Immunolocalization of a class III chitinase in two muskmelon cultivars reacting differently to Fusarium oxysporum f. sp. melonis

Fusarium oxysporum f. sp. melonis is a highly specialized fungus that attacks the root system of melon (Cucumis melo L.). In this work the presence of a class III chitinase was examined by immunological techniques in the root and stem base of a susceptible (cv. Galia) and a resistant (cv. Bredor) melon during the infection process. By immunolocalization it was not possible to detect the constitutive presence of class III chitinase in any of the cultivars. However, the immunolabelling appeared in the root tissues of both cultivars as a consequence of wounding and of infection by F. oxysporum f. sp. melonis. Distinct patterns of chitinase detection were observed in the roots of the two cultivars as the infection progressed. Furthermore, by western blotting distinct class III chitinase isoforms were detected, which responded differently to the F. oxysporum f. sp. melonis infection. Our results strongly indicate that a relationship exists between class III chitinase and melon resistance to Fusarium infection, and that the resistance is associated with certain isoforms of this enzyme.     

两株镰孢菌的鉴定及其粗毒素对甜瓜幼苗的化感作用

从甜瓜枯萎病发病植株及其根际病土中分离到8株真菌,以甜瓜品种“西甜1号”为材料,采用回接试验和种子萌发试验确定真菌分离物对甜瓜的致病性和生长抑制作用;通过测定盆栽幼苗根系诱导酶活性、抗性物质含量及细胞膜相对透性,研究两株有害真菌粗毒素对甜瓜幼苗的化感作用;根据形态学特征及ITS序列分析对两株有害真菌进行初步鉴定.结果表明： 两株有害真菌TF和HF粗毒素对甜瓜种子萌发以及胚轴、胚根生长具有抑制作用,可导致甜瓜幼苗根系丙二醛(MDA)含量、可溶性蛋白含量及细胞膜相对透性增加,其中TF粗毒素原液处理下甜瓜幼苗根系MDA含量和细胞膜相对透性较对照分别增加108.6%和40.6%.两株有害真菌粗毒素可提高甜瓜幼苗根系保护性诱导酶活性,其中,TF粗毒素10倍稀释液处理下根系苯丙氨酸解氨酶(PAL)和过氧化物酶(POD)活性较对照分别增加25.6%和23.2%;HF粗毒素原液处理下根系PAL活性较对照提高30.0%.两株有害真菌TF和HF初步鉴定为木贼镰孢菌(Fusarium equiseti)和层出镰孢菌(F. proliferatum).两株镰孢菌虽然不能通过回接侵染甜瓜,但能通过分泌毒素影响甜瓜正常生长和生理生化代谢,同时提高甜瓜根系保护性诱导酶活性,具有有害和有益双重作用,其化感危害是引起甜瓜连作障碍的主要原因之一.     

3种间作植物对薄皮甜瓜植株生长、营养吸收和果实品质的影响

为解决薄皮甜瓜生产中的连作障碍问题,设置薄皮甜瓜单作(MM)、茴香/薄皮甜瓜间作(FM)、分蘖洋葱/薄皮甜瓜间作(TM)、艾草/薄皮甜瓜间作(WM)4个处理,研究不同间作植物对薄皮甜瓜植株株高、茎粗、根系活力和矿质元素(N、P、K、Ca和Mg)含量,以及果实品质和产量的影响,以期筛选出具有间作优势的露地栽培薄皮甜瓜间作作物组合。结果表明: 随着间作栽培时间的延长,3个间作处理的薄皮甜瓜株高均显著高于单作对照;FM和TM处理的茎粗显著高于单作对照,WM处理的茎粗与单作对照差异不显著;在伸蔓期、坐果期和果实膨大期,FM处理的根系活力最高,显著高于单作对照和TM处理。3个间作处理薄皮甜瓜植株矿质元素含量均不同程度高于单作对照,且FM处理的矿质元素含量均较高。FM和TM间作处理单果重与单作对照差异不显著。FM处理果实品质指标不低于单作对照;WM和TM处理的部分果实品质指标(葡萄糖、果糖和蔗糖含量)低于单作对照。综上,小茴香较适合作为薄皮甜瓜的间作植物。     

Effect of genotype and graft type on the hydraulic characteristics and water relations of grafted melon

Abstract

From the measurements of the profiles of hydraulic conductance and water potential from soil through to the leaf system in fully established melon plants, the limits to water flow set by coupling of hydraulic conductance (k) with water relation parameters was evaluated in the laboratory using high pressure flow device (HPFM) and evaporative flux method (EF). The rootstock Arava was grafted onto self, and onto two genotypes (AR57 and AR82) using side and V graft types, and there was an ungrafted control. Hydraulic transport efficiency was estimated from measurements of evaporative flux (transpiration rate) and leaf water potential (ψL) measured between pre-dawn and sunset during the growth cycle. Measured parameters to characterize the hydraulic efficiency (architecture) of the vascular system of melon were normalized to areas of leaves and stem cross section; this enabled the examination of their physiological and ecological functions. The effects of rootstock genotype were more marked on graft union and scion water relations. Differences in the magnitudes of water relation parameters of hydraulic conductance, water potential (lwp) and evaporative water loss (EF) were detected. AR/RS82 side grafted exhibited high EF and Kh despite its lower leaf water potential compared to AR/RS57 V grafted. Self grafting (Arava/Arava grafts) in melon seems to improve water relations and xylem water transport efficiency. Parameters describing the hydraulic efficiency (architecture) of vascular system of melon plants were described in relation to plant attributes. The expression of hydraulic conductance of the root and shoot system relative to plant attributes did not eliminate differences in the magnitudes of conductance elements in tomato and melon. Differences obtained among the different melon grafts in whole plant leaf and stem area specific hydraulic conductance (Kl) indicate the carbon efficiency and hence the cost of resource allocation to areas of root surface and leaves. The role of plant water relations in root-shoot communications and whole plant regulation of water flux are inferred from this study.     

甜瓜自毒相关基因CmGST的克隆及其对自毒胁迫的响应

植物谷胱甘肽硫转移酶基因(GST)在清除生物和非生物胁迫产生的氧化损伤中扮演着重要的角色,为探究GST在根系分泌物介导的甜瓜自毒胁迫中的响应机制,该实验在转录组测序基础上,以甜瓜叶片cDNA为模板,采用RT PCR技术获得了一个根系分泌物介导的甜瓜自毒作用密切相关的GST基因,命名为CmGST(GenBank 登录号：AYU66762.1);对CmGST基因进行了相关生物信息学分析,同时对自毒胁迫过程中,CmGST的差异表达情况、谷胱甘肽硫转移酶(GST)活性和谷胱甘肽(GSH)含量进行了相关测定。结果显示：(1)CmGST基因含有完整开放阅读框(ORF)654 bp,编码217个氨基酸;CmGST蛋白为酸性亲水蛋白,且较为稳定,具有GST家族Tau亚家族的2个典型的保守结构域;CmGST蛋白与黄瓜的亲缘关系较近。(2)亚细胞定位结果表明CmGST定位于细胞质中。(3)qRT PCR分析结果表明,CmGST在甜瓜根系和幼苗中均有表达,在正常生长的甜瓜幼苗中表达量相对稳定;自毒胁迫后在叶中的表达先降后升,后期迅速增加;在根系中变化趋势类似,但程度较为缓和。(4)GST活性和GSH含量分析表明,叶片中GST活性变化趋势与CmGST基因表达变化基本吻合;在根系中,GST活性呈现先升再降最后又升高的趋势;随胁迫时间增加,GSH含量在根系和叶片中均都有不同程度的增加,但根部含量低于叶片。研究认为,在自毒胁迫过程中,CmGST基因在甜瓜植株根和叶片均有相应表达水平的变化,参与了对根系分泌物介导的自毒胁迫响应。     

Use of Cucumis metuliferus as a Rootstock for Melon to Manage Meloidogyne incognita

Root-knot nematode-susceptible melons (Cantaloupe) were grown in pots with varying levels of Meloidogyne incognita and were compared to susceptible melons that were grafted onto Cucumis metuliferus or Cucurbita moschata rootstocks. In addition, the effect of using melons as transplants in nematode-infested soil was compared to direct seeding of melons in nematode-infested soil. There were no differences in shoot or root weight, or severity of root galling between transplanted and direct-seeded non-grafted susceptible melon in nematode-infested soil. Susceptible melon grafted on C. moschata rootstocks had lower root gall ratings and, at high nematode densities, higher shoot weights than non-grafted susceptible melons. However, final nematode levels were not lower on the grafted than on the non-grafted plants, and it was therefore concluded that grafting susceptible melon on to C. moschata rootstock made the plants tolerant, but not resistant, to the nematodes. Grafting susceptible melons on C. metuliferus rootstocks also reduced levels of root galling, prevented shoot weight losses, and resulted in significantly lower nematode levels at harvest. Thus, C. metuliferus may be used as a rootstock for melon to prevent both growth reduction and a strong nematode buildup in M. incognita-infested soil.     

自毒胁迫对甜瓜种子萌发与幼苗保护酶活性和转录组的影响

甜瓜自毒作用是导致其栽培实践中连作障碍严重的关键原因之一。该实验以甜瓜植株水浸提液处理模拟甜瓜自毒胁迫,通过测定自毒条件下甜瓜种子萌发、幼苗根系保护酶活性和MDA含量的变化及转录组分析,以探讨甜瓜自毒作用机理。结果显示:(1)甜瓜自毒胁迫总体上抑制了甜瓜种子萌发和后续生长,胁迫处理的阈值为0.03 g/mL。(2)甜瓜植株水浸提液处理后其幼苗根系保护酶活性和MDA含量变化非常剧烈,SOD活性表现为降-升-降趋势,POD活性先降后升,CAT活性先升后降,MDA含量持续增加。(3)转录组分析结果共鉴定出2 599个差异表达基因(differentially expressed genes, DEGs),胁迫2 d后共产生2 251个DEGs,显著多于胁迫4 d后的329个DEGs;且2 d与0 d对比有923个DEGs上调,1 328个DEGs下调,表明甜瓜幼苗在基因水平对自毒胁迫产生了积极响应。(4)相关生物信息学分析表明,自毒胁迫导致的DEGs主要与苯丙烷代谢、活性氧代谢、光合作用和植物激素信号转导有关,同时也涉及到渗透调节、膜和蛋白保护等过程;且这些DEGs主要富集于2 d与0 d,说明自毒胁迫条件下幼苗光合作用发生了改变,且这种变化主要发生在胁迫早期。(5)对6个甜瓜自毒胁迫密切相关DEGs(AP2-2、bZIP1、bZIP2、AP2-1、bHLH和HIS)的qRT-PCR分析显示,AP2-1、bHLH和HIS等3个基因在自毒胁迫2 d时表达量出现峰值,对照组AP2-2基因在2 d时表达量达到峰值,结果与转录组测序分析结果一致。研究表明,甜瓜自毒胁迫引起了其植物细胞的异常,进而会对生长和器官结构产生不良影响并诱发了大量与刺激或胁迫相关的基因差异表达,且甜瓜自毒胁迫下短期响应的基因或转录因子数量明显多于长期响应的数量,暗示甜瓜幼苗可以对自毒胁迫做出快速响应。     

Biological Control of Meloidogyne arenaria by Pasteuria penetrans

An obligate parasitic bacterium of nematode, Pasteuria penetrans 98–35 (PP), isolated from oriental melon greenhouse soil in Korea, was evaluated against root-knot nematode, Meloidogyne arenaria (MA), in tomato (Lycopersicon esculentum Mill. var. Youngkwang) and oriental melon (Cucumis melo L. var. Eunchun). Pot experiments were conducted by planting the tomato seedlings in a medium inoculated with 5,000 MA juveniles/pot (J2), J2+100,000 PP endospores/1 g medium, and J2+200,000 PP endospores/1 g medium. After 10 weeks from planting, root gall percentage in J2+100,000 and J2+200,000 PP endospores/1 g medium were significantly lower with 37.5% and 6.7%, respectively, compared with the J2 of 85%. In the second planting of tomatoes in the same pots, root gall numbers were significantly lower in PP treated pots representing 68.8 and 31.4/root in J2+100,000 and J2+200,000 PP endospores/1 g medium, respectively, compared with the J2 of 460.6/plant. In oriental melon experiment, numbers of root galls after 10 weeks from planting were significantly lower in J2+100,000 endospores/1 g medium with 32.5 compared with 64.1 and 87.5 in J2+100,000 endospores/1 g medium and the J2, respectively. However, there were no significant differences in plant growth characteristics among the treatments in the both crops.     

内生拮抗细菌在哈密瓜植株体内的传导定殖和促生作用研究

采用抗生素标记的方法研究了内生拮抗细菌P38和B167菌株在哈密瓜植株体内的定殖动态和对植株生长的影响.结果表明,接种方法显著影响P38菌株在植株体内的定殖和传导,并以浸种处理最佳,蘸根和灌根处理次之,喷叶处理最差;浸种可使P38菌株在根、茎、叶中良好传导和稳定定殖,随着植株的生长,根内菌量呈下降趋势,而茎、叶内的含菌量先上升后下降;P38菌株还具有促进哈密瓜种子萌发和植株生长的作用.B167菌株只在根内定殖,在体内的扩展性较差,不能进入叶片;它对植株的生长表现出一定的抑制作用.     

Plant Growth‐Promoting Bacteria from Solarized Soil with the Ability to Protect Melon Against Root Rot and Vine Decline Caused by Monosporascus cannonballus

This study was undertaken to isolate indigenous plant growth‐promoting (PGP) bacteria from solarized soil effective in the biocontrol of Monosporascus cannonballus, the cause of root rot and vine decline of melon, which is one of the most destructive soilborne diseases of this crop worldwide. The screening strategy resulted in the selection of two interesting PGP bacteria as biocontrol candidates against M. cannonballus belonging to the same microbial community. The two bacterial species, identified according to phenotypic, physiological tests and analysis of the 16S rDNA sequence as Bacillus subtilis/amyloliquefaciens (BsCR) and Pseudomonas putida (PpF4), showed PGP traits and in vitro antagonistic activity towards M. cannonballus. Antagonism by BsCR was characterized by a consistent inhibition of the pathogen in vitro growth; PpF4 strongly inhibited the development of perithecia of the pathogen. Under greenhouse conditions, the selected bacteria were tested for their biocontrol activity in the pathosystem melon‐M. cannonballus. BsCR alone and in combination with PpF4 determined a consistent decrease in the disease symptoms. BsCR and the combination of the bacterial strains significantly increased root biomass in both inoculated and un‐inoculated plant. Upon seed treatment with BsCR, the accumulation and isoenzyme induction of peroxidase in roots as biochemical marker for induction of resistance were found, thus indicating that BsCR may reduce the disease severity also by the activation of the plant defence responses. The study highlights the synergistic biocontrol potential of B. subtilis BsCR and P. putida PpF4 in the integrated management of root rot and vine decline of melon caused by M. cannonballus.     

Pathogenicity of fungi associated with melon vine decline and selection strategies for breeding resistant cultivars

Melone Vine Decline is a severe rot root disease of increasing world‐wide importance. In Eastern Spain it is related to the presence of Acremonium cucurbitacearum and Monosporascus cannonballus. The strong influence of environmental conditions on the progress of this disease has made its study difficult. A field screening of Cucumis melo accessions has been conducted over four years. Simultaneously, the pathogenicity of isolates of the two fungi recovered from the screening field was studied. These were more aggressive than other Spanish and American isolates. Percentage of vine decay was scored, togeter with root damage, the latter being evaluated by using four scoring systems based on root characteristics and disease severity. Root inspection alowed the selection of resistance sources, even when aboveground symptoms did not appear, due to the lack of environmental stresses at time of fruit maturity. The root damage scoring if environmental stresses occur during fruit maturity. The accession C. melo var. agrestis Pat 81 consistently exhibited high field resistance level, expressed as a higher percentage of symptomeless plants, together with a significant delay in symptoms appearance. The F₁ hybrids derived from the cross Pat 81 ×C. melo susceptible varienties showed an intermediate level of resistance between the parents, suggesting a partial dominance gene action. The high resistance level found in Pat 81, and also in its derived hybrids, against the aggressive isolates found in this area, makes it promissing for breeding melon verieties resistant to melon vine decline.     

Root-pressure driven xylem sap flow in greenhouse melon (Cucumis melo L.): diurnal change and the effects of shading, growth stage, rootstock and fruit number

Grafted and ungrafted greenhouse melon were used to investigate the effect of diurnal change, shading, growth stage, rootstock and fruit numbers on melon xylem sap flow rate. A clear diurnal change was observed in xylem sap flow rate. Shading of the plant on the previous day decreased the sap flow rate. An increase in the number of fruit on a plant decreased the sap flow rate, but, grafting to squash plant lessened the effect of fruit number. Consideration of timing and environmental factors is necessary when the xylem sap is collected for root study.     

棉蚜寄主专化型及其形成的行为机理

通过生活在甜瓜和棉花上的棉蚜Aphisgossypii Glover的行为,研究棉蚜的寄主专化型及其形成的行为机理。生物学观察显示： 两类棉蚜在寄主植物相互交换以后,定居数显著减少,棉花蚜型棉蚜的繁殖系数及若虫存活率显著下降,说明棉蚜存在甜瓜蚜型和棉花蚜型两种寄主专化型。通过刺探电位技术研究棉蚜的取食行为,以探索其寄主专化型形成的行为机理。结果表明： 甜瓜蚜型棉蚜在棉花上的取食行为容易被中断,但其口针定位韧皮部的能力并没有显著削弱;而棉花蚜型棉蚜在甜瓜上的取食行为受到更大的影响,口针无法顺利定位至韧皮部,并在2 h内根本无法在筛管内取食。生物学观察和EPG取食行为分析都显示： 与甜瓜蚜型棉蚜相比,棉花蚜型棉蚜对寄主的要求更严格-寄主专化程度更高,对寄主的利用率更高。     

热处理和壳聚糖涂膜对采后接菌哈密瓜生理生化特性的影响

以‘西州蜜25号’哈密瓜果实为试材,分别进行55℃热水浸渍3min、2%壳聚糖涂膜及两者结合处理(55℃热水浸渍3min+2%壳聚糖涂膜),以不进行任何预处理为对照,再对各处理的哈密瓜接种交链孢菌(Alternariaalternata),研究接菌哈密瓜在常温贮藏过程中抗病性和相关生理生化指标的变化情况。结果表明,与对照相比较,3种预处理均能明显抑制哈密瓜细胞膜渗透率、呼吸强度、乙烯释放量上升,显著提高贮藏后期果实几丁质酶、β-1,3-葡聚糖酶、苯丙氨酸解氨酶和过氧化物酶的活性,从而增强了哈密瓜的抗病性,有效降低贮藏过程中接菌哈密瓜病斑直径和病斑深度、防止哈密瓜的腐烂变质,并以55℃热水浸渍3min+2%壳聚糖涂膜结合处理的效果最佳。     

Cotton,cowpea and sesame are alternative crops to cucurbits in soils naturally infested with Monosporascus cannonballus

Monosporascus cannonballus is an important cucurbit root pathogen, which has been reported in the main production areas of melon and watermelon in Brazil and worldwide and potentially capable to colonize roots of different species. Crop rotation is considered an effective management strategy to prevent this disease. The aim of this study was to evaluate the response of different crops, pumpkin, cotton, cowpea, sesame, watermelon, melon, corn, cucumber, sorghum and tomato, to the infection of this pathogen. Seedlings were transplanted into plastic containers with an inoculum concentration of 20 colony‐forming units (CFU) g^?1 of M. cannonballus. Fifty days after transplanting, the variables analysed were the degree of disease severity on the root system and the frequency of reisolation. On cucurbits, the results demonstrated different degrees of susceptibility among crops and cultivars, being melon and watermelon the most sensitive species. In contrast, Cucurbita cultivars were the most tolerant. Regarding non‐cucurbit crops, maize, sorghum and tomato presented root discoloration and M. cannonballus was reisolated from roots. Cotton, cowpea and sesame cultivars were not affected by the pathogen, so they can be considered as alternative crops to be cultivated, or in rotation with cucurbits, in M. cannonballus infested soils.     

抗菌肽基因转化厚皮甜瓜的研究

以厚皮甜瓜FY1的子叶为试材,研究不同激素组合和配比对甜瓜植株再生的影响,并用农杆菌介导法将银杏抗菌肽基因Gnk2-1转化甜瓜。结果表明:(1)甜瓜子叶的最佳愈伤组织及不定芽诱导培养基为MS+2.0mg.L-1 6-BA+0.1mg.L-1 IAA,出愈率和出芽率均达95%以上;最佳芽伸长培养基为MS+0.2mg.L-1 KT;最佳生根培养基为1/2MS+0.1mg.L-1 IAA。(2)卡那霉素对甜瓜外植体的生长和分化有明显的抑制作用,适宜的筛选压力为100mg.L-1。(3)将子叶预培养2d,在农杆菌菌液浓度OD600值为0.5左右时,侵染外植体10min左右,共培养3d,转化效率最高。(4)经PCR鉴定获得了抗性植株,抗病性鉴定显示,转基因植株对枯萎病的抗性有所增强,发病迟缓。     

甜瓜遗传多样性的AFLP分析

利用AFLP(Amplified fragment length polymorphism)分子标记技术对48份甜瓜(Cucumis meloL.)材料遗传多样性和亲缘关系进行了研究.从64对引物中筛选出了10对进行选择性扩增,共扩增出423条带,其中多态性条带172条,多态率为40.66%.聚类分析将供试材料分成了两大类群,即厚皮甜瓜类群和薄皮甜瓜类群,并进一步将厚皮甜瓜类群分成了4个亚类,与甜瓜传统分类结果基本一致.     

福寿螺的寄主植物及其对福寿螺体重的影响

为了明确福寿螺对寄主植物的选择性,进一步掌握其可能的危害及入侵暴发机制,对稻田水域及周边的24科43种植物进行了室外取食试验,分别在取食后3、5d对受试样品及成螺称重,比较其无选择条件下的取食偏好性及其对成螺增长率的影响。结果表明:福寿螺对受试的24科43种植物(样品)的根、茎、叶、果实以及块根(茎)均能取食,对水生(湿生)和陆生植物没有偏好性;在取食过程中,优先取食细嫩的植株和部分,硬老部分需经水浸泡软化后才能被取食;辣椒、苦瓜等果实的辣苦味对福寿螺无驱避作用;取食率与体重增长率呈显著正相关,取食越多生长越快;但体重与植物的总酚含量呈负相关。福寿螺的广食性、取食量、植物总酚含量、植物丰盛度等种群和生态系统特征是影响其快速扩散的重要原因,同时可以借助福寿螺对植物的不同利用类型防范其扩散和危害。     

甜瓜种质资源遗传多样性的SRAP分析

为研究甜瓜材料之间的亲缘关系及其分类, 更有效地利用种质资源, 为培育新品种提供依据, 文章采用 SRAP (Sequence-related amplified polymorphism)技术对61份甜瓜种质资源的遗传多样性进行研究。结果表明: 从42对引物组合中筛选出16对扩增条带清晰、多态性高的引物组合分析供试材料, 共检测出452个位点, 其中265个为多态性位点, 多态性比率达58.63％, 平均每对引物组合产生28.56个位点和16.56个多态性位点。61份材料间的相似系数为0.48～0.93, 平均为0.73。这些结果说明, 供试甜瓜材料具有较为丰富的遗传多样性。聚类分析结果表明, 61个甜瓜品种中首先可分为薄皮甜瓜与厚皮甜瓜两大类, 彼此亲缘关系最远。以遗传相似系数0.74为截值, 可把供试材料分为5个类群。在生态区域中, 新疆厚皮甜瓜的Nei’s基因多样性指数(0.2231)和Shannon’s信息指数(0.3422)最高。