This study aimed to compare a real-time PCR (rPCR) test with improved detection limit to serology and culture for the detection of Mycoplasma gallisepticum (MG) infection in chicken breeder flocks. Six hundred and forty-six blood and tracheal swab samples belonging to 31 grandparent chicken breeder flocks were tested by rPCR. The detection limit of rPCR was 0.9 pg microl(-1), with pure MG S6 strain DNA and 100 colony forming units (CFUs) ml(-1), where both pure culture and tracheal swabs were artificially spiked with the same strain. The seropositive flock rate based on both MG RPA and HI were calculated as 48.4% (15/31) and 32.3% (10/31), respectively, while culture- and rPCR-positive flock rates were 16.1% (5/31) and 29.0% (9/31), respectively. On flock-based analysis, culture method detected 5 out of 10 MG seropositive flocks (sensitivity 50%, specificity 100%), whereas rPCR detected 8 out of 10 flocks (sensitivity 80%, specificity 95%). Agreements between serology and culture, and serology and rPCR were 83.9% and 90.3%, respectively. On individual sample-based analysis, culture method detected 26 out of 78 MG seropositive chicken (sensitivity 33%, specificity 100%), whereas rPCR detected 51 out of 78 MG seropositive chickens (sensitivity 65%, specificity 96%). There was 91.9% and 91.4% agreement between serology and culture, and serology and rPCR, respectively. Results of this study indicate that the rPCR with improved in vitro detection limit could detect MG in seropositive chicken flocks. Therefore, we advise the use of rPCR and/or culture for confirmation of serology results obtained from screening MG infection in chicken flocks.
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