EPA Science Inventory

Integrated assessment of runoff from livestock farming operations: Analytical chemistry, in vitro bioassays, and in vivo fish exposures


Cavallin, J., E. Durhan, N. Evans, K. Jensen, M. Kahl, D. Kolpin, C. LaLone, L. Makynen, S. Seidl, L. Thomas, D. Villeneuve, M. Weberg, V. Wilson, AND G. Ankley. Integrated assessment of runoff from livestock farming operations: Analytical chemistry, in vitro bioassays, and in vivo fish exposures. ENVIRONMENTAL TOXICOLOGY AND CHEMISTRY. Society of Environmental Toxicology and Chemistry, Pensacola, FL, 33(8):1849-1857, (2014).


While the trend toward using concentrated animal feeding operations (CAFOs) has resulted in increased efficiency in food production, this has prompted concern regarding the impact these operations have on the environment. Animal waste from CAFOs can contain varying levels of natural and synthetic androgens and/or estrogens which can pollute surrounding waterways through various transport mechanisms. In these studies, surface stream water was collected from six different CAFO basins. Aqueous concentrations of selected hormones were determined via chemical analyses. Relative androgenic and estrogenic activity was measured using in vitro cell assays (MDA-kb2 and T47D-Kbluc cell assays, respectively). Finally, 48 h static-renewal in vivo exposures, using the same water samples analyzed in vitro, were conducted to examine potential endocrine-disrupting effects in male and female fathead minnows (Pimephales promelas). Mature fish were exposed to four surface water dilutions (0%, 25%, 50%, and 100%) and a 10 ng 17á-ethynylestradiol (EE2)/L or 50 ng 17â-trenbolone (TRB)/L as positive controls. Hepatic expression of vitellogenin (vtg) and estrogen receptor á (esr1) mRNA and gonadal ex vivo testosterone (T) and 17â-estradiol (E2) production were examined after 48 h of exposure. Potentially estrogenic and androgenic steroids were detected in the samples, generally at low ng/L concentrations. Estrogenic activity, as determined by the T47D-Kbluc assays, was detected in all of the samples. In contrast, the MDA-kb2 assay detected androgenic activity in only one of the six CAFO samples. In vivo exposures to surface water from the CAFO basins had no significant dose-dependent effect on gonadal ex vivo T or E2 production in fathead minnows, with the exception of increased male T production in one study. Significant up-regulation of hepatic vitellogenin mRNA expression in male fish exposed to CAFO sample dilutions was also observed at one of the sites, although the response was not dose-dependent. The current study which combines analytical chemistry measurements, in vitro bioassays, and in vivo fish exposures, provides insight into the value of integrating a combination of techniques to obtain a comprehensive representation of an environmental chemical mixture.


The use of concentrated animal feeding operations (CAFOs) has increased over the past several decades as a means to improve food production efficiency. However, the concentration of livestock associated with these operations makes them a potential source of endocrine active pollution of both endogenous and exogenous (e.g., veterinary drugs) to the environment. When assessing the potential risks associated with exposure to endocrine-disrupting chemical mixtures in the environment (e.g., areas of CAFO runoff), combinations of analytical chemistry techniques, in vitro bioassays, and in vivo assays can be useful for predicting responses to such mixtures, where taken alone, a single assay approach may not be adequate. The present study employed an integrated approach that combine the complementary strengths of analytical chemistry, in vitro bioassays, and in vivo testing to characterize surface water from six CAFO basins that was collected following runoff associated with rainfall events. In vitro bioassay approaches detected significant estrogenic activity, even at sites where specific estrogenic analytes were non-detectable. However, employing targeted in vivo endpoints, anchored to key events within endocrine adverse outcome pathways, the results also demonstrated that in vitro estrogenic activity does not necessarily translate into in vivo hazard. Overall, the research provides a useful case-study that supports the value of applying integrated approaches to environmental monitoring. Such approaches are of direct interest to many of EPA’s regions, including Region 5 GLNPO and Region 8. The study also reflects a successful inter-agency collaboration between USGS and US EPA, which leveraged the relative strengths of USGS in environmental sampling and analytical characterization with US EPA MED’s strengths in biological effects characterization.


Integrated assessment of runoff from livestock farming operations: Analytical chemistry, in vitro bioassays, and in vivo fish exposures   Exit

Record Details:

Start Date: 07/21/2014
Completion Date: 07/21/2014
Record Last Revised: 04/27/2015
Record Created: 07/21/2014
Record Released: 07/21/2014
OMB Category: Other
Record ID: 281619