云南芒果采后炭疽病病原菌的鉴定及室内生防菌的筛选

随着对云南芒果需求量的增加,其种植面积日趋扩大,并不断引入新品种,这也导致云南省芒果炭疽病害发生日趋加重。为了有针对性地开展芒果炭疽病的生物防治,采用组织块分离法分离芒果采后炭疽病病原菌,通过形态学观察初步鉴定,并遵循柯赫氏法则对病原菌进行验证。随后,利用rDNA-ITS序列和系统发育树分析明确病原菌的分类学地位。最后,采用5种生防细菌对病原菌进行拮抗试验。通过对芒果采后炭疽病病原菌进行分离鉴定,确定胶孢炭疽菌(Colletotrichum gloeosporioides)是引起云南芒果采后炭疽病的病原菌,该菌株内转录间隔区(internal transcribed spacer,ITS)序列长度为536 bp,登录号为MH744668;5株生防细菌对芒果炭疽病病原菌都有一定的抑菌作用,具有较好的生防开发潜能,其中抗生素溶杆菌L-44的抑菌效果最好,抑制率达53.7%。研究结果为云南省芒果采后病害的生物防治提供了新的思路。     

芒果炭疽病菌侵染过程中致病基因的表达分析

为研究芒果胶孢炭疽菌在侵染过程中致病相关基因的差异表达。采用实时荧光定量PCR的方法,分析病菌侵染芒果叶片和果实过程中PL、PG等11个基因的表达量变化。研究发现,病原菌侵染叶片时,PG和PL基因均持续高效表达,SIN3P基因表达较低,其余基因在侵染6 h时表达量较高,随后下降;病原菌侵染果实时,PL、PG、SDH、ECH等基因高效表达,其余基因则有升有降。说明致病基因在胶孢炭疽菌侵染芒果不同组织时表达有差异,在侵染的不同时段也不同程度地发挥功能。     

芒果胶孢炭疽病菌应答菌丝机械损伤产生无性孢子的分子机制

胶孢炭疽菌(Colletotrichum gloeosporioides)是引发芒果(Mangifera indica)炭疽病的主要病原体。室内平板培养胶孢炭疽菌不产生或产生很少分生孢子的情况时有发生, 但菌丝在机械损伤后24-48小时会产生大量分生孢子。胶孢炭疽菌应答机械损伤诱导产孢的核心基因及关键代谢通路尚未见报道。基于转录组测序(RNA-seq)技术检测了芒果胶孢炭疽菌菌丝在机械损伤处理后2小时内5个时间点的基因表达变化, 对差异表达基因进行GO富集和KEGG代谢通路富集分析, 并对菌丝响应胁迫的基因动态表达数据进行分析。基于常微分方程ODE模型结合变量选择技术, 构建了动态基因调控网络。结果表明, 有417个差异表达基因参与应答胶孢炭疽菌菌丝机械损伤, 分属12个聚类模块, 有4条通路存在显著富集, 分别是丙酮酸代谢、硫代谢、黄曲霉素合成途径和二萜合成途径。结合功能注释筛选出12个应答菌丝损伤胁迫的核心基因。研究结果为后续深入开展芒果胶孢炭疽菌产孢和致病机理研究奠定了重要基础。     

虎耳草炭疽病病原菌鉴定

【目的】从虎耳草炭疽病病斑分离病原菌,并对病原菌进行鉴定。【方法】采用柯赫氏法则对从病斑组织块分离的病原菌进行验证,通过形态学结合多基因分子系统学方法对病原菌进行鉴定。【结果】从虎耳草炭疽病病斑分离到2株菌株(菌株号:LPSU 20120244、LPSU 20120251)。2株菌株孢子均无色,无隔,直、圆柱状,菌株LPSU 20120244孢子(11-25)μm×(5-9)μm,菌株LPSU 20120251孢子(15-25)μm×(5-7)μm。多基因分子系统树中,2株菌株分别与喀斯特炭疽菌(Colletotrichum karstii Y.L.Yang,Zuo Y.Liu,K.D.HydeL.Cai)和江西炭疽菌(C.jiangxiense F.LiuL.Cai)模式菌株聚为一支,支持率均为100%。【结论】形态学及多基因分子系统学分析表明菌株LPSU 20120244为喀斯特炭疽菌,菌株LPSU 20120251为江西炭疽菌。     

浅谈植物炭疽病

自２００１年美国纽约的“９·１１”事件后,又接二连三的发生了邮寄炭疽热病菌孢子的恐怖事件,致使全美国再次陷于十分恐慌的状态。为什么人们对炭疽热病如此惧怕呢？这是由于炭疽菌的致病力极强,感染后潜伏期短,发病急以及病死率高等特点。炭疽菌不仅能感染家畜和人类,而且还可使植物染病,只不过它们感染的病原菌不同而已,其中引起人畜炭疽病的病原菌叫炭疽杆菌（Bacillusanthracis）,而植物炭疽病的病原菌则为一类真菌,即指由黑盘孢目中的炭疽属（Colletotrichum）真菌的有性世代所引起的病害的统称。以往也曾有些人使用侵染植…     

芒果炭疽病菌小柱孢酮脱水酶基因SCD1的克隆与敲除载体构建

芒果炭疽病(Colletotrichum gloeosporioides)是为害芒果的重要病害之一,为了明确小柱孢酮脱水酶基因SCD1与芒果炭疽病菌致病力之间的相互关系,本研究以芒果炭疽病菌DNA为模板,利用同源克隆技术扩增SCD1,分析其序列特征、推测了其蛋白的保守结构域,并借助In-Fusion~ HD Cloning Kit技术进行敲除载体构建。结果表明,该基因DNA和cDNA全长分别为796 bp、564 bp,编码区有两个内含子(大小为52 bp,180 bp),推测编码187个氨基酸,其分子量约为21.52 kD,等电点PI为5.90,与NCBI网站中已公布的基因进行Blastp比对,发现该序列与香蕉炭疽菌(C.musae)、无花果炭疽菌(C.caricae)(登录号:GQ266389.1,GQ266386.1)的小柱孢酮脱水酶基因相似性分别为98%和97%。该基因的敲除载体pSCDGH-1已构建成功,为下一步获得SCD1基因敲除突变体,研究该基因功能打下了材料基础。     

禾谷炭疽菌磷脂酰肌醇转移蛋白生物信息学分析

目的:禾谷炭疽菌侵染玉米、小麦等农作物引起的炭疽病,给各国农业生产造成了巨大经济损失。植物病原菌在对植物侵染、定殖、扩展等致病过程中,细胞信号转导起着重要的作用。磷脂酰肌醇转移蛋白(PITP)参与众多生理生化过程,并在细胞信号转导过程中发挥重要作用。本研究为深入开展禾谷炭疽菌PITP的功能研究打下坚实的理论基础,同时,也为进一步开展其他炭疽菌的研究提供重要的理论指导。方法:基于酿酒酵母中已经报道的5个典型PITP序列,对炭疽菌属蛋白质数据库进行Blastp比对以及关键词搜索,并通过SMART保守结构域分析。结果:明确禾谷炭疽菌存在3个典型的PITP,同时,明确上述蛋白疏水性、细胞信号肽、跨膜区结构、亚细胞定位以及二级结构等情况。结论:与其他物种中9个同源序列进行遗传关系比较分析,该菌与希金斯炭疽菌亲缘关系较近。     

生防细菌T132的鉴定及其对采后柑橘炭疽病的抑制效果

【目的】柑橘(Citri)是世界上重要的果树。由胶孢炭疽菌[Colletotrichum gloeosporioides(Penz.)]引起的柑橘炭疽病是柑橘生产的主要病害之一。为探索对采后柑橘炭疽病有效的生防措施,分离鉴定柑橘根围土壤中一株细菌T132,并研究其特性及生防效果。【方法】根据菌株T132的形态特征、生理生化特性以及16S rDNA序列对其进行鉴定;通过连续8次在人工培养基上传代培养,测定该菌株的遗传稳定性;采用柑橘果实刺伤挑战接种和拮抗菌液直接浸泡健康果实两种方法研究该菌株对柑橘炭疽病的抑菌防病效果;利用洋葱伯克霍尔德氏菌致病因子的特异性引物检测菌株T132是否为潜在的人类致病菌。【结果】菌株T132鉴定为越南伯克霍尔德氏菌(Burkholderia vietnamiensis)。连续8次在人工培养基上传代培养,菌株T132抑制胶孢炭疽病菌生长的能力没有发生明显改变。菌株T132对胶孢炭疽菌C.gloeosporioides引起的柑橘炭疽病有明显的防治作用,刺伤接种的防效为88.2%,自然发病的防效为54.9%。未检测到该拮抗菌株有人体致病相关的洋葱伯克霍尔德氏菌致病因子(BCESM)毒力基因。【结论】首次报道对柑橘采后炭疽病具有生防效果、对人类相对安全的越南伯克霍尔德氏菌生防菌株。     

钩藤炭疽病生物学特性研究

钩藤是常绿藤本,属茜草科药用植物,具有很高的药用价值。文章就剑河钩藤较为严重的炭疽病进行病原菌分离,并对其中分离出的葡萄座腔菌进行生物学特性研究。通过病叶接种、分离纯化、病原菌鉴定、葡萄座腔菌的生物学特性研究,得出最适合萄座腔菌的生长环境,即温度在30℃左右,喜光,偏酸性,pH在5左右,最适碳源为果糖、最适氮源为蛋白胨和酵母浸粉,以为钩藤炭疽病的绿色防控提供理论支撑。     

油茶果生炭疽菌小分子GTP酶Rab7的功能研究

【目的】炭疽病是油茶的一种重要病害,果生炭疽菌是油茶炭疽病的主要致病菌。本文对果生炭疽菌小分子GTP酶Rab7进行研究,为油茶炭疽病的防控治理提供依据。【方法】构建CfRAB7基因敲除载体,通过PEG介导的原生质体转化、抗性筛选和PCR电泳验证获得果生炭疽菌突变体菌株△Cfrab7和互补菌株△Cfrab7/CfRAB7。进一步分析CfRAB7基因敲除突变体△Cfrab7的生长、产孢、附着孢的形成、胁迫应答、液泡融合和致病力等生物学表型。【结果】在PDA和MM培养基上,突变体△Cfrab7的菌落直径显著减小,产孢量和附着孢形成率显著降低,且不能穿透玻璃纸;在10mmol/LH2O2条件下,△Cfrab7生长受到明显抑制;进一步研究发现突变体△Cfrab7液泡无法正常融合,在油茶有伤和无伤的幼叶上均不发病。【结论】CfRAB7基因参与调控果生炭疽菌生长产孢、附着孢形成、H2O2胁迫应答、液泡融合和致病力。     

Effect of hot water treatment on development of anthracnose (Colletotrichum gloeosporioides) and quality of mango fruit at Jimma southwest Ethiopia

Anthracnose is the major postharvest disease of mango and occurs throughout mango producing areas of the world including Ethiopia. Evaluating effect of hot water treatment on development of anthracnose and quality of mango fruit is imperative. A total of three hot water levels 48, 52 and 56 °C at two time interval (5 and 10 min) were tested with factorial arrangement in completely randomised design. The study indicated that hot water treatment at different temperatures and time interval significantly (p < 0.001) affects disease development and shelf life and postharvest quality of mango fruits. Hot water treatments reduced the incidence and severity of anthracnose disease significantly (p < 0.001) in mango fruits as compared to control. There was a highly significant difference (p < 0.0001) on weight loss, total soluble solids, titratable acidity and fruit firmness of mango fruits due to treatment. The present study reviled that hot water treatment has a potential in reducing the postharvest loss due to anthracnose and improving the shelf life and quality of mango fruits. However, the reduction of disease pressure on fruits was not at applicable level, which call ups future effort on developing on integrated disease management strategies for reduction of postharvest loss of mango fruits.     

Advances in understanding the mechanism of resistance to anthracnose and induced defence response in tea plants

The tea plant (Camellia sinensis) is susceptible to anthracnose disease that causes considerable crop loss and affects the yield and quality of tea. Multiple Colletotrichum spp. are the causative agents of this disease, which spreads quickly in warm and humid climates. During plant–pathogen interactions, resistant cultivars defend themselves against the hemibiotrophic pathogen by activating defence signalling pathways, whereas the pathogen suppresses plant defences in susceptible varieties. Various fungicides have been used to control this disease on susceptible plants, but these fungicide residues are dangerous to human health and cause fungicide resistance in pathogens. The problem-solving approaches to date are the development of resistant cultivars and ecofriendly biocontrol strategies to achieve sustainable tea cultivation and production. Understanding the infection stages of Colletotrichum, tea plant resistance mechanisms, and induced plant defence against Colletotrichum is essential to support sustainable disease management practices in the field. This review therefore summarizes the current knowledge of the identified causative agent of tea plant anthracnose, the infection strategies and pathogenicity of C. gloeosporioides, anthracnose disease resistance mechanisms, and the caffeine-induced defence response against Colletotrichum infection. The information reported in this review will advance our understanding of host–pathogen interactions and eventually help us to develop new disease control strategies.     

广西杧果病虫害调查初报

对广西24个市(县)杧果病虫害进行了系统的调查。发现病害有20种,主要为白粉病、炭疽病、细菌性角斑病、蒂腐病等,其中杧果水泡病、杧果畸型病和杧果红点病为国内新发现的杧果病害;发现害虫94种,隶属于8目38科,主要为杧果小齿螟Pseudonoorda minor Munroe、杧果横线尾夜蛾Chlumetia transversa Walker、杧果茶黄蓟马Scirtothrips dorsalis Hood、桔小实蝇Dacus dorsalis Hendel、杧果扁喙叶蝉Idioscopus incertus Baker等。腹足纲有害生物1种。     

Role of Antifungal Gallotannins,Resorcinols and Chitinases in the Constitutive Defence of Immature Mango (Mangifera indica L.) against Colletotrichum gloeosporioides

Conidia of Colletotrichum gloeosporioides germinate and form infection hyphae on inoculated, immature mango but remain quiescent until fruit ripening. Antifungal resorcinols have previously been implicated for quiescence of C. gloesoporioides and Alternaria alternata on mango. This study revealed the presence of a mixture of several gallotannins with glycosidic linkages, including 1,2,3,4,6‐penta‐O‐galloyl‐β‐D‐glucopyranose, with significant antifungal activity in the unripe mango fruit peel. Gallotannin antifungal activity was greater in a cultivar resistant (295.8 mm² inhibition) to anthracnose than in a susceptible (148.4 mm² inhibition) cultivar. In both, the activity decreased with ripening but the decrease was 10% less in the resistant cultivar. Three recorcinols, 5‐pentadecylresorcinol, 5‐(12‐cis‐heptadecenyl)resorcinol, AR 21 and another resorcinol derivative were present in the unripe fruit peel and all declined during ripening, more significantly the 5‐(12‐cis‐heptadecenyl)resorcinol and AR 21. Mango latex, when drained out, separates into an oily and aqueous phase. The aqueous phase showed significant chitinase activity and the ability to digest conidia of C. gloeosporioides. The oily phase has previously been reported to contain resorcinols. Draining fruits of latex soon after harvest resulted in greater incidence and severity of anthracnose at ripe stage. Chitinase activity was less in the peel of fruits from which latex was drained. The evidence suggests that the resistance of unripe mango to C. gloeosporioides is because of an elaborate constitutive defence system comprising antifungal resorcinols, gallotannins and chitinases.     

Evaluation of Water Yam (Dioscorea alata L.) Germplasm for Reaction to Yam Anthracnose and Virus Diseases and their Effect on Yield

Use of genetic resistance is the most practical and economic way to manage major diseases of yams. In a search for sources of resistance, 40 water yam (Dioscorea alata L.) accessions from Benin, Ghana, Nigeria and Puerto Rico were screened under natural disease infection conditions in Ibadan, Nigeria. The accessions were evaluated at 1, 3 and 6 months after planting (MAP) for severity of yam anthracnose and viral diseases. The effect of the pathogens on yield was also evaluated at harvest 9 MAP. There were significant differences (P < 0.001) between accessions for severities of anthracnose and viral diseases. Eight (20%) of them had lower anthracnose area under disease progress curves (AUDPC) values than the resistant check while 10 (25%) had AUDPC values below the trial mean. There were significant variations (P < 0.001) in yield components among the accessions. There was significant negative correlation of anthracnose severity with fresh tuber yield (r = −0.51) and with number of tubers per plot (r = −0.40). Similarly, significant negative correlations were observed of virus disease severity with fresh tuber yield (r = −0.78) and number of tubers per plot (r = −0.65). Linear regression models also showed that the fresh yield had significant negative relationships with anthracnose (R² = 0.26) and viral (R² = 0.62) diseases. The accessions identified as resistant constitute a valuable resource for breeding of resistant germplasm.     

不同油茶无性系炭疽病抗性及主要生理指标分析

为选育对炭疽病高抗的油茶(Camellia oleifera)良种,以5个闽西当地的油茶无性系为材料,采用病情指数法进行调查,评价其炭疽病抗性,并测定各无性系叶片的过氧化物酶(POD)、超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性及可溶性蛋白含量等指标,分析油茶炭疽病抗性与其主要生理指标的相关性。结果表明,参试的5个油茶无性系中,高抗无性系1个(新罗2号)、中抗无性系3个(河田2号、红山1号、漳平2号)、低感无性系1个(武平2号);不同无性系之间的POD、SOD、CAT活性及可溶性蛋白含量差异均达极显著水平(P<0.01)。相关性分析显示,POD活性、SOD活性、可溶性蛋白含量与油茶炭疽病的感病指数成负相关,CAT活性与感病指数的相关性未达显著水平。综上所述,不同油茶无性系对炭疽病的抗性各不相同,且其主要生理指标与抗性有一定的相关性,其中新罗2号无性系对炭疽病抗性强,主要生理指标也相对较高。该结果为油茶抗炭疽病良种选育及抗病机理研究提供依据。     

A PCR-based molecular marker applicable for marker-assisted selection for anthracnose disease resistance in lupin breeding

Selection for anthracnose disease resistance is one of the major objectives in lupin breeding programs. The aim of this study was to develop a molecular marker linked to a gene conferring anthracnose resistance in narrow-leafed lupin (Lupinus angustifolius L.), which can be widely used for MAS in lupin breeding. A F(8)derived RIL population from a cross between cultivar Tanjil (resistant to anthracnose) and Unicrop (susceptible) was used for marker development. DNA fingerprinting was conducted on 12 representative plants by combining the AFLP method with primers designed based on conserved sequences of plant disease resistance genes. A co-dominant candidate marker was detected from a DNA fingerprint. The candidate marker was cloned, sequenced, and converted into a sequence-specific, simple PCR based marker. Linkage analysis based on a segregating population consisting of 184 RILs suggested that the marker, designated as AntjM2, is located 2.3 cM away from the R gene conferring anthracnose resistance in L. angustifolius. The marker has now being implemented for MAS in the Australian national lupin breeding program.     

Mechanisms governing the responses to anthracnose pathogen in Juglans spp

Juglans nigra and Juglans regia are two highly economically important species for wood and fruit production that are susceptible to anthracnose caused by Gnomonia leptostyla. The identification of genotypes resistant to anthracnose could represent a valid alternative to agronomic and chemical management. In this study, we analyzed 72 walnut genotypes that showed a variety of resistance phenotypes in response to natural infection. According to the disease severity rating and microsatellite fingerprinting analysis, these genotypes were divided into three main groups: (40) J. nigra resistant, (1) J. nigra susceptible, and (31) J. regia susceptible. Data on leaf emergence rates and analysis of in vivo pathogenicity indicated that the incidence of anthracnose disease in the field might be partially conditioned by two key factors: the age and/or availability of susceptible leaves during the primary infection of fungus (avoidance by late flushing) and partial host resistance. NBS profiling approach, based on PCR amplification with an adapter primer for an adapter matching a restriction enzyme site and a degenerate primer targeting the conserved motifs present in the NBS domain of NBS-LRR genes, was applied. The results revealed the presence of a candidate marker that correlated to a reduction in anthracnose incidence in 72 walnut genotypes.     

Morphological and molecular screening of French bean (Phaseolus vulgaris L.) germplasm using SCAR markers for Colletotrichum lindemuthianum (Sacc. and Magn.) Scrib. causing anthracnose resistance

This study was carried out to screen Phaseolus vulgaris L. germplasm accessions for anthracnose resistance genes to the fungus Colletotrichum lindemuthianum. (Sacc. and Magn.) Scrib. This fungus is made up of many pathogenic races which poses a challenge in developing resistant plant varieties; however, screening for and selection of resistant plant sources plays an important role in developing resistant plant lines. This screening work involved examining 69 accessions consisting of two resistant lines (D-line, L-line) and two susceptible varieties (Kanchana and Jwala). Fourteen SCAR primers specific to anthracnose disease resistance in the French bean were used. Of these 14 SCAR primers, 5 of them, SAS13, SF10, SC08, SZ04 and SBB14, produced amplification with good monomorphic bands.