Differentiation of Mycoplasma hyopneumoniae, M flocculare, and M hyorhinis on the basis of amplification of a 16S rRNA gene sequence.

Abstract

To differentiate Mycoplasma hyopneumoniae, the cause of mycoplasmal pneumonia in pigs, from M flocculare and M hyorhinis, an assay, using the polymerase chain reaction to amplify a segment of the 16S rRNA gene sequence, was developed. The assay was found to be useful for identification of field isolates, as well as for identification of laboratory-adapted strains. Amplification of DNA from M hyopneumoniae and M flocculare resulted in products of 200 and 400 base pairs, respectively. The DNA from M hyorhinis was not amplified. The assay was sensitive enough to detect as little as 1,000 genome equivalents of M hyopneumoniae and M flocculare DNA. Sensitivity was increased 100-fold by increasing the concentration of magnesium ion in the reaction buffer from 2 to 4 mM; however, DNA from M hyorhinis was also amplified under these conditions. The DNA from several walled bacteria and from other mycoplasmas was also tested, but none of these DNA samples was amplified, suggesting that the assay was specific for porcine mycoplasmas.