Cytochrome P450 Induction and Gene Expression in Channel Catfish (Ictalurus Punctatus) Following Wastewater Treatment Plant Effluent Exposure in Field and Laboratory Settings

DOI: 10.4236/jep.2010.14042   PDF   HTML     6,115 Downloads   12,223 Views   Citations


The objectives of this study were as follows: 1) to establish a baseline ethoxyresorufin-O-deethylase (EROD) activity level in channel catfish (Ictalurus punctatus), 2) to assess changes in induction of cytochrome P450 enzyme in channel catfish following exposure to creek water at the discharge point from the Troy (Alabama) Wastewater Treatment Plant (TWWTP) compared to upstream samples from Walnut Creek, 3) to compare EROD activity in populations maintained in laboratory and field settings, and 4) to quantify cytochrome P450 gene expression. Enzyme activity was measured fluorometrically and CYP1 gene expression was analyzed by quantitative real-time reverse transcription polymerase chain reaction. A mean EROD baseline was established at 0.03 nmol/min/µg of protein. The overall mean field effluent (TF) EROD had a significant 5-fold increase over field upstream (UF) exposed catfish; and overall mean laboratory effluent (TL) exposed catfish EROD had a significant 1.8-fold increase over laboratory upstream (UL) exposed catfish. Field exposures generally showed more robust enzyme induction over laboratory exposures on all sampling days. Ex- pression of the CYP1B gene following TF exposure was 6-fold over UF. Results suggested that in situ exposure to wastewater pollutants using caged test organisms provided a much more sensitive local monitor of pollutant exposure and biological impact than ex situ toxicological studies.

Share and Cite:

A. Whatley, I. Cho, C. Magrath, P. Stewart and R. Li, "Cytochrome P450 Induction and Gene Expression in Channel Catfish (Ictalurus Punctatus) Following Wastewater Treatment Plant Effluent Exposure in Field and Laboratory Settings," Journal of Environmental Protection, Vol. 1 No. 4, 2010, pp. 362-373. doi: 10.4236/jep.2010.14042.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] M. L. Haasch, R. Prince, P. J. Wejksnora, K. R. Cooper and J. J. Lech, “Caged and Wild Fish: Induction of Hepatic Cytochrome P-450 (CYP1A1) as an Environmental Bio- monitor,” Environmental Toxicology and Chemistry, Vol. 12, No. 5, pp. 885-895, 1993.
[2] J. C. Amiard, T. Caquet, and L. Lagadic, “Use of Bio- Markers for Environmental Quality Assessment,” CRC Press, London, 2000.
[3] M. Parveen, Y. Momose, E. Kitagawa, S. Kurita, O. Ko- dama and H. Iwahashi, “Bioassay of Pesticide Lindane Using Yeast-DNA Microarray Technology,” Chem-Bio In- formatics Journal, Vol. 3, No. 1, 2003, pp. 12-29.
[4] Murata, S. Murata-Mizukami, E. Kitagawa, H. Iwahashi and K. Takamizawa, “The Evaluation of Environmental Waters Using Yeast DNA Microarray,” Chem-Bio Infor- matics Journal, Vol. 6, No. 2, 2003, pp. 29-46.
[5] C. M. Brumley, V. S. Haritos, J. T. Ahokas and D. A. Holdway, “Validation of Biomarkers of Marine Pollution Exposure in Sand Flathead Using Aroclor 1254,” Aquatic Toxicology, Vol. 31, No. 3, 1995, pp. 249-262.
[6] P. A. van Veld, D. J. Westbrook, B. R. Woodin, R. C. Hale, C. L. Smith, R. J. Huggett and J. J. Stegeman, “In- duced Cytochrome P-450 in Intestine and Liver of Spot (Leiostomus xanthurus ) from a Polycyclic Aromatic Hy- drocarbon Contaminated Environment,” Aquatic Toxicol- ogy, Vol. 17, No. 2, 1990, pp. 119-132.
[7] B. Hoeger, M. R. van der Heuvel, B. C. Hitzfeld and D. R. Dietrich, “Effects of Treated Sewage Effluent on Immune Function in Rainbow Trout (Oncorhynchus mykiss),” Aquatic Toxicology, Vol. 70, No. 4, 2004, pp. 345-355.
[8] K. R. Munkittrick, G. J. van Der Kraak, M. E. McMaster, C. B. Portt, M. R. van der Heuvel and M. R. Servos, “Survey of Receiving-Water Environmental Impacts Associated with Discharges from Pulp Mills,” Environmental Toxicology and Chemistry, Vol. 13, No. 7, 1994, pp. 1089-1101.
[9] ADEM (Alabama Department of Environmental Man- agement), “Water Quality Program, Admin. Code R. 335-6-6 et seq., ” Montgomery, Alabama, 2007.
[10] ADEM (Alabama Department of Environmental Man- agement), Environmental Indicators Section, Field Opera- tions Division, “Water quality assessment, Walnut Creek, Troy, Alabama, Pike County,” Montgomery, Alabama, 1997.
[11] U.S EPA (Environmental Protection Agency), National Recommended Water Quality Criteria, 2009. http://www.
[12] S. S. Bhattarai, “Induction of Cytochrome P450 in Chan- nel Catfish (Ictalurus punctatus) Following Exposure to Troy (Alabama) Waste Water Treatment Plant Effluent,” M.S. thesis. Troy University, Troy, 2005.
[13] A. Whatley, P. M. Stewart, S. S. Bhattarai and C. Ma- grath, “Cytochrome P450 in Channel Catfish (Ictalurus punctatus) Following Exposure to Troy (Alabama) Waste Water Treatment Plant Effluent,” Aquatic Ecosystem Health and Management, Vol. 12, No. 2, 2009, pp. 206- 214.
[14] R. W. Li and G. C. Waldbieser, “Genomic Organization and Expression of the Natural Killer Cell Enhancing Factor (NKEF) Gene in Channel Catfish, Ictalurus punctatus (Rafinesque),” Fish & Shellfish Immunology, Vol. 20, No. 1, 2006, pp. 72-82.
[15] M. D. Burk and R. T. Mayer, “Ethoxyresorufin: Direct Fluorimetric Assay of a Microsomal O-Dealkylation Which is Preferentially Inducible by 3-Methylcholanthrene,” Drug Metabolism and Disposition, Vol. 2, No. 6, 1974, pp. 583-588.
[16] K. M. Chan, “Kinetic Cytofluor EROD Assay for Micro- Somes in 48-Well Plates,” 2005. http://www.bch.cuhk.
[17] E. Stephensen, M. Adolfsson-Erici, M. Celander, M. Hu- lander, J. Parkkonen, T. Hegelund, J. Sturve, L. Hassel- berg, M. Bengtsson and L. F?rlin, “Biomarker Responses and Chemical Analyses in Fish Indicate Leakage of Poly- Cyclic Aromatic Hydrocarbons and Other Compounds from Car Tire Rubber,” Environmental Toxicology and Chemis- try, Vol. 22, No. 12, 2003, pp. 2926-2931.
[18] C. D. Rice and L. E. Roszell, “Tributyltin Modulates 3,3’,4,4’,5-Pentachlorobiphenyl (PCB-12 6)-Induced He- patic CYP1A Activity in Channel Catfish, Ictalurus punc- tatus,” Journal of Toxicology and Environmental Health A, Vol. 55, No. 3, 1998, pp. 197-212.
[19] K. J. Livak and T. D. Schmittgen, “Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2-ΔΔCT Method,” Methods, Vol. 25, 2001, pp. 402-408.
[20] W. J. Birge, J. A. Blac, T. M. Short and A. G. Wester- man, “A Comparative Ecological and Toxicological Inves- tigation of a Secondary Wastewater Treatment Plant Efflux- ent and Its Receiving Stream,” Environmental Toxicology and Chemistry, Vol. 8, No. 5, 1989, pp. 437- 450.
[21] R. M. Maier, I. L. Pepper and C P. Gerba, “Environ- mental Microbiology,” Academic Press, Burlington, 2009.
[22] A. M. El-Shehawi, F. K. Ali and M. A. Seehy, “Estima- tion of Water Pollution by Genetic Biomarkers in Tilapia and Catfish Species Shows Species-Site Interaction,” African Journal of Biotechnology, Vol. 6, No. 7, 2007, pp. 840-846.
[23] D. Nadeau, S. Corneau, I. Plante, G. Morrow and R. M. Tanguay, “Evaluation for HSP 70 as a Biomarker of Effect of Pollutants on the Earthworm Lumbricus terrestri,” Cell Stress and Chaperones, Vol. 6, No. 2, 2001, pp. 153-163.
[24] D. Schlenk, M. Chelius, S. K. Wolford and K. M. Chan, “Characterization of Hepatic Metallothionein Expression in Channel Catfish (Ictalurus punctatus) by Reverse- Transcriptase Polymerase Chain Reaction,” Biomarkers, Vol. 2, No. 3, 1997, pp. 161-167.
[25] P. J. Kloepper-Sams and E. Benton, “Exposure of Fish to Biologically Treated Bleached-Kraft Effluent. 2. Induction of Hepatic Cytochrome P450 in Mountain Whitefish (Prosopium williamsoni) and Other Species,” Environmental Toxicology and Chemistry, Vol. 13, No. 9, 1994, pp. 1483-1497.
[26] J. M. Grizzle, S. A. Horowitz and D. R. Strength, “Caged Fish as Monitors of Pollution: Effects of Chlorinated Effluxent from a Wastewater Treatment Plant,” Journal of the American Water Resources Association, Vol. 24, No. 5, 1988, pp. 951-959.
[27] M. O. James, A. H. Altman, K. Morris, K. M. Kleinow and Z. Tong, “Dietary Modulation of Phase 1 and Phase 2 Activities with Benzo(A)Pyrene and Related Compounds in the intestine But Not the Liver of the Channel Catfish, Ictalurus punctatus,” Drug Metabolism and Disposition, Vol. 25, No. 3, 1997, pp. 346-354.
[28] D. E. Watson, W. Reichert and R. T. Di Giulio, “Induc- tion of Hepatic CYP1A in Channel Catfish Increases Binding of 2-Aminoanthracene to DNA in Vitro and in Vivo,” Carcinogenesis, Vol. 19, No. 8, 1998, pp. 1495- 1501.
[29] K. L. Willett, P. R. Gardinali, L. A. Lienesch and R. T. Di Giulio, “Comparative Metabolism and Excretion of Benzo(A)Pyrene in 2 Species of Ictalurid Catfish,” Toxi- cological Sciences, Vol. 58, No. 1, 2000, pp. 68-76.
[30] R. Soimasuo, L. Jokinen and J. Kukkonen, “Biomarker Responses along a Pollution Gradient: Effects of Pulp Mill Effluents on Caged Whitefish,” Aquatic Toxicology, Vol. 31, No. 3, 1995, pp. 329-346.
[31] M. Celander, “Impact of Stress on Animal Toxicology,” In: P. H. M. Balm, Ed., Stress Physiology in Animals, Sheffield Academic Press, Sheffield, 1999, pp. 246-279.
[32] T. K. Collier, B. F. Anulacion and J. E. Stein, “A Field Evolution of Cytochrome p4501A as a Biomarker of Contaminant Exposure in Three Species of Flatfish,” Environmental Toxicology and Chemistry, Vol. 14, 1995, pp. 143-152.
[33] F. Gagne and C. Blaise, “Hepatic Metallothionein Level and Mixed Function Oxidase Activity in Fingerling Rainbow Trout (Oncorhynchus mykiss) after Acute Exposure to Pulp Mill Effluents,” Water Resesearch, Vol. 27, No. 11, 1993, pp. 1669-1682.
[34] J. F. Crivello, “Liver-Somatic Index,” 2009. http://estuary.
[35] M. Z. Vosylien?, “Distinctive Features of Changes in Biological Parameters of Rainbow Trout Exposed to Heavy Metal Model Mixture,” Ekologija, Vol. 48, No. 2, 2002, pp. 22-25.
[36] G. Monod, “L’induction du Cytochrome P4501A1 chez les Poissons,” In: L. Lagadic, T. Caquet, J. C. Amiard and F. Ramade, Eds., Biomarqueurs en Ecotoxicologie: Aspects Foundamentaux, Paris, 1997, pp. 33-54.
[37] U. Kammann, T. Lang, M. Vobach, and W. Wosniok, “Ethoxyresorufin-O-deethylase (EROD) Activity in Dab (Limanda limanda) as Biomarker for Marine Monitoring,” Environmental Science and Pollution Research, Vol. 12, No. 3, 2005, pp. 140-145.

comments powered by Disqus

Copyright © 2020 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.