Grantee Research Project Results
2004 Progress Report: Fate and Effects of Fluoroquinolone Antibacterial Agents in Aquatic Ecosystems
EPA Grant Number: R829008Title: Fate and Effects of Fluoroquinolone Antibacterial Agents in Aquatic Ecosystems
Investigators: Graham, David W. , deNoyelles, Frank J. , Lydy, Michael J. , Larive, Cynthia K.
Institution: University of Kansas
EPA Project Officer: Page, Angela
Project Period: August 20, 2001 through August 19, 2004 (Extended to August 19, 2006)
Project Period Covered by this Report: August 20, 2003 through August 19, 2004
Project Amount: $520,976
RFA: Drinking Water (2000) RFA Text | Recipients Lists
Research Category: Drinking Water , Water Quality , Water
Objective:
The primary objective of this research project is to assess the fate, attenuation, and ecotoxicity of selected fluoroquinolone (FQ) antibiotics on surface water quality. These compounds were selected for specific investigation because: (1) they are potent antibacterial agents and possible genotoxins; (2) they are frequently used in agriculture and medicine; and (3) little is known about their environmental fate or impact. Another objective is to assess FQs under both laboratory- and field-scale conditions to permit the ready extension of fundamental results to a practical level. In addition, emphasis is placed on developing new methods for detecting and quantifying FQs and their degradation products at low concentrations in environmental samples and developing new molecular techniques for monitoring antibacterial resistance to FQs in exposed microorganisms.
Progress Summary:
Early work on the research project focused on the development of new methods for quantifying ciprofloxacin (cipro), enrofloxacin (enro), and their breakdown products at low concentrations typical of environmental samples. This task largely was achieved in earlier years of the study, primarily using refined extraction and concentration protocols and liquid chromatography (LC)-nuclear magnetic resonance and LC-mass spectrometry (MS)/MS analytical methods.
Third-year studies focused on verification of laboratory testing and the assessment of cipro and enro fate and effects under field conditions (11.3 m3 aquatic mesocosms). Field results generally confirmed laboratory data, indicating that organic particulate matter (particulate organic carbon) and light conditions were both centrally important to FQ fate in surface waters. FQs readily adsorb onto organic particles, and photodegradation reactions substantially destroy both enro and cipro in full sunlight with half-lives as low as 1.2 hours. Field mesocosm data indicate, however, that low levels of residual cipro (a breakdown product of enro in the environment) sometimes can be detected when sunlight levels are less than approximately 25 percent ambient. Furthermore, residual enro levels still were greater than 20 ppb after 30 days environmental exposure (initial level was 25 ppb) when light levels were very low (0.5% of ambient sunlight), suggesting that FQ residuals can accumulate in surface waters under low light conditions (e.g., deep waters in a thermally stratified lake or in aquatic sediments).
Associated with fieldwork assessing FQ fate in aquatic mesocosms, preliminary tests also were performed on new molecular biological methods for tracking FQ antibacterial resistance in exposed organisms. These methods, which initially were developed by Dr. Elizabeth Wellington’s group at the University of Warwick in the United Kingdom, use density gradient gel electrophoresis (DGGE) for tracking mutations in gyrA gene sequences (the quinolone resistance-determining region [QRDR]) that are known to confer FQ resistance in many bacteria (e.g., Pseudomonas aeruginosa). Early data suggest that after 6 weeks of enro exposure at 20-25 ppb in mesocosms, no major change in significant QRDR mutations were noted among exposed organisms (i.e., no apparent increase in FQ resistance based on exposure). Some changes in overall microbial community composition were noted (as detected by community total extracted DNA DGGE), although it has not been possible to unambiguously state whether such variations resulted from FQ exposure or were a consequence of natural dynamic variability in the microbial communities. Statistical analysis is ongoing on these data to quantify the observed effects.
Toxicity tests were performed on five aquatic organisms with seven different FQs, including cipro, lomefloxacin, ofloxacin, levofloxacin, clinafloxacin, enro, and flumequine. Overall, toxicity varied greatly among organisms with the cyanobacteria, Microcystis aeruginosa, being the most sensitive, followed by duckweed (Lemna minor) and then green algae (Pseudokirchneriella subcapitata). Testing on the crustacean Daphnia magna and fathead minnows (Pimephales promelas) showed little or no toxicity with no observed effect levels at or near 10 mg/L. The “actual hazard” of adverse effects in the test organisms was quantified using hazard quotients. An environmental concentration of 1 µg/L was chosen for assessment based on previously reported levels seen in surface waters. At this level, only M. aeruginosa can be considered “at risk”. It should be noted that this observation only considers direct toxicity of FQs under laboratory trial conditions; indirect effects on communities also are possible. Preliminary mesocosm-scale testing has been performed to assess indirect toxicity, although data are not fully analyzed at this time.
Future Activities:
We will continue the three main tasks still ongoing, and a no-cost extension has been requested and granted on the work. First, we will continue refining the molecular biological protocols for tracking FQ resistance and make the methods more quantitative using “real-time” PCR. Second, laboratory experiments are underway assessing the impact of long-term exposure to FQs in bacteria using bench-scale bioreactors simulating biological wastewater treatment units. These experiments have been undertaken because early results indicated that FQs typically were short-lived in light-exposed surface waters. Although elevated light exposure prevails in many systems, such conditions are not always the case in wastewater facilities and such systems may be breeding grounds for antibacterial resistance development. This experiment, therefore, will monitor FQ antibacterial resistance using molecular and traditional culturing methods under five different FQ loading conditions over 3 months (tentative) to assess chronic FQ exposure effects. The final ongoing task is assessing the fate and impact of tetracyclines in surface waters. The inclusion of this new work was requested and granted as a supplement of the workplan. Real-time PCR methods recently were developed, and previous data had suggested that tetracycline resistance might be transported readily through the environment via surface waters. The work here is assessing tetracycline resistance development in mesocosm-scale tanks similar to the FQ experiments. Simultaneous monitoring also is being performed in two geographically distant watersheds that have variable land use to assess relationships between land use, tetracycline resistance, and migration.
Journal Articles on this Report : 6 Displayed | Download in RIS Format
| Other project views: | All 43 publications | 19 publications in selected types | All 18 journal articles |
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Cardoza LA, Almeida VK, Carr A, Larive CK, Graham DW. Separations coupled with NMR detection. Trends in Analytical Chemistry 2003;22(10):766-775. |
R829008 (2002) R829008 (2003) R829008 (2004) R829008 (2005) R829008 (Final) |
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Cardoza LA, Korir AK, Otto WH, Wurrey CJ, Larive CK. Applications of NMR spectroscopy in environmental science. Progress in Nuclear Magnetic Resonance Spectroscopy 2004;45(3-4):209-238. |
R829008 (2003) R829008 (2004) R829008 (2005) R829008 (Final) |
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Cardoza LA, Cutak BJ, Ketter J, Larive CK. High-performance liquid chromatographic-nuclear magnetic resonance investigation of the isomerization of alachlor-ethanesulfonic acid. Journal of Chromatography A 2004;1022(1-2):131-137. |
R829008 (2004) |
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Cardoza LA, Knapp CW, Larive CK, Belden JB, Lydy M, Graham DW. Factors affecting the fate of ciprofloxacin in aquatic field systems. Water, Air and Soil Pollution 2005;161(1-4):383-398. |
R829008 (2003) R829008 (2004) R829008 (2005) R829008 (Final) |
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Knapp CW, Cardoza LA, Hawes J, Wellington EMH, Larive CK, Graham DW. Fate and effects of enrofloxacin in aquatic systems under different light conditions. Environmental Science & Technology 2005;39(23):9140-9146. |
R829008 (2004) R829008 (2005) R829008 (Final) |
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Robinson AA, Belden JB, Lydy MJ. Toxicity of fluoroquinolone antibiotics to aquatic organisms. Environmental Toxicology and Chemistry 2005;24(2):423-430. |
R829008 (2003) R829008 (2004) R829008 (2005) R829008 (Final) |
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Supplemental Keywords:
fluoroquinolone, FQ, photodegradation, adsorption, density gradient gel electrophoresis, DGGE, ecotoxicology, antibacterial resistance, real-time PCR, tetracyclines, quinolone resistance-determining region, QRDR, ciprofloxacin, cipro, enrofloxacin, ecosystem protection/environmental exposure and risk, waste, water, drinking water, ecological effects/environmental exposure and risk, ecological effects, human health, ecological risk assessment, ecosystem/assessment/indicators, environmental chemistry, health risk assessment, antibacterial agents, antibiotics, antiseptics, aquatic ecosystems, chemical contaminants, drinking water contaminants, ecological exposure, exposure, exposure and effects, fate, fate and transport, human health effects, microbial contamination, microbial effects, microbial risk management, monitoring, risk management, pharmaceuticals, stressors, toxicokinetics,, RFA, Scientific Discipline, Ecosystem Protection/Environmental Exposure & Risk, Water, Waste, Ecological Risk Assessment, Health Risk Assessment, Fate & Transport, Environmental Chemistry, Ecosystem/Assessment/Indicators, Ecological Effects - Environmental Exposure & Risk, Ecological Effects - Human Health, Drinking Water, Ecology and Ecosystems, anticeptics, toxicokinetics, ecological effects, microbial contamination, antibiotics, water quality, stressors, microbial risk management, monitoring, chemical contaminants, fate, microbial effects, drinking water contaminants, fate and transport, exposure, ecological exposure, antibacterial agents, pharmaceuticals, fluoroquinolone, human health effects, aquatic ecosystems, exposure and effectsProgress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.