固氮蓝藻在松嫩平原盐碱土生态修复中作用的研究
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摘要
由于不合理的农田灌溉、过度使用化肥及植被破坏,土地盐碱化越来越严重,盐碱地的改良和修复工作也一直深受重视。本论文从生态工程学和盐碱地生态修复的角度出发,选择松嫩平原典型的盐碱化草地作为研究对象,考察了盐碱化草地土壤的理化特征及土壤微生物的分布规律;对盐碱土中土著蓝藻的形态、种属、生态分布规律以及酶活性进行了研究;考察了固氮蓝藻Nostoc commune和Anabaena azotica Ley在不同浓度Na2CO3培养液环境下的适应能力;研究了含水率、接种量以及土壤深度在Nostoc commune和Anabaena azotica Ley对盐碱土改性作用中的的影响。在此基础上,考察了Anabaena azotica Ley对羊草的促生效果,探讨了作用机理,并初步研究了Anabaena azotica Ley对3种类型盐碱土土质的改善效果。
按照土壤生态学研究方法,在黑龙江省安达市典型盐碱化草地中,分别从重度、中度和轻度盐碱土壤中选择3块100×100m2大方作为研究对象,采用梅花布点法采样,分析了土壤样品的Na+、K+、Ca2+、Mg2+等无机离子含量,SO42-、Cl-、NO3-等阴离子浓度以及EC、pH等。2006年按季节分12个月,采集土壤样品,采用MPN法分析了土壤中细菌、放线菌、真菌、藻类含量。研究结果表明:3种类型土壤的理化特征和微生物构成各有其特点,轻度盐碱土为非碱性钠质土,中度和重度盐碱土属于碱性钠质土,重度盐碱化区土壤氮含量严重缺乏;微生物数量呈现明显的季节变化,随着土壤盐碱化程度的增加,细菌、真菌、放线菌数量逐渐降低,而重度盐碱土中放线菌处于优势地位,微生物数量差异达到显著或极显著的水平。
利用BG110培养基从黑龙江省安达市典型盐碱化草地中分离筛选出了20株具有一定耐盐能力的固氮蓝藻,它们分别属于10个属,轻度盐碱土和重度盐碱土上各有3种蓝藻,而中度盐碱土上有4种蓝藻。其中Nostoc和Anabaena丰度最高,且可耐受15%的NaCl。
采用BG110培养基进行纯培养,分别测定了Nostoc commune和Anabaena azotica Ley在不同生长阶段的叶绿素a含量、胞外多糖含量、氨基酸含量和固氮酶活性。结果表明:两种固氮蓝藻都能在Na2CO3盐胁迫条件下生长,且随着Na2CO3盐胁迫性增加,两种蓝藻的叶绿素a含量、胞外多糖含量、氨基酸含量和固氮酶活性都呈现先增加后减少的趋势;而随着培养天数的增加,两种藻类叶绿素a的变化曲线与微生物的生长曲线很相似,胞外多糖和氨基酸的增长和叶绿素a的增长是同步的,其中Anabaena azotica Ley的固氮酶活性在培养到21天时达到最大值,Nostoc commune的固氮酶活性在培养到28天时达到最大值;比较而言Anabaena azotica Ley的耐盐性好于Nostoc commune的耐盐性,固氮能力也高于Nostoc commune。
考察了含水率对固氮蓝藻Anabaena azotica Ley和Nostoc commune改性盐碱土的作用效果。结果表明:含水率在30%,蓝藻已经可以形成藻结皮,而含水率在50%,蓝藻生长得更好。盆栽试验考察了接种量对固氮蓝藻Anabaena azotica Ley和Nostoc commune改性盐碱土的作用效果。结果表明:接种量为0.03mg干藻/cm2时,更适合固氮蓝藻Anabaena azotica Ley在轻度盐碱土上生长,而接种量为0.18 mg干藻/cm2时,更适合固氮蓝藻Nostoc commune在中度盐碱土上着生;分别测定2种固氮蓝藻在盆栽试验不同培养阶段(每10天为一个阶段)的阴阳离子、pH值、电导率、水溶性总盐、交换性钠、TOC、BAB和固氮酶活性的变化。结果表明:固氮蓝藻提高了土壤有机质530.01%,降低土壤pH 8.0以下,提高了土壤氮含量269.12%,降低了土壤交换性Na+ 86.28%。但当土壤深度达到9cm时,蓝藻对盐碱土的改性效果减弱。
以盐碱土典型植物—羊草作为考查对象,在实验室盆栽条件下分析了菌种的促生效果。接种一定时间后,测定羊草植株高度,发现接种Anabaena azotica Ley后,植株高度比对照增加30%左右,而且这种促进作用具有持续性。分析了接种菌种后羊草根际土壤固氮酶活的变化,接种固氮蓝藻能促进根际菌构成发生变化,进而影响植物的生长。结果表明:Anabaena azotica Ley不仅能适应盐碱土环境,而且能提高盐碱土的氮含量,改变根际微生物构成,促进植物生长。
现场实验表明,Anabaena azotica Ley在三种类型盐碱土上对温度、光照和碱度都有很好的生态适应性,而且可以积累土壤的有机质、转化无机盐、降低土壤的pH,使土壤氮含量提高106.99%。
本论文研究表明,固氮蓝藻可以改善土质,促进植物生长,这对于生态修复盐碱化土壤具有一定的作用。
Soil saline-alkalifying is becoming more and more severe because of the unsuitable irrigation, excess using of fertilizers and vegetation destroying. People have paid more attention on the soil improvement and remediation, including engineering, chemical and biological techniques. The engineering and chemical strategies are effective, but they are usually unefficient with high cost. Nowadays, the bio-remediation is mainly focused on the culture of alkaline-resistant plants and it appeared that the improvement has been work slowly for either the traditional or the genic method.
Songnen alkaline-saline plain was selected as research objective at present paper based on the view of ecological engineering and for the purpose of eco-remediation of alkaline-saline land. The physi-chemical properties and the microbe communities were investigated. Appraise the original cyanobacterias was conducted. Two species of nitrogen-fixing cyanobacteria belonging to two genera Nostoc commune and Anabaena azotica Ley tolerate of different concentrations of Na2CO3 in BG110 medium. Some experiments were carried on to investigate the effects of the sum of inoculation and the ratio of water in the soil towards the remediation of saline-alkaline soil which based on nitrogen-fixing cyanobacteria Nostoc commune and Anabaena azotica Ley. The effects of the Anabaena azotica Ley on the growth of Leymus chinensis were investigated by inoculation of the strains to laboratorial basin experiments. Ecological adaptability and repairing effect of Anabaena azotica Ley were studied in field experiments.
Three blocks of 100×100m2, which represents different saline-alkalifying extent of heavy, medium and common alkaline-saline land were choosed for the present study. The results showed that different physic-chemical characteristics and microbe structure in three types of soil. Light saline soil belongs to non-alkalinity sodium soils, middling saline soil and heavy saline soil belong to alkalinity sodium soil. Soil salinization related to soil physicochemical properties and microbial communities. The soil saline alkaline degree became larger amount of bacterium, fungi, actinomycetes and algae gradually decreased. The difference of microbial quantity reaches extremely significant level or significant level.
Twenty out of 200 isolates of cyanobacteria mainly from saline soils of Songnen Plain of China were successfully grown on BG11 N-free medium. However, at 5 % NaCl only 6 of the isolates exhibited a high rate of nitrogenase activity. The experiment of appraising the original cyanobacterias shows that there are ten kinds of cyanobacterias in the three typical areas. There are four kinds in medium alkaline-saline and three kinds in the other two kinds. We can conclude that medium alkaline-saline is more proper for the growth of cyanobacterias.
Two species of nitrogen-fixing cyanobacteria belonging to two genera Nostoc commune and Anabaena azotica Ley tolerate of different concentrations of Na2CO3 (from 0.2×10-4g/ml to 1.8×10-4g/ml) in BG110 medium. The variation in the chlorophyll (Chl) a content, extracellular saccharide , amino acid and nitrogenase activity are measured respectively. It is found that both of nitrogen-fixing cyanobacteria can thrive under Na2CO3 salt stress. When the concentration of Na2CO3 increase, the chlorophyll (Chl) a content,extracellular saccharide,amino acid and nitrogenase activity increase first and then decrease. With the increase of time of incubation, the change curve of chlorophyll (Chl) a content of Nostoc commune and Anabaena azotica Ley is similar to the growth curve of microorganism, extracellular saccharide and amino acid increase in a synchronism with chlorophyll (Chl) a, the maximum of nitrogenase activity appears at 21 days for Anabaena azotica Ley and appears at 28 days for Nostoc commune. Compare Anabaena azotica Ley with Nostoc commune, the former possesses better resistance to salt and higher nitrogen-fixing ability.
In the experiments which carried on in flowerpots we find that when the ratio of water in the soil is 30%, the cyanobacteria can grow normally, when the ratio is 50%, they can grow much better; when the sum of inoculation is 0.03mg dry cyanobacterias/cm2, it is more proper for Anabaena azotica Ley to be planted in common alkaline-saline land, when the sum is 0.18 mg dry cyanobacterias/cm2,it is better to plant Nostoc commune in the medium alkaline-saline land. We measure cation and antion, pH value, the ratio of conducter、the total sum of dissolved salt, the exchange-able Na+, TOC at every periods (ten days is a period) and measure BAB and nitrogenase activity at the last period. The results showed that the inoculation of nitrogen-fixing cyanobacteria can increase the organic matter by 530.01% in the soil effectively, decreasing soil pH, increasing soil nitrogen content by 269.12% and decreasing soil exchangeable sodium by 86.28%. But when the depth was up to 9cm, the function is becoming weaker. Taking the typical plant of Leymus chinensis as target research objective, the effects of inoculation of Anabaena azotica Ley into laboratorial potted plant were investigated by measuring the heights of Leymus chinensis at different periods. After inoculation, the heights of Leymus chinensis were 30% higher than that of controlled and the function were lasted in the whole experimental periods. The nitrogenase of rhizomes was also determined and results implied the changes of nitrogenase might be owing to inoculation of Anabaena azotica Ley. This might be one reason resulting in the growth of Leymus chinensis. Strains of alkaline-saline resistant cyanobacteria can not only adapt to alkaline-saline soil environment, but also can improve the content of nitrogen.
The experiment results onsite have shown that the Anabaena azotica Ley have a good ecological adaptability with temperature, light and alkalinity, but also accumulate soil organic matter, transform inorganic salts, fixation of atmospheric nitrogen, decrease soil pH and increase soil nitrogen content 106.99%.
The results showed that nitrogen fixing cyanobacteria can improve the soil quality, simultaneouly promote plant growth. Thus, the cyanobacteria were important and effective in ecological restoration of saline soils.
按照土壤生态学研究方法,在黑龙江省安达市典型盐碱化草地中,分别从重度、中度和轻度盐碱土壤中选择3块100×100m2大方作为研究对象,采用梅花布点法采样,分析了土壤样品的Na+、K+、Ca2+、Mg2+等无机离子含量,SO42-、Cl-、NO3-等阴离子浓度以及EC、pH等。2006年按季节分12个月,采集土壤样品,采用MPN法分析了土壤中细菌、放线菌、真菌、藻类含量。研究结果表明:3种类型土壤的理化特征和微生物构成各有其特点,轻度盐碱土为非碱性钠质土,中度和重度盐碱土属于碱性钠质土,重度盐碱化区土壤氮含量严重缺乏;微生物数量呈现明显的季节变化,随着土壤盐碱化程度的增加,细菌、真菌、放线菌数量逐渐降低,而重度盐碱土中放线菌处于优势地位,微生物数量差异达到显著或极显著的水平。
利用BG110培养基从黑龙江省安达市典型盐碱化草地中分离筛选出了20株具有一定耐盐能力的固氮蓝藻,它们分别属于10个属,轻度盐碱土和重度盐碱土上各有3种蓝藻,而中度盐碱土上有4种蓝藻。其中Nostoc和Anabaena丰度最高,且可耐受15%的NaCl。
采用BG110培养基进行纯培养,分别测定了Nostoc commune和Anabaena azotica Ley在不同生长阶段的叶绿素a含量、胞外多糖含量、氨基酸含量和固氮酶活性。结果表明:两种固氮蓝藻都能在Na2CO3盐胁迫条件下生长,且随着Na2CO3盐胁迫性增加,两种蓝藻的叶绿素a含量、胞外多糖含量、氨基酸含量和固氮酶活性都呈现先增加后减少的趋势;而随着培养天数的增加,两种藻类叶绿素a的变化曲线与微生物的生长曲线很相似,胞外多糖和氨基酸的增长和叶绿素a的增长是同步的,其中Anabaena azotica Ley的固氮酶活性在培养到21天时达到最大值,Nostoc commune的固氮酶活性在培养到28天时达到最大值;比较而言Anabaena azotica Ley的耐盐性好于Nostoc commune的耐盐性,固氮能力也高于Nostoc commune。
考察了含水率对固氮蓝藻Anabaena azotica Ley和Nostoc commune改性盐碱土的作用效果。结果表明:含水率在30%,蓝藻已经可以形成藻结皮,而含水率在50%,蓝藻生长得更好。盆栽试验考察了接种量对固氮蓝藻Anabaena azotica Ley和Nostoc commune改性盐碱土的作用效果。结果表明:接种量为0.03mg干藻/cm2时,更适合固氮蓝藻Anabaena azotica Ley在轻度盐碱土上生长,而接种量为0.18 mg干藻/cm2时,更适合固氮蓝藻Nostoc commune在中度盐碱土上着生;分别测定2种固氮蓝藻在盆栽试验不同培养阶段(每10天为一个阶段)的阴阳离子、pH值、电导率、水溶性总盐、交换性钠、TOC、BAB和固氮酶活性的变化。结果表明:固氮蓝藻提高了土壤有机质530.01%,降低土壤pH 8.0以下,提高了土壤氮含量269.12%,降低了土壤交换性Na+ 86.28%。但当土壤深度达到9cm时,蓝藻对盐碱土的改性效果减弱。
以盐碱土典型植物—羊草作为考查对象,在实验室盆栽条件下分析了菌种的促生效果。接种一定时间后,测定羊草植株高度,发现接种Anabaena azotica Ley后,植株高度比对照增加30%左右,而且这种促进作用具有持续性。分析了接种菌种后羊草根际土壤固氮酶活的变化,接种固氮蓝藻能促进根际菌构成发生变化,进而影响植物的生长。结果表明:Anabaena azotica Ley不仅能适应盐碱土环境,而且能提高盐碱土的氮含量,改变根际微生物构成,促进植物生长。
现场实验表明,Anabaena azotica Ley在三种类型盐碱土上对温度、光照和碱度都有很好的生态适应性,而且可以积累土壤的有机质、转化无机盐、降低土壤的pH,使土壤氮含量提高106.99%。
本论文研究表明,固氮蓝藻可以改善土质,促进植物生长,这对于生态修复盐碱化土壤具有一定的作用。
Soil saline-alkalifying is becoming more and more severe because of the unsuitable irrigation, excess using of fertilizers and vegetation destroying. People have paid more attention on the soil improvement and remediation, including engineering, chemical and biological techniques. The engineering and chemical strategies are effective, but they are usually unefficient with high cost. Nowadays, the bio-remediation is mainly focused on the culture of alkaline-resistant plants and it appeared that the improvement has been work slowly for either the traditional or the genic method.
Songnen alkaline-saline plain was selected as research objective at present paper based on the view of ecological engineering and for the purpose of eco-remediation of alkaline-saline land. The physi-chemical properties and the microbe communities were investigated. Appraise the original cyanobacterias was conducted. Two species of nitrogen-fixing cyanobacteria belonging to two genera Nostoc commune and Anabaena azotica Ley tolerate of different concentrations of Na2CO3 in BG110 medium. Some experiments were carried on to investigate the effects of the sum of inoculation and the ratio of water in the soil towards the remediation of saline-alkaline soil which based on nitrogen-fixing cyanobacteria Nostoc commune and Anabaena azotica Ley. The effects of the Anabaena azotica Ley on the growth of Leymus chinensis were investigated by inoculation of the strains to laboratorial basin experiments. Ecological adaptability and repairing effect of Anabaena azotica Ley were studied in field experiments.
Three blocks of 100×100m2, which represents different saline-alkalifying extent of heavy, medium and common alkaline-saline land were choosed for the present study. The results showed that different physic-chemical characteristics and microbe structure in three types of soil. Light saline soil belongs to non-alkalinity sodium soils, middling saline soil and heavy saline soil belong to alkalinity sodium soil. Soil salinization related to soil physicochemical properties and microbial communities. The soil saline alkaline degree became larger amount of bacterium, fungi, actinomycetes and algae gradually decreased. The difference of microbial quantity reaches extremely significant level or significant level.
Twenty out of 200 isolates of cyanobacteria mainly from saline soils of Songnen Plain of China were successfully grown on BG11 N-free medium. However, at 5 % NaCl only 6 of the isolates exhibited a high rate of nitrogenase activity. The experiment of appraising the original cyanobacterias shows that there are ten kinds of cyanobacterias in the three typical areas. There are four kinds in medium alkaline-saline and three kinds in the other two kinds. We can conclude that medium alkaline-saline is more proper for the growth of cyanobacterias.
Two species of nitrogen-fixing cyanobacteria belonging to two genera Nostoc commune and Anabaena azotica Ley tolerate of different concentrations of Na2CO3 (from 0.2×10-4g/ml to 1.8×10-4g/ml) in BG110 medium. The variation in the chlorophyll (Chl) a content, extracellular saccharide , amino acid and nitrogenase activity are measured respectively. It is found that both of nitrogen-fixing cyanobacteria can thrive under Na2CO3 salt stress. When the concentration of Na2CO3 increase, the chlorophyll (Chl) a content,extracellular saccharide,amino acid and nitrogenase activity increase first and then decrease. With the increase of time of incubation, the change curve of chlorophyll (Chl) a content of Nostoc commune and Anabaena azotica Ley is similar to the growth curve of microorganism, extracellular saccharide and amino acid increase in a synchronism with chlorophyll (Chl) a, the maximum of nitrogenase activity appears at 21 days for Anabaena azotica Ley and appears at 28 days for Nostoc commune. Compare Anabaena azotica Ley with Nostoc commune, the former possesses better resistance to salt and higher nitrogen-fixing ability.
In the experiments which carried on in flowerpots we find that when the ratio of water in the soil is 30%, the cyanobacteria can grow normally, when the ratio is 50%, they can grow much better; when the sum of inoculation is 0.03mg dry cyanobacterias/cm2, it is more proper for Anabaena azotica Ley to be planted in common alkaline-saline land, when the sum is 0.18 mg dry cyanobacterias/cm2,it is better to plant Nostoc commune in the medium alkaline-saline land. We measure cation and antion, pH value, the ratio of conducter、the total sum of dissolved salt, the exchange-able Na+, TOC at every periods (ten days is a period) and measure BAB and nitrogenase activity at the last period. The results showed that the inoculation of nitrogen-fixing cyanobacteria can increase the organic matter by 530.01% in the soil effectively, decreasing soil pH, increasing soil nitrogen content by 269.12% and decreasing soil exchangeable sodium by 86.28%. But when the depth was up to 9cm, the function is becoming weaker. Taking the typical plant of Leymus chinensis as target research objective, the effects of inoculation of Anabaena azotica Ley into laboratorial potted plant were investigated by measuring the heights of Leymus chinensis at different periods. After inoculation, the heights of Leymus chinensis were 30% higher than that of controlled and the function were lasted in the whole experimental periods. The nitrogenase of rhizomes was also determined and results implied the changes of nitrogenase might be owing to inoculation of Anabaena azotica Ley. This might be one reason resulting in the growth of Leymus chinensis. Strains of alkaline-saline resistant cyanobacteria can not only adapt to alkaline-saline soil environment, but also can improve the content of nitrogen.
The experiment results onsite have shown that the Anabaena azotica Ley have a good ecological adaptability with temperature, light and alkalinity, but also accumulate soil organic matter, transform inorganic salts, fixation of atmospheric nitrogen, decrease soil pH and increase soil nitrogen content 106.99%.
The results showed that nitrogen fixing cyanobacteria can improve the soil quality, simultaneouly promote plant growth. Thus, the cyanobacteria were important and effective in ecological restoration of saline soils.
引文
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