Evaluation of multiple laboratory performance and variability in analysis of recreational freshwaters by a rapid Escherichia coli qPCR method (Draft Method C)

Tiong Gim Aw; Mano Sivaganesan; Shannon Briggs; Erin Dreelin; Asli Aslan; Samuel Dorevitch; Abhilasha Shrestha; Natasha Isaacs; Julie Kinzelman; Greg Kleinheinz; Rachel Noble; Rick Rediske; Brian Scull; Susan Rosenberg; Barbara Weberman; Tami Sivy; Ben Southwell; Shawn Siefring; Kevin Oshima; Richard Haugland

doi:10.1016/j.watres.2019.03.014

Evaluation of multiple laboratory performance and variability in analysis of recreational freshwaters by a rapid Escherichia coli qPCR method (Draft Method C)

Water Res. 2019 Jun 1:156:465-474. doi: 10.1016/j.watres.2019.03.014. Epub 2019 Mar 16.

Authors

Tiong Gim Aw¹, Mano Sivaganesan², Shannon Briggs³, Erin Dreelin⁴, Asli Aslan⁵, Samuel Dorevitch⁶, Abhilasha Shrestha⁶, Natasha Isaacs⁷, Julie Kinzelman⁸, Greg Kleinheinz⁹, Rachel Noble¹⁰, Rick Rediske¹¹, Brian Scull¹¹, Susan Rosenberg¹², Barbara Weberman¹², Tami Sivy¹³, Ben Southwell¹⁴, Shawn Siefring¹⁵, Kevin Oshima¹⁵, Richard Haugland¹⁶

Affiliations

¹ Department of Global Environmental Health Sciences, School of Public Health and Tropical Medicine, Tulane University, 1440 Canal Street, Suite 2100, New Orleans, LA, 70112, USA.
² U.S. Environmental Protection Agency, National Risk Management Research Laboratory, 26 W. M.L. King Dr, Cincinnati, OH, 45268, USA.
³ Water Resources Division, Michigan Department of Environmental Quality, P. O. Box 30458, 525 West Allegan Street, Lansing, MI, 48909, USA.
⁴ Center for Water Sciences, Michigan State University, 1405 South Harrison Road, East Lansing, MI, 48823, USA.
⁵ Georgia Southern University, Department of Environmental Health Sciences, 501 Forest Drive, Statesboro, GA, 30458, USA.
⁶ University of Illinois at Chicago, School of Public Health, 2121 W. Taylor Street, Chicago, IL, 60612, USA.
⁷ U.S. Geological Survey, Upper Midwest Water Science Center, 6520 Mercantile Way, Ste 5, Lansing, MI, 48911, USA.
⁸ City of Racine Public Health Department, 730 Washington Ave, Racine, WI, 53403, USA.
⁹ University of Wisconsin-Oshkosh, Environmental Research Laboratory, 800 Algoma Boulevard, Oshkosh, WI, 54901, USA.
¹⁰ Institute of Marine Sciences, University of North Carolina at Chapel Hill, 3431 Arendell Street, Morehead City, NC, 28557, USA.
¹¹ Annis Water Resources Institute, Lake Michigan Center, 740 W. Shoreline Dr, Muskegon, MI, 49441, USA.
¹² Oakland County Health Division Laboratory, 1200 N. Telegraph, Pontiac, MI, 48341, USA.
¹³ Saginaw Valley State University, Department of Chemistry, 7400 Bay Road, University Center, MI, 48710, USA.
¹⁴ Lake Superior State University, Environmental Analysis Laboratory, 650 W. Easterday Ave, Sault Ste Marie, MI, 49783, USA.
¹⁵ U.S. Environmental Protection Agency, Office of Research and Development, National Exposure Research Laboratory, 26 W. M.L. King Dr, Cincinnati, OH, 45268, USA.
¹⁶ U.S. Environmental Protection Agency, Office of Research and Development, National Exposure Research Laboratory, 26 W. M.L. King Dr, Cincinnati, OH, 45268, USA. Electronic address: haugland.rich@epa.gov.

Abstract

There is interest in the application of rapid quantitative polymerase chain reaction (qPCR) methods for recreational freshwater quality monitoring of the fecal indicator bacteria Escherichia coli (E. coli). In this study we determined the performance of 21 laboratories in meeting proposed, standardized data quality acceptance (QA) criteria and the variability of target gene copy estimates from these laboratories in analyses of 18 shared surface water samples by a draft qPCR method developed by the U.S. Environmental Protection Agency (EPA) for E. coli. The participating laboratories ranged from academic and government laboratories with more extensive qPCR experience to "new" water quality and public health laboratories with relatively little previous experience in most cases. Failures to meet QA criteria for the method were observed in 24% of the total 376 test sample analyses. Of these failures, 39% came from two of the "new" laboratories. Likely factors contributing to QA failures included deviations in recommended procedures for the storage and preparation of reference and control materials. A master standard curve calibration model was also found to give lower overall variability in log₁₀ target gene copy estimates than the delta-delta Ct (ΔΔCt) calibration model used in previous EPA qPCR methods. However, differences between the mean estimates from the two models were not significant and variability between laboratories was the greatest contributor to overall method variability in either case. Study findings demonstrate the technical feasibility of multiple laboratories implementing this or other qPCR water quality monitoring methods with similar data quality acceptance criteria but suggest that additional practice and/or assistance may be valuable, even for some more generally experienced qPCR laboratories. Special attention should be placed on providing and following explicit guidance on the preparation, storage and handling of reference and control materials.

Keywords: Draft method C; E. coli; QA; Recreational water; Variability; qPCR.

Published by Elsevier Ltd.

MeSH terms

Enterococcus
Escherichia coli*
Fresh Water
Water Microbiology*
Water Quality

Grants and funding

EPA999999/ImEPA/Intramural EPA/United States