Polymerase chain reaction for Mycoplasma hyopneumoniae detection in tracheobronchiolar washings from pigs

We have used the polymerase chain reaction (PCR) to detect Mycoplasma hyopneumoniae in tracheobronchiolar washings collected from experimentally infected piglets. On the basis of the published nucleotide sequence of M. hyopneumoniae \141 probe (accession number U02537), primers were chosen to produce an amplified fragment of 1561 bp. All the M. hyopneumoniae strains tested could be detected by the PCR test. DNA from other mycoplasmal and bacterial species currently isolated from respiratory tract of piglets gave negative result. The detection limit was estimated to be 500 fg of purified DNA corresponding to 4 . 10(2) organisms. The sensitivity of PCR reaction was also evaluated on microorganisms in culture, the limit sensitivity was 2 . 5 10(3) organisms. In the present study, a total of 143 tracheobronchiolar washings collected from experimentally infected piglets were submitted to PCR. For each tracheobronchiolar washing, PCR was performed on crude extracts treated with lysis buffer and on extracted DNA. The PCR results obtained with the two kinds of samples were compared to the immunofluorescence (IF) results. This comparison indicates a good correlation between PCR and IF test in 121/143 cases. The presence of M. hyopneumoniae is revealed in 19/143 of the washing samples only by PCR. In our hand, PCR appears to be the more sensitive test to detect M. hyopneumoniae in experimentally infected piglets. (C) 1996 Academic Press Limited