evaluation of the economic benefits of long-term PM application at various rates (PM2, PM, and PM0.5), and the impact of poultry manure application on soil health and nutrient levels, crop yield, and drainage water quality. Lower NO3-N concentrations were reported in drainage from PM treated plots when compared to UAN fertilizer applied at the same agronomic rate. Of the parameters tested for soil health analysis after twenty years of repeat application, particulate organic matter (POM) present was significantly greater in the PM treated soils (6.1-6.7 g kg soil-1) when compared to UAN plots (4.6 g kg soil-1), showing potential for stabilized soil particles, increased infiltration and water-holding capacity. The results show a consistent positive impact of manure application on corn and soybean yields when compared to yields observed in UAN treated plots. During the CS phase, we estimated the same average revenue per dollar spent for PM and UAN treatments, while the average return rate for PM2 was 1% lower; during CC phase,15% increased return rates were observed when PM0.5 and PM were compared against the UAN treatment. When managed properly, PM application to cropland is a sustainable option for diversifying agroecosystems, improving soil health and improving farm economics.

Investigating the dispersal of antibiotic resistance associated genes from manure application to soil and drainage waters in simulated agricultural farmland systems.

Manure from animals that have been treated with antibiotics is often used to fertilize agricultural soils and its application has previously been shown to enrich for genes associated with antibiotic resistance in agroecosystems. To investigate the magnitude of this effect, we designed a column experiment simulating manure-treated agricultural soil that utilizes artificial subsurface drainage to determine the duration and extent which this type of manure fertilization impacts the set of genes associated with antibiotic resistance in drainage water. We classified ARGs in manure-treated drainage effluent water by its source of origin. Overall, we found that 61% and 7% of the total abundance of ARGs found in drainage water samples could be attributed to manure enrichment and manure addition, respectively. Among these ARGs, we identified 75 genes unique to manure that persisted in both soil and drainage water throughout a drainage season typical of the Upper Midwestern United States. While most of these genes gradually decreased in abundance over time, the IS6100-associated tet(33) gene accrued. These results demonstrate the influence of manure applications on the composition of the resistome observed in agricultural drainage water and highlight the importance of anthropogenic ARGs in the environment.