Real time PCR quantification in groundwater of the dehalorespiring Desulfitobacterium dichloroeliminans strain DCA1.
Quantifying microorganisms responsible for bioremediation can provide insight in their behavior and can help to obtain a better understanding of the physicochemical parameters monitored during bioremediation. A real time PCR (RTm PCR) assay based on the detection with SYBR Green I was optimized in order to quantify the 1,2-dichloroethane dehalorespiring Desulfitobacterium dichloroeliminans strain DCA1. A primer pair targeting unique regions of the 16 S rRNA gene was designed and tested in silico for its specificity. Selectivity was furthermore evaluated and a Limit of Quantification of 1.5 x 10(4) cells/microL DNA extract was obtained for spiked groundwater. Real time measurements of groundwater samples retrieved from a bioaugmented monitoring well and which had an average concentration lying in the range of the Limit of Quantification were evaluated positively with regards to reproducibility. Validation of the RTm PCR assay on groundwater samples originating from different sites confirmed the specificity of the designed primer pair. This RTm PCR assay can be used to survey the abundance and kinetics of strain DCA1 in in situ bioaugmentation field studies.
Van Raemdonck
Hilde
H
Ghent University, Laboratory of Microbial Ecology and Technology, Coupure Links 653, B-9000 Gent, Belgium.
Maes
Ann
A
Ossieur
Wendy
W
Verthé
Kristof
K
Vercauteren
Tom
T
Verstraete
Willy
W
Boon
Nico
N
D052160:Benzothiazoles; D001673:Biodegradation, Environmental; D016000:Cluster Analysis; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D045858:Desulfitobacterium / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D003959:Diamines; D005025:Ethylene Dichlorides / Q000378:metabolism*; D005434:Flow Cytometry; D005618:Fresh Water / Q000382:microbiology*; D008856:Microscopy, Fluorescence; D009930:Organic Chemicals / Q000737:chemistry; D010802:Phylogeny; D016133:Polymerase Chain Reaction / Q000379:methods*; D011804:Quinolines; D012336:RNA, Ribosomal, 16S / Q000737:chemistry / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014874:Water Pollutants, Chemical / Q000378:metabolism*
D052160:Benzothiazoles; D004269:DNA, Bacterial; D003959:Diamines; D005025:Ethylene Dichlorides; D009930:Organic Chemicals; D011804:Quinolines; D012336:RNA, Ribosomal, 16S; D014874:Water Pollutants, Chemical; C098022:SYBR Green I; C024565:ethylene dichloride