目前正在使用许多技术来研究微生物。这些可以用来检测,识别,表征细菌,真菌,和病毒物种。一种适用于高通量分析的技术是基于流式细胞术的荧光原位杂交,或流-鱼。该技术采用(荧光标记的)针对DNA或(m)RNA的探针,例如靶向感兴趣的基因或微生物并提供单细胞水平的信息。此外,通过将Flow-FISH与基于抗体的蛋白质检测相结合,感兴趣的蛋白质可以与遗传物质同时测量。此外,根据Flow-FISH分析的类型,Flow-FISH也可以复用,允许同时测量多个基因靶标和/或微生物。一起,这允许,例如,单细胞基因表达分析或混合培养物中(亚)菌株的鉴定。Flow-FISH已用于哺乳动物细胞,但也已广泛用于研究各种微生物物种。这里,综述了Flow-FISH在微生物研究中的应用。具体来说,(细胞内)病原体的检测,研究微生物生物学和疾病发病机理,和细菌的鉴定,真菌,并讨论了混合培养物中的病毒株,特别关注EBV病毒,HIV-1和SARS-CoV-2。
Many techniques are currently in use to study microbes. These can be aimed at detecting, identifying, and characterizing bacterial, fungal, and viral species. One technique that is suitable for high-throughput analysis is flow cytometry-based fluorescence in situ hybridization, or Flow-FISH. This technique employs (fluorescently labeled) probes directed against DNA or (m)RNA, for instance targeting a gene or microorganism of interest and provides information on a single-cell level. Furthermore, by combining Flow-FISH with antibody-based protein detection, proteins of interest can be measured simultaneously with genetic material. Additionally, depending on the type of Flow-FISH assay, Flow-FISH can also be multiplexed, allowing for the simultaneous measurement of multiple gene targets and/or microorganisms. Together, this allows for, e.g., single-cell gene expression analysis or identification of (sub)strains in mixed cultures. Flow-FISH has been used in mammalian cells but has also been extensively employed to study diverse microbial species. Here, the use of Flow-FISH for studying microorganisms is reviewed. Specifically, the detection of (intracellular) pathogens, studying microorganism biology and disease pathogenesis, and identification of bacterial, fungal, and viral strains in mixed cultures is discussed, with a particular focus on the viruses EBV, HIV-1, and SARS-CoV-2.