中文摘要：公共卫生急需快速、定量、灵敏的多重检测方法。本文报道了一种双重修饰噬菌体传感器，用于同时检测多种致病菌。双重修饰M13KE噬菌体，将目标肽展示在小外壳蛋白pIII上(-5个拷贝)，链霉菌亲和素结合(StrB)肽展示在主外壳蛋白pVIII上(-2700个拷贝)。目标肽特异性识别目标细菌，StrB肽与荧光标记的链霉亲和素结合后，作为有效的信号放大和转导单元。流式细胞术和荧光显微镜均能清晰地分辨出单个靶菌发出的明亮荧光。构建针对大肠杆菌O157:H7、鼠伤寒沙门氏菌和铜绿假单胞菌的三种不同的双修饰噬菌体，并用大量其他非靶标细菌验证了其高特异性。使用40mL的样品量，在存在其他非目标细菌的情况下，通过流式细胞术检测目标细菌的检测限约为102个细胞/mL。将这三种双修饰噬菌体混合，可以同时定量检测三种目标病原菌，并具有良好的线性。可见，构建双修饰噬菌体是一种很有前途的检测手段。
外文摘要：Rapid, quantitative, and sensitive assays for the multiplexed detection of bacterial pathogens are urgently needed for public health. Here, we report the generation of dual-modified phage sensors for the simultaneous detection of multiple pathogenic bacteria. The M13KE phage was dual modified to display the targeting peptide on the minor coat protein pIII (-5 copies) and the streptavidin-binding (StrB) peptide on the major coat protein pVIII (-2700 copies). The targeting peptide specifically recognizes the target bacteria, and the StrB peptide acts as the efficient signal amplification and transduction unit upon binding with fluorescently tagged streptavidin. The bright fluorescence emitted from individual target bacteria can be clearly distinguished from the background via both the flow cytometry and fluorescence microscopy. Three different dual-modified phages targeting E. coli O157:H7, Salmonella Typhimurium, and Pseudomonas aeruginosa were constructed, and high specificity was verified via a large excess of other non-target bacteria. Using a 40 mL sample volume, the target bacteria detection limit was approximately 102 cells/mL via flow cytometry measurement in the presence of other non-target bacteria. By combining these three dual-modified phages into a cocktail, simultaneous detection and quantification of three target bacterial pathogens was demonstrated with good linearity. The strategy of constructing dual-modified phage represents a promising tool in the detection of bacterial pathogens.
外文关键词：Dual-modified phage；Phage cocktail；Multiplexed detection；Bacterial pathogens；Flow cytometry
作者：Wu, LN；Hong, XY；Luan, T；Zhang, YZ；Li, LH；Huang, TT；Yan, XM
作者单位：Xiamen Univ
期刊名称：ANALYTICA CHIMICA ACTA
期刊影响因子：5.977
出版年份：2021
出版刊次：1166
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