Abstract
Mycoplasma gallisepticum (MG) is considered one of the most economically important mycoplasmal pathogens among poultry industry worldwide. This pathogen has various strains, therefore their detection using culture method is not sufficient. From this point of view, this study was designed to develop a novel TaqMan® real-time polymerase chain reaction assay for direct detection of MG using cytadhesin to encode a surface protein (mgc2) containing a TaqMan FAM-labeled minor groove binder probe that targets this gene and to compare it with conventional polymerase chain reaction and immunological methods. Using serial broth dilution methods against identified MG isolates, minimum inhibitory concentrations of various antimicrobial agents were detected. Throughout Al-Qassim region, Kingdom of Saudi Arabia, 208 specimens were collected from 18 commercial chicken farms where respiratory diseases were present. Furthermore, 180 blood serum samples were investigated for serological diagnosis of MG. Serum plate agglutination and enzyme-linked immunosorbent assay techniques detected positive serological identification in 83 (46.11%) and 97 (53.88%) isolates, respectively. The sensitivity recorded for TaqMan RT-PCR was 10–3 CFU/ml for MG template DNA. Results for RT-PCR revealed 169 positive samples (81.25%), while 108 samples (51.92%) were identified as positive through conventional polymerase chain reaction assay. The results of MIC for 11 antimicrobial agents against 60 identified MG isolates indicated that tiamulin gave the lowest level of MIC50 (≤0.032 µg/ml) followed by tylosin and doxycycline (≤0.032 – ≤0.062 µg/ml); unlike erythromycin, gentamycin did not show a higher activity at low concentrations. In conclusion, the developed TaqMan RT-PCR exhibited higher sensitivity and applicable accuracy than other molecular and serological techniques and thus could be highly recommended for the management of MG susceptibility and to facilitate implementation of effective treatment and prophylactic measures.

Key words: Mycoplasma gallisepticum, TaqMan RT-PCR assay, Antimicrobial resistance