泌尿生殖道沙眼衣原體體外單獨和聯(lián)合藥物敏感性檢測
發(fā)布時間：2019-01-19 10:11
【摘要】：沙眼衣原體(Chlamydia trachomatis, Ct)感染是目前國內(nèi)外最常見的性病病原體,可引起尿道炎、宮頸炎、子宮內(nèi)膜炎、附睪炎、前列腺炎、不孕癥多種疾病,且發(fā)病率呈逐年上升趨勢,已經(jīng)成為一個危害極大的公共健康問題。目前,泌尿生殖道Ct感染的發(fā)病率與其流行率密切相關(guān),因此抗菌藥物治療是該病二級預(yù)防的重要策略；同時由于Ct疫苗至今仍不能用于臨床,因此抗菌藥物對于治療Ct感染就顯得更加重要。然而,泌尿生殖道Ct感染患者的臨床表現(xiàn)常無特異性或呈隱匿性,不易引起重視,致使病情遷延、復(fù)發(fā)或反復(fù)感染,導(dǎo)致Ct對多種抗菌藥物的敏感性降低,臨床治療失敗病例急劇增加。 目的：檢測泌尿生殖道沙眼衣原體標(biāo)準(zhǔn)株和臨床株對阿奇霉素、米諾環(huán)素、莫西沙星、多西環(huán)素和利福平五種臨床常用抗菌藥物體外單獨和聯(lián)合藥物敏感性,研究其與藥物臨床療效的相關(guān)性和耐藥的分子生物學(xué)機(jī)制。 方法：收集2005-2009年間到天津市性傳播疾病研究所門診就診且符合采樣標(biāo)準(zhǔn)的患者的男性尿道或女性宮頸上皮細(xì)胞,并記錄其臨床相關(guān)資料。采用McCoy細(xì)胞培養(yǎng)法對臨床株進(jìn)行培養(yǎng),所有臨床株進(jìn)行五代盲傳培養(yǎng),對陽性標(biāo)本繼續(xù)傳代培養(yǎng),直至感染率達(dá)到90%以上,共獲得41株臨床菌株。對標(biāo)準(zhǔn)株和41株臨床株,分別采用微量稀釋法和棋盤稀釋法進(jìn)行五種常用抗菌藥的單獨和聯(lián)合藥敏檢測。核酸擴(kuò)增臨床株的omp1基因并用AluⅠ、MspⅠ雙酶切進(jìn)行基因分型。對臨床治療失敗病例的臨床株,PCR方法擴(kuò)增檢測與大環(huán)內(nèi)酯類耐藥相關(guān)的23S核蛋白體RNA基因,通過產(chǎn)物測序檢測基因突變；限制性片段分析(RFLP)檢測gyrA喹諾酮耐藥決定區(qū)(Quinolone-Resistance Determining Region,QRDR)常見的基因點突變。統(tǒng)計學(xué)方法：用SPSS11.5軟件進(jìn)行單因素方差分析(One-Way ANOVA),在方差分析顯著的情況下,滿足方差齊性要求數(shù)據(jù)兩兩比較采用最下顯著差法即LSD (least-significant different)法進(jìn)行組間多重比較。以P0.05為差異有統(tǒng)計學(xué)意義。 結(jié)果：總共培養(yǎng)臨床標(biāo)本238例,得到41例陽性臨床株,陽性率約為17.23%。對此41株標(biāo)本進(jìn)行單獨藥敏,檢測最低抑菌濃度(MIC)(單位ug/m1)分別是：阿奇霉素0.063-0.5,莫西沙星0.03-0.24,米諾環(huán)素0.008-0.064,多西環(huán)素0.063-0.5,利福平0.002-0.016。聯(lián)合藥敏檢測部分抑菌濃度(FIC)示：阿奇霉素與莫西沙星、多西環(huán)素、利福平在體外聯(lián)合時,分別對51.22%、53.66%和58.54%的菌株為協(xié)同或相加作用,拮抗作用較少；ANOVA檢驗示,三組之間差異無統(tǒng)計學(xué)意義(當(dāng)a=0.05時,P=0.755)。米諾環(huán)素與阿奇霉素、莫西沙星、利福平在體外聯(lián)合時,分別對90.24%、85.37%、92.68%的Ct菌株為拮抗作用,無協(xié)同作用。對臨床治療失敗菌株均未檢測到相應(yīng)的耐藥基因。 結(jié)論：1、成功分離培養(yǎng)Ct臨床株,為進(jìn)一步開展該病原體的致病機(jī)理、免疫機(jī)制、體外藥物敏感性檢測、耐藥機(jī)制和保護(hù)性疫苗等方面的研究奠定基礎(chǔ)。2、Ct對常用抗菌藥的敏感性呈不同程度的下降,MIC值均較以往文獻(xiàn)報道增高。對Ct體外耐藥性與抗菌效果之間關(guān)系的研究并不明確。3、不同抗菌藥物體外聯(lián)合藥敏試驗,在一定程度上能夠彌補單獨藥敏實驗的一些不足,具有顯著的臨床意義,并且將為進(jìn)一步研究抗菌藥物聯(lián)合作用發(fā)生機(jī)制奠定實驗基礎(chǔ)。
[Abstract]:Chlamydia trachomatis (Ct) infection is the most common venereal disease at home and abroad, which can cause urethritis, cervicitis, endometritis, epididymitis, prostatitis, and infertility. has become a public health problem that has a great danger. At present, the incidence of Ct infection in the urogenital tract is closely related to the prevalence rate, so the treatment of the antibacterial drugs is an important strategy of the secondary prevention of the disease, and because the Ct vaccine can not be used for clinical use so far, the antibacterial medicine is more important for treating the Ct infection. However, the clinical manifestation of the patients with the urinary tract Ct infection is often non-specific or latent, which is not easy to cause attention, which leads to the change of the condition, the recurrence or the repeated infection, and the sensitivity of the Ct to a plurality of antibacterial drugs is reduced, and the clinical treatment failure cases are rapidly increased. Objective: To test the sensitivity of the standard strains and clinical strains of Chlamydia trachomatis in the urogenital tract to the five clinical commonly used antibacterials in the treatment of the five clinical commonly used antibacterials, such as the azoxycycline, the minocycline, the moxifloxacin, the polycidin and the rifampin. Molecular biological machine for the study of its correlation and drug-resistance with the clinical curative effect of drugs Methods: To collect the male urethra or female cervical epithelial cells of the patients who visit and meet the sampling criteria in the outpatient department of the Institute of Sexually Transmitted Diseases in Tianjin from 2005 to 2009, and to record their clinical phase The clinical strains were cultured by the McCoy cell culture method. All the clinical strains were cultured for five generations, and the positive specimens were continued to be subcultured until the infection rate was over 90%. The standard strains and 41 clinical strains were used separately and in combination with five common antimicrobial agents by using a microdilution method and a checkerboard dilution method, respectively. Detection of the omp1 gene of the clinical strain of the nucleic acid amplification and the basis of the two-enzyme digestion of Alu I and Msp I For typing, a clinical strain of a failed case of clinical treatment was used to amplify the 23S nucleoprotein RNA gene related to the drug resistance of macrolides, and the gene mutation was detected by product sequencing; restriction fragment analysis (RFLP) was used to detect the drug resistance determining region (Quinn-Resistance Detection Reg.) of the gyrA Common genes of ion (QRDR) Point mutation. Statistical method: The single-factor analysis of variance (One-Way ANOVA) was performed with the SPSS11.5 software. In the case of the variance analysis, the most significant difference method (LSD) method was used to compare the two-to-two comparisons. Re-comparison. The difference of the difference is P0. 05. Results: In total, 238 cases of clinical specimens were cultured, and 41 positive clinical strains were obtained. The positive rate was about 1. The minimum inhibitory concentration (MIC) (unit ug/ ml) of this 41 specimens were respectively: Aichromycin 0. 063-0.5, moxifloxacin 0.03-0.24, minocycline 0. 008-0. 064, polycidin 0. 063-0.5, rifampin 0. 002-0. 016. The inhibitory concentration (FIC) of the combined drug sensitivity test part showed that, in combination with Moxifloxacin, Moxifloxacin, and rifampin in vitro, the antagonistic effect was less than that of the strains of 51. 22%, 53. 66% and 58. 54%, respectively. The VA test showed no statistical significance between the three groups (P = 0 when a = 0.05). No synergistic effect. No corresponding clinical treatment failure strains were detected Conclusion: 1. The successful isolation and culture of the Ct clinical isolates will provide the basis for further research on the pathogenesis, immune mechanism, in vitro drug sensitivity detection, drug resistance mechanism and protective vaccine of the pathogen. The MIC values were lower for different levels of decline The study of the relationship between the in vitro drug resistance and the antibacterial effect of Ct is not clear. Significant clinical significance and will be the case for further study of the combination of antibacterial drugs
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R759
本文編號：2411271
[Abstract]:Chlamydia trachomatis (Ct) infection is the most common venereal disease at home and abroad, which can cause urethritis, cervicitis, endometritis, epididymitis, prostatitis, and infertility. has become a public health problem that has a great danger. At present, the incidence of Ct infection in the urogenital tract is closely related to the prevalence rate, so the treatment of the antibacterial drugs is an important strategy of the secondary prevention of the disease, and because the Ct vaccine can not be used for clinical use so far, the antibacterial medicine is more important for treating the Ct infection. However, the clinical manifestation of the patients with the urinary tract Ct infection is often non-specific or latent, which is not easy to cause attention, which leads to the change of the condition, the recurrence or the repeated infection, and the sensitivity of the Ct to a plurality of antibacterial drugs is reduced, and the clinical treatment failure cases are rapidly increased. Objective: To test the sensitivity of the standard strains and clinical strains of Chlamydia trachomatis in the urogenital tract to the five clinical commonly used antibacterials in the treatment of the five clinical commonly used antibacterials, such as the azoxycycline, the minocycline, the moxifloxacin, the polycidin and the rifampin. Molecular biological machine for the study of its correlation and drug-resistance with the clinical curative effect of drugs Methods: To collect the male urethra or female cervical epithelial cells of the patients who visit and meet the sampling criteria in the outpatient department of the Institute of Sexually Transmitted Diseases in Tianjin from 2005 to 2009, and to record their clinical phase The clinical strains were cultured by the McCoy cell culture method. All the clinical strains were cultured for five generations, and the positive specimens were continued to be subcultured until the infection rate was over 90%. The standard strains and 41 clinical strains were used separately and in combination with five common antimicrobial agents by using a microdilution method and a checkerboard dilution method, respectively. Detection of the omp1 gene of the clinical strain of the nucleic acid amplification and the basis of the two-enzyme digestion of Alu I and Msp I For typing, a clinical strain of a failed case of clinical treatment was used to amplify the 23S nucleoprotein RNA gene related to the drug resistance of macrolides, and the gene mutation was detected by product sequencing; restriction fragment analysis (RFLP) was used to detect the drug resistance determining region (Quinn-Resistance Detection Reg.) of the gyrA Common genes of ion (QRDR) Point mutation. Statistical method: The single-factor analysis of variance (One-Way ANOVA) was performed with the SPSS11.5 software. In the case of the variance analysis, the most significant difference method (LSD) method was used to compare the two-to-two comparisons. Re-comparison. The difference of the difference is P0. 05. Results: In total, 238 cases of clinical specimens were cultured, and 41 positive clinical strains were obtained. The positive rate was about 1. The minimum inhibitory concentration (MIC) (unit ug/ ml) of this 41 specimens were respectively: Aichromycin 0. 063-0.5, moxifloxacin 0.03-0.24, minocycline 0. 008-0. 064, polycidin 0. 063-0.5, rifampin 0. 002-0. 016. The inhibitory concentration (FIC) of the combined drug sensitivity test part showed that, in combination with Moxifloxacin, Moxifloxacin, and rifampin in vitro, the antagonistic effect was less than that of the strains of 51. 22%, 53. 66% and 58. 54%, respectively. The VA test showed no statistical significance between the three groups (P = 0 when a = 0.05). No synergistic effect. No corresponding clinical treatment failure strains were detected Conclusion: 1. The successful isolation and culture of the Ct clinical isolates will provide the basis for further research on the pathogenesis, immune mechanism, in vitro drug sensitivity detection, drug resistance mechanism and protective vaccine of the pathogen. The MIC values were lower for different levels of decline The study of the relationship between the in vitro drug resistance and the antibacterial effect of Ct is not clear. Significant clinical significance and will be the case for further study of the combination of antibacterial drugs
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R759
【引證文獻(xiàn)】
相關(guān)碩士學(xué)位論文 前1條
1 尤聰;沙眼衣原體培養(yǎng)陽性率提高及臨床株熱休克蛋白60的轉(zhuǎn)錄隨傳代的變化[D];天津醫(yī)科大學(xué);2012年
，本文編號：2411271
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