Evaluation of sputum decontamination methods for Mycobacterium tuberculosis using viable colony counts and flow cytometry
Evaluation of sputum decontamination methods for Mycobacterium tuberculosis using viable colony counts and flow cytometry
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DOI：
10.1016/s0732-8893(03)00138-x
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发表时间：
2003-11-01
影响因子：
2.9
通讯作者：
Kabani, A
中科院分区：
文献类型：
--
作者：
Burdz, TVN;Wolfe, J;Kabani, A
据报告，用于检测结核分枝杆菌的连续监测系统的污染率高于传统的辐射测量技术。最近报道了多种去污方法，试图优化这些系统的污染率。在这项研究中，使用活菌落计数和流式细胞术评价了几种痰液净化方法。评价的去污方案包括N-乙酰基-L-半胱氨酸-氢氧化钠（NALC-NaOH），改进的Petroffs方法。和Yamane手术分析了NALC-NaOH方法的几个参数，包括0.5- 3%的最终NaOH浓度、0-30分钟的NaOH暴露时间以及所得沉淀物的重悬介质的变化。所有的净化方法都是在合并的无菌痰液上进行的，这些痰液分别接种了常见污染菌或M。结核病H37 Ra。通过菌落计数和流式细胞术分析评估去污后微生物的活力。流式细胞术活力测定利用活力染料和参考珠的组合来确定活生物体浓度。结果表明，无去污方法显然是上级的，然而，浓度为1-2%的NaOH和NaOH暴露时间增加到30分钟将有效地杀死污染细菌，而不显著影响M.结核病H37 Ra。虽然流式细胞术活力分析不直接对应于活菌落计数，但它是用于快速活力分析M的有用工具。结核(C)2003年爱思唯尔公司All rights reserved.
Continuous monitoring systems for the detection of Mycobacterium tuberculosis are reported to have higher contamination rates than traditional radiometric technologies. Multiple decontamination methods have recently been reported in an attempt to optimize contamination rates for these systems. In this study, several decontamination methods for sputum were evaluated using viable colony count and flow cytometry. The decontamination protocols evaluated include N-Acetyl-L-Cysteine-Sodium Hydroxide (NALC-NaOH), modified Petroffs's method. and the Yamane procedure. Several parameters of the NALC-NaOH method were analyzed including final NaOH concentrations of 0.5-3%, NaOH exposure times of 0-30 min, and variations in resuspension media for the resultant pellet. All decontamination methods were performed on pooled and sterilized sputum seeded separately with either a mixture of common contaminating bacteria or M. tuberculosis H37Ra. Viability of organisms following decontamination was assessed by both colony counts and flow cytometric analysis. Flow cytometry viability assays utilized a combination of viability dyes and reference beads to determine viable organism concentrations. The results indicated that no decontamination method was clearly superior, however a concentration of 1-2% NaOH and an increase in the time of NaOH exposure to 30 min will effectively kill contaminating bacteria without significantly affecting the viability of M. tuberculosis H37Ra. While flow cytometry viability analysis did not directly correspond to viable colony counts, it was a useful tool for rapid viability analysis M. tuberculosis. (C) 2003 Elsevier Inc. All rights reserved.