Evaluation of loop-mediated isothermal amplification method for efficient detection of the periodontopathic bacteria Porphyromonas gingivalis

Abstract: Background: Periodontitis is a multifactorial, polymicrobial oral inflammatory illness brought on by oral pathogens. Porphyromonas gingivalis is a Gram-negative, obligatory anaerobic, black-pigmented coccobacillus and is regarded as a primary etiological factor in the progression of periodontitis. Rapid, highly senstitive and specific detection methods are emerging. The present study aimed to evaluate the loop-mediated isothermal amplification (LAMP) technique for efficiently detecting P . gingivalis from subgingival plaque samples of chronic periodontitis patients. Materials and Methods: This study included 50 subgingival plaque samples from patients suffering from chronic periodontitis. The DNA (Deoxyribonucleic acid) was extracted by the “modified proteinase K” method. A set of six primers, targeting the pepO gene of P . gingivalis , was used for conducting LAMP. The amplification was visualized by naked-eye detection and agarose electrophoresis. Conventional polymerase chain reaction (PCR) and real-time qantitative PCR (qPCR) were carried out by targeting the 16SrRNA (16S ribosomal ribonucleic acid) gene of P . gingivalis . Results: The results showed that LAMP detected P . gingivalis in 40 out of 50 samples (80%). Whereas, qPCR and conventional PCR technique detected P. gingivalis in 38 (76%) and 33 (66%) samples respectively. The sensitivity and specificity of the LAMP method were 94.87% and 90.90%, respectively. With qPCR, the sensitivity and specificity were found to be 92.30% and 81.81%, respectively, whereas, with conventional PCR, it was found to be 76.92% and 72.72%, respectively. Conclusion: LAMP is an efficient technique for quick, accurate, and reliable identification of P . gingivalis from subgingival plaque samples. The technique needs to be validated analytically, and further studies can be conducted by taking saliva and/or gingival crevicular fluid samples from periodontitis patients.