New PCR for rapid identification of Mycoplasma pneumoniae in clinical specimens from children with respiratory tract infections

We designed a set of primers on the 16 S rRNA gene to directly detect Mycoplasma pneumoniae by PCR in clinical specimens from patients with respiratory infections. The sensitivity of the primers was 2 CFU/tube under the following conditions: 35 cycles of 15 sec at 94°C, 15 sec at 53°C and 15 sec at 72°C/cycle. We therefore judged the PCR results to be positive if the CFU for M. pneumoniae in a clinical specimen was 1.1 × 10³. The results of the PCR were obtained in about 2.5 h. We then used the PCR to examine 783 clinical specimens (epipharynx [n = 612], throat [n = 141], ear discharge [n = 12] etc.) collected from pediatric patients between May 2002 and January 2003. The PCR results were positive in 79 cases (10.1%; 58/291 cases (19.9%) corresponded to specimens from patients with pneumonia, 13/207 cases (6.3%) to specimens from patients with acute bronchitis, 2/130 cases (1.5%) to those from patients with acute pharyngitis and 2/45 cases (4.4%) to those from patients with acute upper respiratory infections. The percentage of PCR-positive cases was significantly higher among the patients with lower part respiratory tract infections (χ² = 53.3008, p = 0.0000). Among the patients with pneumonia, 65 were diagnosed with M. pneumoniae based on a significant rise or high antibody titer for M. pneumoniae as determined by the PA or CF test, and 46 of these cases were PCR-positive (70.8%). We also investigated the correlation between the antibiotics used before PCR and the PCR results in these 65 cases. Of the total of 43 cases, 21 not treated with antibiotics before PCR and 22 cases given oral β-lactam agents ineffective against M. pneumoniae, 38 (88.4%) were PCR-positive, whereas only 3 (23.1%) of the specimens from 13 patients treated with macrolides or new quinolones effective against M. pneumoniae were PCR-positive. We concluded from these results, that the PCR is very useful in selecting suitable chemotherapeutic agents for patients not treated with antibiotics, however it may be necessary to consider the influence of days after to develop infection on PCR results.