Bhirud Prasanna, Joshi Ameeta, Hirani Nilma, Chowdhary Abhay
Department of Microbiology, Grant Government Medical College and Sir JJ Group of Hospital, Mumbai, Maharashtra, India.
Int J Mycobacteriol. 2017 Jul-Sep;6(3):296-301. doi: 10.4103/ijmy.ijmy_61_17.
Tuberculosis (TB) ranks as the second leading cause of death from an infectious disease worldwide. Early diagnosis of Mycobacterium tuberculosis in clinical samples becomes important in the control of TB both for the treatment of patients and for curbing of disease transmission to the others in the community. The study objective was to perform Ziehl-Neelsen (ZN) staining, fluorochrome staining, line probe assay (LPA), and loop-mediated isothermal amplification (LAMP) assay for rapid detection of pulmonary TB (PTB) and to compare the results of LPA and LAMP in terms of sensitivity, specificity, and turnaround time.
A total of 891 sputum samples from clinically diagnosed/suspected cases of TB were subjected to ZN and fluorochrome staining. Smear positive samples were subjected to LPA, and smear negative were cultured on Lowenstein-Jensen media. A total of 177 samples were subjected to liquid culture and LAMP. Conventional culture was considered as "gold standard" for calculation of parameters.
Light-emitting diode fluorescence microscopy had the same sensitivity as ZN with similar high specificity. LPA was performed on 548 sputum samples which includes 520 smear positive and 28 smear negative culture positive samples and multidrug-resistant TB was detected in 32.64%. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of TB-LAMP on direct sputum samples was found to be 98.96%, 95%, 96%, and 98.70%, respectively, when compared with ZN smear microscopy. By considering culture as "gold standard," LAMP showed a sensitivity, specificity, PPV, and NPV of 98.94%, 96.34%, 96.90%, and 98.75%, respectively. The sensitivity and PPV of TB-LAMP were 98.97% and 96%, respectively, when compared with LPA.
A successful rapid laboratory diagnosis of PTB is possible when one combines the available methodology of microscopy, culture as well as molecular techniques. The LAMP assay was found to be simple, self-contained, and efficacious for early diagnosis of suspected cases of PTB with advantages of having a high throughput, no requirements of sophisticated equipment, and complex biosafety facilities.
结核病是全球第二大传染病致死病因。对临床样本中的结核分枝杆菌进行早期诊断，对于患者治疗以及控制疾病在社区内传播都至关重要。本研究目的是进行萋-尼（ZN）染色、荧光染色、线性探针分析（LPA）和环介导等温扩增（LAMP）检测，以快速检测肺结核（PTB），并比较LPA和LAMP在敏感性、特异性和周转时间方面的结果。
对891份来自临床诊断/疑似结核病病例的痰液样本进行ZN和荧光染色。涂片阳性样本进行LPA检测，涂片阴性样本接种于罗-琴培养基上培养。177份样本进行液体培养和LAMP检测。计算参数时，将传统培养视为“金标准”。
发光二极管荧光显微镜的敏感性与ZN染色相同，特异性也较高。对548份痰液样本进行了LPA检测，其中包括520份涂片阳性和28份涂片阴性培养阳性样本，检测出耐多药结核病的比例为32.64%。与ZN涂片显微镜检查相比，直接痰液样本中结核LAMP检测的敏感性、特异性、阳性预测值（PPV）和阴性预测值（NPV）分别为98.96%、95%、96%和98.70%。以培养为“金标准”时，LAMP检测的敏感性、特异性、PPV和NPV分别为98.94%、96.34%、96.90%和98.75%。与LPA相比，结核LAMP检测的敏感性和PPV分别为98.97%和96%。
将显微镜检查、培养以及分子技术等现有方法结合起来，有可能实现对PTB的快速实验室诊断。发现LAMP检测对于PTB疑似病例的早期诊断简单、自成体系且有效，具有高通量、无需复杂设备和生物安全设施的优点。
