title: A novel microbial source tracking microarray for pathogen detection and fecal source identification in environmental systems.
abstract: Pathogen detection and the identification of fecal contamination sources are challenging in environmental waters. Factors including pathogen diversity and ubiquity of fecal indicator bacteria hamper risk assessment and remediation of contamination sources. A custom microarray targeting pathogens (viruses, bacteria, protozoa), microbial source tracking (MST) markers, and antibiotic resistance genes was tested against DNA obtained from whole genome amplification (WGA) of RNA and DNA from sewage and animal (avian, cattle, poultry, and swine) feces. Perfect and mismatch probes established the specificity of the microarray in sewage, and fluorescence decrease of positive probes over a 1:10 dilution series demonstrated semiquantitative measurement. Pathogens, including norovirus, Campylobacter fetus, Helicobacter pylori, Salmonella enterica, and Giardia lamblia were detected in sewage, as well as MST markers and resistance genes to aminoglycosides, beta-lactams, and tetracycline. Sensitivity (percentage true positives) of MST results in sewage and animal waste samples (21-33%) was lower than specificity (83-90%, percentage of true negatives). Next generation DNA sequencing revealed two dominant bacterial families that were common to all sample types: Ruminococcaceae and Lachnospiraceae. Five dominant phyla and 15 dominant families comprised 97% and 74%, respectively, of sequences from all fecal sources. Phyla and families not represented on the microarray are possible candidates for inclusion in subsequent array designs.
authors.affiliation: †Department of Civil and Environmental Engineering, West Virginia University, P.O. Box 6103, Morgantown, West Virginia 26506, United States.
authors.forename: Xiang
authors.initials: X
authors.lastname: Li
authors.affiliation: ‡Department of Integrative Biology, University of South Florida, Tampa, Florida 33620, United States.
authors.forename: Valerie J
authors.initials: VJ
authors.lastname: Harwood
authors.affiliation: ‡Department of Integrative Biology, University of South Florida, Tampa, Florida 33620, United States.
authors.forename: Bina
authors.initials: B
authors.lastname: Nayak
authors.affiliation: §BioTechnology Institute, University of Minnesota, St. Paul, Minnesota 55108, United States.
authors.forename: Christopher
authors.initials: C
authors.lastname: Staley
authors.affiliation: ∥Department of Soil, Water, and Climate, BioTechnology Institute, University of Minnesota, St. Paul, Minnesota 55108, United States.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Sadowsky
authors.affiliation: †Department of Civil and Environmental Engineering, West Virginia University, P.O. Box 6103, Morgantown, West Virginia 26506, United States.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Weidhaas
mesh_terms: D000818:Animals; D001419:Bacteria / Q000235:genetics / Q000302:isolation & purification*; D001717:Birds; D002417:Cattle; D004269:DNA, Bacterial / Q000378:metabolism; D004352:Drug Resistance, Microbial / Q000235:genetics; D004783:Environmental Microbiology; D005243:Feces / Q000382:microbiology* / Q000821:virology; D005453:Fluorescence; D016680:Genome, Bacterial; D006094:Gram-Positive Bacteria; D059014:High-Throughput Nucleotide Sequencing; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D013552:Swine; D062065:Wastewater / Q000382:microbiology
chemical_list: D004269:DNA, Bacterial; D062065:Wastewater