Background and objective: The microbial profiles of stimulated saliva samples have been shown to differentiate between patients with periodontitis, patients with dental caries, and orally healthy individuals. Saliva was stimulated to allow for easy and rapid collection; however, microbial composition may not reflect the more natural, unstimulated state. The purpose of this study was to validate whether stimulated saliva is an adequate surrogate for unstimulated saliva in determining salivary microbiomes.

Design: Unstimulated (n=20) and stimulated (n=20) saliva samples were collected from 20 orally and systemically healthy, non-smoking participants. Salivary bacterial profiles were analyzed by means of the Human Oral Microbe Identification using Next Generation Sequencing (HOMINGS), and statistical analysis was performed using Mann-Whitney test with Benjamini-Hochberg's correction for multiple comparison, cluster analysis, principal component analysis, and correspondence analysis.

Results: From a total of 40 saliva samples, 496 probe targets were identified with a mean number of targets per sample of 203 (range: 146-303), and a mean number of probe targets of 206 and 200 in unstimulated and stimulated saliva samples, respectively (p=0.62). Based on all statistical methods used for this study, the microbial profiles of unstimulated and stimulated saliva samples collected from the same person were not statistically significantly different.

Conclusions: Analysis of bacterial salivary profiles in unstimulated and stimulated saliva samples collected from the same individual showed comparable results. Thus, the results verify that stimulated saliva is an adequate surrogate of unstimulated saliva for microbiome-related studies.