Qualitative and quantitative estimation of host-specific fecal pollution using Bacteroides–Prevotella 16S rRNA genetic markers by T-RFLP and real-time PCR analyses

摘要

Rapid and reliable determination of the non-point sources of fecal pollution is a critical issue fornthe environmental microbiologists all over the world. In this work we evaluated the use ofnanaerobic bacterial group Bacteroides–Prevotella as an alternative fecal pollution indicator.nTerminal restriction fragment length polymorphism (T-RFLP) and real-time polymerase chainnreaction (RT-PCR) analyses were used to monitor and quantify human-, cow- and pig-specific fecalncontamination in natural river waters. We also included DNA sequence analysis of the identifiednfecal markers revealed by T-RFLP in order to clarify the specificity of each marker. It wasnsuggested that the most influent peaks for each fecal source could be used to identify the sourcenof fecal pollution. Development of specific probes based on these markers permit to quantifynsource of contamination by quantitative RT-PCR. Therefore, we combined the T-RFLP results andnRT-PCR assay to quantify fecal contamination by certain host. We can conclude that T-RFLP andnRT-PCR analyses showed high reproducibility and sensitivity during analyzing real water samplesnand can be used to identify, track and quantify host-specific bacterial genetic markers in complexnnatural water environments.