Dental biofilm pH is the most important determinant of virulence for the development of caries lesions. Confocal microscopy-based pH ratiometry allows monitoring biofilm pH with high spatial resolution. Experiments performed on simplified biofilm models under static conditions identified steep pH gradients as well as localized acidogenic foci that promote enamel demineralization. The present work used pH ratiometry to perform a comprehensive analysis of the effect of whole saliva flow on the microscale pH in complex, in situ-grown 48-h and 96-h biofilms (n = 54) from 9 healthy participants. pH was monitored in 12 areas at the biofilm bottom and top, and saliva flow with film thicknesses corresponding to those in the oral cavity was provided by an additively manufactured microfluidic flow cell. Biofilm pH was correlated to the bacterial composition, as determined by 16S rRNA gene sequencing. Biofilm acidogenicity varied considerably between participants and individual biofilms but also between different areas inside one biofilm, with pH gradients of up to 2 units. pH drops were more pronounced in 96-h than in 48-h biofilms (P = 0.0121) and virtually unaffected by unstimulated saliva flow (0.8 mm/min). Stimulated flow (8 mm/min) raised average biofilm pH to near-neutral values but it did not equilibrate vertical and horizontal pH gradients in the biofilms. pH was significantly lower at the biofilm base than at the top (P < 0.0001) and lower downstream than upstream (P = 0.0046), due to an accumulation of acids along the flow path. pH drops were positively correlated with biofilm thickness and negatively with the thickness of the saliva film covering the biofilm. Bacterial community composition was significantly different between biofilms with strong and weak pH responses but not their species richness. The present experimental study demonstrates that stimulated saliva flow, saliva film thickness, biofilm age, biofilm thickness, and bacterial composition are important modulators of microscale pH in dental biofilms.

中文翻译：

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原位生长的牙科生物膜中微量 pH 值的决定因素。

牙齿生物膜 pH 值是龋齿病变发展毒力的最重要决定因素。基于共聚焦显微镜的 pH 比率测定法可以以高空间分辨率监测生物膜 pH 值。在静态条件下对简化的生物膜模型进行的实验确定了陡峭的 pH 梯度以及促进牙釉质脱矿的局部产酸灶。目前的工作使用 pH 比率测定法对来自 9 名健康参与者的全唾液流对复杂的原位生长 48 小时和 96 小时生物膜 (n = 54) 的微量 pH 值的影响进行了全面分析。监测生物膜底部和顶部 12 个区域的 pH 值，并通过增材制造的微流体流动池提供膜厚度与口腔中的唾液流相对应的唾液流。通过 16S rRNA 基因测序确定，生物膜 pH 值与细菌组成相关。生物膜的产酸性在参与者和个体生物膜之间存在很大差异，而且在一个生物膜内的不同区域之间也存在很大差异，pH 梯度高达 2 个单位。96 小时生物膜的 pH 值下降比 48 小时生物膜更明显 (P = 0.0121)，并且几乎不受未刺激唾液流 (0.8 mm/min) 的影响。刺激流 (8 毫米/分钟) 将平均生物膜 pH 值提高到接近中性值，但它没有平衡生物膜中的垂直和水平 pH 梯度。由于酸沿流路累积，生物膜底部的 pH 值显着低于顶部 (P < 0.0001)，下游的 pH 值低于上游 (P = 0.0046)。pH值下降与生物膜厚度呈正相关，与覆盖生物膜的唾液膜厚度呈负相关。pH 响应强和弱的生物膜之间的细菌群落组成存在显着差异，但物种丰富度没有差异。目前的实验研究表明，刺激的唾液流动、唾液膜厚度、生物膜年龄、生物膜厚度和细菌组成是牙齿生物膜中微尺度pH值的重要调节剂。