Evaluation of molecular community analysis methods for discerning fecal sources and human waste

Yiping Cao, Laurie C. Van De Werfhorst, Eric A. Dubinsky, Brian D. Badgley, Michael J. Sadowsky, Gary L. Andersen, John F. Griffith, Patricia A. Holden

Research output: Contribution to journalArticlepeer-review

47 Scopus citations


Molecular microbial community analyses provide information on thousands of microorganisms simultaneously, and integrate biotic and abiotic perturbations caused by fecal contamination entering water bodies. A few studies have explored community methods as emerging approaches for microbial source tracking (MST), however, an evaluation of the current state of this approach is lacking. Here, we utilized three types of community-based methods with 64 blind, single- or dual-source, challenge samples generated from 12 sources, including: humans (feces), sewage, septage, dogs, pigs, deer, horses, cows, chickens, gulls, pigeons, and geese. Each source was a composite from multiple donors from four representative geographical regions in California. Methods evaluated included terminal restriction fragment polymorphism (TRFLP), phylogenetic microarray (PhyloChip), and next generation (Illumina) sequencing. These methods correctly identified dominant (or sole) sources in over 90% of the challenge samples, and exhibited excellent specificity regardless of source, rarely detecting a source that was not present in the challenge sample. Sensitivity, however, varied with source and community analysis method. All three methods distinguished septage from human feces and sewage, and identified deer and horse with 100% sensitivity and 100% specificity. Method performance improved if the composition of blind dual-source reference samples were defined by DNA contribution of each single source within the mixture, instead of by Enterococcus colony forming units. Data analysis approach also influenced method performance, indicating the need to standardize data interpretation. Overall, results of this study indicate that community analysis methods hold great promise as they may be used to identify any source, and they are particularly useful for sources that currently do not have, and may never have, a source-specific single marker gene.

Original languageEnglish (US)
Pages (from-to)6862-6872
Number of pages11
JournalWater Research
Issue number18
StatePublished - Nov 15 2013

Bibliographical note

Funding Information:
Funding for this project has been provided in part through an agreement with the California State Water Resources Control Board . The contents of this document do not necessarily reflect the views and policies of the California State Water Resources Control Board, nor does mention of trade names or commercial products constitute endorsement or recommendation for use. A portion of this work was performed under the auspices of the U.S. Department of Energy under contract DE-AC02-05CH11231 to Lawrence Berkeley National Laboratory. We would like to thank staff from the Southern California Coastal Water Research Project, Stanford University, University of California, Los Angeles, and University of California, Santa Barbara for collecting the fecal sources and preparing the blind samples, all the staff members at the community analysis labs for assisting in laboratory work.


  • Microbial community analysis
  • Microbial source tracking
  • Next generation sequencing
  • PhyloChip


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