舌バイオフィルム中の硫化水素産生細菌と口腔内の悪臭との関係

Hydrogen sulfide-producing bacteria in tongue biofilm and their relationship with oral malodour.

抄録

本研究の目的は、舌バイオフィルム微生物叢中の硫化水素（H2S）産生菌を同定し、細菌叢と口腔内空気中のH2S濃度との関係を調べることである。被験者10名（21～56歳）の口腔内の悪臭レベルを、ガスクロマトグラフィー、ブレストロンおよびオルゴレプティックスコアにより評価した。これらの評価に基づいて、被験者を臭気群と無臭/低臭気群の2群に分けた。舌苔を採取し、0.05%システイン、0.12%グルタチオン、0.02%酢酸鉛を含む嫌気性寒天培地上に広げ、37℃で2週間嫌気培養した。黒または灰色のコロニーを形成する細菌をH2S産生表現型として選択した。無臭/低臭群の総菌数（P<0.005）とH2S産生菌数（P<0.05）は有意に多かった。黒色または灰色のコロニーを形成する細菌（におい群からは126株、においなし／低臭気群からは242株）を再培養し、H2S産生を確認し、16S rRNA遺伝子の塩基配列決定により同定した。Veillonella属（臭気群：38.1％；無臭／低臭気群：46.3％）、Actinomyces属（25.4％；17.7％）、Prevotella属（10.3％；7.8％）が、臭気群および無臭／低臭気群の両方で優勢なH2S産生菌であった。これらの結果は、舌バイオフィルム中のH2S産生菌数の増加が口腔内の悪臭の原因であることを示唆しているが、舌バイオフィルムの細菌組成は2群間で類似していた。

The aims of this study were to identify hydrogen sulfide (H2S)-producing bacteria among tongue biofilm microflora and to investigate the relationship between bacterial flora and H2S levels in mouth air. Oral malodour levels in 10 subjects (age 21-56 years) were assessed by gas chromatography, and Breathtron and organoleptic scores. Based on these assessments, subjects were divided into two groups: an odour group and a no/low odour group. Tongue coatings were sampled and spread onto Fastidious Anaerobe Agar plates containing 0.05% cysteine, 0.12% glutathione and 0.02% lead acetate, and were then incubated anaerobically at 37 degrees C for 2 weeks. Bacteria forming black or grey colonies were selected as H2S-producing phenotypes. The numbers of total bacteria (P<0.005) and H2S-producing bacteria (P<0.05) in the odour group were significantly larger than those in the no/low odour group. Bacteria forming black or grey colonies (126 isolates from the odour group; 242 isolates from the no/low odour group) were subcultured, confirmed as producing H2S and identified according to 16S rRNA gene sequencing. Species of Veillonella (38.1% in odour group; 46.3% in no/low odour group), Actinomyces (25.4%; 17.7%) and Prevotella (10.3%; 7.8%) were the predominant H2S-producing bacteria in both the odour and no/low odour groups. These results suggest that an increase in the number of H2S-producing bacteria in the tongue biofilm is responsible for oral malodour, although the bacterial composition of tongue biofilm was similar between the two groups.