Vogelbein Wolfgang K, Lovko Vincent J, Shields Jeffrey D, Reece Kimberly S, Mason Patrice L, Haas Leonard W, Walker Calvin C
Virginia Institute of Marine Science, The College of William and Mary, Gloucester Point, Virginia 23062, USA.
Nature. 2002 Aug 29;418(6901):967-70. doi: 10.1038/nature01008. Epub 2002 Aug 5.
Pfiesteria piscicida and P. shumwayae reportedly secrete potent exotoxins thought to cause fish lesion events, acute fish kills and human disease in mid-Atlantic USA estuaries. However, Pfiesteria toxins have never been isolated or characterized. We investigated mechanisms by which P. shumwayae kills fish using three different approaches. Here we show that larval fish bioassays conducted in tissue culture plates fitted with polycarbonate membrane inserts exhibited mortality (100%) only in treatments where fish and dinospores were in physical contact. No mortalities occurred in treatments where the membrane prevented contact between dinospores and fish. Using differential centrifugation and filtration of water from a fish-killing culture, we produced 'dinoflagellate', 'bacteria' and 'cell-free' fractions. Larval fish bioassays of these fractions resulted in mortalities (60-100% in less than 24 h) only in fractions containing live dinospores ('whole water', 'dinoflagellate'), with no mortalities in 'cell-free' or 'bacteria'-enriched fractions. Videomicrography and electron microscopy show dinospores swarming toward and attaching to skin, actively feeding, and rapidly denuding fish of epidermis. We show here that our cultures of actively fish-killing P. shumwayae do not secrete potent exotoxins; rather, fish mortality results from micropredatory feeding.
据报道，杀鱼费氏藻（Pfiesteria piscicida）和舒氏费氏藻（P. shumwayae）会分泌强效外毒素，据信这些毒素会在美国大西洋中部河口引发鱼类病变事件、急性鱼类死亡以及人类疾病。然而，费氏藻毒素从未被分离或鉴定过。我们采用三种不同方法研究了舒氏费氏藻杀死鱼类的机制。在此我们表明，在装有聚碳酸酯膜插入物的组织培养板中进行的幼鱼生物测定，只有在鱼和动孢子发生物理接触的处理中才出现死亡率（100%）。在膜阻止动孢子与鱼接触的处理中未出现死亡情况。通过对杀鱼培养物的水进行差速离心和过滤，我们得到了“甲藻”、“细菌”和“无细胞”组分。对这些组分进行的幼鱼生物测定显示，只有在含有活的动孢子的组分（“全水”、“甲藻”）中才出现死亡率（在不到24小时内为60 - 100%），而在“无细胞”或富含“细菌”的组分中未出现死亡情况。视频显微镜和电子显微镜显示，动孢子成群地朝鱼皮游动并附着，积极摄食，并迅速剥去鱼的表皮。我们在此表明，我们培养的具有活跃杀鱼能力的舒氏费氏藻不会分泌强效外毒素；相反，鱼类死亡是由微型捕食性摄食导致的。
