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Methods

Use of Bacteroidales Microbial Source Tracking To Monitor Fecal Contamination in Fresh Produce Production

Kruti Ravaliya, Jennifer Gentry-Shields, Santos Garcia, Norma Heredia, Anna Fabiszewski de Aceituno, Faith E. Bartz, Juan S. Leon, Lee-Ann Jaykus
Kruti Ravaliya
aDepartment of Food, Bioprocessing, and Nutrition Sciences, North Carolina State University, Raleigh, North Carolina, USA
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Jennifer Gentry-Shields
aDepartment of Food, Bioprocessing, and Nutrition Sciences, North Carolina State University, Raleigh, North Carolina, USA
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Santos Garcia
bFacultad de Ciencias Biológicas, Universidad Autónoma de Nuevo León, San Nicolás, Nuevo León, Mexico
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Norma Heredia
bFacultad de Ciencias Biológicas, Universidad Autónoma de Nuevo León, San Nicolás, Nuevo León, Mexico
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Anna Fabiszewski de Aceituno
cHubert Department of Global Health, Rollins School of Public Health, Emory University, Atlanta, Georgia, USA
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Faith E. Bartz
cHubert Department of Global Health, Rollins School of Public Health, Emory University, Atlanta, Georgia, USA
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Juan S. Leon
cHubert Department of Global Health, Rollins School of Public Health, Emory University, Atlanta, Georgia, USA
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Lee-Ann Jaykus
aDepartment of Food, Bioprocessing, and Nutrition Sciences, North Carolina State University, Raleigh, North Carolina, USA
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DOI: 10.1128/AEM.02891-13
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  • FIG 1
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    FIG 1

    Box and whisker plot depicting the concentration (in log10 GEC/100 ml) of the AllBac marker on hands, in irrigation water (source and in-field), and on produce samples, sorted by produce type. The lower boundary of the box indicates the 25th percentile, the line within the box represents the median, and the boundary of the box farthest from zero indicates the 75th percentile. Whiskers above the box indicate the 90th percentiles. Points represent outliers. A statistically significant difference was detected in AllBac marker concentration by produce type (P < 0.0001) but not by sample type (P > 0.05).

  • FIG 2
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    FIG 2

    Box and whisker plot depicting the concentration (in log10 GEC/100 ml) of the BFD marker on hands, in irrigation water, and on produce samples, sorted by produce type. The lower boundary of the box indicates the 25th percentile, the line within the box represents the median, and the boundary of the box farthest from zero indicates the 75th percentile. Whiskers above the box indicate the 90th percentiles. Points represent outliers. There was no statistically significant difference in the concentrations of the BFD marker between produce types or between sample types (P > 0.05).

Tables

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  • TABLE 1

    Primers and probes used in this study

    MarkerAssayOligonucleotide name5′→3′ sequenceaReference or source
    General fecal contaminationAllBacAllBac296fGAGAGGAAGGTCCCCCAC25
    AllBacAllBac412rCGCTACTTGGCTGGTTCAG
    AllBacAllBac375Bhqr(FAM)-CCATTGACCAATATTCCTCACTGCTGCCT-(BHQ-1)
    BovineBoBacBoBac367fGAAG(G/A)CTGAACCAGCCAAGTA25
    BoBacBoBac467rGCTTATTCATACGGTACATACAAG
    BoBacBoBac402Bhqr(FAM)-TGAAGGATGAAGGTTCTATGGATTGTAAACTT-(BHQ-1)
    HumanHF183HF183ATCATGAGTTCACATGTCCG16
    HF183BFDRevCGTAGGAGTTTGGACCGTGT
    HF183BFDFAM(FAM)-CTGAGAGGAAGGTCCCCCACATTGGA-(BHQ-1)
    BFDBFDForCGTTCCATTAGGCAGTTGGT26
    BFDBFDRevCGTAGGAGTTTGGACCGTGT
    BFDBFDFAM(FAM)-CTGAGAGGAAGGTCCCCCACATTGGA-(BHQ-1)
    BvulgBVulgF1CATCATGAGTCCRCATGTTCA16
    BvulgBFDRevCGTAGGAGTTTGGACCGTGT
    BvulgBFDFAM(FAM)-CTGAGAGGAAGGTCCCCCACATTGGA-(BHQ-1)
    IACIACProbe(TET)-ATCTCAGTTCGGTGTAGGTCGTTCGCTCC-(BHQ-1)This study
    • ↵a FAM, 6-carboxyfluorescein; BHQ-1, black hole quencher 1; TET, 6-carboxy-2′,4,7,7′-tetrachlorofluorescein succinimidyl ester.

  • TABLE 2

    Percent of samples positive, negative, and uninterpretable for the AllBac Bacteroidales marker

    Produce typeSample type% (no.) of samples
    PositiveNegativeUninterpretableTotal
    Jalapeño pepperProduce rinse24 (4)76 (13)0 (0)100 (17)
    Hand rinse14 (2)86 (12)0 (0)100 (14)
    Irrigation water11 (1)89 (8)0 (0)100 (9)
    All sample types18 (7)83 (33)0 (0)100 (40)
    CantaloupeProduce rinse74 (20)26 (7)0 (0)100 (27)
    Hand rinse47 (8)47 (8)6 (1)100 (17)
    Irrigation water75 (9)25 (3)0 (0)100 (12)
    All sample types66 (37)32 (18)2 (1)100 (56)
    TomatoProduce rinse39 (16)51 (21)10 (4)100 (41)
    Hand rinse37 (7)37 (7)26 (5)100 (19)
    Irrigation water6 (1)89 (16)6 (1)100 (18)
    All sample types31 (24)56 (44)13 (10)100 (78)
    All produceProduce rinse47 (40)48 (41)5 (4)100 (85)
    Hand rinse34 (17)54 (27)12 (6)100 (50)
    Irrigation water28 (11)69 (27)3 (1)100 (39)
    All sample types39 (68)55 (95)6 (11)100 (174)
  • TABLE 3

    Percent of AllBac-positive samples also positive for human-source marker (BFD, HF183, or BVulg)

    Produce typeSample type% (no.) positive
    Jalapeño pepperProduce rinse25 (1)
    Hand rinseNDa
    Irrigation waterND
    All sample types14 (1)
    CantaloupeProduce rinse58 (11)
    Hand rinse67 (6)
    Irrigation water67 (6)
    All sample types62 (23)
    TomatoProduce rinse19 (3)
    Hand rinse57 (4)
    Irrigation waterND
    All sample types29 (7)
    All produceProduce rinse38 (15)
    Hand rinse59 (10)
    Irrigation water55 (6)
    All sample types46 (31)
    • ↵a ND, not detected.

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Use of Bacteroidales Microbial Source Tracking To Monitor Fecal Contamination in Fresh Produce Production
Kruti Ravaliya, Jennifer Gentry-Shields, Santos Garcia, Norma Heredia, Anna Fabiszewski de Aceituno, Faith E. Bartz, Juan S. Leon, Lee-Ann Jaykus
Applied and Environmental Microbiology Jan 2014, 80 (2) 612-617; DOI: 10.1128/AEM.02891-13

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Use of Bacteroidales Microbial Source Tracking To Monitor Fecal Contamination in Fresh Produce Production
Kruti Ravaliya, Jennifer Gentry-Shields, Santos Garcia, Norma Heredia, Anna Fabiszewski de Aceituno, Faith E. Bartz, Juan S. Leon, Lee-Ann Jaykus
Applied and Environmental Microbiology Jan 2014, 80 (2) 612-617; DOI: 10.1128/AEM.02891-13
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