Norwegian Institute for Water Research (NIVA), Gaustadalléen 21, Oslo 0349, Norway.
Norwegian Institute for Water Research (NIVA), Gaustadalléen 21, Oslo 0349, Norway.
Sci Total Environ. 2019 Oct 15;687:827-838. doi: 10.1016/j.scitotenv.2019.06.124. Epub 2019 Jun 12.
Microalgal toxicity tests using integrative endpoints as algal growth are regularly required to analyse the toxicity of potentially hazardous substances in the aquatic environment. However, these do not provide mechanistic information on the toxic mode of action by which contaminants may affect algae. Bottled waters can be used as a substitute for culturing media and should not impose any stress to the cultured organisms. However, certain chemical components can interfere with specific cell targets which are not revealed by general toxicity assays. The present study investigated the sensitivity of flow cytometry (FCM) to analyse sub-lethal effects of different bottled waters to the freshwater microalgae Raphidocelis subcapitata. Several endpoints were analysed including growth rate, natural pigments content, cell size, complexity, viability and cycle, Reactive Oxygen Species (ROS) formation, mitochondrial membrane potential and Lipid Peroxidation (LPO). Additionally, photosystem II (PSII) performance was analysed by PAM fluorometry, to provide further information on the absorption, distribution and use of energy in photosynthesis. Results indicated that the most sensitive endpoints were the oxidative stress related endpoints ROS formation and LPO, pigment content, morphological endpoints as cell size, complexity and cycle, with growth rate being one of the least sensitive. Although being essential macronutrients for algal growth, the chemical elements Ca, Na, Mg, and NH were identified as being primarily responsible for the observed toxicological effects to exposed algae. The applied methodology proved to be of high throughput, simultaneously assembling information on morphological, biochemical, and physiological status of algal cells. FCM also showed potential to reveal mechanistic information on the toxic mode of action of the bottled waters before any effects on algal growth was observed. The used approach demonstrated its potential for being integrated into future microalgal toxicity bioassays for testing chemicals to improve the hazard information obtained from currently approved internationally accepted test guidelines.
微藻毒性测试通常需要使用综合终点（如藻生长）来分析水生环境中潜在有害物质的毒性。然而，这些方法不能提供污染物可能影响藻类的毒性作用模式的机制信息。瓶装水可用作培养介质的替代品，不应给培养的生物带来任何压力。然而，某些化学成分可能会干扰特定的细胞靶标，而这些靶标不会被一般毒性测定所揭示。本研究调查了流式细胞术（FCM）分析不同瓶装水对淡水微藻莱茵衣藻亚致死效应的敏感性。分析了包括生长率、天然色素含量、细胞大小、复杂性、活力和周期、活性氧（ROS）形成、线粒体膜电位和脂质过氧化（LPO）在内的多个终点。此外，通过 PAM 荧光法分析了光合作用系统 II（PSII）的性能，以提供有关光合作用中能量吸收、分布和利用的进一步信息。结果表明，最敏感的终点是与氧化应激相关的终点 ROS 形成和 LPO、色素含量、形态终点如细胞大小、复杂性和周期，而生长率是最不敏感的终点之一。尽管 Ca、Na、Mg 和 NH 是藻类生长所必需的大量营养元素，但研究表明，这些化学元素是导致暴露于藻类的观察到的毒性作用的主要原因。所应用的方法被证明具有高通量的特点，同时提供了藻类细胞形态、生化和生理状态的信息。FCM 还显示出在观察到藻类生长受到影响之前，揭示瓶装水毒性作用模式的机制信息的潜力。所采用的方法展示了其在未来微藻毒性生物测定中的潜力，用于测试化学品，以改善从当前批准的国际公认测试指南中获得的危害信息。
