National Oceanic and Atmospheric Administration, National Ocean Service, National Centers for Coastal Ocean Science, Center for Coastal Fisheries and Habitat Research, 101 Pivers Island Road, Beaufort, NC 28516, USA.
Toxicon. 2013 Apr;65:15-33. doi: 10.1016/j.toxicon.2012.12.016. Epub 2013 Jan 10.
This study examined the toxicity of six Gambierdiscus species (Gambierdiscus belizeanus, Gambierdiscus caribaeus, Gambierdiscus carolinianus, Gambierdiscus carpenteri, Gambierdiscus ribotype 2 and Gambierdiscus ruetzleri) using a human erythrocyte lysis assay. In all, 56 isolates were tested. The results showed certain species were significantly more toxic than others. Depending on the species, hemolytic activity consistently increased by ∼7-40% from log phase growth to late log - early stationary growth phase and then declined in mid-stationary growth phase. Increasing growth temperatures from 20 to 31 °C for clones of G. caribaeus showed only a slight increase in hemolytic activity between 20 and 27 °C. Hemolytic activity in the G. carolinianus isolates from different regions grown over the same 20-31 °C range remained constant. These data suggest that growth temperature is not a significant factor in modulating the inter-isolate and interspecific differences in hemolytic activity. The hemolytic activity of various isolates measured repeatedly over a 2 year period remained constant, consistent with the hemolytic compounds being constitutively produced and under strong genetic control. Depending on species, greater than 60-90% of the total hemolytic activity was initially associated with the cell membranes but diffused into solution over a 24 h assay incubation period at 4 °C. These findings suggest that hemolytic compounds produced by Gambierdiscus isolates were held in membrane bound vesicles as reported for brevetoxins produced by Karenia brevis. Gambierdiscus isolates obtained from other parts of the world exhibited hemolytic activities comparable to those found in the Caribbean and Gulf of Mexico confirming the range of toxicities is similar among Gambierdiscus species worldwide. Experiments using specific inhibitors of the MTX pathway and purified MTX, Gambierdiscus whole cell extracts, and hydrophilic cell extracts containing MTX, were consistent with MTX as the primary hemolytic compound produced by Gambierdiscus species. While the results from inhibition studies require validation by LC-MS analysis, the available data strongly suggest differences in hemolytic activity observed in this study reflect maitotoxicity.
本研究采用人红细胞溶解试验检测了 6 种冈比亚型双鞭藻（冈比亚型双鞭藻、加勒比型双鞭藻、卡罗来纳型双鞭藻、卡彭特氏型双鞭藻、2 型冈比亚型双鞭藻和鲁氏型双鞭藻）的毒性。共检测了 56 个分离株。结果表明，某些种的毒性明显高于其他种。根据种的不同，溶血活性从对数生长期到对数末期-早期静止期持续增加约 7-40%，然后在中期静止期下降。将加勒比型双鞭藻克隆的生长温度从 20°C 升高到 31°C 时，在 20°C 至 27°C 之间仅观察到溶血活性略有增加。在相同的 20-31°C 范围内生长的不同地区的卡罗来纳型双鞭藻分离株的溶血活性保持不变。这些数据表明，生长温度不是调节种间和种内溶血活性差异的重要因素。在 2 年内反复测量的各种分离株的溶血活性保持不变，这与溶血化合物持续产生并受强遗传控制一致。根据种的不同，大于 60-90%的总溶血活性最初与细胞膜相关，但在 4°C 孵育 24 小时的测定过程中扩散到溶液中。这些发现表明，与报道的短凯伦藻产生的布雷韦毒素一样，冈比亚型双鞭藻分离株产生的溶血化合物被束缚在膜结合小泡中。从世界其他地区获得的冈比亚型双鞭藻分离株表现出与加勒比海和墨西哥湾相似的溶血活性，这证实了在全球范围内，冈比亚型双鞭藻的毒性范围相似。使用 MTX 途径的特定抑制剂和纯化的 MTX、冈比亚型双鞭藻全细胞提取物以及含有 MTX 的亲水性细胞提取物进行的实验结果与 MTX 作为冈比亚型双鞭藻产生的主要溶血化合物一致。虽然抑制研究的结果需要通过 LC-MS 分析进行验证，但现有数据强烈表明，本研究中观察到的溶血活性差异反映了麦角毒性。
