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Enhancing Mycoplasma synoviae control: A novel quantitative real-time PCR approach to differentiate MS-H vaccine
Mengyao Chen1, Mengjiao Guo1, Chengcheng Zhang1
1Jiangsu Co-Innovation Center for the Prevention and Control of Animal Infectious Disease and Zoonoses, College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China.
Abstract:
Mycoplasma synoviae (MS) is a prevalent pathogen in poultry, causing substantial economic losses. Vaccination is a crucial strategy for controlling MS infections. In China, a temperature-sensitive live vaccine, Vaxsafe®MS (MS-H strain), was introduced to address the rising prevalence of MS since 2017. Accurate differentiation between the MS-H vaccine strain and field strains is essential for the effective application of the vaccine and the eradication of MS in poultry. In this study, a duplex TaqMan real-time quantitative PCR (qPCR) method was first developed based on a single nucleotide polymorphism (SNP) at position 367 of the obg gene. However, some reisolated strains of the MS-H vaccine strain may exhibit reverse mutations at this site, potentially leading to false-positive results field strains detection. To address this issue, a comprehensive comparison of 24 MS genomes was conducted, identifying 18 SNPs unique to the MS-H lineage. Subsequently, an (A/G) SNP at position 131 of the MSH_02330 gene was selected through sequencing. Based on this SNP, a novel duplex qPCR method was established, characterized by high sensitivity, specificity, and repeatability. This method eliminates the influence of obg revertant strains, enabling accurate differentiation of the MS-H vaccine strain from MS field strains. It can be applied to both pure cultures and clinical samples, serving as a valuable tool for evaluating the immunogenicity of the MS-H vaccine and advancing the prevention and control of MS infections.
Insights
A new qPCR method accurately distinguishes the Mycoplasma synoviae (MS) vaccine strain from field strains. This tool is vital for effective vaccination strategies and controlling MS in poultry populations.
Area of Science:
- Veterinary Microbiology
- Molecular Diagnostics
- Poultry Disease Control
Background:
- Mycoplasma synoviae (MS) causes significant economic losses in poultry farming.
- Effective vaccination strategies are essential for MS control, with the MS-H vaccine strain used in China since 2017.
- Accurate differentiation between MS vaccine and field strains is critical for disease management and eradication efforts.
Purpose of the Study:
- To develop a reliable method for differentiating the MS-H vaccine strain from field strains of Mycoplasma synoviae.
- To overcome limitations of previous methods susceptible to reverse mutations in vaccine strains.
- To provide a tool for monitoring vaccine efficacy and guiding MS control programs.
Main Methods:
- Initial development of a duplex TaqMan real-time quantitative PCR (qPCR) targeting a SNP in the obg gene.
- Genomic comparison of 24 MS strains to identify lineage-specific SNPs.
- Selection of a novel (A/G) SNP in the MSH_02330 gene for assay development.
- Establishment and validation of a new duplex qPCR method based on the selected SNP.
Main Results:
- The initial obg gene-based qPCR method showed potential for MS-H strain differentiation but was susceptible to false positives due to reverse mutations.
- Genomic analysis identified 18 SNPs unique to the MS-H lineage, leading to the selection of a novel SNP in the MSH_02330 gene.
- The newly developed duplex qPCR method demonstrated high sensitivity, specificity, and repeatability.
- This novel method accurately differentiates the MS-H vaccine strain from field strains, unaffected by obg revertant strains.
Conclusions:
- A novel duplex qPCR assay targeting an MSH_02330 gene SNP provides accurate differentiation of the Mycoplasma synoviae MS-H vaccine strain.
- This method overcomes the limitations of previous assays, ensuring reliable detection and preventing misidentification of field strains.
- The assay is applicable to pure cultures and clinical samples, supporting vaccine immunogenicity evaluation and MS control strategies in poultry.
