| Objective:Mycoplasma pneumoniae is a common pathogen causing human mycoplasma pneumonia, In recent years, the infection of mycoplasma pneumoniae is characterized by multiple pulmonary complications and the cases of refractory pneumonia increased. However, the mechanism is unknown. Macrolides are generally considered to be the first-choice agents for treatment of M. pneumoniae infection. The mutations of the M. pneumoniae 23 S rRNA gene result in resistance to macrolides. It is reported that macrolide-resistant (MR) gene mutations were found in 90% samples in Beijing. This study was to analysis the clinical effects of the macrolides for MR M. pneumoniae and the clinical significance of genetic testing. And finally, through a rat model infected with M. pneumoniae, we study the immunological mechanisms of M. pneumoniae,which may offer useful information for more reasonable clinical treatment.Methods:1. Swabs,sputa and Bronchio-alreolar Lavage Fluid (BALF) specimens of 66 patients who fulfilled our criteria were collected. DNA was extracted and then deteced by PCR.The 933bp fragment of the 23S rRNA gene of M. pneumoniae containing mutations was also amplified. Clinical informations of the patients were collected and analyzed. We adopted the number of febrile days,the days of hospitalization and the occurrence of complications and glucocorticoids use as the primary outcome measurement.2.45 healthy male Sprague-Dawley rats were randomly divided into M. pneumoniae infection group and free culture media group (control group). Infection group and control group were inoculated with M. pneumoniae and free pathogen liquid culture for four days, respectively. All rats were sacrificed on days 1,3,5,8,10 after modeling. BALF specimens were measured using real-time PCR and ELISA, and lung histopatholog were assessed by histopathologic score (HPS).Results: 1.46 specimens were positive for M. pneumoniae tested by PCR.69.7%,18.2%, 6.1% of BALF, sputa, swabs were M. pneumoniae DNA positive respectively tested by PCR. BALF had a higher DNA positive rate than sputa and swabs (P＜0.01). During 46 positive specimens for M. pneumoniae,31 (67.4%) specimens had a 2063A→G mutation, not other mutations found. Compared with patients having macrolide-susceptible M. pneumoniae, there were no statistical significance in number of febrile days,the number of in hospital days and the occurrence of complications and glucocorticoids use for patients having MR M. pneumoniae infection.(P value were 0.459,0.632,0.793.0.575, respectively).2. Sprague-Dawley rats in infection group had inflammatory reaction 1 day later,and it reach a peak at the 3 day with the highest HPS 16. No obvious inflammation was identified by HPS in control group. M. pneumoniae can be detected in BAL of infection rats by real-time PCR during the entire process.Thl cytokine IFN-γrise after infection and have a peak on day 3, while Th2 cytokine did not have a significant change. The concentration of IFN-γcorrelated with the histopathological score.Conclusions:This study suggests that the macrolide-resistant geng mutation occurrence of M. pneumoniae was 63.4%. All mutations were 2063A→G mutation. Microlide is effective for both M.R and M.S M. pneumoniae, which maybe benefits from its immunomodulating effects.So the detection of macrolide-resistant gene mutations is not necessary for clinical treatment. After infected with M. pneumoniae, the concentration of IFN-y, which was a proinflamation cytokine, increased. And the concentration of IFN-γcorrelated with the HPS. But the concentration of IL-4, which was a antiinflamation cytokine, did not change significantly. There has a disbalance of Thl/Th2 cytokines. Cell-Mediated Immune Reaction is critical for M. pneumoniae pathology. For the treatment of M. pneumoniae, especially refractory M. pneumoniae infection responding badly to macrolides, immune modulators may be effective.
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