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Comparison of disruption procedures for enumeration of activated sludge floc bacteria by flow cytometry
Author(s) -
Falcioni Tania,
Manti Anita,
Boi Paola,
Caico Barbara,
Balsamo Maria,
Papa Stefano
Publication year - 2006
Publication title -
cytometry part b: clinical cytometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.646
H-Index - 61
eISSN - 1552-4957
pISSN - 1552-4949
DOI - 10.1002/cyto.b.20097
Subject(s) - activated sludge , flow cytometry , sonication , bacteria , chemistry , enumeration , sewage treatment , wastewater , staining , microorganism , microbiology and biotechnology , chromatography , viability assay , pulp and paper industry , biology , cell , environmental engineering , environmental science , biochemistry , mathematics , genetics , combinatorics , engineering
Background: In a wastewater treatment plant, the degradation process is performed by a variable and mixed community of microorganisms in an aerobic aquatic environment. The activated‐sludge process is based on the formation of strong microbial flocs where many bacteria are attached to sludge flocs. Methods: Cytometric analysis requires an homogeneous cell suspension and so detachment of bacteria from flocs is required. In this study, sonication and homogenization were compared to find the most adequate pretreatment method for bacterial cytometric analysis in activated sludge samples. Bacterial viability was tested with a nucleic acid double‐staining (NADS) protocol (Barbesti et al., Cytometry 2000;40:214–218) and on flow cytometry. Results: Each method showed a good efficiency in terms of bacterial detachment; thus finally, the choice of which could be the best treatment method was based on both viability results and analysis rapidity. On the basis of the degree of cell detachment and viability, the maximum value was obtained by sonication (2 × 45″). Conclusions: The use of flow cytometry in conjunction with fluorescent dyes and an adequate pretreatment represents a useful method to rapidly detect and enumerate bacteria in activated sludge samples. © 2006 International Society for Analytical Cytology