Sources of resistance to Pseudocercospora fijiensis,the cause of black Sigatoka in banana

Black Sigatoka, caused by Pseudocercospora fijiensis, is one of the most devastating diseases of banana. In commercial banana-growing systems, black Sigatoka is primarily managed by fungicides. This mode of disease management is not feasible for resource-limited smallholder farmers. Therefore, bananas resistant to P. fijiensis provide a practical solution for managing the disease, especially under smallholder farming systems. Most banana and plantain hybrids with resistance to P. fijiensis were developed using few sources of resistance, which include Calcutta 4 and Pisang Lilin. To broaden the pool of resistance sources to P. fijiensis, 95 banana accessions were evaluated under field conditions in Sendusu, Uganda. Eleven accessions were resistant to P. fijiensis. Black Sigatoka symptoms did not progress past Stage 2 (narrow brown streaks) in the diploid accessions Pahang (AA), Pisang KRA (AA), Malaccensis 0074 (AA), Long Tavoy (AA), M.A. Truncata (AA), Tani (BB), and Balbisiana (BB), a response similar to the resistant control Calcutta 4. These accessions are potential sources of P. fijiensis resistance and banana breeding programmes can use them to broaden the genetic base for resistance to P. fijiensis.     

Genetic variability of Pseudocercospora fijiensis,the black Sigatoka pathogen of banana (Musa spp.) in Mexico

Pseudocercospora (previously known as Mycosphaerella) fijiensis causes black Sigatoka disease in banana (Musa spp.) and is considered to be the most devastating pathogen of this crop worldwide. To improve knowledge of its evolutionary patterns, this study determined the genetic variability of populations from two regions of Mexico: Central Pacific (Colima and Michoacan) and Southern (Chiapas, Tabasco and Oaxaca), using 10 simple sequence repeat (SSR) loci and the MAT-specific PCR assay. Both mating types were present in all regions under study, with frequencies of 63% MAT1-1 and 37% MAT1-2. The SSR markers showed an average of three alleles per locus, resulting in 34 alleles in total. The genetic diversity (H_T) was 0.3308, but at the local level (H_S) ranged from 0.0976 (Colima) to 0.2228 (Oaxaca). However, the genotypic diversity was usually high (H′ > 2.4, S > 0.89). Cluster analysis grouped the isolates into five clusters with high statistical support (au > 80%), suggesting a geographic organization of the genetic variability of P. fijiensis; AMOVA, the minimum spanning tree and the population structure analysis supported this result, and all data indicated that the major genetic differences were between the different populations under analysis. Thus, the high level of genetic variability in P. fijiensis is attributed partly to a high rate of sexual reproduction, and also to a strong evolutionary capacity coupled with isolation due to limited genetic flow between distant populations. Both possibilities could be playing a relevant role in population differentiation of the pathogen.     

Evaluation of strobilurins,acibenzolar and other chemicals,alone and in spray programs for the control of yellow Sigatoka leaf spot (Mycosphaerella musicola) of bananas in far northern Queensland,Australia

Abstract

Several chemicals including the strobilurins (trifloxystrobin, azoxystrobin, pyraclostrobin and DPX KZ 165), a plant activator (acibenzolar), the triazoles (propiconazole, tebuconazole, epoxiconazole, fenbuconazole and JAU 6475) and tridemorph, spiroxamine, pyrimethanil, fenarimol and various formulations of mancozeb were evaluated in three field experiments in northern Queensland, Australia for control of yellow Sigatoka of banana (caused by Mycosphaerella musicola). In all experiments, the strobilurins used alone or in spray programs with mancozeb and acibenzolar were as effective or better than the industry standards mancozeb and propiconazole. Acibenzolar used in spray programs with mancozeb significantly improved the control of Sigatoka compared to mancozeb alone. The triazoles, epoxiconazole, fenbuconazole and JAU 6476 used alone and tebuconazole in a spray program with mancozeb were as effective as the industry standard propiconazole. Tridemorph, pyrimethanil and spiroxamine were as effective as the industry standard mancozeb, and fenarimol failed to effectively control the disease. In 2004, trifloxystrobin, pyraclostrobin and epoxiconazole were registered for control of yellow Sigatoka of banana.     

应用RAPD标记技术鉴定香蕉褐缘灰斑病菌

采用Mycosphaerella fijiiensis和M.musicola的RAPD标记技术鉴定海南香蕉褐缘灰斑病菌,结果表明,分离自海南儋州、乐东、文昌、东方、澄迈、临高、琼海、昌江、琼山、三亚香蕉上的10个菌株均为M.fijiensis,引起香蕉黑叶条斑病。     

Genetic structure of Mycosphaerella fijiensis populations from Australia, Papua New Guinea and the Pacific Islands

Single-copy restriction fragment length polymorphism (RFLP) markers were used to determine the genetic structure of Mycosphaerella fijiensis , the cause of black leaf streak (black Sigatoka) disease of banana and plantain, in the Torres Strait, Papua New Guinea (PNG), and the Pacific Islands. A moderate level of genetic variation was observed in all populations with genotypic diversity values of 60–78% of the theoretical maximum, and gene diversity ( H ) values between 0·269 and 0·336. All populations were at gametic equilibrium, and with the high level of genotypic diversity observed this indicated that sexual reproduction has a major role in the genetic structure of the M. fijiensis populations examined. Population differentiation was tested on several hierarchical scales. No evidence of population differentiation was observed between sites on Mer Island. A moderate level of population differentiation was observed within the Torres Strait, between Badu and Mer Islands ( F _ST = 0·097). On a regional scale, the greatest differentiation was found between the populations of the Torres Strait and the Pacific. Populations from these regions were more closely related to the PNG population than to each other, suggesting they were founded in separate events from the same population.     

广西部分烟区烟草曲叶病病原的分子鉴定

[目的]明确广西西部地区靖西(JX)、凌云(LY)、德保(DB)和乐业(LeY)等4个县市烟草曲叶病的病原。[方法]2010年5-6月分别从广西靖西、凌云、德保和乐业等县市采集具有典型曲叶症状的烟草叶片,用基于双生病毒DNA保守序列设计简并引物Bego-1和Bego-6对病叶组织总DNA抽提物进行PCR扩增和对PCR产物进行序列测定,用BLAST、Vector NTI、MEGA 4.0和Simplot program 3.2软件等进行病毒序列分析、系统进化树构建和病毒重组分析。[结果]从选取的9个表现典型曲叶症状的样品叶组织总DNA抽提物中均可扩增出约1500bp与预期大小相符的DNA片段。测序和序列比对分析显示,9个样品扩增产物核苷酸序列相似性为73.7%～99.2%,与已报道的双生病毒具较高的相似性。其中,JX-2与中国番茄曲叶病毒广西番茄分离物(G32)的相似性最高,达99.2%;JX-3和JX-5与云南胡椒曲叶病毒云南辣椒分离物(YN323)相似性最高,分别为92.5%和93.4%;LeY-1、LY-1、DB-1、JX-1、JX-4和JX-6则与中国番茄黄化曲叶病毒中国番茄分离物(CHI)和广西烟草分离物(G102)的相似性最高,均高于95.0%。基于PCR扩增产物及已报道的双生病毒属代表种相应核苷酸序列构建的系统进化树分析表明,9个广西烟草分离物分属3个簇群:中国番茄曲叶病毒簇、云南辣椒曲叶病毒簇和中国番茄黄化曲叶病毒簇。重组分析结果表明:JX-3是云南辣椒曲叶病毒和中国番茄曲叶病毒的重组病毒,JX-5是云南辣椒曲叶病毒和中国番茄黄化曲叶病毒的重组病毒。[结论]9个广西烟草分离物分属于4种双生病毒:中国番茄曲叶病毒和中国番茄黄化曲叶病毒,以及分别由上述两种病毒与云南辣椒曲叶病毒重组而来的2种重组病毒。其中,中国番茄曲叶病毒自然侵染烟草、云南辣椒曲叶病毒和中国番茄曲叶病毒及中国番茄黄化曲叶病毒的重组病毒等结果此前均未见报道。     

甘肃省红豆草病原真菌鉴定及病害发生动态调查

为明确甘肃省红豆草的病害种类、发生动态和危害状况,采用病原物分离与培养、形态学及分子生物学鉴定和致病性测定确定红豆草病害种类,于2012—2013年在通渭、渭源、榆中和碌曲4县调查各病害的发病率以确定发生动态,观察病害田间发生特点并结合调查数据评价其重要性。结果表明,4县共发生真菌性病害12种,分别为大茎点霉叶斑病(病原为大茎点霉属真菌Macrophoma sp.)、壳针孢叶斑病(病原为歪头菜壳针孢Septoria orobina)、炭疽病(病原为白蜡树刺盘孢Colletotrichum spaethianum)、黑秆病(病原为红豆草壳二孢Ascochyta onobrychis、菠菜刺盘孢C.spinaciae和链格孢Alternaria alternata混合侵染)、壳二孢叶斑病、茎点霉叶斑病、尾孢叶斑病、柱格孢白斑病、匍柄霉叶斑病、链格孢黑斑病、锈病和白粉病,其中大茎点霉属真菌、白蜡树刺盘孢和菠菜刺盘孢在红豆草上首次发现;尾孢叶斑病和壳针孢叶斑病为甘肃新记录病害;大茎点霉叶斑病为世界新病害,仅于碌曲县发现。白粉病、锈病、链格孢黑斑病发生于红豆草生长后期,其它病害则始于6月;6—9月危害加重的为黑秆病和柱格孢白斑病,发病率最高达89.7%和96.0%;危害渐轻的为茎点霉叶斑病、壳二孢叶斑病和壳针孢叶斑病,发病率最高达88.7%、57.4%和45.1%。黑秆病和茎点霉叶斑病在甘肃省目前危害最重。     

Phytoanticipins from banana (<Emphasis Type="Italic">Musa acuminata</Emphasis> cv. Grande Naine) plants,with antifungal activity against <Emphasis Type="Italic">Mycosphaerella fijiensis</Emphasis>, the causal agent of black Sigatoka

The previously detected antifungal activity against Mycosphaerella fijiensis of aqueous infusions of healthy banana (Musa acuminata cv. Grande Naine) leaves, suggested the production of phytoprotectants by the plant. The bioassay-guided VLC-purification of the lyophilized infusion of the leaves of 4-month old healthy banana (M. acuminata cv. Grande Naine) plants, resulted in the purification of a phytoanticipin with strong antifungal activity against M. fijiensis Morelet, the causal agent of black Sigatoka, the most destructive and devastating disease of bananas and plantains in the world. The LC-MS analysis of the purified phytoanticipin suggests a steroidal saponin structure with four sugar units attached to the C-3 position of a diosgenin-like aglycone. This represents the first report of phytoanticipins occurring in M. acuminata.     

香蕉枯萎病菌生理小种鉴定及其SCAR标记

 通过室内人工接种蕉类鉴别寄主,对采集于广东蕉区的18个蕉类枯萎病菌菌株进行鉴定,KP021、KP022、GZ981和JL021 4个菌株属Racel,其余14个菌株属Race4,说明广东蕉区同时存在尖孢镰刀菌古巴专化型Race1和Race4。用RAPD技术对上述18个菌株进行分析,从200条随机引物中筛选出8条引物可产生生理小种RAPD标记12个,其中标记Racel的8个,标记Race4的4个。对这些RAPD标记带分别进行回收、克隆、测序,根据这些特异片段序列分别设计相应的SCAR引物,通过对18个菌株的PCR扩增检验,有4个RAPD标记成功地转化为SCAR标记,其中Race1-SCAR标记1个、Race4-SCAR标记2个、同时能鉴定出2个小种的SCAR标记1个。应用这4个SCAR标记同时对采自田间的9个病菌分离物进行检测,能够准确地鉴定出广东蕉区的尖孢镰刀菌古巴专化型Racel和Race4,这为下一步开展香蕉枯萎病菌生理小种的分子鉴定及各生理小种田间流行动态监测奠定了基础。     

三叶木通叶斑病病原菌鉴定

叶斑病是三叶木通生长过程中一种严重的病害.为明确引起叶斑病的病原,从三叶木通人工栽培基地叶斑病样本上分离病原菌,对病原菌进行致病性测定、显微形态与分子鉴定.结果表明,三叶木通叶斑病病原菌为链格孢属细极链格孢(Alternaria tenuissima).     

Aetiology and causal agents of mango sudden decline disease in the Sultanate of Oman

Mango sudden decline is a recently introduced, economically serious disease in Oman. Affected mango trees have wilting symptoms that usually begin on one side and later spread to involve the entire tree. Trees exude amber-coloured gum from the bark of their trunks or branches and vascular tissues are discoloured. Having entered Oman in the recent past, survey data is presented that shows the disease to have spread throughout the northern part of the country. Evidence is presented that the vascular wilt pathogen Ceratocystis fimbriata causes mango sudden decline disease in Oman, possibly in concert with Lasiodiplodia theobromae and the recently described Ceratocystis omanensis. Isolates of these fungi from affected trees, cause infection and can be recovered from inoculated seedlings. Bark beetles (Hypocryphalus mangiferae) are shown to carry C. fimbriata and L. theobromae and are presumably responsible for transmitting both pathogens to healthy mango trees. Acting as a wounding agent and vector, the bark beetle is likely to have assisted the rapid spread of the disease across Oman.     

TaqMan探针实时荧光PCR快速检测苹果牛眼果腐病菌方法的建立

苹果牛眼果腐病菌(Neofabraea malicorticis、N.perennans、N.alba和N.kienholzii)是我国检疫性植物病原真菌,为建立该病菌的实时荧光PCR检测法,根据病菌及其近缘种的翻译延伸因子(EF-1α)的保守序列设计了特异性探针,分别以苹果牛眼果腐病菌菌株和本研究构建的EF-1α重组质粒DNA为阳性标准品检验探针的特异性和灵敏度。结果显示探针MAL-P、PER-P、ALB-P和KIE-P分别对N.malicorticis、N.perennans、N.alba和N.kienholzii表现特异性阳性扩增,而与近缘种及其他常见的果腐病菌无交叉反应,单重探针和4种探针混合液的灵敏度分别达1 fg/μL和10 fg/μL DNA。总计从美国、智利、新西兰、法国进境截获的29批可疑病果的分离物中检测到PCR阳性荧光信号,包括20批N.perennans、8批N.alba和1批N.kienholzii。该方法在6 h内即可完成整个检测流程,其特异性强、灵敏度高,适用于实际样品中苹果牛眼果腐病菌的快速检测。     

Physiological effects of the hydrophilic phytotoxins produced by <Emphasis Type="Italic">Mycosphaerella fijiensis</Emphasis>, the causal agent of black sigatoka in banana plants

Although Mycosphaerella fijiensis, the causal agent of black sigatoka disease of banana, has been known to produce numerous lipophilic host-selective (HSTs) and nonhost selective phytotoxins (non-HSTs), only recently we have reported that the pathogen also produces hydrophilic phytotoxins. Here we examined the effect of light on the toxicity of the hydrophilic phytotoxins and estimated the electrolyte leakage and H₂O₂ and superoxide generation in detached banana leaves to study their mode of action at the cellular level. Nonhost plant species were also tested to determine whether the toxins are HSTs or non-HSTs. Our results suggest that the hydrophilic phytotoxins are non-HSTs, that their phytotoxicity is not light dependent, and that they may act at the plasma membrane by altering permeability through oxidative damage, by inducing ROS production as part of their mechanism of action.     

<Emphasis Type="Italic">Teratosphaeria (Mycosphaerella) nubilosa,</Emphasis> the causal agent of Mycosphaerella leaf disease (MLD), recently introduced into Uruguay

Mycosphaerella leaf disease on Eucalyptus is well known in Uruguay but none of the more serious Mycosphaerella spp. and Teratosphaeria spp. causing this disease have yet been found. In the autumn of 2007, more severe defoliation than has been known in the past and associated with symptoms resembling Mycosphaerella infections was observed on Eucalyptus globulus. Isolations and identifications based on morphology and DNA sequence comparisons showed that the causal agent of the defoliation is the well known and serious pathogen Teratosphaeria nubilosa (=Mycosphaerella nubilosa). This is the first record of the pathogen in South America. Using ten microsatellite loci previously developed for T. nubilosa, only one multilocus haplotype was found from 46 T. nubilosa collected isolates. Interestingly, this haplotype was the same as one previously found in Portugal and Spain. The results suggest that T. nubilosa has recently been introduced into Uruguay and that it most likely originated from the Iberian Peninsula where E. globulus is widely planted.     

Sources of resistance in Musa to Xanthomonas campestris pv. musacearum,the causal agent of banana xanthomonas wilt

It is claimed that, with the exception of Musa balbisiana, all banana varieties are susceptible to bacterial wilt caused by Xanthomonas campestris pv. musacearum (Xcm). Despite being resistant to Xcm infection, M. balbisiana is not preferred for breeding because it belongs to the BB genome subgroup, while most edible bananas are of the A genome. To identify potential sources of resistance to Xcm, 72 banana accessions representing the Musa genetic diversity were evaluated in an outdoor confined potted trial. The midribs of the youngest leaf of 3-month-old banana plants were inoculated with 10⁸ CFU mL⁻¹ of Xcm isolate USY13P, and symptom development assessed weekly for 4 months. Results confirmed that M. balbisiana genotypes are indeed resistant to Xcm. Varieties within the Musa acuminata subsp. zebrina (AA) set were further identified as potentially useful sources of Xcm resistance. These findings reveal the potential to develop banana and plantain varieties with tolerance to Xcm.     

Assessment of PCR-based tools for the specific identification of some temperate Meloidogyne species including M. chitwoodi,M. fallax and M. minor

Several conventional PCR tests have been developed for the identification of the European quarantine root-knot nematodes Meloidogyne chitwoodi and M. fallax but data are lacking for the evaluation of their performance in terms of sensitivity, repeatability, reproducibility and specificity against a large range of populations. This study evaluated the performance criteria of three conventional PCR tests recommended by the consensus diagnostic protocol for Meloidogyne chitwoodi and Meloidogyne fallax published by the European and Mediterranean Plant Protection Organization (EPPO): a species-specific PCR (IGS target), a SCAR PCR, and a rDNA ITS PCR-RFLP. Evaluation was carried out with DNA extracts from juveniles, males and females according to EPPO recommendations for test validation. A minimum of 34 populations of target and non target nematode species were tested to check the specificity of these three PCR assays. The three PCR tests were ranked according to their specificity (with regard to cross reaction with other nematodes species or genus) and their sensitivity (detection of a single juvenile or mixed with other species). The species-specific PCR proved to be more sensitive but less specific than the SCAR PCR. The PCR-RFLP enables the identification of several Meloidogyne species but profile analysis can be difficult when several species are present in the mixture. Specific PCR products and RFLP profiles were also observed for M. arenaria and M. enterolobii, and described for M. minor and M. artiellia.     

Species-specific DNA markers for identification of two root-knot nematodes of coffee: Meloidogyne arabicida and M. izalcoensis

Seven root-knot nematodes (RKN), including Meloidogyne exigua, M. incognita, M. paranaensis, M. enterolobii, M. arabicida, M. izalcoensis and M. arenaria are major pathogens of coffee crop in the Americas. Species-specific primers for their identification have been developed for five of them and constitute a fast and reliable method of identification. Here we report a PCR-based assay for specific detection of M. arabicida and M. izalcoensis. Random Amplified Polymorphic DNA fragments specific for these two species were converted into sequence characterized amplified region (SCAR) markers. PCR amplification using the SCAR primers produced a specific fragment of 300 bp and 670 bp for M. arabicida and M. izalcoensis, respectively, which were absent in other coffee-associated Meloidogyne spp. tested. SCAR primers also allowed successful amplification of DNA from single second-stage juveniles (J2), males and females. In addition, these primers were able to unambiguously detect the target species in nematode suspensions extracted from soil and roots samples, in different isolates of the same species or when used in multiplex PCR reactions containing mixtures of species. These results demonstrated the effectiveness of these SCAR markers and their multiplex use with those previously developed for M. exigua, M. incognita, M. paranaensis, M. enterolobii and M. arenaria constitute an essential detection tool. This diagnostic kit will contribute for specific J2 identification of the major RKN infecting coffee from field samples in the Americas.     

海南香蕉污斑病病原鉴定及防治试验初报

通过发病蕉叶组织分离培养和回接致病性测定表明,在海南省发生的香蕉污斑病是由半知菌亚门丝核菌(Rhizoctoniaspp.)侵染所致。调查发现,高温多湿、通风透光较差和荫蔽的蕉园发生重。采用清洁蕉园和药剂防治,取得了较好防效。     

广西香蕉煤污病病原菌的分离鉴定及其生物学特性

香蕉是我国南方的重要特色作物,香蕉煤污病的发生降低了果实品质,严重影响其市场价值,给蕉农造成经济损失.为明确广西香蕉上新发生煤污病的病原及其生物学特性,采用单孢分离法获得病原菌株BFMY-2,并依据柯赫氏法则验证其为广西香蕉煤污病的病原菌.通过rDNA-ITS和SSU序列分析并结合形态学鉴定,将病原菌鉴定为枝状枝孢霉(...