TY - JOUR
T1 - Validation of Bacteroidales quantitative PCR assays targeting human and animal fecal contamination in the public and domestic domains in India
AU - Odagiri, Mitsunori
AU - Schriewer, Alexander
AU - Hanley, Kaitlyn
AU - Wuertz, Stefan
AU - Misra, Pravas R.
AU - Panigrahi, Pinaki
AU - Jenkins, Marion W.
N1 - Funding Information:
Funding was provided by the Bill and Melinda Gates Foundation (Grant no. OPP1008084 ) through a supplemental grant via the London School of Hygiene and Tropical Medicine (LSHTM) under the Orissa Rural Sanitation Health Impact Study led by Thomas Clasen. Mitsunori Odagiri was also supported by the Japanese Student Exchange Support Program through a Scholarship for Long-term Study Abroad. Sophie Boisson and Parimita Routray helped recruit anonymous volunteers to donate human stool samples. Dipti Mohanty, Tapoja Swain, Jyoti Mohanty, Alok Patra and field staff from AIPH assisted with animal, volunteer human, and diarrhea patient sample collection and processing. Miles Daniel assisted with animal sample collection. The authors acknowledge Mrutyunjay Suar and Priyadarshi Sahu for providing access to the qPCR thermocycler and laboratory space at KIIT.
Publisher Copyright:
© 2014 Elsevier B.V.
PY - 2015/1/1
Y1 - 2015/1/1
N2 - We compared host-associated Bacteroidales qPCR assays developed in the continental United States and Europe for the purpose of measuring the effect of improved sanitation on human fecal exposure in rural Indian communities where both human and animal fecal loading are high. Ten candidate Bacteroidales qPCR assays were tested against fecal samples (human, sewage, cow, buffalo, goat, sheep, dog and chicken) from a test set of 30 individual human, 5 sewage, and 60 pooled animal samples collected in coastal Odisha, India. The two universal/general Bacteroidales assays tested (BacUni, GenBac3) performed equally well, achieving 100% sensitivity on the test set. Across the five human-associated assays tested (HF183 Taqman, BacHum, HumM2, BacH, HF183 SYBR), we found low sensitivity (17 to 49%) except for HF183 SYBR (89%), and moderate to high cross-reactivity with dog (20 to 80%) and chicken fecal samples (60 to 100%). BacHum had the highest accuracy (67%), amplified all sewage samples within the range of quantification (ROQ), and did not cross-react with any fecal samples from cows, the most populous livestock animal in India. Of the ruminant- and cattle-associated assays tested (BacCow, CowM2), BacCow was more sensitive in detecting the full range of common Indian livestock animal fecal sources, while CowM2 only detected cow sources with 50% sensitivity. Neither assay cross-reacted with human sources. BacCan, the dog-associated assay tested, showed no cross-reactivity with human sources, and high sensitivity (90%) for dog fecal samples. Overall, our results indicate BacUni, BacHum, HumM2, BacCan and BacCow would be the most suitable MST assays to distinguish and quantify relative amounts of human-associated and livestock/domestic animal-associated contributions to fecal contamination in Odisha, India.
AB - We compared host-associated Bacteroidales qPCR assays developed in the continental United States and Europe for the purpose of measuring the effect of improved sanitation on human fecal exposure in rural Indian communities where both human and animal fecal loading are high. Ten candidate Bacteroidales qPCR assays were tested against fecal samples (human, sewage, cow, buffalo, goat, sheep, dog and chicken) from a test set of 30 individual human, 5 sewage, and 60 pooled animal samples collected in coastal Odisha, India. The two universal/general Bacteroidales assays tested (BacUni, GenBac3) performed equally well, achieving 100% sensitivity on the test set. Across the five human-associated assays tested (HF183 Taqman, BacHum, HumM2, BacH, HF183 SYBR), we found low sensitivity (17 to 49%) except for HF183 SYBR (89%), and moderate to high cross-reactivity with dog (20 to 80%) and chicken fecal samples (60 to 100%). BacHum had the highest accuracy (67%), amplified all sewage samples within the range of quantification (ROQ), and did not cross-react with any fecal samples from cows, the most populous livestock animal in India. Of the ruminant- and cattle-associated assays tested (BacCow, CowM2), BacCow was more sensitive in detecting the full range of common Indian livestock animal fecal sources, while CowM2 only detected cow sources with 50% sensitivity. Neither assay cross-reacted with human sources. BacCan, the dog-associated assay tested, showed no cross-reactivity with human sources, and high sensitivity (90%) for dog fecal samples. Overall, our results indicate BacUni, BacHum, HumM2, BacCan and BacCow would be the most suitable MST assays to distinguish and quantify relative amounts of human-associated and livestock/domestic animal-associated contributions to fecal contamination in Odisha, India.
KW - Bacteroidales
KW - Fecal pollution
KW - India
KW - Microbial source tracking
KW - Quantitative PCR
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U2 - 10.1016/j.scitotenv.2014.09.040
DO - 10.1016/j.scitotenv.2014.09.040
M3 - Article
C2 - 25285421
AN - SCOPUS:84908354746
VL - 502
SP - 462
EP - 470
JO - Science of the Total Environment
JF - Science of the Total Environment
SN - 0048-9697
ER -