Halitosis Biofilm: A Study Combining Culture And Cloning Technique

Oral malodor is a very discomforting condition deriving from the presence of volatile Sulphur compounds in the expired air. In halitosis of intra-oral etiology, the volatile Sulphur compounds are metabolic products of the oral microorganisms within the biofilm coating the tongue dorsum. The aim of this clinical study was to characterize and compare the microbial biofilm in halitosis and healthy volunteers by means of both the culture method and culture-independent 16S-rDNA cloning technique. The study was approved by the ethics committee of the University of Freiburg (Nr. 74/15). The data were statistically assessed by the Wilcoxon Rank-Sum and the Fishers’ exact test. A high bacterial variety (more than 80 different species) resulted from the combination of the two methods. The culture method allowed to identify 47 species in the halitosis group samples among them Veillonella rogosae, never isolated from the tongue surface biofilm so far. High concentrations of Actinomyces graevenitzii were revealed only in the tongue biofilm samples from halitosis patients. The culture-independent cloning method detected more than 50 species and a statistically significant association was found between the presence of Streptococcus mitis, Streptococcus oralis and Streptococcus pseudopneumeniae and the halitosis condition. In conclusion, the combination of the culture-dependent and culture-independent methods provided new insights into the tongue biofilm in halitosis patients highlighting the associations of the aforementioned bacteria with halitosis tongue biofilm. These etiological agents should be considered for further pharmacological research and halitosis therapy strategies.