A novel approach for measuring the bacterial composition of oral plaques, using optical scattering (""""""""Raman"""""""") spectroscopy, is proposed. Raman spectroscopy measures the characteristic vibrational frequencies of a molecule and is, therefore, well-suited for discriminating between similar bacterial species. The proposed method has the potential to be faster, simpler, and more accurate in analyzing the major components of microbial communities than plating and immunofluorescence techniques. The ability to measure bacterial concentrations within minutes by placing samples directly in a laser beam, without preparation or labeling, would be attractive in many laboratory settings. ? ? Experiments will be performed on a series of increasingly complex bacterial samples, from titrated mixtures of two streptococci to bacterial ensembles harvested from the oral plaque of human volunteers. In each phase of the study, spectra of at least twenty samples will be obtained using state-of-the-art Raman spectroscopy. Due to the use of a confocal microscope, only very small sample volumes (0.1 microliters) will be required, and spectral scans will last on the scale of minutes. Spectral differences from sample to sample, caused by variations in chemical content, will be detected and exploited to speciate and enumerate bacterial samples. The target of each experiment will be Raman-based prediction of bacterial concentrations, compared to reference values. Accuracy of prediction will be characterized in each step, providing benchmarks for subsequent studies on more complex samples. ? ? At the conclusion of these studies, the potential for Raman spectroscopy to analyze bacterial content in oral plaques will be thoroughly explored, and the feasibility of developing practical techniques for widespread usage in oral biology research will be clarified. In addition, the methodologies developed during this investigation will be broadly applicable to other problems requiring quantitative discrimination between two or more similar organisms in biological mixtures. ? ?
| Beier, Brooke D; Quivey, Robert G; Berger, Andrew J (2010) Identification of different bacterial species in biofilms using confocal Raman microscopy. J Biomed Opt 15:066001 |
| Beier, Brooke D; Berger, Andrew J (2009) Method for automated background subtraction from Raman spectra containing known contaminants. Analyst 134:1198-202 |
| Smith, Zachary J; Berger, Andrew J (2008) Integrated Raman- and angular-scattering microscopy. Opt Lett 33:714-6 |
| Zhu, Qingyuan; Quivey Jr, Robert G; Berger, Andrew J (2007) Raman spectroscopic measurement of relative concentrations in mixtures of oral bacteria. Appl Spectrosc 61:1233-7 |
