Ali Shimaa E, Thoen Even, Evensen Øystein, Wiik-Nielsen Jannicke, Gamil Amr A A, Skaar Ida
Norwegian Veterinary Institute, Oslo, Norway.
Norwegian Veterinary Institute, Oslo, Norway; Norwegian University of Life Sciences, Oslo, Norway.
PLoS One. 2014 Oct 29;9(10):e110343. doi: 10.1371/journal.pone.0110343. eCollection 2014.
There has been a significant increase in the incidence of Saprolegnia infections over the past decades, especially after the banning of malachite green. Very often these infections are associated with high economic losses in salmonid farms and hatcheries. The use of boric acid to control the disease has been investigated recently both under in vitro and in vivo conditions, however its possible mode of action against fish pathogenic Saprolegnia is not known. In this study, we have explored the transformation in Saprolegnia spores/hyphae after exposure to boric acid (1 g/L) over a period 4-24 h post treatment. Using transmission electron microscopy (TEM), early changes in Saprolegnia spores were detected. Mitochondrial degeneration was the most obvious sign observed following 4 h treatment in about 20% of randomly selected spores. We also investigated the effect of the treatment on nuclear division, mitochondrial activity and function using confocal laser scanning microscopy (CLSM). Fluorescence microscopy was also used to test the effect of treatment on mitochondrial membrane potential and formation of reactive oxygen species. Additionally, the viability and proliferation of treated spores that correlated to mitochondrial enzymatic activity were tested using an MTS assay. All obtained data pointed towards changes in the mitochondrial structure, membrane potential and enzymatic activity following treatment. We have found that boric acid has no effect on the integrity of membranes of Saprolegnia spores at concentrations tested. It is therefore likely that mitochondrial dysfunction is involved in the toxic activity of boric acid against Saprolegnia spp.
在过去几十年里，水霉感染的发生率显著上升，尤其是在孔雀石绿被禁用之后。这些感染常常给鲑鱼养殖场和孵化场带来巨大的经济损失。最近，人们在体外和体内条件下研究了使用硼酸来控制这种疾病的情况，然而其对鱼类致病性水霉的可能作用方式尚不清楚。在本研究中，我们探究了在处理后4至24小时的时间段内，水霉孢子/菌丝在接触硼酸（1克/升）后的变化。使用透射电子显微镜（TEM），检测到了水霉孢子的早期变化。在随机选择的约20%的孢子中，处理4小时后观察到最明显的迹象是线粒体退化。我们还使用共聚焦激光扫描显微镜（CLSM）研究了该处理对核分裂、线粒体活性和功能的影响。荧光显微镜也被用于测试处理对线粒体膜电位和活性氧形成的影响。此外，使用MTS测定法测试了与线粒体酶活性相关的处理后孢子的活力和增殖情况。所有获得的数据都指向处理后线粒体结构、膜电位和酶活性的变化。我们发现，在所测试的浓度下，硼酸对水霉孢子的膜完整性没有影响。因此，线粒体功能障碍很可能参与了硼酸对水霉属的毒性作用。
