A modified decontamination and storage method for sputum from patients with tuberculosis.

Q1 Medicine Wellcome Open Research Pub Date : 2025-02-28 eCollection Date: 2023-01-01 DOI:10.12688/wellcomeopenres.18888.2

Nguyen Le Quang, Do Dang Anh Thu, Le Pham Tien Trieu, Nguyen Hong Hanh, Nguyen Huu Lan, Dang Thi Minh Ha, Guy Thwaites, Nguyen Thuy Thuong Thuong, Timothy M Walker

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Abstract

Background: Collecting and storing large number of sputum samples with a view to culturing these in the future requires an efficient initial handling method. We devised a modified sputum digestion and decontamination method that maximised storage capacity and Mycobacterium tuberculosis (M.tb) recovery from culture while minimising laboratory workload and risk of contamination.

Methods: We collected smear microscopy positive sputum samples from patients with pulmonary tuberculosis (TB). The sputum samples were split and processed using both the standard N-Acetyl-L-cysteine and sodium hydroxide (NALC-NaOH) method and our modified method before freezing and later culturing in BD BACTEC 960 Mycobacterium Growth Indicator Tubes (MGIT) system. We assessed the Time to Positivity (TPP) and Growth Unit (GU) data.

Results: We selected 22 sputum samples to compare two digestion and decontamination methods. The samples that underwent the modified method had longer TTP (p < 0.05) but similar GU in comparison to standard method. Overall, 1/22 samples failed to grow in MGIT after being processed by the modified method. We then applied the modified method to 348 sputum samples with Rifampicin resistance detected by GeneXpert MTB/RIF assay, which were frozen for between 1-25 months. The overall MGIT positive, negative, and contamination rate was 90.5%, 7.8%, and 1.7%, respectively. There was no significant difference in MGIT result when samples were grouped by duration of storage or positive smear grade.

Conclusions: Our modified method yielded acceptable M.tb recovery rate and low contamination risk while allowing us to collect and store thousands of sputum samples over a long period of time for future tests.

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肺结核患者痰液的改良净化和储存方法。

背景：收集和储存大量痰液样本以便将来进行培养，需要一种高效的初始处理方法。我们设计了一种改良的痰液消化和净化方法，既能最大限度地提高储存能力和结核分枝杆菌（M.tb）培养回收率，又能最大限度地减少实验室工作量和污染风险：我们收集了肺结核（TB）患者涂片镜检阳性的痰液样本。方法：我们采集了肺结核（TB）患者涂片镜检阳性的痰液样本，采用标准的 N-乙酰-L-半胱氨酸和氢氧化钠（NALC-NaOH）法和我们改进的方法对痰液样本进行分割和处理，然后冷冻并在 BD BACTEC 960 分枝杆菌生长指示管（MGIT）系统中进行培养。我们评估了阳性时间（TPP）和生长单位（GU）数据：我们选择了 22 份痰样本，对两种消化和净化方法进行了比较。与标准方法相比，采用改良方法的样本的阳性时间更长（p < 0.05），但生长单位相似。总体而言，1/22 的样本在使用改良方法处理后未能在 MGIT 中生长。然后，我们将改良方法应用于经 GeneXpert MTB/RIF 检测出对利福平耐药的 348 份痰样本，这些样本被冷冻了 1-25 个月。总体 MGIT 阳性率、阴性率和污染率分别为 90.5%、7.8% 和 1.7%。按储存时间或阳性涂片等级对样本进行分组后，MGIT 结果无明显差异：我们改进后的方法可获得可接受的结核杆菌回收率和较低的污染风险，同时允许我们长期收集和储存数千份痰标本，以备将来检测之用。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Wellcome Open Research Biochemistry, Genetics and Molecular Biology-Biochemistry, Genetics and Molecular Biology (all)

CiteScore

5.50

自引率

0.00%

发文量

426

审稿时长

1 weeks

期刊介绍： Wellcome Open Research publishes scholarly articles reporting any basic scientific, translational and clinical research that has been funded (or co-funded) by Wellcome. Each publication must have at least one author who has been, or still is, a recipient of a Wellcome grant. Articles must be original (not duplications). All research, including clinical trials, systematic reviews, software tools, method articles, and many others, is welcome and will be published irrespective of the perceived level of interest or novelty; confirmatory and negative results, as well as null studies are all suitable. See the full list of article types here. All articles are published using a fully transparent, author-driven model: the authors are solely responsible for the content of their article. Invited peer review takes place openly after publication, and the authors play a crucial role in ensuring that the article is peer-reviewed by independent experts in a timely manner. Articles that pass peer review will be indexed in PubMed and elsewhere. Wellcome Open Research is an Open Research platform: all articles are published open access; the publishing and peer-review processes are fully transparent; and authors are asked to include detailed descriptions of methods and to provide full and easy access to source data underlying the results to improve reproducibility.