Isolation ssDNA aptamers specific for both live and viable but nonculturable state Vibrio vulnificus using whole bacteria-SEILEX technology.
Isolation ssDNA aptamers specific for both live and viable but nonculturable state Vibrio vulnificus using whole bacteria-SEILEX technology.
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使用全细菌-SEILEX 技术分离活的和活的但不可培养状态的创伤弧菌特异性的 ssDNA 适体
作者：
创伤弧菌是一种普遍存在的海洋细菌，可能导致快速和致命的感染，威胁生活在自然水体周围的人们的生命，特别是在沿海地区。然而，传统的基于培养的方法是耗时的，并且无法检测活的但不可培养的（VBNC）创伤弧菌细胞。在本工作中，我们分离了一批在所有培养状态下特异性结合创伤弧菌的检测适体。通过传统的全菌-SELEX（指数富集配体的系统进化）、流式细胞仪分析和成像，我们鉴定了18个候选物，并验证了其中两个（V8和V13）为适用的适体。它们的截短序列也显示出相当的性能。V8的解离常数（KD）值显示为低至11.22 ± 1.32 nM。最佳适体V8和V13也被验证为使用流式细胞术在不同结合环境下有效检测不同创伤弧菌菌株。对于检测参数，V8细胞计数的LOD为29.96 CFU mL−1，线性范围为102-5 × 105 CFU mL−1。这是第一例证明适体可以检测VBNC细菌以及活细菌的存在。
Vibrio vulnificus is a ubiquitous marine bacterium that may cause rapid and deadly infection, threatening lives of people living around natural bodies of water, especially in coastal regions. However, traditional culture-based methods are time-consuming and unable to detect Viable But Non-Culturable (VBNC) V. vulnificus cells. In this work, we isolated a batch of detection aptamers specifically binding to V. vulnificus in all culture status. With traditional whole bacteria-SELEX (Systematic Evolution of Ligands by EXponential enrichment), flow cytometer analysis and imaging, we identify 18 candidates and validated two of them (V8 and V13) as applicable aptamers. Their truncated sequences also showed comparable performance. The dissociation constant (KD) value of V8 is shown to be as low as 11.22 ± 1.32 nM. Optimal aptamers V8 and V13 are also validated to be effective to detect different Vibrio vulnificus strains under different binding environments using flow cytometry. As for detection parameters, the LOD of the V8 from cytometry is 29.96 CFU mL−1, and the linear range is 102–5 × 105 CFU mL−1. This is the first case demonstrating that aptamers can detect the existence of VBNC bacteria as well as live bacteria.
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影响因子：
1.5
作者：
Baker-Austin C;Oliver JD
通讯作者：
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影响因子：
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通讯作者：
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影响因子：
2.9
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12.6
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