title: Flow Cytometric Assessment of Bacterial Abundance in Soils, Sediments and Sludge.
abstract: Bacterial abundance is a fundamental measure in microbiology, but its assessment is often tedious, especially for soil, and sediment samples. To overcome this limitation, we adopted a time-efficient flow-cytometric (FCM) counting method involving cell detachment and separation from matrix particles by centrifugation in tubes receiving sample suspensions and Histodenz(®) solution. We used this approach to assess bacterial abundances in diverse soils (natural and agricultural), sediments (streams and lakes) and sludge from sand-filters in a drinking water treatment plant and compared the results to bacterial abundances determined by two established methods, epifluorescence microscopy (EM) and adenosine triphosphate (ATP) quantification. Cell abundances determined by FCM and EM correlated fairly well, although absolute cell abundances were generally lower when determined by FCM. FCM also showed significant relations with cell counts converted from ATP concentrations, although estimates derived from ATP determinations were typically higher, indicating the presence of ATP sources other than bacteria. Soil and sediment organic matter (OM) content influenced the goodness of fit between counts obtained with EM and FCM. In particular, bacterial abundance determined by FCM in samples containing less than 10% OM, such as stream sediment, was particularly well correlated with the cell counts assessed by EM. Overall, these results suggest that FCM following cell detachment and purification is a useful approach to increase sample throughput for determining bacterial abundances in soils, sediments and sludge. However, notable scatter and only partial concordance among the FCM and reference methods suggests that protocols require further improvement for assessments requiring high precision, especially when OM contents in samples are high.
authors.affiliation: Forest Soils and Biogeochemistry, Swiss Federal Institute for Forest, Snow and Landscape Research (WSL)Birmensdorf Switzerland; Department of Aquatic Ecology, Swiss Federal Institute of Aquatic Science and Technology (Eawag)Dübendorf Switzerland; Department of Experimental Limnology, Leibniz Institute of Freshwater Ecology and Inland Fisheries (IGB), StechlinGermany; Institute of Integrative Biology (IBZ), ETH ZürichZürich Switzerland.
authors.forename: Aline
authors.initials: A
authors.lastname: Frossard
authors.affiliation: Department of Environmental Microbiology, Swiss Federal Institute of Aquatic Science and Technology (Eawag) Dübendorf Switzerland.
authors.forename: Frederik
authors.initials: F
authors.lastname: Hammes
authors.affiliation: Department of Aquatic Ecology, Swiss Federal Institute of Aquatic Science and Technology (Eawag)Dübendorf Switzerland; Department of Experimental Limnology, Leibniz Institute of Freshwater Ecology and Inland Fisheries (IGB), StechlinGermany; Institute of Integrative Biology (IBZ), ETH ZürichZürich Switzerland; Department of Ecology, Berlin Institute of Technology (TU Berlin)Berlin Germany.
authors.forename: Mark O
authors.initials: MO
authors.lastname: Gessner
keywords: ATP; bacterial abundance; drinking water treatment plant sand-filters; epifluorescence microscopy; flow cytometry; sediment; soil