Department of Biological Sciences, Seton Hall University, South Orange, NJ 07079, USA.
Toxins (Basel). 2020 Jan 30;12(2):92. doi: 10.3390/toxins12020092.
Cyanobacteria harmful algal blooms (CHABs) are primarily caused by man-made eutrophication and increasing climate-change conditions. The presence of heavy metal runoff in affected water systems may result in CHABs alteration to their ecological interactions. Certain CHABs produce by-products, such as microcystin (MC) cyanotoxins, that have detrimentally affected humans through contact via recreation activities within implicated water bodies, directly drinking contaminated water, ingesting biomagnified cyanotoxins in seafood, and/or contact through miscellaneous water treatment. Metallothionein (MT) is a small, metal-sequestration cysteine rich protein often upregulated within the stress response mechanism. This study focused on zinc metal resistance and stress response in a toxigenic cyanobacterium, UTEX LB 2385, by monitoring cells with (0, 0.1, 0.25, and 0.5 mg/L) ZnCl treatment. Flow cytometry and phase contrast microscopy were used to evaluate physiological responses in cultures. Molecular assays and an immunosorbent assay were used to characterize the expression of MT and MC under zinc stress. The results showed that the half maximal inhibitory concentration (IC) was 0.25 mg/L ZnCl. Flow cytometry and phase contrast microscopy showed morphological changes occurred in cultures exposed to 0.25 and 0.5 mg/L ZnCl. Quantitative PCR (qPCR) analysis of selected cDNA samples showed significant upregulation of through all time points, significant upregulation of at a later time point. ELISA MC-LR analysis showed extracellular MC-LR (µg/L) and intracellular MC-LR (µg/cell) quota measurements persisted through 15 days, although 0.25 mg/L ZnCl treatment produced half the normal cell biomass and 0.5 mg/L treatment largely inhibited growth. The 0.25 and 0.5 mg/L ZnCl treated cells demonstrated a ~40% and 33% increase of extracellular MC-LR(µg/L) equivalents, respectively, as early as Day 5 compared to control cells. The 0.5 mg/L ZnCl treated cells showed higher total MC-LR (µg/cell) quota yield by Day 8 than both 0 mg/L ZnCl control cells and 0.1 mg/L ZnCl treated cells, indicating release of MCs upon cell lysis. This study showed this strain is able to survive in 0.25 mg/L ZnCl concentration. Certain morphological zinc stress responses and the upregulation of and genes, as well as periodical increased extracellular MC-LR concentration with ZnCl treatment were observed.
蓝藻有害藻华（CHABs）主要是由人为富营养化和不断变化的气候变化引起的。受重金属径流影响的水系中 CHABs 的生态相互作用可能会发生变化。某些 CHABs 会产生副产物，例如微囊藻毒素（MC）蓝藻毒素，这些毒素会通过在受影响水体中的娱乐活动直接接触、直接饮用受污染的水、摄入海鲜中的生物放大蓝藻毒素以及/或通过各种水处理接触对人类造成不利影响。金属硫蛋白（MT）是一种小分子、富含半胱氨酸的金属结合蛋白，通常在应激反应机制中上调。本研究通过监测（0、0.1、0.25 和 0.5mg/L）ZnCl 处理下的细胞，研究了产毒蓝藻 UTEX LB 2385 的锌金属抗性和应激反应。流式细胞术和相差显微镜用于评估培养物的生理反应。分子测定和免疫吸附测定用于表征锌胁迫下 MT 和 MC 的表达。结果表明，半最大抑制浓度（IC）为 0.25mg/L ZnCl。流式细胞术和相差显微镜显示，暴露于 0.25 和 0.5mg/L ZnCl 的培养物发生形态变化。对选定 cDNA 样本的定量 PCR（qPCR）分析显示，在所有时间点均显著上调 ，在稍后的时间点显著上调 。ELISA MC-LR 分析显示，尽管 0.25mg/L ZnCl 处理产生了正常细胞生物量的一半，而 0.5mg/L 处理则大大抑制了生长，但细胞外 MC-LR（µg/L）和细胞内 MC-LR（µg/细胞）配额测量仍持续 15 天。0.25 和 0.5mg/L ZnCl 处理的细胞在第 5 天与对照细胞相比，细胞外 MC-LR（µg/L）当量分别增加了约 40%和 33%。第 8 天，0.5mg/L ZnCl 处理的细胞的总 MC-LR（µg/细胞）产量比 0mg/L ZnCl 对照细胞和 0.1mg/L ZnCl 处理的细胞都高，表明细胞裂解时释放了 MC。这项研究表明，这种 菌株能够在 0.25mg/L ZnCl 浓度下存活。观察到某些形态锌应激反应以及 和 基因的上调，以及随着 ZnCl 处理周期性增加细胞外 MC-LR 浓度。
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