Differential diagnosis between ts-11 vaccine strain and field Mycoplasma gallisepticum isolates in clinical samples by PCR-RFLP

Abstract

Summary Mycoplasma gallisepticum ( MG) is an important avian pathogen causing significant economic losses within the poultry industry. The aim of the present study was to investigate the prevalence of MG in poultry by PCR of mgc2 and 16S rRNA and to set a differential diagnosis between vaccine strain and field MG. We examined the differentiating potential of diagnostic polymerase chain reaction (PCR) primers targeted to the 16S rRNA and mgc2 genes present in MG. For the 16S rRNA and mgc2 PCR assays, 109 samples were taken from 10 RSAT positive farms, including: lung, air sacs and tracheal swabs. For differential diagnosis between the ts-11 vaccine strain and other field isolates, PCR-RFLP with HaeII restricted enzyme was applied. PCR products of 530Â bp and 300Â bp, respectively, appeared on electrophoresis gel with 16S rRNA and mgc2 PCR diagnostic primers specific for MG. Differential diagnosis of MG can be achieved within 1 day of submission of tracheal swab samples by application of the mgc2-PCR–restriction fragment length polymorphism (mgc2-PCR-RFLP) assay with HaeII, giving a 270-bp fragment for ts-11 or no restriction for other MG strains. The test was successfully applied in vivo for detection of MG, and RFLP-PCR can be used for rapid differentiation of the ts-11 vaccine strain from field isolates by direct amplification from clinical samples without the need for isolation by culture.