黄瓜连作障碍微生物修复研究
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摘要
黄瓜是设施蔬菜中的主要作物,但是随着种植年限的增加,黄瓜连作障碍问题日益显著,已成为黄瓜产业发展的主要限制因素之一。在黄瓜连作障碍中,最重要的就是黄瓜枯萎病的发生。本文主要研究利用微生态修复解决黄瓜连作障碍问题。本研究主要包括4个部分:1以连作三年的黄瓜为研究对象,采取根部土壤,进行微生物区系研究,观察连作后土壤微生物种群的变化,以及优势菌的类型,为微生物修复黄瓜连作障碍提供理论基础。2以本研究室筛选的抑菌效果优良的3株放线菌为生防菌,研究其对黄瓜枯萎菌的抑制作用和机理,通过盆栽试验和大田试验研究了生防菌对黄瓜连作障碍的防治效果,并探讨了不同处理土壤微生物变化。3利用本研究室筛选保藏的放线菌资源,以黄瓜枯萎病病原菌为靶标菌,筛选新的对黄瓜枯萎病具有拮抗作用的放线菌,研究拮抗机理,并对性状优良放线菌菌进行生理生化和分子鉴定。4以对黄瓜枯萎菌具有良好拮抗效果的放线菌为研究对象,研究对黄瓜连作障碍有主要影响作用的化感物质肉桂酸和对羟基苯甲酸降解效果好的菌种。主要得到以下结果:
1对温室连作黄瓜根区根表土壤微生物区系研究表明:
①连作黄瓜根区土壤细菌数量大幅度下降、真菌数量大幅度增加,其中病株细菌数量较健株减少16.9%,真菌数量增加56.1%;病株根区病原真菌茄病镰刀菌(Fusarium solani)及尖孢枝孢菌(Cladosporium oxysporum)的数量分别较健株增加了366.15%和2201.85%;B/F、A/F值病株较健株分别降低了46.8%和36.8%,微生物由“细菌型”向“真菌型”过渡。②黄瓜根系染病溃烂导致黄瓜根表土壤细菌、真菌和放线菌数量急剧增加,病株根表细菌、真菌及放线菌数量分别是健株的56.3倍、7.9倍及2.7倍。③黄瓜病健株根区土壤养分存在差异,病株根区速效磷钾含量较健株低,说明速效磷钾含量与黄瓜枯萎病发生有一定关系。
对优势菌通过形态和分子手段鉴定最终为:B1为荧光假单胞菌(Pseudomonas fluorescens),B3为绿针假单胞菌(Pseudomonas chlororaphis),F1为茄病镰刀菌(Fusarium solani),F2为毛壳属菌(Chaetomium),F4为尖孢枝孢(Cladosporium oxysporum),F7为波兰青霉(Penicillium.polonicum),F9为甜瓜枯萎病(Fusarium oxysporum f. sp. Melonis),F11为刀孢轮枝菌(Lecanicillium psalliotae),A1为绿色产色链霉菌(Streptomyces viridochromogenes),A3为多产色链霉菌(Streptomyces polychromogenes),A4为加德那链霉菌(Streptomyces gardneri),A 5为高加索山链霉菌(Streptomyces ciscaucasicu)。
2对研究室筛选的3株优秀放线菌对黄瓜枯萎病防治研究表明:拮抗和机理试验表明:拮抗试验表明,3株放线菌对2种病原菌都有拮抗作用,而且拮抗性强弱为Act12>Act11>Act1,其中Act12的拮抗性效果最好;抑菌试验表明,
3株放线菌发酵液对黄瓜枯萎病病原菌生长具有抑制作用,抑菌强弱也表现为Act12>Act11>Act1。并且发现不同浓度的发酵液对黄瓜的发芽率几乎没有影响,但是对胚根的生长有明显的促进作用。对3株放线菌发酵液的活性物质进行HPLC测定,结果表明,放线菌Act12、Act11的指纹色谱比较相近,而且通过与生长素、细胞分裂素和赤霉素3种生长因子的色谱图比较,可以发现,3种放线菌都含有这些物质,这为3株放线菌能够促进黄瓜生长和抑制枯萎病提供了理论基础。
盆栽试验研究了3种生防放线菌剂单接和与病原菌混接对黄瓜的促生作用及对黄瓜叶片PPO酶活性的影响。结果表明:①供试放线菌制剂对黄瓜幼苗有显著的促生作用,在接种量为1.5 g/kg、20d时,菌剂Act12(密旋链霉菌,Streptomyces pactum)单独接种处理后,黄瓜植株总重量、根系鲜重、根系长度、根系活力及叶面积分别较对照CK提高18.8 %、225.0 %、57.3 %、482.0 %及45.3 %;Act12与黄瓜枯萎菌HK混合接种时,其各对应指标测值分别较对照HK提高78.9 %、114.6 %、162.6 %、443.4 %及50.0 %。②施用菌剂后对叶片诱导酶PPO活性有一定提高,45d时,单接Act12(1.50 g/kg)较对照增加了54.4 %。③接种放线菌剂对黄瓜幼苗叶片绿色度SPDA无明显影响。
大田试验研究了生防菌剂Act11和Act12结合有机肥,通过育苗和定植两种试验方案对连作黄瓜的影响,为应用放线菌剂防治黄瓜连作障碍提供理论依据。结果表明,黄瓜生长前期:向营养基质中接种ARMER或移栽时穴施接种ARMER及育苗移栽两次接种ARMER可以显著促进黄瓜幼苗生长,提高产量和黄瓜Vc含量,改善连作黄瓜的生长状况,对霜霉病也有一定控制作用,有机肥与ARMER混合使用也有类似效果;黄瓜的生物量、主蔓长度、单节长度、结果数和产量几乎都明显大于对照,尤其对黄瓜的根部生长有明显的促进作用,而且在生长后期有机肥施用后作用更加明显,施入菌剂处理后,黄瓜根区、根表内的微生物区系组成正常,处于健康状态。
3新的拮抗放线菌的筛选研究表明:
通过拮抗试验从600多株放线菌中筛选出72株拮抗效果较好的放线菌,在此基础上通过抑菌率试验筛选出12株放线菌,其中对黄瓜枯萎病的中的其中一株(从黄瓜枯萎病病株中分离的,已经过分子鉴定),效果突出,放线菌编号为D141,拮抗试验中D141对病原菌的抑菌圈直径达21mm;生长速率法试验中,D141无菌发酵液的抑菌作用最强,在72 h内抑菌率保持在92 %以上;搭片试验也表明D141对病原菌菌丝具有溶解作用。经过生理生化试验和分子鉴定确定为:球孢链霉菌球孢亚种(Streptomyces globisporus subsp. globisporus)。
编号为D220,D154和D141的3株放线菌,对黄瓜枯萎病两种病原菌效果均明显,而且是从连作黄瓜土壤中分离出的,具有很好的开发利用前景。经过菌落形态、生理生化、C源利用和分子鉴定最终确定此3株菌分别为:D220为绿淀粉酶链霉菌(Streptomyces viridodiastaticus),D154为孔雀石刺链霉菌(Streptomycesmalachitospinus),D141为球孢链霉菌球孢亚种(S. globisporus subsp. globisporus)。
4黄瓜化感物质降解菌的筛选试验表明:
以对羟基苯甲酸和肉桂酸作为黄瓜的2种主要化感物质,筛选对黄瓜化感物质具有降解能力的优良菌。结果表明:不同浓度的对羟基苯甲酸和肉桂酸对黄瓜枯萎病病原菌生长都有促进作用。其中菌株49、D163、ZY114和8的对对羟基苯甲酸和肉桂酸的降解能力比较好,尤其是两种酚酸类物质在浓度较高时,降解率达到90%左右。
Cucumber is a main cultivation vegetable, but as the increasing of cultiving time, continuous cropping obstacles of cucumber is becoming serious problem and limiting the development of cucumber industry. The incidence of Fusarium oxysporum of cucumber is the most serious one. This paper mainly discussed the solution to solve the problem by microbial way. The paper included four parts, the first part studied the microbial ecology mechanism including the change of microbial populations and the type of dominant microbes from rhizosphere of 3 years’cultivation to provide a theoretical basis of control of F. oxysporum by microorganisms. The second part studied the mechanism of three excellent kinds of actionmyces against F. oxysporum , the practical effect of actionmyces by pot and field experiment and the change of microbial populations by using actionmyces. Third: to select new actionmyces which against F. oxysporum from the laboratory actinomycetes resources, and studied the antagonistic mechanism. Also, effective actionmyces were identificated by physiological and biochemical as well as molecular method. Four: searching better actionmyces which can degradate cinnamic acid and P-hydroxy benzoic acid.The result is as following:
1 the study of microbial ecology showed :
The analysis of microbial ecology showed :①The number of bacteria decreased and the number of fungi increased. The number of bacteria decreased by 16.9% while fungi increased by 56.1%, compared with healthy cucumber plants, of which Fusarium solani and Cladosporium oxysporum increased by 366.15 % and 2 201.85 % in infected cucumber plants. The B/F、A/F value of the healthy cucumber decreased by 46.8 %、36.8 % compared with the infected.②The microorgnism quantities in root surface were increased for the ulcreation of cucumber root in infected cucumber plants, the number of bacteria、fungi and actinomycetes were 56.3 times、7.9 times and 2.7 times respectively as many as the number of healthy cucumber plants.③the content of available P and K in rhizosphere of infected cucumber plants decreased by 16.3 %、16.8 % compared with the healthy cucumber. There are no significant difference between healthy and infected cucumber on the value of available N, organic matter, soluble salt and pH. It indicated the content of available P and K had relationship with incidence of cucumber F. oxysporum.
The identified result of dominant microbies: the strain B1 was similar to the Pseudomonas fluorescens, B3 was the Pseudomonas chlororaphis, F1 was the Fusarium solani,F2 was the Chaetomium,F4 was the Cladosporium oxysporum,F7 was the Penicillium polonicum,F9 was the Fusarium oxysporum f. sp. Melonis,F11 was the Lecanicillium psalliotae,A1 was the Streptomyces viridochromogenes,A3 was the Streptomyces polychromogenes,A4 was the Streptomyces gardneri and A5 was the Streptomyces ciscaucasicu。
2 the study of the three excellent kinds of actionmyces against F. wilt showed: Antagonistic test showed that 3 strains of actinomycetes all had antagonistic effects on the two kinds of fusarium wilt, and antagonistic effects of the strain Act12 was the best, strain Act11 was better than strain Act1.
Inhibition test showed that the three actinomycetes had fermentation on the F. oxysporum; the effect of strain Act12, Act11 and Act1 was as the same as Antagonistic test. Different concentrations of fermented extracts can obviously promote radicle growth, but almost no effect on germination rate of cucumber seed. Determined activity of the substances of fermented extracts by HPLC instrument and compared to the chromatograms of auxin, cytokinin, and gibberellin, strain Act12 and Act11 had the similar chromatograms ,and the 3 actinomyces strains chromatograms inclued the three kinds of growth factors. That may be the reason that the three actinomycetes can promote cucumber growth..
The pot experiment showed the effect of three strains of antimicrobial actinomyces on growth of cucumber and PPO (Polyphenol Oxidase) activity of cucumber leaves. The results were:①The growth of cucumber was promoted by three strains of antimicrobial actinomyces, specifically the biomass, root fresh weight, root length, PPO activity and leaf area increased respectively by 18.8%, 225.0%, 57.3%, 482.0% and 45.3% compared with the control treatment when only Act12 was inoculated 20 days with the inoculation concentration 1.5g/kg. Meanwhile, the biomass, root fresh weight, root length, PPO activity and leaf area rose respectively by 78.9%, 114.6%, 162.6%, 443.4% and 50.0% compared with the HK treatment when Act12 was inoculated with the pathogen.②The PPO activity of cucumber leaves was promoted significantly by the three strains of antimicrobial actinomyces, to be more exact, the PPO activity increased 54.4 % compared with the control treatment when only Act12 was inoculated 45 days with the inoculation concentration 1.5g/kg.③The three actinomyces had no significantly effect on SPAD of cucumber leaves.
The field experiment studied the effect of the treatment of strain Act11 and Act12 combined with organic fertilizer on cropping cucumber, by the nursery and planting pilot treatment, in order to provide the theory of actinomycetes control cucumber continuous cropping obstacle. The results showed that inoculated ARMER (Actinomycetes of Rhizosphere Micro Ecosystem Restoration) can significantly promote cucumber seedling growth, productivity and content of Vc, when nurserying and transplanting cucumber in the early growth of cucumber. And also ARMER had a positive effect on downy mildew. The treatment of organic fertilizer mixed with ARMER also had the similar effect. In the later growth of cucumber, ARMER and fertilizer also played a significant role. Cucumber biomass, main vine length, single-length, fruit number and yield of almost all significantly higher than the control, especially root, the effect of fertilizer was more significant.
3 New actinomycetes selecting showed :
72 strains of actinomycetes had selected from more than 600 strains by antagonizing test, based on this test 12 strains of actinomycetes were selected again by inhibition test. One of the strain, the strain D141, had prominent effect on one of F. oxysporum in which isolated from diseased cucumber and has been molecular identificated. Using D141 strain, the diameter of pathogen inhibition zone was 21mm in antagonistic test and inhibitory rate remained more than 92% in inhibition test, the patch test also showed that D141 strain can dissolve the mycelia. According to the physiological and biochemical properties and 16SrRNA sequence analysis , D141 strain was similar to the Streptomyces globisporus subsp. Globisporus.
3 actinomycetes strains, D220, D154 and D141 were seleted for having obviously effects to F. oxysporum pathogen and were isolated from contiouns cropping cucumber soil, which had positive prospects for the development and utilization. According to the morphological, cultural characteristics, the physiological and biochemical properties, C source utilization and 16SrRNA sequence analysis, the strain D220 was similar to the Streptomyces viridodiastaticus,D154 was similar to the Streptomyces malachitospinus,D141 was similar to the S. globisporus subsp. Globisporus.
4 Cucumber allelochemicals degrading actinomycetes test showed:
By cinnamic acid and P-hydroxy benzoic acid, two main types of cucumber allelochemicals, the degradation capability was tested. The results showed that: different concentrations of the cinnamic acid and P-hydroxy benzoic acid can inhibited the growth of F. oxysporum. The result showed: the strains 49, D163, ZY114 and 8 played better than other strains, especially the two phenolic acids were in high concentrations, the degradation rate can reach to 90%.
1对温室连作黄瓜根区根表土壤微生物区系研究表明:
①连作黄瓜根区土壤细菌数量大幅度下降、真菌数量大幅度增加,其中病株细菌数量较健株减少16.9%,真菌数量增加56.1%;病株根区病原真菌茄病镰刀菌(Fusarium solani)及尖孢枝孢菌(Cladosporium oxysporum)的数量分别较健株增加了366.15%和2201.85%;B/F、A/F值病株较健株分别降低了46.8%和36.8%,微生物由“细菌型”向“真菌型”过渡。②黄瓜根系染病溃烂导致黄瓜根表土壤细菌、真菌和放线菌数量急剧增加,病株根表细菌、真菌及放线菌数量分别是健株的56.3倍、7.9倍及2.7倍。③黄瓜病健株根区土壤养分存在差异,病株根区速效磷钾含量较健株低,说明速效磷钾含量与黄瓜枯萎病发生有一定关系。
对优势菌通过形态和分子手段鉴定最终为:B1为荧光假单胞菌(Pseudomonas fluorescens),B3为绿针假单胞菌(Pseudomonas chlororaphis),F1为茄病镰刀菌(Fusarium solani),F2为毛壳属菌(Chaetomium),F4为尖孢枝孢(Cladosporium oxysporum),F7为波兰青霉(Penicillium.polonicum),F9为甜瓜枯萎病(Fusarium oxysporum f. sp. Melonis),F11为刀孢轮枝菌(Lecanicillium psalliotae),A1为绿色产色链霉菌(Streptomyces viridochromogenes),A3为多产色链霉菌(Streptomyces polychromogenes),A4为加德那链霉菌(Streptomyces gardneri),A 5为高加索山链霉菌(Streptomyces ciscaucasicu)。
2对研究室筛选的3株优秀放线菌对黄瓜枯萎病防治研究表明:拮抗和机理试验表明:拮抗试验表明,3株放线菌对2种病原菌都有拮抗作用,而且拮抗性强弱为Act12>Act11>Act1,其中Act12的拮抗性效果最好;抑菌试验表明,
3株放线菌发酵液对黄瓜枯萎病病原菌生长具有抑制作用,抑菌强弱也表现为Act12>Act11>Act1。并且发现不同浓度的发酵液对黄瓜的发芽率几乎没有影响,但是对胚根的生长有明显的促进作用。对3株放线菌发酵液的活性物质进行HPLC测定,结果表明,放线菌Act12、Act11的指纹色谱比较相近,而且通过与生长素、细胞分裂素和赤霉素3种生长因子的色谱图比较,可以发现,3种放线菌都含有这些物质,这为3株放线菌能够促进黄瓜生长和抑制枯萎病提供了理论基础。
盆栽试验研究了3种生防放线菌剂单接和与病原菌混接对黄瓜的促生作用及对黄瓜叶片PPO酶活性的影响。结果表明:①供试放线菌制剂对黄瓜幼苗有显著的促生作用,在接种量为1.5 g/kg、20d时,菌剂Act12(密旋链霉菌,Streptomyces pactum)单独接种处理后,黄瓜植株总重量、根系鲜重、根系长度、根系活力及叶面积分别较对照CK提高18.8 %、225.0 %、57.3 %、482.0 %及45.3 %;Act12与黄瓜枯萎菌HK混合接种时,其各对应指标测值分别较对照HK提高78.9 %、114.6 %、162.6 %、443.4 %及50.0 %。②施用菌剂后对叶片诱导酶PPO活性有一定提高,45d时,单接Act12(1.50 g/kg)较对照增加了54.4 %。③接种放线菌剂对黄瓜幼苗叶片绿色度SPDA无明显影响。
大田试验研究了生防菌剂Act11和Act12结合有机肥,通过育苗和定植两种试验方案对连作黄瓜的影响,为应用放线菌剂防治黄瓜连作障碍提供理论依据。结果表明,黄瓜生长前期:向营养基质中接种ARMER或移栽时穴施接种ARMER及育苗移栽两次接种ARMER可以显著促进黄瓜幼苗生长,提高产量和黄瓜Vc含量,改善连作黄瓜的生长状况,对霜霉病也有一定控制作用,有机肥与ARMER混合使用也有类似效果;黄瓜的生物量、主蔓长度、单节长度、结果数和产量几乎都明显大于对照,尤其对黄瓜的根部生长有明显的促进作用,而且在生长后期有机肥施用后作用更加明显,施入菌剂处理后,黄瓜根区、根表内的微生物区系组成正常,处于健康状态。
3新的拮抗放线菌的筛选研究表明:
通过拮抗试验从600多株放线菌中筛选出72株拮抗效果较好的放线菌,在此基础上通过抑菌率试验筛选出12株放线菌,其中对黄瓜枯萎病的中的其中一株(从黄瓜枯萎病病株中分离的,已经过分子鉴定),效果突出,放线菌编号为D141,拮抗试验中D141对病原菌的抑菌圈直径达21mm;生长速率法试验中,D141无菌发酵液的抑菌作用最强,在72 h内抑菌率保持在92 %以上;搭片试验也表明D141对病原菌菌丝具有溶解作用。经过生理生化试验和分子鉴定确定为:球孢链霉菌球孢亚种(Streptomyces globisporus subsp. globisporus)。
编号为D220,D154和D141的3株放线菌,对黄瓜枯萎病两种病原菌效果均明显,而且是从连作黄瓜土壤中分离出的,具有很好的开发利用前景。经过菌落形态、生理生化、C源利用和分子鉴定最终确定此3株菌分别为:D220为绿淀粉酶链霉菌(Streptomyces viridodiastaticus),D154为孔雀石刺链霉菌(Streptomycesmalachitospinus),D141为球孢链霉菌球孢亚种(S. globisporus subsp. globisporus)。
4黄瓜化感物质降解菌的筛选试验表明:
以对羟基苯甲酸和肉桂酸作为黄瓜的2种主要化感物质,筛选对黄瓜化感物质具有降解能力的优良菌。结果表明:不同浓度的对羟基苯甲酸和肉桂酸对黄瓜枯萎病病原菌生长都有促进作用。其中菌株49、D163、ZY114和8的对对羟基苯甲酸和肉桂酸的降解能力比较好,尤其是两种酚酸类物质在浓度较高时,降解率达到90%左右。
Cucumber is a main cultivation vegetable, but as the increasing of cultiving time, continuous cropping obstacles of cucumber is becoming serious problem and limiting the development of cucumber industry. The incidence of Fusarium oxysporum of cucumber is the most serious one. This paper mainly discussed the solution to solve the problem by microbial way. The paper included four parts, the first part studied the microbial ecology mechanism including the change of microbial populations and the type of dominant microbes from rhizosphere of 3 years’cultivation to provide a theoretical basis of control of F. oxysporum by microorganisms. The second part studied the mechanism of three excellent kinds of actionmyces against F. oxysporum , the practical effect of actionmyces by pot and field experiment and the change of microbial populations by using actionmyces. Third: to select new actionmyces which against F. oxysporum from the laboratory actinomycetes resources, and studied the antagonistic mechanism. Also, effective actionmyces were identificated by physiological and biochemical as well as molecular method. Four: searching better actionmyces which can degradate cinnamic acid and P-hydroxy benzoic acid.The result is as following:
1 the study of microbial ecology showed :
The analysis of microbial ecology showed :①The number of bacteria decreased and the number of fungi increased. The number of bacteria decreased by 16.9% while fungi increased by 56.1%, compared with healthy cucumber plants, of which Fusarium solani and Cladosporium oxysporum increased by 366.15 % and 2 201.85 % in infected cucumber plants. The B/F、A/F value of the healthy cucumber decreased by 46.8 %、36.8 % compared with the infected.②The microorgnism quantities in root surface were increased for the ulcreation of cucumber root in infected cucumber plants, the number of bacteria、fungi and actinomycetes were 56.3 times、7.9 times and 2.7 times respectively as many as the number of healthy cucumber plants.③the content of available P and K in rhizosphere of infected cucumber plants decreased by 16.3 %、16.8 % compared with the healthy cucumber. There are no significant difference between healthy and infected cucumber on the value of available N, organic matter, soluble salt and pH. It indicated the content of available P and K had relationship with incidence of cucumber F. oxysporum.
The identified result of dominant microbies: the strain B1 was similar to the Pseudomonas fluorescens, B3 was the Pseudomonas chlororaphis, F1 was the Fusarium solani,F2 was the Chaetomium,F4 was the Cladosporium oxysporum,F7 was the Penicillium polonicum,F9 was the Fusarium oxysporum f. sp. Melonis,F11 was the Lecanicillium psalliotae,A1 was the Streptomyces viridochromogenes,A3 was the Streptomyces polychromogenes,A4 was the Streptomyces gardneri and A5 was the Streptomyces ciscaucasicu。
2 the study of the three excellent kinds of actionmyces against F. wilt showed: Antagonistic test showed that 3 strains of actinomycetes all had antagonistic effects on the two kinds of fusarium wilt, and antagonistic effects of the strain Act12 was the best, strain Act11 was better than strain Act1.
Inhibition test showed that the three actinomycetes had fermentation on the F. oxysporum; the effect of strain Act12, Act11 and Act1 was as the same as Antagonistic test. Different concentrations of fermented extracts can obviously promote radicle growth, but almost no effect on germination rate of cucumber seed. Determined activity of the substances of fermented extracts by HPLC instrument and compared to the chromatograms of auxin, cytokinin, and gibberellin, strain Act12 and Act11 had the similar chromatograms ,and the 3 actinomyces strains chromatograms inclued the three kinds of growth factors. That may be the reason that the three actinomycetes can promote cucumber growth..
The pot experiment showed the effect of three strains of antimicrobial actinomyces on growth of cucumber and PPO (Polyphenol Oxidase) activity of cucumber leaves. The results were:①The growth of cucumber was promoted by three strains of antimicrobial actinomyces, specifically the biomass, root fresh weight, root length, PPO activity and leaf area increased respectively by 18.8%, 225.0%, 57.3%, 482.0% and 45.3% compared with the control treatment when only Act12 was inoculated 20 days with the inoculation concentration 1.5g/kg. Meanwhile, the biomass, root fresh weight, root length, PPO activity and leaf area rose respectively by 78.9%, 114.6%, 162.6%, 443.4% and 50.0% compared with the HK treatment when Act12 was inoculated with the pathogen.②The PPO activity of cucumber leaves was promoted significantly by the three strains of antimicrobial actinomyces, to be more exact, the PPO activity increased 54.4 % compared with the control treatment when only Act12 was inoculated 45 days with the inoculation concentration 1.5g/kg.③The three actinomyces had no significantly effect on SPAD of cucumber leaves.
The field experiment studied the effect of the treatment of strain Act11 and Act12 combined with organic fertilizer on cropping cucumber, by the nursery and planting pilot treatment, in order to provide the theory of actinomycetes control cucumber continuous cropping obstacle. The results showed that inoculated ARMER (Actinomycetes of Rhizosphere Micro Ecosystem Restoration) can significantly promote cucumber seedling growth, productivity and content of Vc, when nurserying and transplanting cucumber in the early growth of cucumber. And also ARMER had a positive effect on downy mildew. The treatment of organic fertilizer mixed with ARMER also had the similar effect. In the later growth of cucumber, ARMER and fertilizer also played a significant role. Cucumber biomass, main vine length, single-length, fruit number and yield of almost all significantly higher than the control, especially root, the effect of fertilizer was more significant.
3 New actinomycetes selecting showed :
72 strains of actinomycetes had selected from more than 600 strains by antagonizing test, based on this test 12 strains of actinomycetes were selected again by inhibition test. One of the strain, the strain D141, had prominent effect on one of F. oxysporum in which isolated from diseased cucumber and has been molecular identificated. Using D141 strain, the diameter of pathogen inhibition zone was 21mm in antagonistic test and inhibitory rate remained more than 92% in inhibition test, the patch test also showed that D141 strain can dissolve the mycelia. According to the physiological and biochemical properties and 16SrRNA sequence analysis , D141 strain was similar to the Streptomyces globisporus subsp. Globisporus.
3 actinomycetes strains, D220, D154 and D141 were seleted for having obviously effects to F. oxysporum pathogen and were isolated from contiouns cropping cucumber soil, which had positive prospects for the development and utilization. According to the morphological, cultural characteristics, the physiological and biochemical properties, C source utilization and 16SrRNA sequence analysis, the strain D220 was similar to the Streptomyces viridodiastaticus,D154 was similar to the Streptomyces malachitospinus,D141 was similar to the S. globisporus subsp. Globisporus.
4 Cucumber allelochemicals degrading actinomycetes test showed:
By cinnamic acid and P-hydroxy benzoic acid, two main types of cucumber allelochemicals, the degradation capability was tested. The results showed that: different concentrations of the cinnamic acid and P-hydroxy benzoic acid can inhibited the growth of F. oxysporum. The result showed: the strains 49, D163, ZY114 and 8 played better than other strains, especially the two phenolic acids were in high concentrations, the degradation rate can reach to 90%.
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