本文關(guān)鍵詞： 肺炎支原體 耐藥 亞急性咳嗽　出處：《中國(guó)人民解放軍軍醫(yī)進(jìn)修學(xué)院》2012年博士論文　論文類型：學(xué)位論文

【摘要】：【目的】深入研究我國(guó)肺炎支原體（Mp）對(duì)大環(huán)內(nèi)酯類的耐藥機(jī)制和預(yù)測(cè)Mp對(duì)四環(huán)素類將可能產(chǎn)生的耐藥機(jī)制；調(diào)研亞急性咳嗽與Mp感染的關(guān)系，以助于分析亞急性咳嗽病因?qū)W。 【方法】（1）收集患者咽拭子標(biāo)本，采用實(shí)時(shí)定量PCR法和液-固-液交替培養(yǎng)獲取純化的Mp菌株。（2）采用96孔板肉湯微量稀釋法，測(cè)定大環(huán)內(nèi)酯類（紅霉素、阿奇霉素、麥迪霉素）、喹諾酮類（氧氟沙星、左氧氟沙星、莫西沙星）、四環(huán)素類（四環(huán)素、米諾環(huán)素）和氨基糖苷類（阿米卡星、硫酸依替米星）10種抗生素的MIC值。（3）采用巢氏PCR和多個(gè)引物一步PCR法對(duì)耐藥Mp進(jìn)行基因分型，并測(cè)定p1基因RepMP4和RepMP2/3序列，進(jìn)行同源性分析。（4）對(duì)大環(huán)內(nèi)酯類耐藥株進(jìn)行23SrRNA和5SrRNA基因測(cè)序，，觀察外排泵抑制劑（利血平）對(duì)MIC的影響，篩查mef、msr、mph(C)等基因，進(jìn)行靶位改變、外排泵和滅活酶耐藥機(jī)制的研究。（5）臨床篩查和體外藥物誘導(dǎo)獲取耐四環(huán)素類Mp，進(jìn)行16SrRNA基因測(cè)序、篩查tet(M)基因和外排泵。（6）收集2010年7月至2011年6月軍事醫(yī)學(xué)科學(xué)院附屬醫(yī)院亞急性咳嗽患者和健康受試者咽拭子標(biāo)本進(jìn)行Mp檢測(cè)，同時(shí)進(jìn)行Mp分型、MIC測(cè)定和療效觀察，分析Mp在亞急性咳嗽中的地位和作用。 【結(jié)果（】1）共獲得72株Mp。（2）MIC結(jié)果顯示Mp大環(huán)內(nèi)酯類耐藥率為41.7%（30/72）；未發(fā)現(xiàn)喹諾酮類耐藥株；發(fā)現(xiàn)一株Mp對(duì)四環(huán)素類耐藥；氨基糖苷類對(duì)Mp抑制作用微乎其微。（3）30株耐藥株p1基因分型29株I型，1株IIa型；I型耐藥菌株間存在明顯同源性。（4）30株耐大環(huán)內(nèi)酯Mp都存在23SrRNA2063位點(diǎn)突變，29株由A→G,1株為雜合子突變。一株耐藥Mp中存在外排泵msrA/B基因，該基因與屎腸球菌TX2465獲得的耐大環(huán)內(nèi)酯類基因具有高度相似性，該基因編碼的蛋白具有P-loop＿NTPase超家族相似的保守序列。未發(fā)現(xiàn)滅活酶mph(C)基因。（5）四環(huán)素誘導(dǎo)耐藥株存在16SrRNA A1173T點(diǎn)突變，所有耐藥株均未發(fā)現(xiàn)tet(M)基因，外排泵抑制劑（利血平）對(duì)耐藥株MIC無(wú)影響。(6)亞急性咳嗽組入選83例患者，對(duì)照組80例。兩組Mp檢出率分別為55.4%和5%（P0.001）�；颊呓M46株Mp基因分型I型38株（82.6%），II型6株(13.0%)，IIa型2株（4.3%）；MIC值示23株（50%）對(duì)大環(huán)內(nèi)酯類耐藥。Mp檢測(cè)率與性別、年齡無(wú)相關(guān)，與季節(jié)有關(guān)（P=0.004），冬秋季節(jié)更易檢測(cè)到Mp。觀察12例耐藥和18例非耐藥Mp感染患者，服用阿奇霉素后咳嗽緩解天數(shù)無(wú)明顯差別(P=0.671)。 【結(jié)論】我國(guó)Mp對(duì)大環(huán)內(nèi)酯類耐藥存在靶點(diǎn)突變和外排泵兩種機(jī)制；四環(huán)素類可體外誘導(dǎo)產(chǎn)生Mp耐藥株，耐藥機(jī)制可能為16SrRNA靶點(diǎn)突變所致； Mp感染很可能在亞急性咳嗽中發(fā)揮了重要作用。
[Abstract]:[objective] to study the drug resistance mechanism of mycoplasma pneumoniae (MP) to macrolides and to predict the possible drug resistance of MP to tetracycline in China, and to investigate the relationship between subacute cough and MP infection in order to help to analyze the etiology of subacute cough. [methods] the throat swabs of patients were collected and purified strains of MP were obtained by real-time quantitative PCR and liquid-solid-liquid alternate culture. The microdilution method of 96 hole broth was used to determine macrolides (erythromycin, azithromycin). Medelomycin, quinolones (ofloxacin, levofloxacin, moxifloxacin, tetracycline (tetracycline, minocycline) and aminoglycoside (amikacin), The MIC value of Etimixine Sulfate (10 antibiotics) was determined by nested PCR and one-step PCR with multiple primers, and the RepMP4 and RepMP2/3 sequences of p1 gene were determined. The 23s rRNA and 5s rRNA genes of macrolide resistant strains were sequenced. The effects of drain pump inhibitor (reserpine) on MIC were observed. Studies on the mechanisms of efflux pump and inactivated enzyme resistance. 5) Clinical screening and drug induction in vitro to obtain tetracycline resistant MP and sequence 16s rRNA gene. From July 2010 to June 2011, the samples of subacute cough patients and healthy subjects were collected from subacute cough patients and healthy subjects to detect MP, and the mics of MP typing and curative effect were observed. To analyze the role of MP in subacute cough. [results] A total of 72 strains of Mp.(2)MIC were obtained. The results showed that the resistance rate of MP macrolides was 41.775%, no quinolone resistant strain was found, one strain was found to be tetracycline resistant. The inhibitory effect of aminoglycosides on MP was minimal. 30 strains of drug resistant strains p1 genotyping 29 strains of type I and 1 strain of IIa type I showed obvious homology. 30 strains of macrolide resistant MP all had 23s rRNA2063 mutation and 29 strains were isolated from A. 鈫扥ne strain was heterozygous. There was an efflux pump msrA/B gene in one resistant strain, which was highly similar to the macrolide resistant gene obtained from Enterococcus faecium TX2465. The protein encoded by this gene has a conserved sequence similar to that of P-loop-NTPase superfamily. No inactivated enzyme mphCase gene, mphC5) has been found in tetracycline induced drug resistance strains. There is a 16s rRNA A1173T point mutation in all resistant strains. Extracorporeal pump inhibitor (reserpine) had no effect on drug-resistant strain MIC. In the control group, the detection rate of MP was 55.4% and 5p 0.001, respectively. There were 46 strains of MP genotyping I (38 strains) in the patient group. There was no correlation between the detection rate and sex, age, and the MIC value of 23 strains of macrolide resistance to macrolide. It was easier to detect MP in winter and autumn than that in winter and autumn. 12 cases of drug resistance and 18 cases of non-drug-resistant MP infection were observed. There was no significant difference in cough remission days after taking azithromycin (Azithromycin). [conclusion] there are two mechanisms for MP resistance to macrolides in China, such as target mutation and efflux pump, and tetracycline can induce Mp-resistant strains in vitro. The mechanism of drug resistance may be caused by 16s rRNA target mutation, and MP infection may play an important role in subacute cough.
【學(xué)位授予單位】：中國(guó)人民解放軍軍醫(yī)進(jìn)修學(xué)院
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2012
【分類號(hào)】：R375

【引證文獻(xiàn)】

相關(guān)期刊論文 前1條

1 潘芬;張泓;;肺炎支原體耐藥性及分子流行病學(xué)研究進(jìn)展[J];上海交通大學(xué)學(xué)報(bào)(醫(yī)學(xué)版);2014年08期

本文編號(hào)：1533925