Genome-wide association mapping and genomic prediction of stalk rot in two mid-altitude tropical maize populations

Maize stalk rot reduces grain yield and quality.Information about the genetics of resistance to maize stalk rot could help breeders design effective breeding strategies for the trait.Genomic prediction may be a more effective breeding strategy for stalk-rot resistance than marker-assisted selection.We performed a genome-wide association study(GWAS)and genomic prediction of resistance in testcross hybrids of 677 inbred lines from the Tuxpe?o and non-Tuxpe?o heterotic pools grown in three environments and genotyped with 200,681 single-nucleotide polymorphisms(SNPs).Eighteen SNPs associated with stalk rot shared genomic regions with gene families previously associated with plant biotic and abiotic responses.More favorable SNP haplotypes traced to tropical than to temperate progenitors of the inbred lines.Incorporating genotype-by-environment(G×E)interaction increased genomic prediction accuracy.

Screening of antagonistic Trichoderma strains and their application for controlling stalk rot in maize

Maize is one of the major crops in China, but maize stalk rot occurs nationwide and has become one of the major challenges in maize production in China. In order to find an environment-friendly and feasible technology to control this disease, a Trichoderma-based biocontrol agent was selected. Forty-eight strains with various inhibition activities to Fusarium graminearum, and Fusarium verticillioides were tested. A group of Trichoderma strains(DLY31, SG3403, DLY1303 and GDFS1009) were found to provide an inhibition rate to pathogen growth in vitro of over 70%. These strains also prevented pathogen infection over 65% and promoted the maize seedling growth for the main root in vivo by over 50%. Due to its advantage in antifungal activity against pathogens and promotion activity to maize, Trichoderma asperellum GDSF1009 was selected as the most promising strain of the biocontrol agent in the Trichoderma spectrum. Pot experiments showed that the Trichoderma agent at 2–3 g/pot could achieve the best control of seedling stalk rot and promotion of maize seedling growth. In the field experiments, 8–10 g/hole was able to achieve over 65% control to stalk rot, and yield increased by 2–11%. In the case of natural morbidity, the control efficiency ranged from 27.23 to 48.84%, and the rate of yield increase reached 11.70%, with a dosage of Trichoderma granules at 75 kg ha^-1. Based on these results, we concluded that the Trichoderma agent is a promising biocontrol approach to stalk rot in maize.

Effect of Potassium on Ultrastructure of Maize Stalk Pith and Young Root and Their Relation to Stalk Rot Resistance

To study the mechanism of potassium （K） application on improvement of maize resistance to stalk rot at cellular level, scanning electron microscope and transmission electron microscope were used to observe the effect of K on the ultrastructure of maize stalk pith tissue and young root tip cell influenced by K and pathogen. In K deficient treatment, parenchyma cells of stalk pith had abnormal structure, and the cell wall between upper and lower adjacent cell was damaged, resulting in the loss of connections between vascular cells and insufficient supporting capacity. However, an improved K nutrition helped to keep a quite tight arrangement of root cell with thick cell wall, and prevent the invasion of pathogen effectively. Moreover, K treated root cell had abundant golgi apparatus, which could excrete large amount of secretions to degrade mycelium. Papillary and highly electronic intensity dot were accumulated at the invading point to prevent the deveJopment of the mycelium. Improved K nutrition could increase the resistant ability of maize plant to stalk rot, through keeping cell structure stability, preventing the expansion of intracellular space to reduce the chances of pathogen invasions, and through reinforcing cell wall and formation of intercellular and intracellular material to restrict further development of pathogen in host cell.

The antioxidant protein ZmPrx5 contributes resistance to maize stalk rot

Maize(Zea mays L.)stalk rot is a devastating disease worldwide,causing severe yield losses.Although previous studies have focused on the genetic dissection of maize resistance to stalk rot,the mechanisms of resistance remain largely unknown.We used a comparative proteomics approach to identify candidate proteins associated with stalk rot resistance.Statistical analyses revealed 763 proteins differentially accumulated between Fusarium graminearum and mock-inoculated plants.Among them,the antioxidant protein ZmPrx5,which was up-accumulated in diseased plants,was selected for further study.ZmPrx5 transcripts were present in root,stalk,leaf,ear,and reproductive tissues.The expression of ZmPrx5 in three inbred lines increased significantly upon F.graminearum infection.ZmPrx5 was localized in the cytoplasm.Compared to control plants,maize plants overexpressing ZmPrx5 showed increased resistance to F.graminearum infection,and ZmPrx5 mutant plants were more susceptible than wild-type plants.Defense-associated pathways including plant–pathogen interactions,phenylalanine metabolism,and benzoxazinoid and flavonoid biosynthesis were suppressed in ZmPrx5 homozygous mutant plants compared with wild-type plants.We suggest that ZmPrx5 positively regulates resistance against stalk rot in maize,likely through defense-oriented transcriptome reprogramming.These results lay a foundation for further research on the roles of Prx5 subfamily proteins in resistance to plant fungal diseases,and provide a potential genetic resource for breeding disease-resistance maize lines.

Resistance Analysis of 25 Corn Varieties to Stalk Rot and Evaluation of Yield Loss

Due to the meteorological factors, Henan Province had a large-scale outbreak of corn stalk rot in 2014, and the incidence status of 25 varieties was investigated. According to the relevant resistance identification standards, Xundan.509 was classified as resistance variety; Yudan606, Jinsai38 and Xundan3136 were classified as moderate resistance ones, while the other 21 varieties were highly susceptible varieties. Using the yield loss rate of 5% as the critical value of disease tolerance standard, XundanS09, Yudan606, Jinsai38, Xundan3136, Huaiyu5288, QiaoyuS, XY046, Zhengyu10, Lile66, WeikeT02 and Xundan29 were classified into tolerance varieties of corn stalk rot. The concept between disease resistance and disease tolerance was distinguished. The diseased plant rate was used as the classification basis of disease resistance and the yield loss rate was used as the evaluation standard of disease tolerance. The relationship between both was showed by a Venn diagram.

Pyramiding the disease resistant genes to southern rust and stalk rot in maize(Zea mays L.) with marker-assisted selection

Southern corn rust(SCR) caused by Puccinia polysora Underw and maize stalk rot caused by Pythium inflatum Matthews(MSR-2) are two destructive diseases of maize(Zea mays L.) in China.Our previous studies indicated that maize inbred line Qi319 is highly resistant to SCR but susceptible to MSR-2,while inbred line 1145 is highly resistant to MSR-2 but susceptible to SCR.The SCR resistant gene(RppQ) in Qi319 and MSR-2 resistant gene(Rpi1) in 1145 have been mapped on chromosome 10 and 4 respectively.In this research,through marker-assisted selection(MAS) with the molecular markers,bnlg1937 tightly linked to Rpi1 and phi041 tightly linked to RppQ,pyramid breeding of the two kinds of disease resistant genes were carried out from the year of 2003 to 2007.Two homozygotic inbred lines of F5 generation,DR94-1-1-1 and DR36-1-1-1 were identified.MAS result suggested DR94-1-1-1 and DR36-1-1-1 contained the two resistance genes RppQ and Rpi1.Field inoculation tests confirmed their high resistance to the two diseases.In addition,field investigation indicated that the two selected inbred lines,particularly DR94-1-1-1,had excellent agronomic traits such as plant height,ear height and yield-relating traits including ear length,ear diameter,ear weight,kernels per ear,kernels per row and kernel weight per ear.The two selected inbred lines DR94-1-1-1 and DR36-1-1-1 can either be directly developed into commercial variety or used as immediate donors of SCR and MSR resistance breeding programs in maize.

Stalk Rot Resistance Analysis and Yield Loss Evaluation on 25 Maize Varieties and Utilization of Resistant Germplasm Jun M9

Stalk rot outbroke in Henan Province in 2014 on maize,and the disease conditions of 25 varieties were investigated.According to national maize variety disease resistance identification standards,Jundan 509 was a resistant variety; Yudan 606,Jinsai 38 and Jundan 3136 were moderately-resistant varieties; and other 21 varieties were all highly susceptible.Based on the yield loss rate of 5% determined as the critical value for division of disease tolerance,disease-tolerant varieties included Jundan 509,Yudan 606,Jinsai 38,Jundan 3136,Huaiyu 5288,Qiaoyu 8,XY046,Zhengyu 10,lile 66,weike 702 and Jundan 29.Varietal disease resistance and tolerance were mainly divided according to incidence and yield loss rate,respectively,and the correlation between them was showed by a Venn diagram.With the male parent Jun M9 of Jundan 509 as a backbone material,multiple maize combinations resistant to stalk rot were formed,and Yongyou 618 among them was approved by Henan Provincial Crop Variety Approval Committee.

木霉微生物菌肥的制备及对玉米茎腐病的应用评价

以前期分离鉴定的生防菌株非洲哈茨木霉Tr35为研究对象,通过载体的筛选,制备木霉菌剂和木霉微生物菌肥。通过测定不同含量的木霉菌剂和木霉有机肥的株高、根长和鲜重,筛选促生效果明显的处理,并通过室内盆栽试验,评价其对玉米苗期茎腐病的防效。结果显示,玉米秸秆粉具有吸水性强、对Tr35无毒和高释放率的优点,与Tr35共同发酵制备木霉菌剂,当土与菌剂为5∶1时,促生效果明显,并对玉米苗期茎腐病防效达51.77%。菌剂土与腐熟羊粪为7∶1时的木霉有机肥促生效果最好,以该比例制备木霉微生物菌肥,对玉米苗期茎腐病防效达61.31%,该研究结果为Tr35菌剂和菌肥的应用开发提供了一定的前期基础。

玉米镰孢茎腐病致病机制及其防治的研究进展

玉米作为主要的农作物,它的生产对保障粮食安全具有重要意义,而茎腐病多年来都是严重威胁世界玉米安全生产的主要病害。本文将针对玉米镰孢茎腐病侵染机制和致病机制进行总结,进一步阐述玉米致病后的抗性遗传机制和免疫机制,同时,结合近年来国内外的相关研究进展概述玉米镰孢茎腐病的科学防治措施,旨在为玉米镰孢茎腐病的有关研究提供参考思路。

Natural variation in maize gene ZmSBR1 confers seedling resistance to Fusarium verticillioides

Maize seedling blight caused by Fusarium verticillioides is a widely occurring maize disease,but the genetics and mechanisms of resistance are not well understood.In this study,GWAS performed by MLM and 3VmrMLM identified 40 and 20 QTNs,associated with seedling blight resistance.These methods identified 49 and 36 genes,respectively.Functional verification of candidate gene ZmSBR1 identified by both methods showed that the resistance of a mutant line to seedling blight decreased by 0.37 grade points after inoculation with F.verticillioides,compared with the WT.The length of the stem rot lesion caused by F.verticillioides increased by 86%in mutant seedlings,and the relative length of the adult plant stalk rot increased by 35%in mutant plants compared to the wild type after inoculation with Fusarium graminearum.Transcriptome analysis showed that expression of defense-related genes after inoculation was down-regulated in the mutant compared to the wild type,synthesis of secondary metabolites associated with resistance was reduced,and the immune response triggered by PAMP decreased,resulting in decreased resistance of mutant maize seedlings.Candidate gene association analysis showed that most maize inbred lines carried the susceptible haplotype.A functional PCR marker was developed.The results demonstrated that ZmSBR1 conferred resistance to multiple Fusarium diseases at the seedling and adult growth stages and had important application value in breeding.

木霉菌对玉米茎腐病的田间防治效果及玉米产量的影响

玉米是我国主要的粮食作物之一。近年来,玉米茎腐病的危害面积和危害程度逐年增长,造成玉米产量降低和品质下降,严重影响了玉米产业的可持续发展。为筛选出防控玉米镰孢茎腐病的优势生防菌株,为玉米茎腐病的绿色防控提供备选材料,以前期室内筛选到的6株木霉菌株为研究对象,将玉米种子分别通过木霉发酵液包衣处理和孢子悬浮液浸种处理后,采用田间随机区组试验测定不同木霉菌株对玉米茎腐病的田间防治效果,并分析其对玉米产量的影响。结果表明,木霉菌株种类、处理方式和病原菌种类不同,对玉米茎腐病的防治效果和玉米产量的影响也不同;其中绿色木霉T43和康氏木霉KS包衣处理对禾谷镰孢茎腐病的防治效果分别达到54.21%、32.71%;绿色木霉T43浸种处理后,接种拟轮枝镰孢菌的前提下,玉米产量为6 760.24 kg/hm^(2),促生率达48.63%;康氏木霉KS浸种处理后,接种禾谷镰孢菌的前提下,玉米产量为6 353.74 kg/hm^(2),促生率达61.26%。从供试的菌株中,筛选出绿色木霉T43和康氏木霉KS作为优势生防菌株,绿色木霉T43的包衣处理对禾谷镰孢茎腐病表现出较好的防治效果,绿色木霉T43和康氏木霉KS浸种处理对玉米产量具有较好的增产效果。

玉米镰孢菌复合病原茎腐病抗性种质资源鉴定及抗性基因挖掘

【目的】玉米茎腐病是我国玉米主产区普遍发生的重要病害之一,自然条件下,玉米茎腐病多由多种病原菌共同侵染造成。筛选抗禾谷镰孢菌(Fusarium graminearum)和拟轮枝镰孢菌(Fusarium verticillioides)复合侵染的优异玉米种质,鉴定与复合病原引起的茎腐病抗性相关数量性状核苷酸(quantitative trait nucleotide,QTN),挖掘抗性候选基因,为玉米抗复合病原抗性分子育种提供基因资源和理论借鉴。【方法】以一个玉米自然群体为试验材料,接种F.graminearum和F.verticillioides复合病原菌,鉴定茎腐病表型;利用全基因组关联分析,筛选显著抗性SNP位点,预测抗病候选基因。【结果】通过田间和室内2个环境下接种复合病原后的茎腐病表型鉴定试验,发现不同来源和亚群的自交系对复合病原侵染的表型差异显著。田间表型鉴定结果表明,收集于中国的自交系普遍抗性较高,而美国的自交系普遍感病性较高;热带及亚热带亚群的自交系抗性较高,混合型亚群的自交系感病性较高。室内表型鉴定结果表明,收集于美国的自交系材料抗性较高,国际玉米小麦改良中心的自交系材料感病性高;硬秆亚群的玉米种质表现较高抗性,混合型亚群的种质表现较高感病性。于田间和室内2个环境下分别筛选出29份和16份对复合病原侵染具有较高抗性水平的种质,2个环境共同筛选到6份抗病种质。基于田间表型GWAS分析,鉴定到18个与复合病原茎腐病抗性显著相关的QTN,挖掘出93个抗病候选基因;有4个基因表现出单倍型变异,且接种后基因表达水平在抗病材料中呈上调趋势。【结论】利用遗传背景丰富的玉米自然群体,在2个环境中共同鉴定出6份玉米镰孢菌复合病原茎腐病抗性材料,可作为潜在的玉米抗茎腐病育种种质资源;挖掘出4个可能参与复合病原菌抗性的候选基因,为玉米镰孢菌复合病原茎腐病抗性育种提供基因资源。

马铃薯细菌性茎腐病综合防治技术研究初探

榆林是中国马铃薯优生区之一,靖边是榆林马铃薯的核心产区,适宜马铃薯规模化生产的资源要素目前已发挥到极致,在马铃薯产业迅猛发展过程中,连年重茬种植在所难免,加之种薯地病原基数呈逐年上升的态势,导致马铃薯黑胫病、细菌性茎腐病、疮痂病、软腐病、环腐病和青枯病等细菌性病害同趋势加重。特别是马铃薯细菌性茎腐病(气生性茎腐病、茎秆腐烂病),目前己成为制约榆林马铃薯生产的主要细菌性病害之一,常年田间平均发病率为10%~20%,严重时发病率高达60%~70%,减产损失达10%~40%,对榆林马铃薯产业发展极具威胁。

玉米禾谷镰孢茎腐病和穗腐病抗性鉴定接种方法的比较

田间采用不同人工接种鉴定方法对92份玉米种质同步进行玉米禾谷镰孢茎腐病和穗腐病抗性评价与筛选,明确了东北春玉米产区玉米禾谷镰孢茎腐病、穗腐病抗性鉴定的最佳接种方法:茎节注射法在玉米禾谷镰孢茎腐病抗性精准鉴定中可更好地评价玉米种质的抗病性,综合考虑接种方法的实用性和可操作性,菌土覆盖法对大体量的规模化种质进行田间抗病性鉴定更具有优势;双牙签法对玉米穗腐病抗性鉴定中在种质接种后发病的严重度、田间进行接种的可操作性和实用性等方面均优于花丝通道法。同时筛选出对玉米禾谷镰孢茎腐病抗性稳定的种质52份,对穗腐病抗性稳定的种质66份,对2种病害均表现稳定抗性的兼抗种质33份。

玉米腐霉茎腐病生防木霉菌株的筛选、鉴定及防治效果分析

【目的】筛选对玉米腐霉茎腐病病原菌具有抑制作用的木霉(Trichodermaspp.)菌株,明确其分类地位以及对腐霉茎腐病的防治效果和抑菌机理,为腐霉茎腐病生防制剂的研发提供菌种资源。【方法】采用菌丝生长速率法测试候选木霉菌株对肿囊腐霉(Pythiuminflatum)、强雄腐霉(P.arrhenomanes)和芒孢腐霉(P.aristosporum)的抑制作用,筛选拮抗菌株;通过形态学和分子生物学特性确定Tr21菌株的分类地位;采用常规抑菌方法观察Tr21对腐霉菌丝形态的影响;采用溴化丙啶(PI)染液检测法及对不同处理时间菌丝上清液中蛋白和核酸吸光值的检测,分析Tr21发酵液对腐霉菌细胞膜通透性的影响;通过不同浓度Tr21发酵滤液浸种试验,检测Tr21发酵滤液对玉米种子发芽性状的影响;通过温室盆栽试验和田间人工接种试验,明确Tr21对腐霉茎腐病的防治效果。【结果】从实验室保存的109株木霉菌中,筛选到7株木霉菌对肿囊腐霉、强雄腐霉和芒孢腐霉具有拮抗活性,抑制率均>60%,其中Tr21菌株对3种腐霉菌的抑制率达到100%,其5×、10×和20×稀释液对3种腐霉菌的抑制率均达到100%。50×稀释液对3种腐霉菌的最低抑制率也达到55.56%。经形态学和分子生物学鉴定,Tr21为非洲哈茨木霉(T.afroharzianum)。显微镜观察显示Tr21发酵滤液能够造成腐霉菌菌丝变粗、菌丝分枝增多、节点缩短、断裂、内含物溢出等畸形现象。PI荧光染色试验显示Tr21发酵滤液导致3种腐霉菌的细胞膜受损,PI染液更易穿透受损的细胞膜进入到菌丝体内,使菌丝染成红色。核酸、蛋白泄露试验发现发酵滤液处理过的菌丝吸光值变化较大,处理5 h后,肿囊腐霉、芒孢腐霉和强雄腐霉菌丝的OD260均增加了0.08,OD280分别增加了0.10、0.11和0.10,表明腐霉菌菌丝细胞膜受损或其完整性被破坏,导致菌丝内含物外溢。不同浓度Tr21发酵滤液对玉米种子的发芽性状无影响,且当Tr21发酵滤液浓度为20×稀释液时对玉米种子的萌发和生长促进效果最好。盆栽试验结果表明,Tr21发酵滤液浓度为5×稀释液时,对3种腐霉茎腐病的室内防治效果最佳,分别为60.67%、63.15%和59.66%。用Tr21的5×稀释液进行种子处理,当药种质量比例为1﹕100时,对腐霉茎腐病的防治效果最高,达82.25%。【结论】获得一株有效防治玉米腐霉茎腐病的木霉菌株Tr21,经鉴定该菌为非洲哈茨木霉,该菌株发酵滤液可导致腐霉菌菌丝畸形、断裂、细胞膜受损、内含物溢出等,是一株具有开发前景的生防微生物。

东北玉米杂交种对茎腐病的抗性鉴定

采用田间人工接种鉴定方法,对50份玉米杂交种进行抗性鉴定。结果表明:玉米杂交种接种病原菌后,均有不同程度的茎腐病发生;表现高抗、抗、中抗、感、高感的杂交种分别为9份、11份、6份、11份、13份,占比分别为18%、22%、12%、22%、26%;抗性水平较好的杂交种共有26份,占比52%。

国外种质对玉米禾谷镰孢茎腐病与穗腐病的抗性鉴定与分析

为了筛选抗玉米茎腐病、穗腐病的玉米种质资源,2020—2022年连续3年对225份国外玉米种质资源禾谷镰孢菌引起的玉米茎腐病、穗腐病进行抗性鉴定和评价。结果表明,有12份资源对玉米镰孢菌腐病抗性较好,25份资源对玉米镰孢穗腐病抗性较好,分别占总数的5.3%和11.1%,玉米茎腐病在3年间综合抗性的r值依次为-0.056、-0.230、-0.110,说明不同环境对镰孢菌茎腐病的抗性反应有差异;玉米穗腐病在3年间综合抗性的r值依次为0.065、0.024、-0.187,表明玉米禾谷镰孢穗腐病的抗性受环境因素影响相对较大。

102份玉米地方种质的茎基腐病抗性评价

采用田间人工接种,对102份河南玉米地方种质进行了镰孢菌茎基腐病的田间表型抗性鉴定,试验结果表明,表现1级高抗的种质材料有16份,3级抗病的种质材料有17份,5级中抗的种质材料有30份,感病和高度感病的种质材料有39份。其中抗性以上的材料占全部材料的62%,且以中抗抗性为主。对病原菌进行分离鉴定,结果表明,禾谷镰孢菌致病力最强,拟轮枝镰孢菌对玉米植株生长影响次之,芒孢腐霉致病率最弱。地方种质抗性鉴定试验最终筛选出的33份抗病性强的地方种质,丰富和拓宽了镰孢菌茎基腐病的抗性基因遗传基础,可作为开展选育抗性新种质和抗病育种的重要基础材料,对镰孢菌茎基腐病选育和抗病育种产生利用价值,为玉米生产上有效地进行抗性基因轮换提供参考。

基于巢式PCR技术对玉米种子携带禾谷镰刀菌的检测体系

禾谷镰刀菌(Fusariumgraminearum)是真菌性玉米茎基腐病的主要致病菌之一,为了减少种子带菌对该病害的传播,本研究采用巢式PCR技术构建玉米种子检测体系,并对其进行特异性、灵敏度和可行性检测。结果表明,该体系可有效检测到玉米(Zeamays)种子携带禾谷镰刀菌的情况,灵敏度达4.224 pg·μL^(-1),是一种快速、灵敏、特异性强的检测体系,为玉米茎基腐病种子带菌检测、早期诊断和及时防治提供技术支持和理论依据。本研究是运用巢式PCR技术检测玉米茎基腐病主要致病菌-禾谷镰刀菌在国内的首次报道。

多粘类芽孢杆菌NPDY05-8对玉米茎基腐病的防治效果及对土壤微生物的影响

【目的】探明多粘类芽孢杆菌NPDY05-8灌施土壤后对玉米茎基腐病的防治效果,及该菌在土壤中定植情况和对土壤的细菌和真菌等微生物群落的影响。【方法】利用Illumina MiSeq高通量测序技术,以未灌施菌株NPDY05-8和种植花生轮作的土壤为对照,对灌施多粘类芽孢杆菌NPDY05-8试验地土壤的细菌及真菌群落组成结构进行研究分析。【结果】该菌株对玉米茎基腐病的田间防治效果在2021年达90.92%,2022年达95.68%,显著优于化学药剂的防效。土壤菌群分析结果显示菌株NPDY05-8能够在土壤中定植,且长期存在于土壤当中。施用菌株NPDY05-8一年Y1和30日Y2的土壤细菌多样性OTUS分别为1 096个和1 149个,均高于未施用菌株的土壤Y3的973个及轮作土壤的941个;赤霉菌属的丰度Y1为4.15%、Y2为8.76%,均低于未施用菌株的土壤Y3的17.18%;镰刀菌属的丰度Y1为0.80%、Y2为1.12%,均低于未施用菌株的土壤Y3的2.17%;施用菌株一年的土壤与施用菌株30日的土壤在细菌和真菌水平的相关性指数高,距离最近。【结论】灌施菌株NPDY05-8能改变土壤的真菌及细菌的种群结构,控制有害真菌,达到持久改良土壤和防治土传性真菌病害的目的。