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Published on: February 24, 2023
Detection of Mycoplasma pneumoniae P1 subtype variations by denaturing gradient gel electrophoresis
Jinhong Xiao1, Yang Liu2, Minggui Wang2
1Department of Microbiology and Immunology, University of South China, Hengyang, 421001, P.R. China; Clinical Laboratory, Hunan Provincial People's Hospital, Changsha, 410005, P. R. China.
Abstract:
There were several methods to detect p1 gene variations in Mycoplasma pneumoniae. In this study polymerase chain reaction (PCR)-based denaturing gradient gel electrophoresis (DGGE) assay was performed to establish a rapid and precise detection method for identifying M. pneumoniae p1 gene variations. We detected p1 gene variations in 109M. pneumoniae clinical isolates from Shanghai, China, which were collected from 2009 to 2011 by DGGE, and compared this method with the PCR-based restriction fragment length polymorphism assay and sequencing. By PCR-DGGE method, among the 109M. pneumoniae isolates, 101 (92.7%) isolates were classified into type I, and 8 (7.3%) were classified into type II. Seven (6.9%) type I variations and 8 (100%) type II variations were identified. The match rate of p1 gene variation detected by DGGE reached 100% when compared to DNA sequencing and was more sensitive than restriction fragment length polymorphism. One new type II variant, designated as V2d, was found in this study. The sequence of the new variant was characterized. Our results indicated that PCR-DGGE is a rapid and reliable bio-technique for direct detection of p1 gene variations.
Insights
This study introduces a rapid polymerase chain reaction (PCR)-based denaturing gradient gel electrophoresis (DGGE) method for detecting Mycoplasma pneumoniae p1 gene variations. The PCR-DGGE assay effectively identified variations in clinical isolates, proving highly reliable.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Mycoplasma pneumoniae is a significant respiratory pathogen.
- Accurate detection of p1 gene variations is crucial for understanding M. pneumoniae epidemiology.
- Existing detection methods have limitations in speed and precision.
Purpose of the Study:
- To develop and validate a rapid and precise method for detecting Mycoplasma pneumoniae p1 gene variations.
- To analyze the genetic diversity of p1 gene in clinical isolates from Shanghai.
Main Methods:
- Polymerase chain reaction (PCR)-based denaturing gradient gel electrophoresis (DGGE) assay.
- Analysis of 109 M. pneumoniae clinical isolates collected between 2009-2011.
- Comparison with PCR-based restriction fragment length polymorphism (RFLP) and DNA sequencing.
Main Results:
- PCR-DGGE classified 109 isolates into Type I (92.7%) and Type II (7.3%).
- Seven Type I variations (6.9%) and eight Type II variations (100%) were identified.
- PCR-DGGE demonstrated 100% concordance with DNA sequencing and higher sensitivity than RFLP, identifying a new Type II variant (V2d).
Conclusions:
- PCR-DGGE is a rapid, reliable, and sensitive biotechnique for direct detection of M. pneumoniae p1 gene variations.
- The method aids in characterizing the genetic diversity of M. pneumoniae.
- This technique can be valuable for epidemiological surveillance and diagnostics.
