Quantitative molecular assay for fingerprinting microbial communities of wastewater and estrogen-degrading consortia.
A quantitative fingerprinting method, called the real-time terminal restriction fragment length polymorphism (real-time-t-RFLP) assay, was developed for simultaneous determination of microbial diversity and abundance within a complex community. The real-time-t-RFLP assay was developed by incorporating the quantitative feature of real-time PCR and the fingerprinting feature of t-RFLP analysis. The assay was validated by using a model microbial community containing three pure strains, an Escherichia coli strain (gram negative), a Pseudomonas fluorescens strain (gram negative), and a Bacillus thuringiensis strain (gram positive). Subsequently, the real-time-t-RFLP assay was applied to and proven to be useful for environmental samples; the richness and abundance of species in microbial communities (expressed as the number of 16S rRNA gene copies of each ribotype per milliliter) of wastewater and estrogen-degrading consortia (enriched with 17alpha-estradiol, 17beta-estradiol, or estrone) were successfully characterized. The results of this study strongly suggested that the real-time-t-RFLP assay can be a powerful molecular tool for gaining insight into microbial communities in various engineered systems and natural habitats.
Yu
Chang-Ping
CP
Department of Civil & Environmental Engineering, 110B Perkins Hall, University of Tennessee, Knoxville, TN 37996, USA.
Ahuja
Rajiv
R
Sayler
Gary
G
Chu
Kung-Hui
KH
D001413:Bacillus thuringiensis / Q000235:genetics / Q000378:metabolism; D001673:Biodegradation, Environmental; D016172:DNA Fingerprinting / Q000379:methods*; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D017753:Ecosystem; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D004967:Estrogens / Q000378:metabolism*; D008954:Models, Biological; D012150:Polymorphism, Restriction Fragment Length; D011551:Pseudomonas fluorescens / Q000235:genetics / Q000378:metabolism; D012329:RNA, Bacterial / Q000235:genetics; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D012722:Sewage / Q000382:microbiology; D014871:Water Microbiology
D004269:DNA, Bacterial; D004967:Estrogens; D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S; D012722:Sewage