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Data Acceptance Criteria for Standardized Human-Associated Fecal Source Identification Quantitative Real-Time PCR
Orin C Shanks1, Catherine A Kelty2, Robin Oshiro3
1U.S. Environmental Protection Agency, Office of Research and Development, Cincinnati, Ohio, USA shanks.orin@epa.gov.
Applied and Environmental Microbiology
|February 28, 2016
Summary
This study establishes data acceptance criteria for human fecal source identification quantitative real-time PCR (qPCR) to improve water quality management. Standardized protocols and interlaboratory data ensure reliable results for public health protection.
Area of Science:
- Environmental microbiology
- Molecular biology
- Water quality assessment
Background:
- Growing interest in human fecal source identification using quantitative real-time PCR (qPCR) for water management.
- Need for standardized protocols and data quality confidence across laboratories for qPCR applications.
- Lack of consensus on evaluating human fecal source identification qPCR experiments hinders widespread adoption.
Purpose of the Study:
- To establish data acceptance criteria for human fecal source identification qPCR experiments.
- To provide benchmarks for evaluating technical quality based on interlaboratory data.
- To facilitate the transition of qPCR from a research tool to a standardized protocol.
Main Methods:
- Generated data from HF183/BacR287 and HumM2 human-associated qPCR methods across 14 laboratories using standardized protocols.
- Utilized nested analysis of variance (ANOVA) to establish proficiency metrics.
- Assessed extraneous DNA contamination and calibration model performance.
Main Results:
- Established benchmarks for lab-to-lab variability, within-lab replicate testing, and random error.
- Defined criteria for assessing DNA contamination and calibration model performance (correlation coefficient, amplification efficiency, lower limit of quantification).
- Demonstrated implementation of proposed criteria with additional laboratory data.
Conclusions:
- Proposed data acceptance criteria enhance the technical quality evaluation of human fecal source identification qPCR.
- Standardized protocols and defined benchmarks improve confidence in water quality management data.
- Facilitates consistent and reliable application of qPCR for public health protection.
