Author(s)

Koji Umezawa^1*, Sachiyo Hayashi¹, Osamu Tsuzukibashi², Akira Fukatsu², Satoshi Uchibori³, Mana Fuchigami², Chiaki Komine², Takashi Asano³, Masanobu Wakami³, Taira Kobayashi³, Masahiko Fukumoto², Takato Nomoto¹

¹Department of Special Needs Dentistry, Nihon University School of Dentistry at Matsudo, Chiba, Japan.
²Department of Laboratory Medicine for Dentistry for the Compromised Patient, Nihon University, School of Dentistry at Matsudo, Chiba, Japan.
³Department of Fixed Prosthodontics, Nihon University School of Dentistry at Matsudo, Chiba, Japan.

Purpose: Fusobacterium nucleatum is an opportunistic pathogen involved in periodontal diseases, extraoral infections, and colorectal cancer. Fusobacterium necrophorum causes a variety of necrotic infections. F. nucleatum and F. necrophorum are classified into five and two subspecies, respectively. Conventional identification methods were technically hard to distinguish each subspecies of two Fusobacterium species accurately. The purpose of the present study was to design primers to identify two medically important Fusobacterium species at the subspecies level, using one-step multiplex PCR. Methods: Polymerase chain reaction (PCR) primers were designed based on partial sequences of the 16S ribosomal RNA (16S rRNA) gene, RNA polymerase B (rpoB) gene, and DNA gyrase subunit B (gyrB) of each subspecies of F. nucleatum and F. necrophorum. Results: These primers were able to distinguish each subspecies of F. nucleatum and F. necrophorum and did not display cross-reactivity with representative Fusobacterium species other than F. nucleatum and F. necrophorum. Conclusion: Our developed one-step multiplex PCR method is accurate, specific, cost-effective, time-saving, and worked without requiring DNA extraction.

Fusobacterium nucleatum, Fusobacterium necrophorum, One-Step Multiplex PCR

Umezawa, K. , Hayashi, S. , Tsuzukibashi, O. , Fukatsu, A. , Uchibori, S. , Fuchigami, M. , Komine, C. , Asano, T. , Wakami, M. , Kobayashi, T. , Fukumoto, M. and Nomoto, T. (2022) One Step Multiplex PCR for Identifications at Subspecies Level of Fusobacterium nucleatum and Fusobacterium necrophorum. Open Journal of Stomatology, 12, 183-195. doi: 10.4236/ojst.2022.126018.