Application of polymerase chain reaction (PCR)-based denaturing gradient gel electrophoresis for analysis of microbiota on the tongue dorsa of subjects with halitosis

Abstract

The purpose of this study was to compare the microbial profiles on the tongue dorsa of healthy subjects and subjects with halitosis using polymerase chain reaction-based denaturing gradient gel electrophoresis (PCR-DGGE). Tongue dorsum scrapings from five healthy subjects and six subjects with halitosis were analyzed by PCR-DGGE using primers specific for the V6 to V8 region of the eubacterial 16S rRNA gene. A dendrogram was derived from the PCR-DGGE fingerprints and the prominent DGGE bands of interest were identified through DNA sequencing. The similarity of the PCR-DGGE fingerprints was determined using the unweighted pair group method with an arithmetic mean dendrogram derived using Dice's Coefficient of Similarity. The sequence of PCR amplicons indicated that the microbial species most associated with halitosis wereHaemophilus parainfluenzae and a phylotype of Lachnospiraceae (Lachnospiraceaegenomosp. C1), whereas Streptococcus salivarius subsp. salivarius, Neisseria mucosaand Neisseria cinerea were species that did not appear to be associated with halitosis and are likely part of the healthy tongue flora. These results suggest that the presence ofH. parainfluenzae and Lachnospiraceae genomosp. C1 may be associated with a shift in the balance of oral microbes in subjects with halitosis. Key words: Polymerase chain reaction-based denaturing gradient gel electrophoresis (PCR-DGGE), 16S rRNA, halitosis, microbial profiles.