| Title |
Investigators | Department | Objectives | Approach Keywords | Progress Reports | Impact Statements | Publications | |
Project * COL00709 | |
| Title | Minimizing the Release of Animal Antibiotics from Aerobic and Anaerobic Lagoons |
| Investigator(s) | Carlson, KH; |
| Department | Civil and Environmental Engineering |
| Objectives | Recent work by our research team has shown the existence of several animal-only antibiotics both on-farm and in streams. We have recently sampled 25 waste lagoons in Northern Colorado and have studied the occurrence of 17 antibiotic compounds (6 tetracyclines including chlortetracycline, 5 sulfonamides, 3 macrolides including tylosin and 3 ionophores including monensin). The waste stream from 25 animal feed operations has been sampled for either manure, lagoon water or both. The individual antibiotic results varied widely. For example, tylosin (a macrolide) was found in all but four waste lagoons sampled (an occurrence rate of 84%) but erythromycin was found in only 36% of the lagoons sampled. Tetracycline (occurrence rate of 96%) and sulfadimethoxane (occurrence rate of 44%) were the most common compounds in their classes. The concentrations of the antibiotic compounds also varied significantly. For example, chlortetracycline ranged from less than 100 mg/L in some swine lagoons to over 9000 mg/L in others. Since the occurrence and concentration of antibiotic concentrations varies so widely among operations that appear similar, it is likely biodegradation is more important in some lagoons than others. We hope to be able to characterize these differences with this research. The goal of the proposed research is to optimize the removal of several classes of antibiotics in concentrated animal feed operations (CAFO) waste lagoons. The objectives that will be completed to achieve this goal are as follows: (1)Simulate in the lab aerobic and anaerobic lagoons and determine the fate of the 4 most common antibiotics under each of these conditions. (2)Construct demonstration scale lined waste lagoons operated both in aerobic and anaerobic modes. Determine the impact on odor and antibiotic fate. (3)Utilizing results from this and concurrent projects, provide guidance on the design and operation of waste lagoons for minimizing release of antibiotics and reducing odors. |
| Approach | The proposed research can be described as three tasks. Task 1: Determine the Fate of Antibiotics in Simulated Anaerobic and Aerobic Lagoons This task will determine the biodegradation kinetics of 4 antibiotics in anaerobically and aerobically operated bioreactors that are fed with an actual animal waste that is amended with additional antibiotics that may not be present. Six bioreactors are operated in parallel for fill and draw periods of time ranging from a few days to a few months. When the animal waste is to be fed to a reactor, it is circulated through a closed-to-the atmosphere system that allows filling of a reactor without oxygen contamination. The aerobic experimental apparatus will be similar except that the individual bioreactors will be aerated with air-fed diffusers. These tests will be conducted without light to minimize the growth of phototrophic organisms. The first stage of analysis for manure or lagoon water will be to extract the antibiotics from the solid phase to the liquid phase. After extraction, SPE and concentration procedures will be conducted in the same manner as for aqueous samples. HPLC analyses will be performed using an HP 1100 Series Liquid Chromatograph equipped with an Agilent 1100 Series Thermostatted Auto Sampler and a variable wavelength UV detector. A ThermoFinnigan LCQ Duo ion trap mass spectrometer equipped with a heated capillary interface and electrospray ionization (ESI) will be used to perform the mass spectrometric analysis. Nitrogen will be used for drying and nebulizing. Task 2: Construct demonstration scale lined waste lagoons operated both in aerobic and anaerobic modes. Determine the impact on odor and antibiotic fate. This task will result in the construction and characterization of demonstration scale animal waste lagoons operated in both aerobic and anaerobic modes. The demonstration scale lagoons will be constructed outdoors at the Engineering Research Center. The lined lagoons will be sized to provide adequate simulation of actual lagoons. The lagoons will be filled with animal waste collected from local farms where we have been monitoring the antibiotic concentrations in the lagoons. We may supplement the waste with additional antibiotics to assure that concentrations representative of what we have measured in the field are present. One lagoon will be fitted with aeration diffusers to assure aerobic conditions across either the entire depth or just the surface, depending on the results from the laboratory testing. We will monitor the lagoons for the concentrations of at least 4 antibiotics during a two-year period. During this period we may need to add additional waste and supplement the antibiotic concentration . Task 3: Provide Guidance for Design and Operation of Waste Lagoons for Minimizing Antibiotics and Odors The results of the project will be documented in a report that can be distributed to the CAFO industry. We will attempt to interpret the results in a context that will provide guidance to the CAFO industry on the operation of lagoons to minimize antibiotics along with an assessment of the impact of this on odor control. |
| Keywords | animal, antibiotics, lagoons, aerobic, anaerobic, biodegradation |
| Progress Reports | |
| 2002 | The project began in July of 2002 and therefore this progress report represents about six months of activity. The project is being conducted with two parallel tracks. The first track is the field survey component and the second track is the laboratory experiments and analytical methods. For the first track, a Research Associate in the Soil and Crop Sciences department has been identified to develop the survey and identify animal feed operations throughout the state to collect management practice information and samples from their animal waste facilities. We expect to begin conducting this survey in late Spring 2003. The other project activity that is occurring in parallel is the development of analytical methods to measure the antibiotic compounds at environmentally relevant concentrations (50-100 ng/L). A PhD student was hired to complete analytical method development. For this project, we need to be able to quantify these compounds in two complex matrices, supernatant from animal waste lagoons and surface water. Using these more complex matrices, existing methods were modified to measure seven tetracyclines (TCs); tetracycline, oxytetracycline, chlortetracycline, doxycycline, meclocycline, minocycline, and democlocycline, and six sulfanoamides (SAs); sulfathiazole, sulfamerazine, sulfamethazine, sulfachloropyridazine, sulfamethoxazole, and sulfadimethoxane, and three macrolides (MLs); erythromycin, roxithromycin and tylosin. Although additional refinements for the solid phase extraction/HPLC-MS analytical procedures are needed for the animal waste lagoon matrix, we are ready to begin measuring field survey samples from actual animal feed operations. The survey and analysis of actual animal waste lagoons will provide data on the occurrence of antibiotics at the animal feed facility. We also want to understand if any of these antibiotic compounds are persisting through the lagooning and land application operations and being transported to nearby surface or ground waters. We have begun a study to examine the concentration of these antibiotic compounds in nearby rivers that are influenced by urban and agricultural land uses. We have studied the Poudre River at five sites that represent pristine, urban and agricultural influences. The only site at which no antibiotics were detected was the pristine site in the mountains before the river had encountered urban or agricultural landscapes. By the time the river had exited Fort Collins, 6 of the 11 antibiotics that were monitored were found in the samples. At the site in Greeley with a landscape dominated by agricultural uses, all five of the TCs monitored were present indicating both urban and agricultural influences. Although 3/6 SAs were detected in the river leaving Fort Collins, only 1/6 was found at Greeley. This result indicates that SAs were not originating from agricultural sources and that significant natural attenuation mechanisms were active in the river. We have submitted a manuscript to Water Research describing these and other results of this study of antibiotic occurrence in the Poudre River. |
| 2003 | The focus of the project in 2003 was the development of analytical methods for measuring veterinary antibiotics in animal waste lagoon water. We developed methods to measure four classes of antibiotics at environmentally relevant concentrations: (1) tetracyclines (tetracycline-TC, oxytetracycline-OTC, chlortetracycline-CTC, doxycycline-DXC, demeclocycline-DMC), (2) sulfonamides (sulfathiazole-STZ, sulfamethoxazole-SMX, sulfamethazine-SMT, sulfamerazine-SMR), (3) macrolides (tylosin-TLS, erythromycin-ETM, roxythromycin-RTM) and (4) ionophores (monensin-MON, salinomycin-SAL, narasin-NAR). A method using solid-phase extraction (SPE) combined with high-performance liquid chromatography/ion trap tandem mass spectrometry (LC/MS-MS) has been developed for determination of trace concentrations of these antibiotics in lagoon water. Considerable effort was expended to achieve acceptable compound recoveries in this difficult water matrix. The average recovery was 85%, 83%, and 86% for ETM, RTM and TLS, respectively. Method detection limits (MDL) were 0 .07, 0.03, and 0.05 mg/L for these compounds. The average recovery for the sulfonamides ranged from 89% for SMX to 106% for SMT with a MDL of 0.05 mg/L for all of the compounds. Tetracycline recovery averages ranged from 78% for DXC to 93% for TC and the MDL was between 0 .05 and 0.07 mg/L for all compounds in this class. For the ionophores, the average recovery ranged from 84% for NAR to 105% for MON with an MDL of less than 0.05 mg/L for all compounds. In parallel with the development of these analytical methods, we contacted and collected waste lagoon water samples at a range of concentrated animal feed operations (CAFOs) throughout the state of Colorado. At this point we have collected samples at 7 hog feed operations, 5 beef operations, 7 dairys, 4 sheep farms and 2 two turkey feed operations. We are currently analyzing these lagoon samples for the antibiotics discussed above. We have also developed a waste management survey for each of these types of operations and have begun working with the CAFO operators to collect this information. We have also used the analytical methods that have been developed to conduct an occurrence survey of all four classes of antibiotics in the Cache la Poudre watershed. At the five sites along the Poudre River that were monitored, the only site at which no antibiotics was detected was the pristine site in the mountains before the river had encountered urban or agricultural landscapes. By the time the river had exited Fort Collins (Site 3), 9 of the 15 compounds that were monitored were found in the samples. At Site 5 in Greeley, CO where the river converges with the South Platte River, all five of the TCs and all three of the ionophores monitored were present indicating both urban and agricultural influences. In particular, monensin only occurred in samples from sites 4 and 5, the two sites that represented an agricultural landscape. Since monensin is not used for human purposes and is not used therapeutically for animals (only as a feed additive), the results indicate that CAFOs are likely releasing this compound to the environment. |
| 2004 | The focus of the project in 2004 was the continued development of analytical methods for measuring veterinary antibiotics in animal waste lagoon water and manure. We have developed methods to measure five classes of antibiotics at environmentally relevant concentrations: (1) tetracyclines (tetracycline-TC, oxytetracycline-OTC, chlortetracycline-CTC, doxycycline-DXC, demeclocycline-DMC), (2) sulfonamides (sulfathiazole-STZ, sulfamethoxazole-SMX, sulfamethazine-SMT, sulfamerazine-SMR), (3) macrolides (tylosin-TLS, erythromycin-ETM, roxithromycin-RTM) (4) ionophores (monensin-MON, salinomycin-SAL, narasin-NAR) and beta-lactams (penicillin G, penicillin V, amoxicillin). A method using solid-phase extraction (SPE) combined with high-performance liquid chromatography/ion trap tandem mass spectrometry (LC/MS-MS) has been developed for determination of trace concentrations of these antibiotics in lagoon water and manure. We have published 4 papers on the methods and have several more in development. In parallel with the development of these analytical methods, we contacted and collected waste lagoon water samples at a range of concentrated animal feed operations (CAFOs) throughout the state of Colorado. The waste stream from 25 animal feed operations was sampled for either manure, lagoon water or both. This includes 7 hog feed operations, 5 beef operations, 7 dairys, 4 sheep farms and 2 two turkey feed operations. The results of this occurrence survey indicate that a wide range of antibiotics are present in most animal waste streams, either runoff ponds, waste lagoons or manure. The individual antibiotic results varied widely. For example, tylosin (a macrolide) was found in all but four waste lagoons sampled (an occurrence rate of 84%) but erythromycin and roxythromycin were found in only 36% and 10% of the lagoons sampled, respectively. Tetracycline (occurrence rate of 96%) and sulfadimethoxane (occurrence rate of 44%) were the most common compounds in their classes. The concentrations of the antibiotic compounds also varied significantly. For example, chlortetracycline ranged from less than 100 mg/L is some swine lagoons to over 9000 mg/L in others. Other ranges for compound classes included non-detect (ND) to 52 mg/L for tetracyclines at beef operations and ND-17 mg/L at dairies. Macrolides were detected at the highest concentrations in beef operation runoff ponds (up to 103 mg/L). The wide range of concentrations seems to indicate that feed practices along with waste management procedures can significantly impact the fate of antibiotics in the waste streams. The next phase of the project will attempt to correlate these practices with residual antibiotic concentration. We hope to be able to provide guidance and BMPs to the animal agriculture industry on how to minimize the release of antibiotics to the environment within. Since the data indicates that a significant amount of antibiotic compounds can concentrate in the manure, we will also study how manure handling techniques influence the transport of antibiotics to the environment. |
| 2005 | The goal of this project that began in 2005 is to determine the occurrence and assess the fate of several classes of antibiotics in concentrated animal feed operation(CAFO) waste lagoons. The objectives include developing a database on the occurrence of at least three classes of antibiotics (10 individual) in cattle and swine CAFO waste lagoons. We will also simulate in the lab aerobic and anaerobic lagoons and determine the fate of at least 10 individual antibiotics under each of these conditions. In addition, we will measure antibiotic resistance genes that correspond with the antibiotics and provide guidance on the design and operation of a waste lagoon for minimizing release of antibiotics. Measurement of relatively low concentrations of complex organic compounds in manure and lagoon water matrices is challenging. Methods that combined liquid-liquid extraction (LLE), solid-phase extraction (SPE), liquid chromatography (LC) and tandem mass spectrometry were developed for quantifying three classes of antibiotics in manure and lagoon water matrices. The antibiotics that were measured included five tetracyclines (tetracycline, oxytetracycline, chlortetracycline, doxycycline, and democlocycline, six sulfonamides (sulfathiazole, sulfamerazine, sulfamethazine, sulfachloropyridazine, sulfamethoxazole, and sulfadimethoxane and three macrolides (tylosin, erythromycin and roxythromycin). The waste stream from 25 animal feed operations (swine, beef, dairy, sheep, poultry) was sampled for either manure, lagoon water or both. The results of the occurrence survey indicates that a wide range of antibiotics are present in most animal waste streams, either runoff ponds, waste lagoons or manure. The individual antibiotic results varied widely. For example, tylosin was found in all but four waste lagoons sampled (an occurrence rate of 84%) but erythromycin and roxythromycin were found in only 36% and 10% of the lagoons sampled. Tetracycline (occurrence rate of 96%) and sulfadimethoxane (occurrence rate of 44%) were the most common compounds in their classes. The concentrations of the antibiotic compounds also varied significantly (from non-detect to over 23 mg/L of a tetracycline in a swine lagoon). The wide range of concentrations seems to indicate that feed practices along with waste management procedures can significantly impact the fate of antibiotics in the waste stream. Since the data indicates that a significant amount of antibiotic compounds can concentrate in the manure, we will also study how manure handling techniques influence the transport of antibiotics to the environment. |
| 2006 |
Analytical methods for quantifying three ionophore antibiotics, monensin (MNS), salinomycin
(SAL), and narasin (NAR), were developed for water and sediment matrices. Samples were
prepared with solid phase extraction (SPE) and HPLC tandem mass spectrometry was used to
measure extracts. Since they are not used for human purposes, these compounds can act as
markers for the transport of animal pharmaceuticals to the watershed.The occurrence of three
ionophore compounds was determined at five sampling sites in the Poudre River watershed. MNS
and SAL were only detected at the sample points in the agriculture-influenced section with a
range of 0.002 to 0.011 g/L and 1.8 to 14.6 g/kg for water and sediment
respectively. NAR was detected at sites 2 and 3 (urban landscape) but showed the highest
concentration in agriculture areas. All three ionophores were found in the sediments at
significantly higher concentrations than in water. The occurrence and fate of veterinary
antibiotics in lagoon water and animal manure are a critical and emerging environmental issue
. One of the objectives of this study was to understand the disappearance of oxytetracycline
(OTC), sufamethoxazole (SMX), tylosin (TYL), and monensin (MON) through biodegradation in
waste lagoons. Analytical methods using SPE and LC-MS-MS with ESI(+) and SRM were used to
quantify the compounds in this matrix. Four lagoon conditions were compared: aerobic-20
C, aerobic-4 C, anaerobic-20 C and anaerobic-4 C. OTC, SMX,
TYL and MON were spiked at a concentration of 20 mg/L in each experiment and 50 g/L of sodium
azide was added in abiotic control experiments to eliminate biological activity. Sodium azide
(50 g/L) and mercuric chloride (0.1 g/L) were added for the abiotic anaerobic control
experiments. OTC, SMX, TYL, and MON showed measurable biodegradation in all experiments. OTC,
SMX, TYL, and MON at 20 C followed first-order degradation kinetics with half-lives
of 12, 6.5, 16, and 9.2 days, respectively, under aerobic treatment and 17, 59, 19, and 71
days, respectively, under anaerobic treatment. The half-lives of OTC, SMX, TYL, and MON at 4
C were found to be 80, 204, 46, and 128 days, respectively, under aerobic treatment
and 110, >242, 103, and 193 days, respectively, under anaerobic treatment. Biodegradation
half-lives in the antibiotic spiked at 20 C followed the order of TYL |
| Impact | |
| 2002 | Animal pharmaceutical compounds have been measured in surface and ground water supplies and this has increasingly caused concern in the environmental community. This project will provide the first occurrence survey of these compounds in animal waste lagoons in the state of Colorado. The project will also provide a survey of waste handling practices in the hope that Best Management Practices (BMPs) can be developed for minimizing the impact of these compounds on the environment. The waste handling practice data will be combined with data from controlled laboratory experiments to provide guidance for operating these facilities, proactively addressing this potentially significant environmental issue. |
| 2003 | This research will provide needed information on the occurrence of veterinary antibiotics in waste lagoons around the state of Colorado. This information coupled with occurrence data in streams and groundwater and knowledge of use patterns (human or animal) will allow us to estimate the degree that animal feed operations contribute antibiotics to the environment. |
| 2004 | This research will provide needed information on the occurrence of veterinary antibiotics in waste lagoons around the state of Colorado. This information coupled with occurrence data in streams and groundwater and knowledge of use patterns (human or animal) will allow us to estimate the degree that animal feed operations contribute antibiotics to the environment. |
| 2005 | This project will provide information on what concentrations of veterinary antibiotics are present in the CAFO lagoons found in Northeastern Colorado. Our research group has already documented the occurrence of these compounds in rivers in the area and we are attempting to locate the ultimate source. The second part of the project will determine the optimal approaches for operating waste lagoons to maximize biodegradation of these compounds. These results will lead to developing guidance in the form of Best Management Practices. |
| 2006 | The objective of the project is to determine what veterinary antibiotic compounds are commonly found in animal waste lagoons and what practices can maximize their biodegradation. The project has identified ionophores in the environment and since these antibiotics are only used for animals, the source appears to be animal feed operations. We have developed and published methods for measuring antibiotics in lagoon water. We are now simulating the operation of animal waste lagoons in the lab and defining Best Management Practices. This will allow producers to minimize the release of these compounds to the environment helping support sustainable operations. |
| Publications | |
| 2003 |
Yang, S., Carlson, K.H., 2003 Evolution of Antibiotic Occurrence in a River through Pristine, Urban and Agricultural Landscapes, Water Research, 37:19:4645. |
| 2004 |
Cha, J.M., Yang, S., Carlson, K.H., 2005. Rapid analysis of trace levels of antibiotic polyether ionophores in surface water by solid-phase extraction and liquid chromatography with ion trap tandem mass spectrometric detection. J. Chromatogr. A 1065: 187-198. Yang, S., Carlson, K.H., 2004 Utility of Radioimmunoassay Methods for Measuring Antibiotics in Water, Water Res. 38: 3155 Yang, S., Cha, J.M., Carlson, K.H., 2004. Quantitative determination of trace concentrations of tetracycline and sulfonamide antibiotics in surface water using solid-phase extraction and liquid chromatography/ion trap tandem mass spectrometry, Rapid Commun. Mass Spectrom. 18: 2455 . Yang. S, Carlson, K.H., 2004. Solid-phase extraction-high performance liquid chromatography-ion trap mass spectrometry for analysis of trace concentrations of macrolide antibiotics in natural and wastewater matrices, J. Chromatogr. A 1038: 141. |
| 2006 |
Kim, S.C., Carlson, K.H., 2006, Occurrence and Partitioning of Ionophore Antibiotics in Water and Sediments of a Mixed-Landscape Watershed, Wat. Res., 40:2549. Kim, S.C., Carlson, K.H., 2006, Quantification of Human and Veterinary Antibiotics in Water and Sediment Using SPE/LC/MS/MS, Anal. Bioanal. Chem., Published on-line 18 July 2006. Kim, S.C., Carlson, K.H., 2007, Temporal and Spatial Trends in the Occurrence of Human and Veterinary Antibiotics in Aqueous and River Sediment Matrices,Environ. Sci. Technol.;41(1); 50-57 |