Microorganisms have been very useful in environmental monitoring due to their constant sensing of the surrounding environment, their easy maintenance and low cost. Some freeze-dried toxicity kits based on naturally bioluminescent bacteria are commercially available and commonly used to assess the toxicity of environmental samples such as Microtox (Aliivibrio fischeri) or ToxScreen (Photobacterium leiognathi), however, due to the marine origin of these bacteria, they could not be the most appropriate for fresh-waters monitoring. Cyanobacteria are one of the most representative microorganisms of aquatic environments, and are well suited for detecting contaminants in aqueous samples. This study presents the development and application of the first freeze-dried cyanobacterial bioassay for fresh-water contaminants detection. The effects of different cell growth phases, cryoprotectant solutions, freezing protocols, rehydration solutions and incubation conditions methods were evaluated and the best combination of these parameters for freeze-drying was selected. The study includes detailed characterization of sensitivity towards reference pollutants, as well as, comparison with the standard assays. Moreover, long-term viability and sensitivity were evaluated after 3 years of storage. Freeze-dried cyanobacteria showed, in general, higher sensitivity than the standard assays and viability of the cells remained after 3 years of storage. Finally, the validation of the bioassay using a wastewater sample was also evaluated. Freeze-drying of cyanobacteria in 96-well plates presents a simple, fast and multi-assay method for environmental monitoring.

中文翻译：

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微孔板冻干蓝藻生物测定法用于淡水环境监测

微生物由于对周围环境的不断感应，易于维护且成本低廉，因此在环境监测中非常有用。一些基于天然生物发光细菌的冻干毒性试剂盒可商购获得，通常用于评估环境样品的毒性，例如Microtox（Aliivibrio fischeri）或ToxScreen（Photobacter leiognathi）但是，由于这些细菌的海洋起源，它们可能不是最适合淡水监测的细菌。蓝细菌是水生环境中最具代表性的微生物之一，非常适合检测水性样品中的污染物。这项研究介绍了用于淡水污染物检测的第一个冷冻干燥的蓝细菌生物测定法的开发和应用。评估了不同细胞生长阶段，冷冻保护剂溶液，冷冻方案，补液溶液和孵育条件方法的影响，并选择了这些参数进行冷冻干燥的最佳组合。该研究包括对参考污染物敏感性的详细表征，以及与标准测定法的比较。而且，储存3年后评估其长期生存能力和敏感性。通常，冷冻干燥的蓝细菌显示出比标准测定法更高的灵敏度，并且在保存3年后仍保留了细胞的活力。最后，还评估了使用废水样品进行生物测定的有效性。在96孔板上冷冻干燥蓝藻细菌提供了一种简单，快速，多种测定的环境监测方法。