Pavankumar Asalapuram R, Engström Anna, Liu Jie, Herthnek David, Nilsson Mats
Science for Life Laboratory, Department of Biochemistry and Biophysics, Stockholm University , Stockholm, Sweden.
Anal Chem. 2016 Apr 19;88(8):4277-84. doi: 10.1021/acs.analchem.5b04312. Epub 2016 Apr 4.
Tuberculosis is a major communicable disease. Its causative agent, Mycobacterium tuberculosis, becomes resistant to antibiotics by acquisition of point mutations in the chromosome. Multi-drug-resistant tuberculosis (MDR-TB) is an increasing public health threat, and prompt detection of such strains is of critical importance. As rolling circle amplification of padlock probes can be used to robustly distinguish single-nucleotide variants, we combined this technique with a sensitive lateral flow nucleic acid biosensor to develop a rapid molecular diagnostic test for MDR-TB. A proof-of-concept test was established for detection of the most common mutations [rpoB 531 (TCG/TTG) and katG 315 (AGC/ACC)] causing MDR-TB and verification of loss of the respective wild type. The molecular diagnostic test produces visual signals corresponding to the respective genotypes on lateral flow strips in approximately 75 min. By detecting only two mutations, the test can detect about 60% of all MDR-TB cases. The padlock probe-lateral flow (PLP-LF) test is the first of its kind and can ideally be performed at resource-limited clinical laboratories. Rapid information about the drug-susceptibility pattern can assist clinicians to choose suitable treatment regimens and take appropriate infection control actions rather than prescribing empirical treatment, thereby helping to control the spread of MDR-TB in the community.
结核病是一种主要的传染病。其病原体结核分枝杆菌通过在染色体上获得点突变而对抗生素产生耐药性。耐多药结核病（MDR-TB）对公共卫生的威胁日益增加，因此及时检测出此类菌株至关重要。由于锁式探针的滚环扩增可用于可靠地区分单核苷酸变体，我们将该技术与灵敏的侧向流动核酸生物传感器相结合，开发了一种用于耐多药结核病的快速分子诊断测试。建立了一个概念验证测试，用于检测导致耐多药结核病的最常见突变[rpoB 531（TCG/TTG）和katG 315（AGC/ACC）]，并验证相应野生型的缺失情况。该分子诊断测试在约75分钟内在侧向流动条上产生与各自基因型相对应的视觉信号。通过仅检测两个突变，该测试可检测约60%的所有耐多药结核病例。锁式探针-侧向流动（PLP-LF）测试尚属首次，非常适合在资源有限的临床实验室进行。关于药物敏感性模式的快速信息可帮助临床医生选择合适的治疗方案并采取适当的感染控制措施，而不是进行经验性治疗，从而有助于控制耐多药结核病在社区中的传播。
