Population Genetic Structure and Host Specificity of Alternaria spp. Causing Brown Spot of Minneola Tangelo and Rough Lemon in Florida

ABSTRACT Alternaria spp. were sampled from two rough lemon (RL) and two Minneola tangelo (MIN) groves in a limited geographic area in central Florida to test for host-specialized forms of the pathogen. Isolates of Alternaria spp. were scored for variation at 16 putative random amplified polymorphic DNA (RAPD) loci and for pathogenicity on both hosts. Subpopulations on each host were differentiated genetically and pathogenically, which was consistent with the hypothesis of host specialization. Highly significant genetic differentiation was detected among all four subpopulations (Nei's coefficient of gene differentiation [G(ST)] = 0.292, P = 0.000); most of the differentiation occurred between hosts (G(ST) = 0.278, P = 0.000). Phenograms of qualitative similarities among isolates within subpopulations revealed two or three distinct clusters of isolates within each subpopulation. The majority of isolates sampled from RL were pathogenic on RL and not on MIN, although a few RL isolates were able to induce disease on MIN, and 44% were nonpathogenic on either host. In contrast, isolates from MIN were pathogenic only on MIN, never on RL, and only 3% of the isolates were nonpathogenic. Overall, three genetically distinct clusters of isolates were detected on both hosts. One of the clusters (cluster A) sampled from RL was pathogenic on RL and not on MIN and consisted almost entirely of one RAPD genotype. This cluster also contained two isolates that were 93% similar to the majority genotype but were pathogenic on MIN and not RL. In isolates from MIN, two distinct clusters of isolates were found in one subpopulation (clusters B and C), and three distinct clusters were found in another subpopulation (clusters A, B, and C). Clusters A and B were found on both hosts, while cluster C was limited to MIN. Populations of Alternaria spp. sampled from RL and MIN showed a high degree of host specificity; however, the specificity obscured a high level of genetic variation within subpopulations.     

Genetic Differentiation and Host Specificity Among Populations of Alternaria spp. Causing Brown Spot of Grapefruit and Tangerine x Grapefruit Hybrids in Florida

ABSTRACT Alternaria spp. were sampled from brown spot lesions in several geographically separated citrus groves and different grapefruit and tangerine x grapefruit hybrid cultivars in Florida and screened for variation at 16 putative random amplified polymorphic DNA loci. Populations of the pathogen on two hybrids, Minneola and Orlando, in five locations throughout Florida were moderately differentiated (Nei's coefficient of gene differentiation [G(ST)] = 0.12) among locations. The hypothesis that host-specialized forms of Alternaria spp. cause brown spot on different Citrus spp. and cultivars was tested by estimating genetic differentiation among isolates sampled from different hosts and by pathogenicity assays. Isolates sampled from grapefruit and the hybrid cv. Nova were genetically distinct from isolates sampled from other hybrid cultivars including Robinson, Sunburst, Minneola, Orlando, and Murcott. No differentiation could be detected among isolates sampled from this latter group of hybrids. Quantitative pathogenicity assays on leaves using spray inoculation revealed that 'Nova' isolates were not significantly more pathogenic on 'Nova' compared with isolates from 'Minneola' and 'Orlando'. Similarly, grapefruit isolates were not significantly more pathogenic on grapefruit compared with isolates from 'Minneola'. Isolates from all hosts had similar disease rankings on each inoculated cultivar, with 'Minneola' the most susceptible, followed in decreasing order of susceptibility by 'Orlando', 'Sunburst', 'Nova', and 'Duncan' grapefruit. Rough lemon was generally immune to all isolates tested; however, occasional brown spot lesions were observed on leaves of this host with isolates from grapefruit. No evidence was found to support the hypothesis that unique genotypes of the pathogen, which are more virulent on 'Sunburst' or grapefruit, have been introduced to Florida. Populations of Alternaria spp. causing brown spot of citrus on grapefruit and 'Nova' in Florida are genetically distinct from isolates on other cultivars, and we speculate that these populations are in the early stages of adaptation to and possible speciation on these hosts.     

Pathogenicity of Aphanomyces spp. from Different Leguminous Crops in Sweden

Host range and pathogenicity of a range of Aphanomyces spp. isolates obtained from pea roots but also from a range of other field-grown leguminous crops in southern Sweden was investigated. The Aphanomyces euteiches isolates originating from pea and the few obtained isolates originating from alfalfa, green bean and yellow sweet-clover were highly pathogenic only to pea. The A. euteiches isolated from common vetch differed from these isolates by being weakly pathogenic to pea and other legumes, but highly pathogenic to common vetch. Vetch isolates also formed a well-defined separate cluster based on principal component analysis of pathogenicity pattern on tested crops. Oospores of A. euteiches were observed in root tissue of pea as well as common vetch, alfalfa, green bean, broad bean, red clover and yellow sweet-clover in the greenhouse pathogenicity tests. An Aphanomyces sp. that morphologically differed from A. euteiches, was frequently isolated from several leguminous plants, but was non-pathogenic to all tested crops in the pathogenicity tests. In isozyme analysis the banding pattern of these isolates resembled the pattern of A. cladogamus. Another, different and so far unidentified Aphanomyces sp. from roots of green bean and broad bean, was also non-pathogenic to the tested legume species. Based on the isolates tested, the results obtained suggest that the population of Aphanomyces spp. infecting legume roots in Sweden consists of a pea-specific and a vetch-specific group of A. euteiches. Two other groups comprised (i) Aphanomyces sp. isolates that resembled A. cladogamus, and (ii) isolates, which resembled neither A. euteiches nor A. cladogamus. In addition, the host range of Swedish A. euteiches isolates was not as broad as reported for A. euteiches isolates from other countries.     

Comparison of Colletotrichum orbiculare and Several Allied Colletotrichum spp. for mtDNA RFLPs, Intron RFLP and Sequence Variation, Vegetative Compatibility, and Host Specificity

ABSTRACT Based on spore morphology, appressorium development, sequence similarities of the rDNA, and similarities in amplified restriction fragment length polymorphism (AFLP), it has been proposed that Colletotrichum orbiculare, C. trifolii, C. lindemuthianum, and C. malvarum represent a single phylogenetic species, C. orbiculare. In the current study, the phylogenetic relationship among isolates in the C. orbiculare species complex was reassessed. In all, 72 isolates of C. orbiculare from cultivated cucurbit or weed hosts, C. trifolii from alfalfa, C. lindemuthianum from green bean, and C. malvarum from prickly sida (Sida spinosa) were examined for mitochondrial DNA (mtDNA) restriction fragment length polymorphisms (RFLPs), RFLPs and sequence variation of a 900-bp intron of the glutamine synthetase gene and a 200-bp intron of the glyceraldehyde-3-phosphate dehydrogenase gene, and vegetative compatibility. In addition, host specificity was examined in foliar inoculations on cucurbit, bean, and alfalfa hosts. Inoculations also were conducted on cucumber fruit. Genetically distinct isolates, based on vegetative compatibility, within the species complex (C. orbiculare, C. trifolii, and C. malvarum) had an identical mtDNA haplotype (haplotype A) when examined with each of three different restriction enzymes. Isolates of C. lindemuthianum had a very similar mtDNA haplotype to haplotype A, with a single polymorphism detected with the enzyme HaeIII. The four species represent a phylogenetically closely related group based on a statistical analysis of the 900- and 200-bp intron sequences. However, distinct RFLPs in the 900-bp intron were consistently associated with each species and could be used to qualitatively and quantitatively distinguish each species. Furthermore, each of the species showed distinct host specificity, with isolates of C. orbiculare (from cucurbits), C. lindemuthianum, and C. trifolii being pathogenic only on cucurbits, green bean, and alfalfa, respectively. Consequently, distinct and fixed nucleotide, or genotypic (intron sequences and RFLPs) and phenotypic (host specificity) characteristics can be used to distinguish C. orbiculare, C. lindemuthianum, and C. trifolii from one another; therefore, they should be recognized as distinct species. This species delineation is consistent with the most current species concepts in fungi. More isolates and further characterization is needed to determine whether C. orbiculare from cocklebur and C. malvarum represent distinct species. RFLPs of the 900-bp intron may represent a relatively inexpensive, reliable, and useful diagnostic tool for general species differentiation in the genus Colletotrichum.     

Identification and Pathogenicity of Rhizoctonia spp. Isolated from Apple Roots and Orchard Soils

ABSTRACT Rhizoctonia spp. were isolated from the roots of apple trees and associated soil collected in orchards located near Moxee, Quincy, East Wenatchee, and Wenatchee, WA. The anastomosis groups (AGs) of Rhizoctonia spp. isolated from apple were determined by hyphal anastomosis with tester strains on 2% water agar and, where warranted, sequence analysis of the rDNA internal transcribed spacer region and restriction analysis of an amplified fragment from the 28S ribosomal RNA gene were used to corroborate these identifications. The dominant AG of R. solani isolated from the Moxee and East Wenatchee orchards were AG 5 and AG 6, respectively. Binucleate Rhizoctonia spp. were recovered from apple roots at three of four orchards surveyed and included isolates of AG-A, -G, -I, -J, and -Q. In artificial inoculations, isolates of R. solani AG 5 and AG 6 caused extensive root rot and death of 2- to 20-week-old apple transplants, providing evidence that isolates of R. solani AG 6 can be highly virulent and do not merely exist as saprophytes. The effect of binucleate Rhizoctonia spp. on growth of apple seedlings was isolate-dependent and ranged from growth enhancement to severe root rot. R. solani AG 5 and AG 6 were isolated from stunted trees, but not healthy trees, in an orchard near Moxee, WA, that exhibited severe symptoms of apple replant disease, suggesting that R. solani may have a role in this disease complex.     

Pathogenicity of Diaporthe spp. on two blueberry cultivars (Vaccinium corymbosum)

Diaporthe vaccinii causes twig blight and fruit rot and is currently listed as a quarantine organism for the European Union. In the Netherlands, two species from the same genus, Diaporthe eres and Diaporthe rudis, are regularly isolated from blighted twigs of Vaccinium corymbosum. This study compared the pathogenicity of these two species to D. vaccinii. To develop a pathogenicity test a field experiment was performed at an experimental station, testing isolates of D. rudis and D. eres. Most of the isolates tested did not cause statistically significantly larger lesions compared to the control treatment (P > 0.05). In a greenhouse experiment a similar test was conducted, comparing D. vaccinii with the closely related species D. eres. Two cultivars of blueberry, Duke and Liberty, were inoculated using two D. vaccinii and two D. eres isolates. Stem canker lesions caused by D. vaccinii were larger than these caused by D. eres isolates. The majority of the lesions caused by D. vaccinii did not pass the first node of the inoculated shoot, which might be an explanation for the ‘mild’ symptoms of D. vaccinii in V. corymbosum, which appear not to cause crop losses. The results of this study suggest that D. vaccinii may not be a major threat to blueberry production in Europe. Further data is required to be able to conclude whether the quarantine status of D. vaccinii is still appropriate.     

不同种昆虫病原线虫对同一寄主个体的竞争及对线虫繁殖的影响

采用注射侵染和自然侵染法，测试了长尾斯氏线虫X-7品系、小卷蛾斯氏线虫All品系和嗜菌异小杆线虫H06品系对同一寄主大蜡螟幼虫的相互作用，及其对线虫繁殖的影响。结果表明，无论是注射侵染还是自然侵染法，对大蜡螟幼虫个体的致死竞争能力最强的是长尾斯氏线虫，其次是小卷蛾斯氏线虫，嗜菌异小杆线虫的致死能力最弱。在同一大蜡螟幼虫体内时。异小杆线虫的繁殖受斯氏线虫的不利影响较大，而两种斯氏线虫的繁殖受异小杆线虫的不利影响较小。两种斯氏线虫对各自的繁殖影响不大。     

Pathogenicity,host specificity and anastomosis groups ofRhizoctonia spp. isolated from soils in Israel

One hundred and twenty-nine isolates ofRhizoctonia spp. were obtained from soil samples in Israel and from culture collections in the U.S.A. and Japan. The isolates varied in host range and disease severity when tested on six to eleven different host plants. Approximately 30% of the isolates were nonpathogenic to all the host plants tested. Mycelial growth rate of the nonpathogenic strains did not differ significantly from that of the virulent isolates. The 107 Israeli isolates represented anastomosis groups (AG) 1, 2, 3, 4, 5 and 6 ofR, solani, two groups ofR. zeae, and three groups of binucleateRhizoctonia AG: A, F and K.     

偃麦草属E染色体组特异SCAR标记对Lr19的特异性和稳定性研究

 小麦抗叶锈基因Lr19来源于长穗偃麦草,表现优良的抗叶锈性,国内外发现对Lr19有毒性的小麦叶锈菌菌株的报道较少。本研究以TcLr19和Thatcher亲本以及TcLr19×Thatcher F₂代单株构建的分离群体为材料,建立了与Lr19共分离的稳定的SCAR分子标记,命名为Y₁₉SCAR₉₈₂。对49个小麦抗叶锈近等基因系材料的稳定性检测结果表明,其重复性好且为Lr19特异的分子标记。对120个小麦品种检测的结果表明,该标记可有效应用于小麦抗叶锈分子辅助选择育种。     

Pathogenicity and fungicide sensitivity of Pythium and Phytopythium spp. associated with soybean in the Huang-Huai region of China

Pythium and Phytopythium spp. cause seed decay, damping off, and root rot in soybean, wheat, and many other crops. However, their diversity and importance as pathogens, particularly in different crop rotation systems, are largely unknown. A survey was conducted in the Huang-Huai region, one of the main areas of soybean–wheat rotation farming in China. In 2016–2018, we collected 300 soybean seedlings and 150 field soil samples from several representative locations, and identified 26 Pythium and 6 Phytopythium spp. from 212 isolates, based on internal transcribed spacer 2 (ITS2) and cytochrome oxidase subunit 1 sequences. The pathogenicity of these isolates was evaluated by growing soybean and wheat seeds in dishes and pots containing oomycete cultures. We found that 12 Pythium spp. (but no Phytopythium spp.) showed high pathogenicity on soybean and/or wheat, and nine of them (75%) were highly pathogenic on both crops. Among the nine species, Pythium spinosum, Pythium ultimum, Pythium species 1 (tentatively designated as ‘Candidatus Pythium huanghuaiense’), Pythium aphanidermatum, and Pythium myriotylum were highly abundant among all isolates (15%, 10%, 9%, 8%, and 5%, respectively). Nine species were selected for testing of sensitivity to the fungicides metalaxyl and mefenoxam. The EC₅₀ values were all less than 10 μg/ml, indicating little resistance. Minimum inhibitory concentration values indicated isolates were about twice as sensitive to mefenoxam as to metalaxyl. These results provide a systematic understanding of Pythium and Phytopythium species associated with soybean in the Huang-Huai region, which is important for disease management and breeding programmes.     

真菌病毒减弱寄主致病性机理及其生防应用

真菌病毒是在真菌体内复制和增殖的病毒,在真菌中广泛存在。农业生产中70%~80%的植物病害由病原真菌侵染导致,真菌病毒能够减弱植物病原真菌的致病性,还可以将其转换为内生真菌从而促进植物生长和抗病,因此真菌病毒是一类重要的生防资源。本文综述了真菌病毒减弱寄主致病性机理及其在病害防治中的应用,同时提出了真菌病毒用于生物防治的前景和面临的问题,为植物真菌病害的生物防治提供参考。     

Pathogenicity of isolates of Magnaporthe spp. from wheat and grasses infecting seedlings and mature wheat plants in Argentina

Wheat blast of wheat (Triticum aestivum), caused by Magnaporthe oryzae pathotype triticum (MoT; anamorph Pyricularia oryzae) is a destructive disease in the South American countries of Brazil, Paraguay and Bolivia. In Argentina, the fungus was recently recorded on wheat and barley plants in the northeast part of the country, Buenos Aires and Corrientes Provinces, with a potential for spreading. This work aimed to study, for the first time, the morphocultural and pathogenic characteristics of Magnaporthe isolates collected from wheat and other herbaceous species in Argentina and three neighbouring countries (Paraguay, Brazil and Bolivia) and determine their aggressiveness on wheat varieties. Statistical differences among isolates, culture media, and development conditions were found for conidia colour, growth rate, size and sporulation rate. Pathogenicity tests performed on seedlings with 19 isolates of Magnaporthe spp. under greenhouse conditions showed a maximum disease severity of 55.3% and 66.7% for varieties BIOINTA 3004 and Baguette 18, respectively. Weed and grass isolates were infectious on wheat, demonstrating their potential epidemiological role on the disease. Spike disease severity was 34.6% for the host × pathogen interaction of BIOINTA 3004 × PY22. Observed symptoms included partial or total spike bleaching, and glume and rachis discolouration. The 1000‐grain weight was significantly reduced to 38.5% and 63.1% for cultivars BIOINTA 3004 and Baguette 18, respectively. The disease affected grain germination, which fell to 65.9% for seeds infected with the PYAR22 isolate. Symptoms observed in infected grains were partial spotting, grain softening, and rot symptoms with the presence of a greyish mould.     

Characterization, Genetic Structure, and Pathogenicity of Rhizoctonia spp. Associated with Rice Sheath Diseases in India

ABSTRACT Isolates of Rhizoctonia spp. were obtained from rice in India during 2000-2003. Characterization by conventional techniques and polymerase chain reaction showed that from 110 isolates, 99 were R. solani and 11 were R. oryzae-sativae. Of 99 isolates identified as R. solani, 96 were AG1-IA, 1 was AG1-IB, and 2 were AG1-IC. Amplified fragment length polymorphism (AFLP) analyzes were used to determine genetic relationships in Rhizoctonia pathogen populations collected from different geographic regions. Cluster analysis based on the AFLP data separated isolates belonging to the three different intraspecific groups of R. solani AG1 and differentiated R. solani from R. oryzae-sativae. Analysis of molecular variance (AMOVA) revealed that geographic region was the dominant factor determining population structure of R. solani AG1-1A; host cultivar had no significant effect. Pathogenicity tests on Oryza sativa cv. Zenith revealed that isolates of R. solani AG1-1A and AG1-1B were more virulent than R. solani AG1-IC and R. oryzae-sativae isolates.     

Genetic Analysis of Host Species Specificity of Magnaporthe oryzae Isolates from Rice and Wheat

ABSTRACT A Triticum isolate (pathogenic on wheat) of Magnaporthe oryzae was crossed with an Oryza isolate (pathogenic on rice) to elucidate mechanisms of their parasitic specificity on wheat. When the pathogenicity of their F (1) cultures (hybrids between a Triticum isolate and an Oryza isolate) was tested on wheat, avirulent and virulent cultures segregated in a 7:1 ratio. This result suggests that three loci are involved in avirulence of the Oryza isolate on wheat. One of the three loci conditioned papilla formation, whereas the others conditioned the hypersensitive reaction. Allelism tests revealed that the locus conditioning papilla formation is Pwt2 while one of the two loci conditioning the hypersensitive reaction is Pwt1. The other locus conditioning the hypersensitive reaction was different from any other known loci and, therefore, was designated as Pwt5.     

Interactions between Fusarium oxysporum f. sp. racheiphilum and Meloidogyne spp. in Vigna unguculata. 2. Specificity of different taxa

A California isolate of Meloidogyne javanica increased Fusarium wilt symptoms in cowpea cultivars California Blackeye No. 3 (CB3) (resistant to wilt) and Grant (tolerant) inoculated with each of the three races of Fusarium oxysporum f. sp. tracheiphilum. The same isolate of M. javancia did not similarly increase wilt in wilt-resistant cultivar CB7977 inoculated with two isolates of race 3 of F. o. tracheiphilum. Six of seven isolates of M. javanica caused similar increases in vascular discoloration in cultivar CB3. but one isolate of M. javanica and seven of M. incognita did not. Vascular discoloration rating was positively correlated with galling severity. However, increasing the initial inoculum density, and thus galling index, of one isolate of M. incognita did not increase vascular discoloration. The vascular discoloration ratings for the wilt-susceptible CB5 controls in each experiment were higher than those for the wilt-resistant cultivars infected with M. javanica. It is hypothesized that M. javanica but not M. incognita reduces, but does not eliminate, resistance to all races of F. o. tracheiphilum in cultivars CB3 and Grant.     

薇甘菊柄锈菌生物学及其寄主专一性

在检疫温室条件下对外来入侵植物薇甘菊的寄生菌——薇甘菊柄锈菌的生物学及其寄主专化性进行了研究。结果表明，薇甘菊柄锈菌可以侵染植物叶片和叶柄等营养器官。受侵染部位起始出现褪绿斑，12～15d，自叶背产生黄色冬孢子堆，并逐步坏死和脱落，最终致寄主死亡。冬孢子黄至暗褐色，包埋在寄主组织中，无明显休眠期。高湿条件下，冬孢子易萌发产生担孢子进行再侵染。采用悬挂法对29个科、62个属的72种植物进行了寄主专化性测定，结果表明，薇甘菊柄锈菌在菊科植物天门冬、紫茎泽兰、向日葵、地胆草上形成褪绿斑，但未发现菌丝和吸器。薇甘菊柄锈菌可成功侵染薇甘菊和同属植物假泽兰。     

Genetic Constitution and Pathogenicity of Lolium Isolates of Magnaporthe oryzae in Comparison with Host Species-Specific Pathotypes of the Blast Fungus

ABSTRACT Fungal isolates from gray leaf spot on perennial ryegrass (prg isolates) were characterized by DNA analyses, mating tests, and pathogenicity assays. All of the prg isolates were interfertile with Triticum isolates and clustered into the crop isolate group (CC group) on a dendrogram constructed from rDNA-internal transcribed spacer 2 sequences. Since the CC group corresponded to a newly proposed species, Magnaporthe oryzae, all of the prg isolates were designated M. oryzae. However, DNA fingerprinting with MGR586, MGR583, and Pot2 showed that the prg isolates are divided into two distinct populations, i.e., TALF isolates and WK isolates. The TALF isolates were virulent only on Lolium species, whereas the WK isolates were less specific, suggesting that gray leaf spot can be caused not only by Lolium-specific isolates but also by less specific isolates. We designated the TALF isolates as Lolium pathotype. The TALF isolates showed diverse karyotypes in spite of being uniform in DNA fingerprints, suggesting that theyare unstable in genome organization.     

Systemic Resistance Induced by Trichoderma spp.: Interactions Between the Host, the Pathogen, the Biocontrol Agent, and Soil Organic Matter Quality

ABSTRACT Several factors affect the ability of Trichoderma spp. to provide systemic disease control. This paper focuses on the role of the substrate in which plants are grown, resistance of the host to disease, and the ability of introduced Trichoderma inoculum to spread under commercial conditions. Several reports reveal that foliar disease control provided by Trichoderma spp. is more effective on plants grown in compost-amended media compared with in lower-in-microbial-carrying-capacity sphagnum peat media. In Rhododendron spp., host resistance affects control of Phytophthora dieback provided by Trichoderma spp. For example, T. hamatum 382 (T382) significantly (P = 0.05) suppressed the disease on susceptible cv. Roseum Elegans while plant vigor was increased. The disease was not suppressed, however, on highly susceptible cvs. Aglo and PJM Elite even though the vigor of these plants was increased. Using a strain-specific polymerase chain reaction assay under commercial conditions, it was demonstrated that introduced inoculum of T382 did not spread frequently from inoculated to control compost-amended media. Other Trichoderma isolates typically are abundant in control media within days after potting unless inoculated with a specific Trichoderma isolate. Thus, the low population of isolates that can induce systemic resistance in composting and potting mix environments may explain why most compost-amended substrates do not naturally suppress foliar diseases.