Resistance toward CHX and cross-resistances toward antibiotics in oral bacteria

抗生素耐药性在过去的二十年中引起了强烈的医学兴趣，从而刺激了这一领域的研究。相反，对防腐剂和杀生物剂的耐药性问题的关注要少得多。在牙科领域，洗必泰（CHX）被认为是金标准防腐剂，因此常用于诊所，也包括在广泛的口腔护理消费品中。由于CHX也常用于其他医学领域（例如用于MRSA的去殖民化），对这种防腐剂的耐药性将对整个医疗行业产生巨大影响。尽管有报道称CHX与医学相关抗生素之间存在交叉耐药性的风险，但迄今为止尚未对此问题进行系统研究。关于对CHX的抗性，人们必须清楚地区分表型适应和获得性抗性，获得性抗性是遗传定义的，因此是稳定的。在口腔生物膜中，使用含CHX的产品时，可能导致细菌产生耐药性的亚抑制浓度CHX区域很容易出现。由于潜在的耐药机制与抗生素相似（如外排泵），可能会进一步产生交叉耐药。因此，里根斯堡和弗赖堡大学医学中心保守牙科和牙周病系的这一联合提案旨在研究口腔细菌在重复暴露于CHX表型和遗传后的耐药性发展。为此，应在体外将口腔分离株反复暴露于CHX，并研究其随后的表型适应。应评估这些分离株的表型和分子遗传学交叉耐药性的发展，并评价其生物膜形成能力的变化。选择对CHX具有明确表型耐药的分离株，通过全基因组测序和mRNA测序，与其各自的野生型菌株进行比较，进一步表征其对CHX的耐药性，此外，还应在牙周手术后需要每天多次使用CHX至少4周的患者的口腔生物膜中，在体内研究对CHX的耐药性的发展。将采用鸟枪-宏基因组分析来评估潜在的微生物组变化以及带有耐药基因的设备。该项目的结果将允许评估对CHX的耐药性传播的风险以及伴随的对抗生素的交叉耐药性，因此可能与牙科和医学领域的所有领域相关。

Antibiotic resistance has attracted strong medial interest during the last two decades, thus stimulating research in this area. Conversely, much less attention has been given to the problem of resistance toward antiseptics and biocides. In the field of dentistry, chlorhexidine (CHX) is considered as gold-standard antiseptic, thus being commonly used in clinics and also included in a wide range of oral care consumer products. As CHX is also commonly used in other medical fields (e.g. for decolonization of MRSA), resistance towards this antiseptic would have tremendous consequences for the whole medical sector.Although there have been reports on the risk of cross-resistances between CHX and medically relevant antibiotics, this issue has not been investigated systematically so far. With regard to resistance towards CHX, one has to clearly distinguish between phenotypic adaptation and acquired resistance, which is genetically defined and therefore stable. In oral biofilms areas with sub-inhibitory concentrations of CHX that potentially lead to development of resistances in bacteria can easily arise upon application of CHX-containing products. As potential resistance mechanisms are similar to those for antibiotics (e.g. efflux pumps), further development of cross-resistances is possible.Therefore, the aim of this joint proposal of the Departments of Conservative Dentistry and Periodontology at the University Medical Centers of Regensburg and Freiburg is to investigate the development of resistance in oral bacteria upon repeated exposure to CHX phenotypically and genetically. For this purpose, oral isolates shall be repeatedly exposed to CHX in vitro and investigated for a consequent phenotypic adaptation. These isolates shall be assessed for development of cross-resistances phenotypically as well as molecular-genetically and changes in their biofilm-forming capacities shall be evaluated. Selected isolates with clear phenotypic resistance towards CHX shall further be characterized by whole genome sequencing and mRNA sequencing as compared to their respective wildtype strains.Furthermore, development of resistance towards CHX shall also be investigated in vivo in oral biofilms of patients that need to use CHX multiple times per day for at least 4 weeks upon periodontal surgery. Shotgun-metagenome analyses shall be employed to assess potential microbiome shifts as well as the equipment with resistance genes.The findings from this project shall allow for assessing the risk of a spread of resistance towards CHX and concomitant cross-resistances towards antibiotics, thus potentially being relevant for the all fields within the sectors of dentistry and medicine.