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评估痰液处理的微生物学变异及分枝杆菌浓度,以优化结核病的显微镜检查和影像学诊断。

Evaluation of microbiological variants of sputum processing and concentration of mycobacteria to optimize the microscopic and imaging diagnosis of tuberculosis.

作者信息

Coronel Jorge E, Del Carpio Christian C, Dianderas Erwin J, Florentini Edgar A, Kemper Guillermo L, Sheen Patricia, Zimic Mirko J

机构信息

Laboratorio de Bioinformática Y Biología Molecular, Faculty of Science, Universidad Peruana Cayetano Heredia, Lima, Perú.

INICTEL Institute, Universidad Nacional De Ingeniería, Lima, Perú.

出版信息

Int J Mycobacteriol. 2019 Jan-Mar;8(1):75-82. doi: 10.4103/ijmy.ijmy_172_18.

Abstract

BACKGROUND

Direct sputum smear is still the first-choice tool for screening of tuberculosis worldwide. Variants of this technique, to improve the sensitivity are desired.

METHODS

Two microbiological variants of the standard sputum smear ("pellet" and "diluted-pellet") for both Ziehl-Neelsen (ZN) and auramine fluorescence (AF) staining were evaluated. In addition, two methods for concentration of mycobacteria in sputum, using positive and negative pressure filtration, were tested and compared. The evaluation of the microbiological variants was performed on 98 culture positive sputum samples from different TB patients. The diagnostics sensitivity and the level of detritus in the processed sputum were determined. Bacilli load in the smear variants was determined by microscopic observation and by manual inspection of microscopic digital images. The comparison of the mycobacteria filtration methods was performed on 76 smear positive sputum samples. Filters retaining the concentrated mycobacteria were stained with AF and compared with the direct smear. Bacilli load, detritus level, filtered volume, filtration time and background noise level, were determined.

RESULTS

The sensitivity of microscopy with the microbiological variants was 7.1% and 2% higher in ZN and AF respectively, compared to direct smear. The sensitivity of AF in diluted pellet was significantly higher than all ZN variants (P < 0.05). Detritus level observed in slides was significantly lower in the diluted pellet than the pellet and direct smear in ZN and AF (P < 0.001). A significant increase in the bacilli load in microscopic observation and digital images analysis was observed in pellet and diluted pellet than the direct method (P <0.0001). The concentration of mycobacteria using positive-pressure filtration showed a trend to produce a higher bacilli load compared to the negative-pressure filtration and direct smear, although it was not significant. Detritus levels were significantly higher in both variants of filtration (P < 0.0001). Filtered volumes were higher in positive-pressure compared to negative-pressure filtration. Filtration times were significantly higher in negative-pressure compared to positive-pressure filtration (P < 0.0001).

CONCLUSION

The proposed variants improved the performance of the standard sputum smear, making it an important test for settings with high rates of smear-negative TB cases.

摘要

背景

直接痰涂片仍然是全球结核病筛查的首选工具。人们期望改进该技术的变体以提高其敏感性。

方法

对用于萋 - 尼(ZN)染色和金胺荧光(AF)染色的标准痰涂片的两种微生物学变体(“沉淀法”和“稀释沉淀法”)进行了评估。此外,测试并比较了两种使用正压和负压过滤的痰液中分枝杆菌浓缩方法。对来自不同结核病患者的98份培养阳性痰标本进行了微生物学变体评估。测定了诊断敏感性和处理后痰液中的碎屑水平。通过显微镜观察和显微镜数字图像人工检查确定涂片变体中的杆菌载量。对76份涂片阳性痰标本进行了分枝杆菌过滤方法的比较。保留浓缩分枝杆菌的滤器用AF染色并与直接涂片进行比较。测定了杆菌载量、碎屑水平、过滤体积、过滤时间和背景噪声水平。

结果

与直接涂片相比,微生物学变体显微镜检查的敏感性在ZN染色和AF染色中分别提高了7.1%和2%。稀释沉淀法中AF的敏感性显著高于所有ZN变体(P < 0.05)。在ZN染色和AF染色中,稀释沉淀法玻片上观察到的碎屑水平显著低于沉淀法和直接涂片(P < 0.001)。与直接方法相比,沉淀法和稀释沉淀法在显微镜观察和数字图像分析中杆菌载量显著增加(P <0.0001)。与负压过滤和直接涂片相比,使用正压过滤浓缩分枝杆菌显示出产生更高杆菌载量的趋势,尽管不显著。两种过滤变体中的碎屑水平均显著更高(P < 0.0001)。正压过滤的过滤体积高于负压过滤。负压过滤的过滤时间显著高于正压过滤(P < 0.0001)。

结论

所提出的变体改进了标准痰涂片的性能,使其成为涂片阴性结核病病例高发地区的一项重要检测方法。

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