Real-time Loop-mediated Isothermal Amplification (LAMP) of mgc2 Gene of Mycoplasma Gallisepticum

INTRODUCTION: Mycoplasma gallisepticum is considered the most pathogenic and economically significant avian Mycoplasma spp. for the worldwide poultry industry. The aim of this study was to develop a novel and sensitive real-time loop-mediated isothermal amplification (LAMP) assay based on the amplification of its mgc2 gene sequence for its rapid molecular detection in poultry. MATERIAL AND METHODS: Blood samples from 300 broiler and layer chickens were screened using a rapid serum agglutination (RSA) test. A real-time LAMP reaction was conducted with seropositive swab samples at 60ºC for 90 min in an ESEQuant tube scanner using 6-carboxyfluorescein as the reporting dye. RESULTS: The sensitivity of the developed assay was 10 fg/µL of DNA. The assay was found 100% specific, showing no cross-reactivity with other avian Mycoplasma species. The proportion found of the positive samples by the real-time LAMP was 58%. In comparison, the RSA was found to detect 52% of positive cases. CONCLUSION: The mgc2 real-time LAMP emerged as a more sensitive and accurate method for molecular detection of M. gallisepticum than RSA. Robustness and precision give it applicability as a potential field diagnostic tool for M. gallisepticum control. The study will be beneficial in reducing economic losses that M. gallisepticum inflicts on the poultry industry. This is the first reported development of a real-time LAMP assay based on the amplification of the mgc2 gene sequence using an ESEQuant tube scanner for galline M. gallisepticum detection.