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SYBR green as a fluorescent probe to evaluate the biofilm physiological state of Staphylococcus epidermidis, using flow cytometry

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Abstract:

Staphylococcus epidermidis biofilms with different proportions of viable but nonculturable bacteria were used to show that SYBR green (SYBR) may be used as a probe to evaluate the bacterial physiological state using flow cytometry. Biofilms grown in excess glucose presented significantly higher proportions of dormant bacteria than biofilms grown in excess glucose with buffered pH conditions or with exponential-phase planktonic cultures. Bacteria obtained from biofilms with high or low proportions of viable but nonculturable cells were further cultured in broth medium and stained with SYBR at different time points. An association between bacterial growth and SYBR staining intensity was observed. In addition, bacteria presenting higher SYBR fluorescence intensity also stained more intensely with cyanoditolyl tetrazolium chloride, used as a probe to evaluate cellular metabolism. Accordingly, planktonic bacteria treated with rifampicin, an inhibitor of bacterial RNA transcription, presented lower SYBR and cyanoditolyl tetrazolium chloride staining intensity than nontreated bacteria. Overall, our results indicate that SYBR, in addition to being used as a component of LIVE/DEAD stain, may also be used as a probe to evaluate the physiological state of S. epidermidis cells.

Keywords: BVNC; CTC; LIVE/DEAD; SYBR green; Staphylococcus epidermidis; VBNC; bactéries viables mais non cultivables; biofilm; chlorure de cyanoditolyl tétrazolium; colorant vital LIVE/DEAD; cyanoditolyl tetrazolium chloride; cytométrie en flux; flow cytometry; viable but nonculturable bacteria

Document Type: Research Article

DOI: http://dx.doi.org/10.1139/w11-078

Affiliations: 1: Laboratório de Imunologia, Departamento de Imunofisiologia e Farmacologia (IMFF), Instituto de Ciências Biomédicas de Abel Salazar – Universidade do Porto (ICBAS–UP), Largo do Professor Abel Salazar 2, 4099-003, Porto, Portugal. 2: Centro de Biologia Molecular e Ambiental (CMBA), Departamento de Biologia, Universidade do Minho, Campus de Gualtar, 4710-057 Braga, Portugal. 3: Centro de Engenharia Biológica – Instituto de Biotecnologia e Bioengenharia (CEB–IBB), Campus de Gualtar, Universidade do Minho, Braga, Portugal. 4: Laboratório de Imunologia, Departamento de Imunofisiologia e Farmacologia (IMFF), Instituto de Ciências Biomédicas de Abel Salazar – Universidade do Porto (ICBAS–UP), Largo do Professor Abel Salazar 2, 4099-003, Porto, Portugal.

Publication date: October 27, 2011

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  • Published since 1954, this monthly journal contains new research in the field of microbiology including applied microbiology and biotechnology; microbial structure and function; fungi and other eucaryotic protists; infection and immunity; microbial ecology; physiology, metabolism and enzymology; and virology, genetics, and molecular biology. It also publishes review articles and notes on an occasional basis, contributed by recognized scientists worldwide.
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