Development and evaluation of a multi-locus sequence typing scheme for <i>Mycoplasma synoviae</i>

Reproducible molecular <i>Mycoplasma synoviae</i> typing techniques with sufficient discriminatory power may help to expand knowledge on its epidemiology and contribute to the improvement of control and eradication programmes of this mycoplasma species. The present study describes the development and validation of a novel multi-locus sequence typing (MLST) scheme for <i>M. synoviae</i>. Thirteen <i>M. synoviae</i> isolates originating from different poultry categories, farms and lesions, were subjected to whole genome sequencing. Their sequences were compared to that of <i>M. synoviae</i> reference strain MS53. A high number of single nucleotide polymorphisms (SNPs) indicating considerable genetic diversity were identified. SNPs were present in over 40 putative target genes for MLST of which five target genes were selected (<i>nanA</i>, <i>uvrA</i>, <i>lepA</i>, <i>ruvB</i> and <i>ugpA</i>) for the MLST scheme. This scheme was evaluated analysing 209 <i>M. synoviae</i> samples from different countries, categories of poultry, farms and lesions. Eleven clonal clusters and 76 different sequence types (STs) were obtained. Clustering occurred following geographical origin, supporting the hypothesis of regional population evolution. <i>M. synoviae</i> samples obtained from epidemiologically linked outbreaks often harboured the same ST. In contrast, multiple <i>M. synoviae</i> lineages were found in samples originating from swollen joints or oviducts from hens that produce eggs with eggshell apex abnormalities indicating that further research is needed to identify the genetic factors of <i>M. synoviae</i> that may explain its variations in tissue tropism and disease inducing potential. Furthermore, MLST proved to have a higher discriminatory power compared to variable lipoprotein and haemagglutinin A typing, which generated 50 different genotypes on the same database.