Resistance profiles of fastidious oral anaerobes and other periodontal species by multipoint inoculator: an in vitro study

Periodontal disease is one of the most prevalent chronic infections worldwide. Associations with systemic conditions, including endocarditis, type 2 diabetes mellitus, cardiovascular disease, and tumor progression, have been extensively investigated in recent studies. Furthermore, it involves pathogens that are often anaerobic, biofilm-associated, and difficult to treat owing to inherent resistance mechanisms and the protective nature of biofilms. As dentists prescribe a substantial proportion of antibiotics and antibiotic resistance is increasing globally, innovative diagnostic methods are urgently needed to ensure effective and sustainable antibiotic stewardship. In this study, anaerobic stock isolates from 30 to 35 frozen samples per genera or species were subjected to minimum inhibitory concentration testing. A multipoint inoculator was used to seed agar plates containing serial antibiotic dilutions, allowing assessment of five commonly used antibiotics: amoxicillin, amoxicillin/clavulanic acid, imipenem, clarithromycin, and levofloxacin. This enabled simultaneous testing of multiple strains under strictly anaerobic conditions, reducing the handling time and material consumption. Among the tested oral pathogens, high resistance levels were observed for clarithromycin. Amoxicillin (±clavulanic acid) remains the most reliable first-line option, whereas levofloxacin may serve as an alternative for certain gram negative anaerobes, when standard treatment protocols are unsuccessful. Mixed-inoculum experiments further showed altered antibiotic responses under biofilm-like conditions. The multipoint inoculator enables reliable testing of oxygen-sensitive anaerobes, including mixed biofilms. Clinical relevance The presented method enables dental practitioners and laboratories to obtain resistance data from subgingival pathogens, especially in low- and middle-income countries or remote regions where access to centralized microbiology laboratories is limited. This benefits the wider healthcare system by supporting sustainable antibiotic stewardship through reliable, cost-effective resistance testing and the patients by facilitating the selection of an effective targeted agent, particularly in complex cases where multiple empirical antibiotic therapies have failed.