TITLE:
Study on Distribution of Four Pseudomonas Species in Living Environment Using Multiplex PCR
AUTHORS:
Sachiyo Hayashi, Koji Umezawa, Osamu Tsuzukibashi, Akira Fukatsu, Mana Fuchigami, Chiaki Komine, Hiroshi Yamamoto, Mio Hagiwara-Hamano, Yukiko Iizuka, Satoshi Uchibori, Masanobu Wakami, Hiroshi Murakami, Taira Kobayashi, Masahiko Fukumoto, Takato Nomoto
KEYWORDS:
Genus Pseudomonas, Multiplex PCR, Living Environment
JOURNAL NAME:
Open Journal of Stomatology,
Vol.14 No.2,
February
9,
2024
ABSTRACT: Purpose: The genus Pseudomonas is a ubiquitous microorganism frequently detected from immunocompromised patients. The inherent resistance to numerous antimicrobial agents contributes to the opportunistic character of this pathogen exhaustive monitoring of this pathogen is considered of critical importance to public health organizations. The reliable identification method able to distinguish genetic close Pseudomonas species is needed, because these organisms are difficult to differentiate by phenotypic or biochemical methods. The purpose of the present study was to design species-specific primers in order to identify and detect four Pseudomonas species which are frequently detected from the human oral cavities, and to investigate the distribution of these organisms in the living environment using a multiplex PCR. Methods: Polymerase chain reaction (PCR) primers were designed based on partial sequences of the rpoD gene of four Pseudomonas species. Swab samples were collected from fifty washstands, and the distribution of Pseudomonas species was investigated using a conventional PCR at genus level and a multiplex PCR at species level. Results: Multiplex PCR method developed in this study was able to distinguish four Pseudomonas species clearly. The genus Pseudomonas was detected from all samples (100%), whereas P. putida, P, aeruginosa, P. stutzeri and P. fluorescens were detected at 44%, 8%, 4% and 2% in fifty swab samples, respectively. Conclusion: Our developed one-step multiplex PCR method is accurate, specific, cost-effective, time-saving, and works without requiring DNA extraction. It was indicated that washstands were the uninhabitable environment for P. putida, P, aeruginosa, P. stutzeri and P. fluorescens.