Laser raman spectroscopy as a potential chair-side microbiological diagnostic device

Culture-dependent and -independent techniques are time-consuming processes requiring highly trained personnel to identify microorganisms contained within a sample. Rapid chair-side identification of microorganisms could reduce the lag time between patient presentation and ideal treatment. As a first step toward this goal, this study aims to determine if laser Raman spectroscopy (LRS) can discern uniqueness among 10 different species of bacteria contained within a medium in unprocessed and processed samples.

Ten bacterial species were individually grown on blood agar plates for 3 days. Checkerboard DNA-DNA hybridization was used for species verification. For the unprocessed samples, a 1.0-cm diameter agar sample, with undisturbed bacterial growth, was transferred for each species to a barium fluoride crystal (BaF(2)) slide and laser scanned for a total of 15 seconds per sample. For the processed samples, bacterial cells were harvested, washed, and resuspended in phosphate-buffered saline buffer at 10(9) cells/mL concentration. Each suspension was laser scanned for 15 seconds on a BaF(2) slide. Select regions of Raman spectra for each species/agar and species/suspension combination were processed using a two-sided t test.

For the 10 bacterial species, 45 bacteria pair combinations were tested for each group. In both groups, LRS was capable of statistically distinguishing among a majority of bacterial pairings based on RS signature differences of means.

Results show each bacterial species generated restricted ranges of unique spectral signatures that were not masked by their containing medium. Chair-side LRS is a promising technique that differentiates among oral bacterial species with a high degree of specificity.