Abstract
The Raman spectra, water content, and biomass density of wild-type (WT) Pseudomonas aeruginosa PAO1, small colony variant (SCV) PAO1, and Pseudoalteromonas sp. NCIMB 2021 biofilms were compared in order to determine their variation with strain and species. Living, fully submerged biofilms were analyzed in situ by confocal Raman microspectroscopy for up to 2 weeks. Water to biomass ratios (W/BRs), which are the ratios of the O–H stretching vibration of water at 3,450 cm−1 to the C–H stretching band characteristic of biomass at 2,950 cm−1, were used to estimate the biomass density and cell density by comparison with W/BRs of protein solutions and bacterial suspensions, respectively, on calibration curves. The hydration within SCV biofilm colonies was extremely heterogeneous whereas W/BRs were generally constant in young WT biofilm colonies. The mean biomass in biofilm colonies of WT or colony cores of SCV was typically equivalent to 16% to 27% protein (w/v), but was 10% or less for NCIMB 2021. The corresponding cell densities were 7.5 to >10 × 1010 cfu mL−1 for SCV, while the maximum cell density for NCIMB biofilms was 2.8 × 1010 cfu mL−1.
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Acknowledgements
We would like to thank Werner Schnepf for the contributions to the design of the flow cell and for fabricating it. We also thank Leslie Klapstein for her experimental work and Sarah Schooling for her expert advice. We thank the Atlantic Innovation Fund–Atlantic Canada Opportunities Agency, the Natural Sciences and Engineering Research Council, and the St Francis Xavier University Council for Research for the funding.
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Sandt, C., Smith-Palmer, T., Comeau, J. et al. Quantification of water and biomass in small colony variant PAO1 biofilms by confocal Raman microspectroscopy. Appl Microbiol Biotechnol 83, 1171–1182 (2009). https://doi.org/10.1007/s00253-009-2072-y
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DOI: https://doi.org/10.1007/s00253-009-2072-y