title: Establishment of quantitative PCR methods for the quantification of geosmin-producing potential and Anabaena sp. in freshwater systems.
abstract: Geosmin has often been associated with off-flavor problems in drinking water with Anabaena sp. as the major producer. Rapid on-site detection of geosmin-producers as well as geosmin is important for a timely management response to potential off-flavor events. In this study, quantitative polymerase chain reaction (qPCR) methods were developed to detect the levels of Anabaena sp. and geosmin, respectively, by designing two PCR primer sets to quantify the rpoC1 gene (ARG) and geosmin synthase one (GSG) in Anabaena sp. in freshwater systems. The ARG density determined by qPCR assay is highly related to microscopic cell count (r(2) = 0.726, p < 0.001), and the limit of detection (LOD) and limit of quantification (LOQ) of the qPCR method were 0.02 pg and 0.2 pg of DNA, respectively. At the same time, the relationship between geosmin concentrations measured by gas chromatography-mass spectrometry (GC-MS) and GSG copies was also established (r(2) = 0.742, p < 0.001) with similar LOD and LOQ values. Using the two qPCR protocols, we succeeded in measuring different levels of ARG and GSG copies in different freshwater systems with high incidence environmental substrata and diverse ecological conditions, showing that the methods developed could be applied for environmental monitoring. Moreover, comparing to the microscopic count and GC-MS analytical methods, the qPCR methods can reduce the time-to-results from several days to a few hours and require considerably less traditional algal identification and taxonomic expertise.
authors.affiliation: State Key Laboratory of Environmental Aquatic Chemistry, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, 18 Shuangqing Rd., Haidian, Beijing 100085, China. ming.su@live.com
authors.forename: Ming
authors.initials: M
authors.lastname: Su
authors.forename: Virginie
authors.initials: V
authors.lastname: Gaget
authors.forename: Steven
authors.initials: S
authors.lastname: Giglio
authors.forename: Michael
authors.initials: M
authors.lastname: Burch
authors.forename: Wei
authors.initials: W
authors.lastname: An
authors.forename: Min
authors.initials: M
authors.lastname: Yang
mesh_terms: D017033:Anabaena / Q000235:genetics* / Q000378:metabolism*; D017931:DNA Primers; D004784:Environmental Monitoring / Q000379:methods; D005618:Fresh Water / Q000382:microbiology*; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D057230:Limit of Detection; D009284:Naphthols / Q000032:analysis / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D015203:Reproducibility of Results; D013997:Time Factors; D014871:Water Microbiology
chemical_list: D017931:DNA Primers; D009284:Naphthols; C001278:geosmin