Abstract
Mycoplasma synoviae is an important pathogen of poultry, causing significant economic losses in this industry. Analysis of the unique genes and shared genes among different M. synoviae strains and among related species is helpful for studying the molecular pathogenesis of M. synoviae and provides valuable molecular diagnostic targets to facilitate the identification of M. synoviae species. We selected a total of 46 strains, including six M. synoviae strains, from 25 major animal (including avian) Mycoplasma species/subspecies that had complete genome sequences and annotation information published in GenBank, and used them for comparative genomic analysis. After analysis, 16 common genes were found in the 46 strains. Thirteen single-copy core genes and the 16s rRNA genes were used for genetic evolutionary analysis. M. synoviae was found to have a distant evolutionary relationship not only with other arthritis-causing mycoplasmas, but also with another major avian pathogen, Mycoplasma gallisepticum, that shares the major virulence factor vlhA with M. synoviae. Subsequently, six unique coding genes were identified as shared among these M. synoviae strains that are absent in other species with published genome sequences. Two of the genes were found to be located in the genetically stable regions of the genomes of M. synoviae and were determined to be present in all M. synoviae isolated strains (n = 20) and M. synoviae-positive clinical samples (n = 48) preserved in our laboratory. These two genes were used as molecular diagnostic targets for which SYBR green quantitative PCR detection methods were designed. The two quantitative PCR methods exhibited good reproducibility and high specificity when tested on positive plasmid controls and genomic DNA extracted from different M. synoviae strains, other major avian pathogenic bacteria/mycoplasmas, and low pathogenic Mycoplasma species. The detection limit for the two genes was 10 copies or less per reaction. The clinical sensitivity and specificity of the quantitative PCR methods were both 100% based on testing chicken hock joint samples with positive or negative M. synoviae infection. This research provides a foundation for the study of species-specific differences and molecular diagnosis of M. synoviae.
Highlights
Mycoplasma synoviae is an important poultry pathogen
The number of phylogenetic branches in multiple M. synoviae strains and the distinction between different M. capricolum subspecies and M. mycoides species indicate that the phylogenetic tree constructed with 13 core genes has higher resolution and more accuracy than the phylogenetic tree constructed with 16S rRNA alone at least within the same species when a large number of different species/subspecies of Mycoplasma are analyzed together
Target genes from single bacteria species/subspecies or different species/subspecies of bacteria with close homology can be used for phylogenetic tree analysis [36]
Summary
Mycoplasma synoviae is an important poultry pathogen. it does not directly lead to death in poultry, it causes infectious synovitis, respiratory disease, growth retardation, decrease in egg production, and the production of deformed eggs, leading serious economic losses in the poultry industry [1,2,3]. Far, the best way to prevent diseases caused by M. synoviae is pathogen eradication; this is very expensive and may be cost-prohibitive for farms in many countries and regions. One of the most widely used in the world is the temperature-sensitive MS-H strain, which was developed by the chemical mutagenesis of an Australian field strain 86079/7NS [7]. Comparative genome analysis of the vaccine strain and its wildtype parent strain can be used to find possible pathogenesisrelated genes. The obg gene, which may affect the temperature sensitivity of the MS-H strain, as well as the possible virulence-related gene oppF, were found via comparative genome analysis of the vaccine strain MS-H and its parent strain 86079/7NS [8,9,10]. A comparative genomic analysis of M. synoviae and other major Mycoplasma has not yet been systematically conducted and is the focus of this study
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