OBJECTIVES We have developed a portable system for the rapid determination of bacterial composition for the diagnosis of infectious diseases. Our system comprises of a nanopore technology-based sequencer, MinION, and two laptop computers. To examine the accuracy and time efficiency of our system, we provided a proof-of-concept for the detection of the causative bacteria of 11 meningitis patients in Zambia. METHODS We extracted DNA from cerebrospinal fluid samples of each patient and amplified the 16S rRNA gene regions. The sequencing library was prepared, and the sequenced reads were simultaneously processed for bacterial composition determination using the minimap2 software and the representative prokaryote genomes. RESULTS The sequencing results of four of the six culture-positive samples were consistent with those of conventional culture-based methods. The dominant bacterial species in each of these samples were identified from the sequencing data within only 3 min. Although the major bacterial species were also detected from the other two culture-positive samples and five culture-negative samples, their presence could not be confirmed. Moreover, as a whole, although the number of sequencing reads obtained within a short sequencing run was small, there was no change in the major bacterial species over time with prolonged sequencing. In addition, the processing time strongly correlated with the number of sequencing reads used for the analysis. CONCLUSION Our results suggest that time-effective analysis could be achieved by determining the number of sequencing reads required for the rapid diagnosis of infectious bacterial species depending on the complexity of bacterial species in a sample.

中文翻译：

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基于测序的赞比亚脑膜炎患者传染性细菌种类的快速诊断。

目标 我们开发了一种便携式系统，用于快速测定细菌成分以诊断传染病。我们的系统由基于纳米孔技术的测序仪 MinION 和两台笔记本电脑组成。为了检验我们系统的准确性和时间效率，我们为赞比亚 11 名脑膜炎患者的致病细菌检测提供了概念验证。方法我们从每位患者的脑脊液样本中提取DNA并扩增16S rRNA基因区域。制备测序文库，并使用 minimap2 软件和代表性原核生物基因组同时处理测序读数以进行细菌组成测定。结果 6个培养阳性样本中有4个的测序结果与传统的基于培养的方法的测序结果一致。只需 3 分钟即可从测序数据中识别出每个样本中的优势细菌种类。尽管从其他两个培养阳性样本和五个培养阴性样本中也检测到了主要细菌种类，但无法证实它们的存在。此外，总体而言，尽管短测序运行内获得的测序读数数量较少，但随着时间的推移，主要细菌种类并没有发生变化。此外，处理时间与用于分析的测序读数数量密切相关。结论我们的结果表明，根据样本中细菌种类的复杂性，确定快速诊断感染性细菌种类所需的测序读数数量，可以实现时效分析。