【作者】 赵光强；

【导师】 唐学玺；

【作者基本信息】 中国海洋大学 ， 生态学， 2009， 博士

【摘要】 利用藻间相互作用进行赤潮的防治工作是目前赤潮研究工作的热点之一,而研究藻间相互作用对CO2加富的响应变化对揭示全球气候变化对海洋生态系统的影响有重要意义。本文在实验室条件下研究了大型海藻:石莼(Ulva pertusa)和江蓠(Gracilaria lemaneiformis)与赤潮微藻间的相互作用及其对CO2加富的响应变化,初步探讨了藻间相互作用及其对CO2加富响应的可能机理,阐明了利用这两种大型海藻进行赤潮防控的可能性和局限性;另一方面,微藻间的相互作用在赤潮的生消过程中同样具有重要作用,本文同时研究了微藻间的相互作用,并利用数学模型对其相互作用进行了实验室条件下的模拟。实验结果表明:1.石莼和江蓠对东海原甲藻、塔玛亚历山大藻、锥状斯氏藻和强壮前沟藻生长的影响石莼和江蓠的新鲜组织、培养液滤液能明显影响共培养体系中四种赤潮甲藻:东海原甲藻,塔玛亚历山大藻,锥状斯氏藻以及强壮前沟藻的生长。与培养液滤液相比,新鲜组织的作用更明显,它能有效抑制甚至在短时间内完全灭杀与之共培养的赤潮微藻,石莼的作用强于江蓠的作用。结果初步表明,石莼和江蓠可能分泌某种相生相克类化合物影响赤潮微藻的生长。2.石莼和江蓠对赤潮异弯藻生长的影响以赤潮异弯藻为目标微藻,在实验室条件下研究了大型海藻对它的作用,结果发现:在共培养体系中(一种大藻和一种微藻组成)石莼和江蓠的新鲜组织、培养液滤液、鲜组织匀浆和干粉末均能显著抑制甚至灭杀共培养体系中的赤潮异弯藻,其中新鲜组织的作用最强。环境因子,如温度、光照、环境微生物、培养体系pH以及碳限制等都不能导致上述现象的发生。在对共培养体系中营养盐的跟踪调查结果显示,当共培养体系中的赤潮异弯藻被完全灭杀时,石莼体系中还剩有足够的营养盐,不会限制微藻的生长;而江蓠培养体系中的营养盐几乎消耗殆尽。当向江蓠体系中补充f/2营养盐后发现,充足的营养盐能够减轻但不能完全消除江蓠对赤潮异弯藻生长的影响。另外,不同浓度的石莼新鲜组织、匀浆和培养液滤液与其对赤潮异弯藻的生长影响之间存在明显的正相关性,即起始浓度越高,对赤潮异弯藻的影响越明显。因此,石莼可能通过相生相克作用影响共培养体系中赤潮异弯藻的生长,而相生相克和营养竞争的共同作用是导致江蓠作用的根本原因。3.石莼和江蓠与5种赤潮甲藻的相互作用对CO2加富的响应变化CO2加富培养后,石莼和江蓠新鲜组织对5种赤潮微藻的影响作用分别作出了不同的响应。有的没有明显变化,有的抑制作用或杀灭作用增强,有的抑制作用或杀灭作用减弱。石莼正常新鲜组织对5种赤潮微藻的杀灭或抑制效应由强到弱的顺序是东海原甲藻>赤潮异弯藻>亚历山大藻>锥状斯氏藻>强壮前沟藻,而CO2加富培养后,其对5种赤潮微藻的杀灭或抑制效应由强到弱的顺序变化为锥状斯氏藻>赤潮异弯藻>亚历山大藻>东海原甲藻>强壮前沟藻。江蓠正常新鲜组织对5种赤潮微藻的杀灭或抑制效应由强到弱的顺序是赤潮异弯藻>东海原甲藻>亚历山大藻和锥状斯氏藻>强壮前沟藻,而CO2加富培养后,其对5种赤潮微藻的杀灭或抑制效应由强到弱的顺序变化为锥状斯氏藻>赤潮异弯藻、亚历山大藻和东海原甲藻>强壮前沟藻。CO2加富培养后,石莼和江蓠培养液滤液对5种赤潮微藻的影响作用分别作出了不同的响应。其响应变化规律与新鲜组织的变化规律相类似。这进一步充分证实,石莼和江蓠都可以通过分泌相生相克类化合物来影响共培养体系中赤潮微藻的生长。石莼正常培养液滤液对5种赤潮微藻的抑制效应由强到弱的顺序是亚历山大藻和锥状斯氏藻>东海原甲藻和赤潮异弯藻>强壮前沟藻,而CO2加富培养后,其对5种赤潮微藻的抑制效应由强到弱的顺序变化为锥状斯氏藻>亚历山大藻>强壮前沟藻和赤潮异弯藻>东海原甲藻。江蓠正常培养液滤液对5种赤潮微藻的抑制效应由强到弱的顺序是锥状斯氏藻和强壮前沟藻>赤潮异弯藻、东海原甲藻和亚历山大藻,而CO2加富培养后,其对5种赤潮微藻的抑制效应由强到弱的顺序变化为锥状斯氏藻和强壮前沟藻>亚历山大藻>赤潮异弯藻和东海原甲藻。4.东海原甲藻和塔玛亚历山大藻的相互作用研究在东海原甲藻—塔玛亚历山大藻的双藻培养体系中,当二者的初始密度/体积比为1:1时,东海原甲藻的生长被完全抑制,最终被完全灭杀,而塔玛亚历山大藻的生长几乎未受影响;当密度/体积比升至10:1时,东海原甲藻和塔玛亚历山大藻的生长均受到抑制,但是没有灭杀现象发生。相同条件下的微藻的培养液滤液试验显示了与双藻培养体系类似的试验结果,说明微藻同样能分泌相生相克类化合物影响培养体系中其它微藻的生长。对于塔玛亚历山大藻滤液的相生相克试验进一步证明了塔玛亚历山大藻能够通过分泌克生类化合物影响体系中东海原甲藻的生长;这种物质的分泌与塔玛亚历山大藻的生长阶段之间有密切关系,稳定期的塔玛亚历山大藻对东海原甲藻的抑制作用强于指数生长期的作用。同时在实验室条件下模拟了共培养体系中两种微藻的生长。当东海原甲藻与塔玛亚历山大藻的密度/体积比分别为1:1和10:1时,塔玛亚历山大藻对东海原甲藻的抑制作用分别是其被后者抑制作用的17倍和8倍。与东海原甲藻相比,塔玛亚历山大藻在双藻培养体系中具有竞争优势。实验同时说明,塔玛亚历山大藻可能分泌毒素以外的其它物质影响共培养体系中其它微藻的生长。5.东海原甲藻和塔玛亚历山大藻的相互作用对CO2加富的响应变化共培养体系中2种赤潮微藻的竞争关系对CO2的加富培养作出了响应性变化。CO2加富培养后,使东海原甲藻对塔玛亚历山大藻的相对竞争能力增强,而使塔玛亚历山大藻对东海原甲藻的相对竞争能力减弱。在培养液滤液中的实验结果也充分证实了这一结论。通过培养液滤液对海原甲藻和塔玛亚历山大藻生长的影响作用证实2种赤潮微藻间的相互作用是通过分泌克生物质产生克生效应来完成的。CO2加富培养后,培养液滤液对海原甲藻和塔玛亚历山大藻生长影响作用的变化也进一步验证了克生物质及其产生的克生效应的存在,同时表明2种赤潮微藻克生活性对CO2加富培养的响应变化是引起二者相对竞争能力发生变化的主要原因。更多还原

【Abstract】 Species interactions between bloom microalga and other alga play essential roles in affecting phytoplankton sequence, and it is important to study responses of species interactions to CO2 enrichment for illustrating the influence of global climate changes on marine ecosystem. Macroalga has long been suspected of suppressing phytoplankton growth through the excretion of chemical substances that inhibit phytoplankton growth. The production and excretion of allelochemcias by aquatic macroalga could be an effective defense strategy against other photosynthetic organisms competing for light and nutrients and therefore, macroalga might be a potentially biological method in bloom mitigating and controlling. However, the potential for this strategy to be successful in practice is exceedingly difficult mainly due to the lack of the information about the possible macroalgal species involved in and lack of the experimental data supporting its practical application. Moreover, the growth interaction between the phytoplankton species mediated by extracellular organic substances that are released by one or both interacting species has been considered to be essential to the development of phytoplankton community, and the increased population of one phytoplankton species might affect the growth of the other one or several species, influence bloom, pulses and seasonal succession. We carry out a serious of experiments under controlled conditions, and the purpose of the study is to identify if Ulva pertusa Kjellm (Chlorophyta) and Gracilaria lemaneiformis (Bory) Dawson (Rhodophyta) are the potential candidates in bloom controlling and mitigating, to investigate the response of algal interaction to CO2 enrichment, and to study the possibly effective mechanism of the interaction and the response. Moreover, the interactions between different microalgae are also studied and their potential role in bloom development has been discussed. Results show that: 1. Influence of U. pertusa and G. lemaneiformis on the growth of Prorocentrum donghaiense, Alexandrium tamarense, Scrippsiella trochoide and Amphidinium carteraeThe fresh tissues of either U. pertusa or G. lemaneiformis significantly suppressed the microalgal growth, even killed them by the end of the experiment. However, their culture filtrates exhibited different effects (inhibitory, stimulatory or none) on the co-cultured microalgae according to different microalgal species. It seemed that the macroalgae could release some allelopathic compounds into culture medium to affect the co-cultured microalgae.2. Influence of U. pertusa or G. lemaneiformis on the growth of Heterosigma akashiwoH. akashiwo was taken as an example to study the possible effect of macroalga in the co-culture. Both U. pertusa and G. lemaneiformis, and especially their fresh tissues, significantly impeded the growth of H. akashiwo, and quickly outcompeted co-cultured cells of this microalga. Carbonate limitations and the presence of bacteria are not necessary for the negative effects of U. pertusa and G. lemaneiformis on H. akashiwo. Nutrient assays showed that nitrate and phosphate were almost exhausted in the G. lemaneiformis co-culture system, but remained at acceptable levels in the U. pertusa system, when all cells of H. akashiwo were dead. When f/2 medium is supplied daily to G. lemaneiformis culture, the growth of H. akashiwo was greatly inhibited but not completely terminated. The allelopathic effects of U. pertusa may be essential for negative effects on H. akashiwo; however, the combined roles of allelopathy and nutrient competition may be responsible for the negative effect of G. lemaneiformis. Different amounts of fresh seaweed tissue, homogenate, and culture medium filtrate prepared from different macroalgal concentrations were analyzed to determine their effects on the growth of H. akashiwo. The results further suggested the release of allelochemicals by U. pertusa.3. Response of the interaction between macroalgae and microalgae to CO2 enrichmentUnder CO2 enrichment condition, the fresh tissues of either U. pertusa or G. lemaneiformis showed different effects on the five microalgae, such as no significant effect, enhanced effect of suppressing or killing, or decreased effect of suppressing or killing. The sensitivity of microalgae to the fresh tissue of U. pertusa was P. donghaiense > H. akashiwo > A. tamarense > S. trochoide > A. carterae. Under CO2 enrichment condition, the sensitivity of microalgae to the fresh tissue of U. pertusa was S. trochoide > H. akashiwo > A. tamarense > P. donghaiense > A. carterae. The sensitivity of microalgae to fresh tissue of G. lemaneiformis was H. akashiwo > P. donghaiense > A. tamarense and S. trochoide > A. carterae. Under CO2 enrichment condition, the sensitivity of microalgae to the fresh tissue of G. lemaneiformis was S. trochoide > H. akashiwo, A. tamarense and P. donghaiense > A. carterae.The five microalgae showed different response to culture filtrate of U. pertusa and G. lemaneiformis with and without CO2 enrichment. The change law of the response of culture filtrate was similar to the response of the fresh tissue with and without CO2 enrichment. This result confirmed that both U. pertusa and G. lemaneiformis could release some allelopathic compounds into culture medium to affect the co-cultured microalgae. Without CO2 enrichment, the sensitivity of microalgae to the culture filtrate of U. pertusa was A. tamarense and S. trochoide > P. donghaiense and H. akashiwo > A. carterae, while under CO2 enrichment condition, the sensitivity was S. trochoide > A. tamarense > A. carterae and H. akashiwo > P. donghaiense. And without CO2 enrichment, the sensitivity of microalgae to the culture filtrate of G. lemaneiformis was S. trochoide and A. carterae > H. akashiwo, P. donghaiense and A. tamarense, while under CO2 enrichment condition, the sensitivity was S. trochoide and A. carterae > A. tamarense > H. akashiwo and P. donghaiense.4. Interaction between P. donghaiense and A. tamarenseInteractions between P. donghaiense and A. tamarense were investigated using bialgal cultures. P. donghaiense was completely killed but A. tamarense was almost not affected by the end of the experiment when the initial density/size ratio was set at 1:1. However, significant growth suppression occurred on either species when the ratio was increased to 10:1, but no outcompetement was observed. The simultaneous assay on the culture filtrates showed that P. donghaiense filtrate prepared at a lower initial density (1.0×104 cells mL-1) induced stimulatory effect on growth of the co-cultured A. tamarense, but filtrate at a higher initial density (1.0×105cells mL-1) depressed its growth. The filtrate of A. tamarense at a density of 0.28×104 cells mL-1 completed killed P. donghaiense at a lower density, but only exhibited inhibitory effect on it at a higher density. It was likely that these two species of microalgae interfered with each other mainly by releasing allelochemical substance(s) into the culture medium, and the direct cell-cell contact was not necessary for their mutual interaction. Moreover, the strain of A. tramarense we used in this experiment was non-toxic species and therefore, there might another allelochemical(s) except the PSP toxin that affected the co-cultured microalga. The allelopathic test further proved that A. tamarense could affect the growth of co-cultured P. donghaiense by producing allelochemical(s); moreover, A. tamarense culture filtrate at the stationary growth phase (SP) had strongly inhibitory effect on P. donghaiense compared to that at the exponential phase (EP). The growth of P. donghaiense and A. tamarense in the bialgal cultures was simulated using a mathematical model to quantify the interaction. The estimated parameters from the model showed that the degree that P. donghaiense inhibited by A. tamarense was respectively about 17 and 8 times stronger than that P. donghaiense exerted on A. tamarense, when the initial density/size ratio were set at 1:1 and 10:1, respective. A. tamarense seemed to have a superior survival strategy to P. donghaiense in the bialgal cultures under controlled laboratory conditions. The results also indicate that A. tamarense can interfere other microalgae by releasing allelochemical substance(s) into the culture medium.5. Interactions between P. donghaiense and A. tamarense with CO2 enrichmentThe competition relation between P. donghaiense and A. tamarense changed with CO2 enrichment in co-culture. Under CO2 enrichment condition, the relative competitive ability of P. donghaiense enhanced, while the relative competitive ability of A. tamarense was weakened. The result from culture experiment in filtrate confirmed the conclusion above.Culture medium filtrate of P. donghaiense and A. tamarense could affected each other, and therefore confirmed that these microalgae could release some allelopathic compounds into culture medium to affect the another co-cultured microalgae. Under CO2 enrichment conditions, effects of culture filtrate of P. donghaiense and A. tamarense changed, and hence these phenomenon confirmed existences of alleopathic compounds and their effect, and indicated that the change of allopathic activity of the two microalgae with CO2 enrichment was the main reason why their relative competitive ability changed.更多还原