溶藻细菌L7对铜绿微囊藻的溶藻效应及机理
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摘要
随着水体富营养化现象的日趋严重及藻类水华的频繁暴发,探索行之有效的抑制藻类水华暴发的途径极为迫切,其中利用溶藻细菌控藻成为目前生物控藻的研究热点。
作者所在课题组从广州城区某富营养化池塘筛选获得一株溶藻细菌L7(简称L7),属蜡状芽孢杆菌(Bacillus cereus),在已有研究基础上,作者以常见的水华藻类——铜绿微囊藻(Microcystis aeruginosa)为研究对象,研究了L7对铜绿微囊藻的溶藻效应,并探讨了其溶藻机理。主要研究结果如下:
1、L7对铜绿微囊藻的溶藻效应
为了探索L7溶藻的适宜环境条件,研究了菌藻浓度及环境因子对L7溶藻效应的影响。结果表明:L7对铜绿微囊藻的溶藻效应呈“剂量-效应”关系,表现出“低浓度促进,高浓度抑制”效应;在连续7 d的实验周期内,在不同温度条件下的溶藻效应强弱依次为25℃>35℃>15℃;12 h:12 h光暗周期条件下的溶藻效应强于全光照和全黑暗;在不同pH值条件下的溶藻效应强弱依次为:pH 9.0>pH 6.0>pH 7.0>pH 8.0>pH 10.0。可见,L7对铜绿微囊藻的溶藻效应与菌藻浓度及环境因子密切相关。
2、L7对铜绿微囊藻生理特性的影响
研究L7对铜绿微囊藻生理特性的影响,从宏观层面初步探讨溶藻机理。结果表明:在高浓度L7(ρL7>107 cfu/mL)作用下,藻细胞脱氢酶活性、可溶性蛋白质含量、可溶性碳水化合物含量和光合色素(叶绿素a、类胡萝卜素和藻胆蛋白)含量均下降,丙二醛含量升高。可见,L7对藻细胞的生理特性影响的途径为:L7通过抑制藻细胞的光合系统活性,进而导致光合作用减弱,使得藻细胞生化反应减慢,细胞膜氧化损伤程度升高,最终影响到藻细胞分裂,使藻细胞生长受到抑制。
3、L7对铜绿微囊藻细胞生物学特征的影响
使用流式细胞仪,以PI染色和叶绿素自发荧光检测L7溶藻过程中藻细胞生物学特征的变化,在微观层面深入探讨溶藻机理。结果表明:L7与藻细胞接触后,先从破坏藻细胞膜完整性开始,然后透过破损的细胞膜破坏藻体的光合色素,从而抑制藻细胞的生理功能的正常发挥,从而造成藻体的非自然死亡;在L7的持续作用下,藻细胞的核酸物质也遭受到严重破坏。可见,L7对藻细胞生物学特征的影响途径在于破坏细胞膜完整性进而破坏光合色素。
4、L7溶藻过程中磷的转移
为明确磷在菌藻共培养体系中的转移过程,研究了L7溶藻过程中磷的转移。结果表明:L7尽管抑制了藻细胞的增殖,但藻细胞对水体磷的吸收并不受其抑制,在后期铜绿微囊藻能在藻细胞内积聚磷。可见,溶藻过程中磷部分积聚在藻细胞内。
5、藻细胞破碎液对L7的促进
为了进一步明晰藻类与溶藻细菌相互影响的作用机理,通过在L7的培养过程中添加铜绿微囊藻细胞破碎液,试图探究藻细胞破碎液对L7的促进作用。结果表明:藻细胞破碎液的添加有利于提高L7溶藻活性物质的溶藻效果,但不同生长阶段藻细胞破碎液的促进作用无显著差异;随着添加的藻细胞破碎液体积浓度的提高,L7溶藻活性物质的溶藻效果提高,呈现一定的“剂量-效应”关系。可见,藻细胞破碎液能提高L7的溶藻效果。
With the increasingly serious eutrophication and frequent outbreaks of algal bloom, it is imperative to explore effective methods to prevent algal bloom. The use of algicidal bacteria is considered as a new method of controlling algae has become the hotspot of biological control of algae.
One algicidal bacteria, strain L7, was isolated from a eutrophic pond in the town of Guangzhou, which belongs to Bacillus cereus. Based on the existing research, the author studied the algicidal effect of L7 on Microcystis aeruginosa, and explored its algicidal mechanism. The main results were as follows:
1. The algicidal effect of L7 on Microcystis aeruginosa
The algicidal effect of L7 on Microcystis aeruginosa was consistents with the relationship of "concentration-effect", showing“low concentration promoted, high concentration inhibited”. During 7d’s consecutive experiment, the algicidal effect under different temperature conditions were 25℃>35℃>15℃. The algicidal effect in the 12h: 12h light and dark cycle was stronger than in illumination and in darkness; At different pH level, the algicidal effect were pH 9.0>pH 7.0>pH 6.0>pH 8.0>pH 10.0.
2. The effect of L7 on the physiological characteristics of Microcystis aeruginosa
Under the treatment of high concentration L7 (ρL7>107cfu/mL), the dehydrogenase activity, soluble protein content, soluble carbohydrate content of Microcystis aeruginosa decreased. The chlorophyll, carotenoids and phycocyanin of Microcystis aeruginosa decreased significantly. The MDA content increased. We conclude that the algicidal mechanism of L7 on Microcystis aeruginosa was: L7 inhibited the photosynthesis system activity of Microcystis aeruginosa, which led to reducing the photosynthesis, and then reducing the dehydrogenase activity, soluble protein, soluble carbohydrate content, making bilchenical reations slow down, ultimately affecting the cell division of algae.
3. The effect of L7 on the cell biological characteristics of Microcystis aeruginosa
The biological characteristics of the algae in the algicidal cycle by FCM analysis showed that the algicidal trends were in steps. Firstly, the Microcystis aeruginosa cell membrane was destroyed, then the chlorophyll was damaged, resulting in the non-natural death of the algae cells at the last step. In the continued effect of L7, the nucleic acids of the algae cells was also damaged severely.
4. The transfer of phosphorus during the algicidal process
Although L7 suppressed the proliferation of the algae cells, the absorption of phosphorus in water was not subject to its suppression, and the phosphorus could be accumulated in the algae in the latter stages.
5. The feedback and induction of intact algal solution to L7
Adding intact algal solution of Microcystis aeruginosa into the culture process of L7 would help improve the aligicidal effect of algicidal components to Microcystis aeruginosa, but there was no significant difference among different growth cycles of algae; With the addition of the intact algal solution of Microcystis aeruginosa increased, the algicidal effect of the algicidal components to Microcystis aeruginosa increased, showing the relationship of“dose-effect”.
作者所在课题组从广州城区某富营养化池塘筛选获得一株溶藻细菌L7(简称L7),属蜡状芽孢杆菌(Bacillus cereus),在已有研究基础上,作者以常见的水华藻类——铜绿微囊藻(Microcystis aeruginosa)为研究对象,研究了L7对铜绿微囊藻的溶藻效应,并探讨了其溶藻机理。主要研究结果如下:
1、L7对铜绿微囊藻的溶藻效应
为了探索L7溶藻的适宜环境条件,研究了菌藻浓度及环境因子对L7溶藻效应的影响。结果表明:L7对铜绿微囊藻的溶藻效应呈“剂量-效应”关系,表现出“低浓度促进,高浓度抑制”效应;在连续7 d的实验周期内,在不同温度条件下的溶藻效应强弱依次为25℃>35℃>15℃;12 h:12 h光暗周期条件下的溶藻效应强于全光照和全黑暗;在不同pH值条件下的溶藻效应强弱依次为:pH 9.0>pH 6.0>pH 7.0>pH 8.0>pH 10.0。可见,L7对铜绿微囊藻的溶藻效应与菌藻浓度及环境因子密切相关。
2、L7对铜绿微囊藻生理特性的影响
研究L7对铜绿微囊藻生理特性的影响,从宏观层面初步探讨溶藻机理。结果表明:在高浓度L7(ρL7>107 cfu/mL)作用下,藻细胞脱氢酶活性、可溶性蛋白质含量、可溶性碳水化合物含量和光合色素(叶绿素a、类胡萝卜素和藻胆蛋白)含量均下降,丙二醛含量升高。可见,L7对藻细胞的生理特性影响的途径为:L7通过抑制藻细胞的光合系统活性,进而导致光合作用减弱,使得藻细胞生化反应减慢,细胞膜氧化损伤程度升高,最终影响到藻细胞分裂,使藻细胞生长受到抑制。
3、L7对铜绿微囊藻细胞生物学特征的影响
使用流式细胞仪,以PI染色和叶绿素自发荧光检测L7溶藻过程中藻细胞生物学特征的变化,在微观层面深入探讨溶藻机理。结果表明:L7与藻细胞接触后,先从破坏藻细胞膜完整性开始,然后透过破损的细胞膜破坏藻体的光合色素,从而抑制藻细胞的生理功能的正常发挥,从而造成藻体的非自然死亡;在L7的持续作用下,藻细胞的核酸物质也遭受到严重破坏。可见,L7对藻细胞生物学特征的影响途径在于破坏细胞膜完整性进而破坏光合色素。
4、L7溶藻过程中磷的转移
为明确磷在菌藻共培养体系中的转移过程,研究了L7溶藻过程中磷的转移。结果表明:L7尽管抑制了藻细胞的增殖,但藻细胞对水体磷的吸收并不受其抑制,在后期铜绿微囊藻能在藻细胞内积聚磷。可见,溶藻过程中磷部分积聚在藻细胞内。
5、藻细胞破碎液对L7的促进
为了进一步明晰藻类与溶藻细菌相互影响的作用机理,通过在L7的培养过程中添加铜绿微囊藻细胞破碎液,试图探究藻细胞破碎液对L7的促进作用。结果表明:藻细胞破碎液的添加有利于提高L7溶藻活性物质的溶藻效果,但不同生长阶段藻细胞破碎液的促进作用无显著差异;随着添加的藻细胞破碎液体积浓度的提高,L7溶藻活性物质的溶藻效果提高,呈现一定的“剂量-效应”关系。可见,藻细胞破碎液能提高L7的溶藻效果。
With the increasingly serious eutrophication and frequent outbreaks of algal bloom, it is imperative to explore effective methods to prevent algal bloom. The use of algicidal bacteria is considered as a new method of controlling algae has become the hotspot of biological control of algae.
One algicidal bacteria, strain L7, was isolated from a eutrophic pond in the town of Guangzhou, which belongs to Bacillus cereus. Based on the existing research, the author studied the algicidal effect of L7 on Microcystis aeruginosa, and explored its algicidal mechanism. The main results were as follows:
1. The algicidal effect of L7 on Microcystis aeruginosa
The algicidal effect of L7 on Microcystis aeruginosa was consistents with the relationship of "concentration-effect", showing“low concentration promoted, high concentration inhibited”. During 7d’s consecutive experiment, the algicidal effect under different temperature conditions were 25℃>35℃>15℃. The algicidal effect in the 12h: 12h light and dark cycle was stronger than in illumination and in darkness; At different pH level, the algicidal effect were pH 9.0>pH 7.0>pH 6.0>pH 8.0>pH 10.0.
2. The effect of L7 on the physiological characteristics of Microcystis aeruginosa
Under the treatment of high concentration L7 (ρL7>107cfu/mL), the dehydrogenase activity, soluble protein content, soluble carbohydrate content of Microcystis aeruginosa decreased. The chlorophyll, carotenoids and phycocyanin of Microcystis aeruginosa decreased significantly. The MDA content increased. We conclude that the algicidal mechanism of L7 on Microcystis aeruginosa was: L7 inhibited the photosynthesis system activity of Microcystis aeruginosa, which led to reducing the photosynthesis, and then reducing the dehydrogenase activity, soluble protein, soluble carbohydrate content, making bilchenical reations slow down, ultimately affecting the cell division of algae.
3. The effect of L7 on the cell biological characteristics of Microcystis aeruginosa
The biological characteristics of the algae in the algicidal cycle by FCM analysis showed that the algicidal trends were in steps. Firstly, the Microcystis aeruginosa cell membrane was destroyed, then the chlorophyll was damaged, resulting in the non-natural death of the algae cells at the last step. In the continued effect of L7, the nucleic acids of the algae cells was also damaged severely.
4. The transfer of phosphorus during the algicidal process
Although L7 suppressed the proliferation of the algae cells, the absorption of phosphorus in water was not subject to its suppression, and the phosphorus could be accumulated in the algae in the latter stages.
5. The feedback and induction of intact algal solution to L7
Adding intact algal solution of Microcystis aeruginosa into the culture process of L7 would help improve the aligicidal effect of algicidal components to Microcystis aeruginosa, but there was no significant difference among different growth cycles of algae; With the addition of the intact algal solution of Microcystis aeruginosa increased, the algicidal effect of the algicidal components to Microcystis aeruginosa increased, showing the relationship of“dose-effect”.
引文
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