Abstract

Autoinducer-2 (AI-2) is a cell-to-cell signaling molecule that is commonly detected by an AI-2 bioassay with the Vibrio harveyi reporter strain BB170. However, the assay is affected by certain factors that can lead to inaccurate test results. Thus, the objective of this study was to establish the optimum method for detecting AI-2 by evaluating the effect of culture conditions for BB170, the preparation of the positive control and test samples, and the time point for AI-2 detection. The results showed that: (1) the ability of BB170 cultured in MB medium to specifically bioluminate in response to AI-2 was significantly higher than in LB medium; (2) the cell-free culture supernatant of V. harveyi BB152 served as a positive control, which was prepared from MB medium containing kanamycin; (3) an optimal time point of 4 to 6 h after inoculation of the bacterial samples was determined when using freshly cultured BB170 bacteria (OD₆₀₀ = 2.0) to detect AI-2 signal molecules; (4) the optimum time point to detect AI-2 activity in the culture supernatant was the logarithmic growth phase. In summary, our findings will improve the screening of AI-2 molecules among microorganisms, thereby aiding the functional analysis of this cell-to-cell signaling molecule.

中文翻译：

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哈维弧菌 BB170 生物测定法检测 AI-2 信号分子的优化方法

摘要

Autoinducer-2 (AI-2) 是一种细胞间信号分子，通常通过使用哈维弧菌报告菌株 BB170的 AI-2 生物测定法检测到。但是，该测定受某些可能导致测试结果不准确的因素的影响。因此，本研究的目的是通过评估 BB170 培养条件、阳性对照和测试样品的制备以及 AI-2 检测时间点的影响，建立检测 AI-2 的最佳方法。结果表明：（1）MB培养基中培养的BB170特异性生物发光响应AI-2的能力明显高于LB培养基；(2)哈维弧菌无细胞培养上清液BB152作为阳性对照，由含卡那霉素的MB培养基制备；(3) 用新鲜培养的BB170菌(OD ₆₀₀ = 2.0)检测AI-2信号分子时，确定最佳时间点为接种菌样后4～6 h ；(4)培养上清液中检测AI-2活性的最佳时间点为对数生长期。总之，我们的发现将改进微生物中 AI-2 分子的筛选，从而有助于这种细胞间信号分子的功能分析。