Phenotypic and Genotypic Identification of Efflux Pump Resistance in Pathogenic Bacteria Isolated from Gingivitis

Antibiotic resistance in pathogenic bacteria is a growing concern in clinical dentistry, particularly in the management of gingivitis and periodontal diseases. Porphyromonas gingivalis, a key periodontal pathogen, has demonstrated increasing resistance to commonly used antibiotics, partly due to efflux pump mechanisms. This study aimed to investigate the phenotypic and genotypic evidence of efflux pump-mediated resistance in P. gingivalis isolates obtained from gingivitis patients. A total of 48 P. gingivalis isolates were collected from 150 gingivitis patients and subjected to antibiotic susceptibility testing. High resistance rates were observed for tetracycline (70.8%) and erythromycin (58.3%), while lower resistance was noted for ciprofloxacin (31.3%) and amoxicillin-clavulanate (25.0%). Phenotypic detection of efflux pump activity was performed using the ethidium bromide (EtBr) cartwheel assay, revealing that 62.5% (30/48) of isolates exhibited efflux activity. The addition of the efflux pump inhibitor carbonyl cyanide m-chlorophenyl hydrazone (CCCP) significantly reduced the minimum inhibitory concentrations (MICs) in 73.3% of these isolates, confirming efflux-mediated resistance. Genotypic analysis via real-time PCR (qPCR) quantified the expression levels of two major efflux pump gene systems, acrAB-tolC and mexAB-oprM, in resistant isolates. High expression (≥5-fold increase) of acrAB-tolC was detected in 60% of isolates, while 40% exhibited high expression of mexAB-oprM. Statistical analysis revealed a strong positive correlation between efflux activity and acrAB-tolC expression (Pearson’s r = 0.82, p < 0.001), and a moderate correlation with mexAB-oprM expression (r = 0.65, p = 0.002). Overexpression of acrAB-tolC was significantly associated with tetracycline (p = 0.003) and erythromycin resistance (p = 0.01), whereas mexAB-oprM overexpression correlated with ciprofloxacin resistance (p = 0.02). These findings underscore the critical role of efflux pumps in antibiotic resistance among P. gingivalis isolates from gingivitis patients. The study highlights the need for alternative therapeutic strategies, such as efflux pump inhibitors, to combat resistance. Further research should explore the clinical applicability of targeting efflux mechanisms to improve treatment outcomes in periodontal infections.