IEEE Trans Biomed Eng. 2018 Dec;65(12):2731-2741. doi: 10.1109/TBME.2018.2814597. Epub 2018 Mar 9.
Inflammation of the meninges is a source of severe morbidity and therefore is an important health concerns worldwide. The conventional clinical microbiology approaches used today to identify pathogens suffer from several drawbacks and frequently provide false results. This research describes a fast method to detect the presence of pathogens using the frequency domain (FD) fluorescence lifetime (FLT) imaging microscopy (FLIM) system.
The study included 43 individuals divided into 4 groups: 9 diagnosed with different types of bacteria; 16 diagnosed with different types of viruses; 5 healthy samples served as a control; and 12 samples were negative to any pathogen, although presenting related symptoms. All samples contained leukocytes that were extracted from the cerebrospinal fluid (CSF) and were subjected to nuclear staining by 4', 6-diamidino-2-phenylindole (DAPI) and FLT analyses based on phase and amplitude crossing point (CRPO).
Using notched boxplots, we found differences in 95% probability between the first three groups through different notch ranges (NR). Pathogen samples presented a longer median FLT (3.28 ns with NR of 3.24-3.32 ns in bacteria and 3.18 ns with NR of 3.16-3.21 ns in viruses) compared to the control median FLT (2.65 ns with NR of 2.63-2.67 ns). Furthermore, we found that the undetected forth group was divided into two types: a relatively normal median FLT (2.72 ns with NR of 2.68-2.76 ns) and a prolonged FLT (3.22 ns with NR of 3.17-3.27 ns).
FLT measurements can differentiate between control and pathogen by the CRPO method.
The FD-FLIM system can provide a high throughput diagnostic technique that does not require a physician.
脑膜炎症是严重发病和致残的根源，也是全世界关注的重要健康问题。目前用于识别病原体的常规临床微生物学方法存在一些缺陷，并且经常提供错误的结果。本研究描述了一种使用频域（FD）荧光寿命（FLT）成像显微镜（FLIM）系统快速检测病原体的方法。
该研究共纳入 43 名个体，分为 4 组：9 名被诊断为不同类型细菌感染；16 名被诊断为不同类型病毒感染；5 名健康样本作为对照；12 名样本虽然有相关症状，但对任何病原体均呈阴性。所有样本均含有白细胞，从脑脊液（CSF）中提取出来，并用 4'，6-二脒基-2-苯基吲哚（DAPI）进行核染色，并根据相位和幅度交点（CRPO）进行 FLT 分析。
通过 notch 箱线图，我们发现前三个组之间存在 95%概率的差异，通过不同的 notch 范围（NR）。病原体样本的中位 FLT 较长（细菌为 3.28 纳米，NR 为 3.24-3.32 纳米；病毒为 3.18 纳米，NR 为 3.16-3.21 纳米），与对照中位 FLT（2.65 纳米，NR 为 2.63-2.67 纳米）相比。此外，我们发现第四个未检出组可分为两类：相对正常的中位 FLT（2.72 纳米，NR 为 2.68-2.76 纳米）和延长的 FLT（3.22 纳米，NR 为 3.17-3.27 纳米）。
CRPO 方法可以通过 FLT 测量来区分对照和病原体。
FD-FLIM 系统可以提供一种不需要医生的高通量诊断技术。
