Point-of-Care Amenable Detection of Mycoplasma genitalium and Its Antibiotic Resistance Mutations.
Point-of-Care Amenable Detection of Mycoplasma genitalium and Its Antibiotic Resistance Mutations.
复制标题
生殖支原体及其抗生素耐药性突变的护理点检测。
DOI：
10.1021/acssensors.2c02630
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发表时间：
2023
期刊：
影响因子：
8.9
通讯作者：
Wang,Tza-Huei
中科院分区：
文献类型：
--
作者：
Chen,Fan-En;Wang,Jonathan;Nambiar,AnjuHaridas;Hardick,Justin;Melendez,Johan;Trick,AlexanderY;Wang,Tza-Huei
生殖支原体（MG）是一种新兴的性传播细菌。由于其挑剔和生长缓慢的性质，MG很难通过基于培养的诊断来检测。与淋病奈瑟菌（Neisseria gonorrheae）一样，另一种与性传播感染（STI）有关的细菌病原体，MG对用于治疗STI的大环内酯类和氟喹诺酮类抗生素产生了耐药性。同时检测MG和鉴定与抗生素耐药性相关的基因组突变的能力可以实现抗生素管理并减轻耐药性MG的传播。为此，我们首先开发了一种基于多重探针的PCR-melt检测方法，用于检测MG和23 S rRNA基因中大环内酯类耐药突变以及parC基因中氟喹诺酮类耐药突变的存在。通过荧光标记和解链温度的独特组合鉴定每个靶标。这种方法允许在感兴趣的基因处的不同类型的突变之间进行区分。在初始测定优化之后，将测定整合到便携式平台中使用的液滴磁流体盒中，以整合自动化样品提取、PCR扩增和检测。最后，我们证明了集成的测定和液滴磁流体平台可以在40分钟内检测掺入尿液样品中的临床分离株中的MG和抗生素耐药性相关突变。
Mycoplasma genitalium(MG) is an emerging sexually transmitted bacterium. Due to its fastidious and slow-growing nature, MG is difficult to detect through culture-based diagnostics. LikeNeisseria gonorrheae, another bacterial pathogen linked to sexually transmitted infections (STIs), MG has developed resistance to macrolide and fluoroquinolone antibiotics used to treat STIs. The ability to detect MG and identify genomic mutations associated with antibiotic resistance simultaneously can enable antibiotic stewardship and mitigate the spread of antibiotic-resistant MG. Toward this end, we first developed a multiplexed probe-based PCR–melt assay that detects MG and the presence of macrolide resistance mutations in the 23S rRNA gene and fluoroquinolone resistance mutations in theparCgene. Each target was identified via its unique combination of fluorescence label and melting temperature. This approach allowed differentiation between the different types of mutations at the genes of interest. Following initial assay optimization, the assay was integrated into a droplet magnetofluidic cartridge used in a portable platform to integrate automated sample extraction, PCR amplification, and detection. Lastly, we demonstrated that the integrated assay and droplet magnetofluidic platform could detect MG and antibiotic resistance-associated mutations in clinical isolates spiked into urine samples in 40 min.