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High-Resolution Melting-Curve Analysis of obg Gene to Differentiate the Temperature-Sensitive Mycoplasma synoviae Vaccine Strain MS-H from Non-Temperature-Sensitive Strains
- Source :
- PLoS ONE, PLoS ONE, Vol 9, Iss 3, p e92215 (2014)
- Publication Year :
- 2014
- Publisher :
- Public Library of Science (PLoS), 2014.
-
Abstract
- Temperature-sensitive (ts +) vaccine strain MS-H is the only live attenuated M. synoviae vaccine commercially available for use in poultry. With increasing use of this vaccine to control M. synoviae infections, differentiation of MS-H from field M. synoviae strains and from rarely occurring non-temperature-sensitive (ts –) MS-H revertants has become important, especially in countries where local strains are indistinguishable from MS-H by sequence analysis of variable lipoprotein haemagglutinin (vlhA) gene. Single nucleotide polymorphisms (SNPs) in the obg of MS-H have been found to associate with ts phenotype. In this study, four PCRs followed by high-resolution melting (HRM)-curve analysis of the regions encompassing these SNPs were developed and evaluated for their potential to differentiate MS-H from 36 M. synoviae strains/isolates. The nested-obg PCR-HRM differentiated ts + MS-H vaccine not only from field M. synoviae strains/isolates but also from ts – MS-H revertants. The mean genotype confidence percentages, 96.9±3.4 and 8.8±11.2 for ts + and ts – strains, respectively, demonstrated high differentiating power of the nested-obg PCR-HRM. Using a combination of nested-obg and obg-F3R3 PCR-HRM, 97% of the isolates/strains were typed according to their ts phenotype with all MS-H isolates typed as MS-H. A set of respiratory swabs from MS-H vaccinated specific pathogen free chickens and M. synoviae infected commercial chicken flocks were tested using obg PCR-HRM system and results were consistent with those of vlhA genotyping. The PCR-HRM system developed in this study, proved to be a rapid and reliable tool using pure M. synoviae cultures as well as direct clinical specimens.
- Subjects :
- Veterinary Microbiology
lcsh:Medicine
Nucleic Acid Denaturation
Polymerase Chain Reaction
Poultry
law.invention
law
Genotype
Medicine and Health Sciences
lcsh:Science
Polymerase chain reaction
Multidisciplinary
Attenuated vaccine
Temperature
Agriculture
Animal Models
Vaccination and Immunization
Bacterial Typing Techniques
Bacterial vaccine
Veterinary Diseases
Bacterial Vaccines
Research Article
DNA, Bacterial
Livestock
Sequence analysis
Molecular Sequence Data
Immunology
Mycoplasma synoviae
Biology
Research and Analysis Methods
Microbiology
High Resolution Melt
Veterinary Epidemiology
Model Organisms
Bacterial Proteins
GTP-Binding Proteins
Diagnostic Medicine
Genetics
Animals
Mycoplasma Infections
Genotyping
Poultry Diseases
DNA Primers
Evolutionary Biology
Base Sequence
lcsh:R
Immunity
Biology and Life Sciences
Computational Biology
Virology
Genetic Polymorphism
lcsh:Q
Veterinary Science
Chickens
Population Genetics
Subjects
Details
- ISSN :
- 19326203
- Volume :
- 9
- Database :
- OpenAIRE
- Journal :
- PLoS ONE
- Accession number :
- edsair.doi.dedup.....3c68dd5792d9f844897b42f9d32e4ea9