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

在过去的二十年里，抗生素耐药性引起了医学界的强烈兴趣，从而刺激了这一领域的研究。相反，对防腐剂和杀菌剂的抗药性问题关注较少。在牙科领域，洗必泰(CHX)被认为是黄金标准的防腐剂，因此在临床上得到了广泛的应用，并被广泛地包括在口腔护理消费品中。由于CHX还被广泛用于其他医学领域(例如用于MRSA的非殖民化)，对这种防腐剂的抗药性将对整个医疗部门造成巨大后果。尽管有报道称CHX与医学上相关的抗生素之间存在交叉耐药性的风险，但到目前为止，这个问题还没有得到系统的研究。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，后者是遗传定义的，因此是稳定的。在口腔生物被膜区，CHX的亚抑制浓度可能会导致细菌产生耐药性，而使用含CHX的产品很容易出现这种情况。由于潜在的耐药机制类似于抗生素的耐药机制(如外排泵)，交叉耐药的进一步发展是可能的。因此，雷根斯堡大学医学中心和弗赖堡大学医学中心保守牙科和牙周病学系的这项联合提案的目的是调查口腔细菌在反复暴露于CHX表型和遗传时的耐药性发展。为此，口腔分离株应在体外反复暴露于CHX，并研究其随后的表型适应。应从表型和分子遗传学的角度评估这些分离株的交叉耐药性发展情况，并评估其生物被膜形成能力的变化。选择对CHX有明显表型耐药性的菌株，应进一步进行全基因组测序和mRNA测序，以与其各自的野生型菌株进行比较。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研究对CHX的耐药性的发展。应使用散弹枪-元基因组分析来评估潜在的微生物组变化以及具有耐药基因的设备。该项目的结果将允许评估对CHX的耐药性扩散的风险以及伴随而来的对抗生素的交叉耐药性，因此潜在地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.