Raman signatures of Cnm-positive Streptococcus mutans: III, clinical validation

This study sought to confirm the possibility of using Raman spectroscopy for oral health management by investigating molecular fingerprints of Streptococcus mutans bacteria obtained from clinical samples. A randomized, double blind study was conducted on 30 samples obtained from clinical subjects (n = 30) in search for Cnm-positive strains, while concurrently and independently performing genomic analyses on the same samples. Samples were characterized after biofilm purification and following a 24 h cycle of in vitro stabilization in absence or in presence of sucrose. Following the basic spectroscopic assessments described in two companion papers, the molecular structures of the isolates were analyzed according to quantitative spectroscopic parameters related to the presence of oxysulfur molecules, the degree of disorder of protein secondary structure, and the level of simplification of N-acetylglucosamine in membrane structure in favor of disordered proteins, based on a previously established spectroscopic assessment framework. Spectroscopic and genomic approaches converged in locating (the same) 4 Cnm-positive isolates over a total sampling of 30 (~13%). Clinical validations confirmed that Cnm-positive strains all had higher protein disorder, higher degree of sulfoxidation, and higher levels of peptidoglycan structural simplification as compared to the Cnm-negative ones. These structural characteristics contribute to an increased level of virulence in the isolate, regardless of its bacterial serotype. Besides matching the results of Cnm genomic analyses, Raman assessments added important information on the pathophysiological stress state of the bacterial isolate and opened the way to fast and insightful diagnostics of the oral flora. The present investigation clearly demonstrated the advantages of Raman technology as compared to traditional culture/molecular detection, in terms of speed, label-free, state sensitivity, and application to specific scenarios, such as high-risk caries risk screening and rapid strain virulence typing.