Specificity and sensitivity evaluation of novel and existing Bacteroidales and Bifidobacteria-specific PCR assays on feces and sewage samples and their application for microbial source tracking in Ireland.
Three novel ruminant-specific PCR assays, an existing ruminant-specific PCR assay and five existing human-specific PCR assays, which target 16S rDNA from Bacteroidales or Bifidobacteria, were evaluated. The assays were tested on DNA extracted from ruminant (n=74), human (n=59) and non-ruminant animal (n=44) sewage/fecal samples collected in Ireland. The three novel PCR assays compared favourably to the existing ruminant-specific assay, exhibiting sensitivities of 91-100% and specificities of 95-100% as compared to a sensitivity of 95% and specificity of 94%, for the existing ruminant-specific assay. Of the five human-specific PCR assays, the assay targeting the Bifidobacterium catenulatum group was the most promising, exhibiting a sensitivity of 100% (with human sewage samples) and a specificity of 87%. When tested on rural water samples that were naturally contaminated by ruminant feces, the three novel PCR assays tested positive with a much greater percentage (52-87%) of samples than the existing ruminant-specific assay (17%). These novel ruminant-specific assays show promise for microbial source tracking and merit further field testing and specificity evaluation.
Dorai-Raj
Siobhán
S
Environmental Microbiology Research Unit, Microbiology, School of Natural Sciences, National University of Ireland, Galway, University Road, Galway, Ireland. siobhan.dorairaj@nuigalway.ie
O' Grady
Justin
J
Colleran
Emer
E
D041963:Bacteroidetes / Q000235:genetics / Q000302:isolation & purification*; D001483:Base Sequence; D001644:Bifidobacterium / Q000235:genetics / Q000302:isolation & purification*; D004269:DNA, Bacterial / Q000235:genetics; D004784:Environmental Monitoring / Q000379:methods*; D005243:Feces / Q000382:microbiology*; D006801:Humans; D007494:Ireland; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012680:Sensitivity and Specificity; D012722:Sewage / Q000382:microbiology*; D014873:Water Pollutants / Q000302:isolation & purification*
D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S; D012722:Sewage; D014873:Water Pollutants