Transmission and selection of macrolide resistant Mycoplasma genitalium infections detected by rapid high resolution melt analysis.
Transmission and selection of macrolide resistant Mycoplasma genitalium infections detected by rapid high resolution melt analysis.
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DOI：
10.1371/journal.pone.0035593
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Tabrizi SN
中科院分区：
文献类型：
--
作者：
Twin J;Jensen JS;Bradshaw CS;Garland SM;Fairley CK;Min LY;Tabrizi SN
生殖道支原体（MG）引起尿道炎、宫颈炎和盆腔炎。2007 - 2009年，澳大利亚性健康诊所使用1g阿奇霉素治疗MG的失败率为31%（95%可信区间23-40%）。我们开发了一种针对MG 23S rRNA基因耐药突变的快速高分辨率熔体分析（HRMA）方法，并通过检测MG感染患者治疗前和治疗后的存档样本，对其DNA测序进行了验证。筛选来自临床治疗失败个体（n = 20）的mg阳性治疗前（n = 82）和治疗后样本（n = 20）的23S rRNA基因突变。16份（20%）治疗前样品存在耐药突变（A2058G、A2059G、A2059C），在症状性阿奇霉素治疗失败患者中（12/26;44%）明显高于临床治愈患者（4/56;7%），p<0.001。所有经历阿奇霉素失效的20例患者在治疗后的样本中都有可检测到的突变。在其中9例病例中，处理前和处理后样本中均存在相同的突变类型，表明传播耐药性，而在其中11例病例（55%）中，处理前样本中没有突变，表明可能发生了耐药分离株的选择。通过DNA测序，HRMA能够检测到本研究中确定的所有突变变化。还开发了一种附加的HRMA检测方法，其中包含未标记探针，用于检测在其他人群中发现的4型单核苷酸多态性，灵敏度略低，为90%。治疗失败与检测到大环内酯类耐药突变有关，这似乎几乎同样是由于暴露于1克阿奇霉素和先前存在的传播性耐药后选择耐药分离株所致。应用快速分子测定法在最初发现感染时检测耐药性，使临床医生能够缩短开始有效的二线治疗的时间。这有可能减少耐药菌株的传播，并避免与持续未经治疗的感染相关的后遗症。
Mycoplasma genitalium (MG) causes urethritis, cervicitis and pelvic inflammatory disease. The MG treatment failure rate using 1 g azithromycin at an Australian Sexual Health clinic in 2007–9 was 31% (95%CI 23–40%). We developed a rapid high resolution melt analysis (HRMA) assay targeting resistance mutations in the MG 23S rRNA gene, and validated it against DNA sequencing by examining pre- and post-treatment archived samples from MG-infected patients. Available MG-positive pre-treatment (n = 82) and post-treatment samples from individuals with clinical treatment failure (n = 20) were screened for 23S rRNA gene mutations. Sixteen (20%) pre-treatment samples possessed resistance mutations (A2058G, A2059G, A2059C), which were significantly more common in patients with symptomatic azithromycin-treatment failure (12/26; 44%) than in those clinically cured (4/56; 7%), p<0.001. All 20 patients experiencing azithromycin-failure had detectable mutations in their post-treatment samples. In 9 of these cases, the same mutational types were present in both pre- and post-treatment samples indicating transmitted resistance, whilst in 11 of these cases (55%), mutations were absent in pre-treatment samples indicating likely selection of resistant isolates have occurred. HRMA was able to detect all mutational changes determined in this study by DNA sequencing. An additional HRMA assay incorporating an unlabelled probe was also developed to detect type 4 single-nucleotide polymorphisms found in other populations, with a slightly lower sensitivity of 90%. Treatment failure is associated with the detection of macrolide resistance mutations, which appear to be almost equally due to selection of resistant isolates following exposure to 1 g azithromycin and pre-existing transmitted resistance. The application of a rapid molecular assay to detect resistance at the time of initial detection of infection allows clinicians to shorten the time to initiate effective second line treatment. This has the potential to reduce transmission of resistant strains and to avoid sequelae associated with persistent untreated infection.
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