Podosphaera pannosa (syn. Sphaerotheca pannosa) on Rosa and Prunus spp.: Characterization of Pathotypes by Differential Plant Reactions and ITS Sequences

The powdery mildew fungus Podosphaera pannosa (Wallr.: Fr.) de Bary (syn. Sphaerotheca pannosa) is a major problem on roses worldwide. Twenty‐six monoconidial isolates of Podosphaera collected on roses and Prunus spp. in Belgium, Germany, France, Denmark, Israel and The Netherlands were characterized on the basis of differential reactions on in vitro rose genotypes and Prunus avium L. and by DNA sequence analysis of the rDNA ITS (internal transcribed spacer) region. Twenty‐four isolates were determined as P. pannosa. Amongst these, different groups could be distinguished. A first group of 18 isolates was highly virulent on rose and avirulent or very weakly virulent on P. avium. A second group of four isolates was highly virulent on both rose and P. avium. Analysis of the ITS sequence could discriminate these two groups of P. pannosa strains by a one base pair difference. Finally, two isolates of powdery mildew collected on Prunus sp. could be classified as P. pannosa based on their ITS sequence, which was identical to the ITS sequence of the isolates only highly virulent on roses. However, these two isolates were not able to infect roses. These results indicate that different strains of P. pannosa exist with varying host specificity. We demonstrated by ITS sequencing and plant reactions that the host range of P. pannosa comprises roses and Prunus spp.     

Powdery mildew resistance in roses: QTL mapping in different environments using selective genotyping

Podosphaera pannosa, the causal agent of rose powdery mildew, hampers the production of cut roses throughout the world. A major tool to control this disease is the use of resistant plant material. Single resistance genes, like Rpp1, may be overcome within a few years by high risk pathogens like powdery mildews. Durable resistance could be achieved using quantitative resistances. Here we describe mapping of QTLs for resistance to P. pannosa in six different environments (artificial and natural infections in the greenhouse over 3 years and natural infections in the field over 2 years). AFLPs, RGAs and other marker types were used to construct an integrated linkage map for the diploid population 97/7 containing 233 markers. In a selective genotyping procedure, marker segregation was analysed for 170 of the up to 270 phenotyped individuals. We identified seven linkage groups with an average length of 60 cM, corresponding to seven rose chromosomes in the haploid set. Using an LOD significance threshold of 3.9 we detected a total of 28 QTLs for the nine powdery mildew disease scores under analysis. Using the data from artificial inoculations with powdery mildew race 9, three resistance QTLs explaining about 84% of the variability were mapped. Twelve and 15 QTLs were detected for resistance to naturally occurring infections in the greenhouse and in the field, respectively, over several years.     

The inhibitory effects of rose powdery mildew infection on the oviposition behaviour and performance of beet armyworms

In this study, we investigated whether the oviposition behaviour and performance of the beet armyworm, Spodoptera exigua (Hübner) (Lepidoptera: Noctuidae), on the rose cultivar Rosa chinensis Jacq. (Rosaceae) were affected when the plants were infected by rose powdery mildew, Podosphaera pannosa (Wallr.: Fr.) de Bary (Erysiphales). The bioassays revealed that the moths significantly avoided ovipositing on mildew‐infected rose leaves when compared to healthy leaves. Pupal weights, emergence rates, and fecundity decreased when the caterpillars were fed mildewed rose leaves. Further laboratory bioassays aimed to elucidate the effects of two volatile headspace extracts (separately collected from healthy and mildewed rose plants) on the oviposition behaviour and performance of the moths. The moths clearly preferred to oviposit on healthy rose leaves that were not sprayed with additional volatiles rather than on healthy leaves sprayed with the volatile extracts from mildewed plants. The mean number of eggs laid on the former leaves was more than six times higher than that laid on the latter leaves. Olfactory bioassays demonstrated that ovipositing moths were significantly more attracted to volatiles emitted by healthy rose leaves than to those emitted by mildew‐infected leaves. Similar results were obtained when comparisons were made between the volatile extracts collected from healthy and mildewed rose plants. Thus, volatiles from mildew‐infected roses have a strong inhibitory effect against the moths. These results indicated that rose volatiles play a role in the oviposition behaviour of the moths, and that the volatiles induced by powdery mildew might be used for insect control.     

QTLs for powdery mildew resistance in peach ×Prunus davidiana crosses: consistency across generations and environments

Powdery mildew, caused by Sphaerotheca pannosa var. persicae is one of the most important diseases in European peach orchards. Quantitative trait loci controlling powdery mildew resistance were detected using three related F1, F2 and BC2 populations derived from the cross between the resistant parent P. davidiana clone P1908 and the susceptible peach cultivar Summergrand. Powdery mildew resistance of each population was evaluated under natural exposure, in several locations and over several years. Thirteen QTLs were detected. For nine of them, the favourable allele came from the resistant parent. Five QTLs were consistently detected across the three populations. The F1 hybrid used to produce F2 and BC2 populations had not inherited the favourable allele from P1908 for QTL detected on LG3 and LG8 in F1 population. QTLs were not detected in the corresponding regions in F2 and BC2 populations. In two other genomic areas, significant substitution effects between P1908 alleles were evidenced in the F1 population, but the favourable allele came from Summergrand in the F2 and BC2 populations. Analysis of phenotypic data suggested an important qualitative change in the distribution of powdery mildew resistance after 1996, confirmed by QTL analysis. Indeed, a dramatic decrease of the effect of the major QTL previously detected on LG6 was observed after 1996, while the QTL on LG8 was increasingly involved in the control of powdery mildew resistance. Consequences for peach breeding strategies to improve powdery mildew resistance are discussed.     

Powdery Mildew on Mulberry in Yunnan,China is Distinct from Other Phyllactinia spp. on Morus

The occurrence of an epidemic outbreak of a powdery mildew disease on mulberry in Yunnan province, China, is reported. Its symptoms are characteristic for powdery mildews and visible as white pathches covering the abaxial surfaces of leaves leading to chlorosis and necrosis. The pathogen is morphologically barely distinguishable from Phyllactinia moricola. However, it exhibits several new morphological characteristics which 2–3 conidia could be formed in short chains at the apex of the conidiophores and the conidia could produce two germ tubes in any position. Phylogenetic analyses of ITS sequences show that the pathogen has a close genetic relationship with P. moricola and Ph. broussonetiae‐kaempferi, two species on hosts belonging to family Moraceae. However, the ITS differences between Japanese sequences and the Chinese sequence derived from mulberry are greater than expected for a single species and suggest a cryptic species in China, but the present data are not sufficient for a final conclusion. Therefore, the Morus powdery mildew in Yunnan can currently only be classified as Phyllactinia sp. Morphological features, including conidial germination pattern of this powdery mildew are described in detail, and the local climatic conditions of the disease are analysed, which will provide the base for finding an effective method, including bio‐control, to control the disease under local conditions.     

Genetic dissection of a TIR‐NB‐LRR locus from the wild North American grapevine species Muscadinia rotundifolia identifies paralogous genes conferring resistance to major fungal and oomycete pathogens in cultivated grapevine

The most economically important diseases of grapevine cultivation worldwide are caused by the fungal pathogen powdery mildew (Erysiphe necator syn. Uncinula necator) and the oomycete pathogen downy mildew (Plasmopara viticola). Currently, grapegrowers rely heavily on the use of agrochemicals to minimize the potentially devastating impact of these pathogens on grape yield and quality. The wild North American grapevine species Muscadinia rotundifolia was recognized as early as 1889 to be resistant to both powdery and downy mildew. We have now mapped resistance to these two mildew pathogens in M. rotundifolia to a single locus on chromosome 12 that contains a family of seven TIR‐NB‐LRR genes. We further demonstrate that two highly homologous (86% amino acid identity) members of this gene family confer strong resistance to these unrelated pathogens following genetic transformation into susceptible Vitis vinifera winegrape cultivars. These two genes, designated r esistance to P lasmopara v iticola (MrRPV1) are the first resistance genes to be cloned from a grapevine species. Both MrRUN1 and MrRPV1 were found to confer resistance to multiple powdery and downy mildew isolates from France, North America and Australia; however, a single powdery mildew isolate collected from the south‐eastern region of North America, to which M. rotundifolia is native, was capable of breaking MrRUN1‐mediated resistance. Comparisons of gene organization and coding sequences between M. rotundifolia and the cultivated grapevine V. vinifera at the MrRUN1/MrRPV1 locus revealed a high level of synteny, suggesting that the TIR‐NB‐LRR genes at this locus share a common ancestor.     

Oleozon: A novel control strategy against powdery mildew in cucumber

Ozonized sunflower oil (oleozon) is an effective agent for controlling powdery mildew in cucumber. In this study, the mechanisms of oleozon in the control of powdery mildew were determined. The development of Podosphaera xanthii on cucumber leaves treated with oleozon (2%) and water was investigated at different times after inoculation. The germinating conidia, hyphae and conidiophores of the pathogen were severely damaged by oleozon. No visible phytotoxic effect was observed on cucumber after the application of oleozon. This compound had highly preventive effects as well as curative effects against powdery mildew based on in vivo potted seedling assays. The control effects of oleozon were further confirmed in a greenhouse trial. These results may provide a basis for further development of a natural fungicide against cucumber powdery mildew.     

Rye Pm8 and wheat Pm3 are orthologous genes and show evolutionary conservation of resistance function against powdery mildew

The improvement of wheat through breeding has relied strongly on the use of genetic material from related wild and domesticated grass species. The 1RS chromosome arm from rye was introgressed into wheat and crossed into many wheat lines, as it improves yield and fungal disease resistance. Pm8 is a powdery mildew resistance gene on 1RS which, after widespread agricultural cultivation, is now widely overcome by adapted mildew races. Here we show by homology‐based cloning and subsequent physical and genetic mapping that Pm8 is the rye orthologue of the Pm3 allelic series of mildew resistance genes in wheat. The cloned gene was functionally validated as Pm8 by transient, single‐cell expression analysis and stable transformation. Sequence analysis revealed a complex mosaic of ancient haplotypes among Pm3‐ and Pm8‐like genes from different members of the Triticeae. These results show that the two genes have evolved independently after the divergence of the species 7.5 million years ago and kept their function in mildew resistance. During this long time span the co‐evolving pathogens have not overcome these genes, which is in strong contrast to the breakdown of Pm8 resistance since its introduction into commercial wheat 70 years ago. Sequence comparison revealed that evolutionary pressure acted on the same subdomains and sequence features of the two orthologous genes. This suggests that they recognize directly or indirectly the same pathogen effectors that have been conserved in the powdery mildews of wheat and rye.     

Artificial elevation of glutathione contents in salicylic acid‐deficient tobacco (Nicotiana tabacum cv. Xanthi NahG) reduces susceptibility to the powdery mildew pathogen Euoidium longipes

• The effects of elevated glutathione levels on defence responses to powdery mildew (Euoidium longipes) were investigated in a salicylic acid‐deficient tobacco (Nicotiana tabacum cv. Xanthi NahG) and wild‐type cv. Xanthi plants, where salicylic acid (SA) contents are normal.
• Aqueous solutions of reduced glutathione (GSH) and its synthetic precursor R‐2‐oxothiazolidine‐4‐carboxylic acid (OTC) were injected into leaves of tobacco plants 3 h before powdery mildew inoculation.
• SA‐deficient NahG tobacco was hyper‐susceptible to E. longipes, as judged by significantly more severe powdery mildew symptoms and enhanced pathogen accumulation. Strikingly, elevation of GSH levels in SA‐deficient NahG tobacco restored susceptibility to E. longipes to the extent seen in wild‐type plants (i.e. enhanced basal resistance). However, expression of the SA‐mediated pathogenesis‐related gene (NtPR‐1a) did not increase significantly in GSH or OTC‐pretreated and powdery mildew‐inoculated NahG tobacco, suggesting that the induction of this PR gene may not be directly involved in the defence responses induced by GSH.
• Our results demonstrate that artificial elevation of glutathione content can significantly reduce susceptibility to powdery mildew in SA‐deficient tobacco.

     

Genetic linkage map of melon (<Emphasis Type="Italic">Cucumis melo</Emphasis> L.) and localization of a major QTL for powdery mildew resistance

Powdery mildew caused by Podosphaera xanthii has become a major problem in melon since it occurs all year round irrespective of the growing system. The TGR-1551 melon genotype was found to be resistant to several melon diseases, among them powdery mildew. However, the corresponding resistance genes have been never mapped. We constructed an integrated genetic linkage map using an F2 population derived from a cross between the multi-resistant genotype TGR-1551 and the susceptible Spanish cultivar ‘Bola de Oro’. The map spans 1,284.9 cM, with an average distance of 3.6 cM among markers, and consists of 354 loci (188 AFLP, 39 RAPD, 111 SSR, 14 SCAR/CAPS/dCAPS, and two phenotypic traits) distributed in 14 linkage groups. QTL analysis identified one major QTL (Pm-R) on LG V for resistance to races 1, 2, and 5 of powdery mildew. The PM4-CAPS marker is closely linked to the Pm-R QTL at a genetic distance of 1.9 cM, and the PM3-CAPS marker is located within the support interval of this QTL. These codominant markers, together with the map information reported here, could be used for melon breeding, and particularly for genotyping selection of resistance to powdery mildew in this vegetable crop species.     

Inoculum production and long-term conservation methods for cucurbits and tomato powdery mildews

The behaviour of cucurbit powdery mildews (Podosphaera xanthii and Golovinomyces cichoracearum) and tomato powdery mildew (Oidium neolycopersici) infesting detached cotyledons of Lagenaria leucantha cv. ‘Minibottle’ was studied in order to develop an easy culture method for pure inoculum production. High spore production was found with a combination of mannitol (0.1 m), sucrose (0.02 m) and agar (8 g l⁻¹) in the cotyledon survival medium. Sporulation on cotyledons and viability of conidia were affected by the age of culture for the three species of powdery mildew tested. The age of cotyledons had also an impact of the spore production. This method was used to produce large amounts of inoculum for P. xanthii, G. cichoracearum and O. neolycopersici and enable the development of other species of powdery mildew like Leveillula taurica. Freezing conidia in liquid nitrogen enabled the long-term conservation of P. xanthii without any loss of virulence. The same method was unsuccessful with G. cichoracearum, and L. taurica and partly successful with O. neolycopersici.     

Temporal isolation explains host-related genetic differentiation in a group of widespread mycoparasitic fungi

Understanding the mechanisms responsible for divergence and specialization of pathogens on different hosts is of fundamental importance, especially in the context of the emergence of new diseases via host shifts. Temporal isolation has been reported in a few plants and parasites, but is probably one of the least studied speciation processes. We studied whether temporal isolation could be responsible for the maintenance of genetic differentiation among sympatric populations of Ampelomyces, widespread intracellular mycoparasites of powdery mildew fungi, themselves plant pathogens. The timing of transmission of Ampelomyces depends on the life cycles of the powdery mildew species they parasitize. Internal transcribed spacer sequences and microsatellite markers showed that Ampelomyces populations found in apple powdery mildew (Podosphaera leucotricha) were genetically highly differentiated from other Ampelomyces populations sampled from several other powdery mildew species across Europe, infecting plant hosts other than apple. While P. leucotricha starts its life cycle early in spring, and the main apple powdery mildew epidemics occur before summer, the fungal hosts of the other Ampelomyces cause epidemics mainly in summer and autumn. When two powdery mildew species were experimentally exposed to Ampelomyces strains naturally occurring in P. leucotricha in spring, and to strains naturally present in other mycohost species in autumn, cross‐infections always occurred. Thus, the host‐related genetic differentiation in Ampelomyces cannot be explained by narrow physiological specialization, because Ampelomyces were able to infect powdery mildew species they were unlikely to have encountered in nature, but instead appears to result from temporal isolation.     

First report of powdery mildew caused by Podosphaera xanthii on Luffa acutangula in Odisha State,India

Abstract

Ridge gourd (Luffa acutangula) is an herbaceous perennial twining vine cultivated globally as vegetable and medicinal plant. During October to January in 2014 and 2015, 40% powdery mildew disease incidence was observed in different areas of Odisha state, India. The pathogenicity experiments confirmed the powdery mildew disease symptoms on artificially inoculated L. acutangula seedlings. Causal organism was identified as Podosphaera xanthii on the basis of morphological and molecular studies. This is the first report of powdery mildew disease on L. acutangula caused by P. xanthii.     

Fungicidal activity and chemical constitution

Six 4-(1-phenylalkyl)-, eight 4-(1-cyclohexylalkyl)-, seven 4-(1-cyclo-pentylalkyl)- and seven 4-(1-cyclobutylalkyl)-2,6-dinitrophenols, together with some derivatives, were tested against the powdery mildews of apple (caused by Podosphaera leucotricha) and blackcurrant (caused by Sphaerotheca mors-uvae) in the greenhouse and in field trials, against rose mildew (caused by S. pannosa) in the field, and against Venturia inaequalis in spore germination tests. Highest activity in the greenhouse was shown by members of the cyclobutyl series with 4–6 carbon atoms in the alkyl group. No phytotoxicity on healthy foliage occurred with any compound in these series.     

The effect of chemical and biological treatment on root and crown rot disease caused by Phytophthora cryptogea Pethybr. & Lafferty in Gerbera

Phytophthora root and crown rot (Phytophthora cryptogea) on gerbera is difficult to manage because most gerbera cultivars are susceptible to P. cryptogea. This study was conducted in order to determine the in vivo (pot experiment) efficacy of some fungicides and biofungicides. In pot experiments, fungicides were applied 7 days after inoculation with P. cryptogea, while biofungicide was applied 7 days before inoculation. In this study, soil drenches of five fungicides were tested. “Ametoctradin+dimethomorph (100 ml/day),” “mandipropamid+difenoconazole (60 ml/day),” “propamocarb+fosetyl‐Al (200 ml/day),” “mancozeb+metalaxyl‐M (250 g/day)” and “azoxystrobin+difenoconazole (100 ml/day)” active substances were used. Similarly, one biofungicide Bacillus amyloliquefaciens syn. MBI 600 (50 g/100 L) was applied by soil drenching. Efficacy of treatments was assessed according to the percentage of the root system which was visibly rotten at the end of the experiment. Root and crown rot severity was rated on a scale of 0 = 0% root system necrotic, 1 = 1%‐25% necrotic, 2 = 26%‐50% necrotic, 3 = 51%‐75% necrotic and 4 = 76%‐100% necrotic from 12 to 21 days. In this experiment, “azoxystrobin 200 g/L + difenoconazole 125 g/L” exhibited the highest efficacy against P. cryptogea with a ratio of 43.75%. The other fungicides and biofungicides ametoctradin 300 g/L + dimethomorph 225 g/L, mandipropamid 250 g/L + difenoconazole 250 g/L, propamocarb 530 g/L + fosety‐Al 310 g/L, mancozeb 64%+metalaxyl‐M 4% and Bacillus amyloliquefaciens syn. MBI 600 11% were ineffective. Importance should be given to management strategies of P. cryptogea of and more experiments should be carried out for a better understanding of the use of registered fungicides and biofungicides.     

Characterization of Reactions to Powdery Mildew (Podosphaera pannosa) in Resistant and Susceptible Rose Genotypes

Fungal development of powdery mildew Podosphaera pannosa (Wallr.: Fr.) de Bary on rose leaves depends on constitutive or induced resistance mechanisms present in attacked rose genotypes. The relationship between fungal development and plant resistance was investigated microscopically on young greenhouse leaves of four rose genotypes with different levels of resistance: Rosa wichuraiana, R. laevigata anemoides and R. hybrida cultivars ‘Excelsa’ and ‘Gomery’. Induced plant reactions, hydrogen peroxide production and cross sections through infected leaves were examined. The variation in development of the fungus on these rose genotypes depended on the relative presence of normal haustoria, abnormal haustoria, induced cell reactions, papilla formation or physical barriers. Formation of papillae could arrest up to one third of the successful penetrations. Papillae formation was often succeeded by total cell reaction. Abnormal haustoria were detected as rudimentary haustoria, haustoria with abnormal shape or haustoria without extra haustorial matrix. Post‐haustorial cell reactions, with and without cell collapse, were detected. In non‐collapsed cells, appositions were directed to both cell wall and haustorium. This was followed by accumulation of non‐identified, probably antifungal compounds. Both single and multicell reactions occurred. Hydrogen peroxide was detected during papilla formation and induced cell reactions.     

Effect of Yangtze River on population genetic structure of the relict plant Parrotia subaequalis in eastern China

Parrotia subaequalis (Hamamelidaceae) is a Tertiary relic species endemic in eastern China. We used inter‐simple sequence repeat (ISSR) markers to access genetic diversity and population genetic structure in natural five populations of P. subaequalis. The levels of genetic diversity were higher at species level (H = 0.2031) but lower at population level (H = 0.1096). The higher genetic diversity at species levels might be attributed to the accumulation of distinctive genotypes which adapted to the different habitats after Quaternary glaciations. Meanwhile, founder effects on the early stage, and subsequent bottleneck of population regeneration due to its biological characteristics, environmental features, and human activities, seemed to explain the low population levels of genetic diversity. The hierarchical AMOVA revealed high levels (42.60%) of among‐population genetic differentiation, which was in congruence with the high levels of Nei's genetic differentiation index (G_ST = 0.4629) and limited gene flow (N_m = 0.5801) among the studied populations. Mantel test showed a significant isolation‐by‐distance, indicating that geographic isolation has a significant effect on genetic structure in this species. Unweighted pair‐group method with arithmetic average clustering, PCoA, and Bayesian analyses uniformly recovered groups that matched the geographical distribution of this species. In particular, our results suggest that Yangtze River has served as a natural barrier to gene flow between populations occurred on both riversides. Concerning the management of P. subaequalis, the high genetic differentiation among populations indicates that preserving all five natural populations in situ and collecting enough individuals from these populations for ex situ conservation are necessary.     

Effects of elevated atmospheric CO2 and temperature on the management of powdery mildew of zucchini

The impact of combined environmental factors, such as temperature and CO₂, on the control of the powdery mildew of zucchini, caused by Podosphaera xanthii, and of different control measures has been studied on plants grown in phytotrons. Five experimental trials were conducted, and the powdery mildew severity of both treated and untreated zucchini plants was found to be significantly affected by the interaction between temperature (three different regimes: 16–18; 18–22; 22–26°C), CO₂ (two concentrations: 400–450 and 800–850 ppm) and the treatments. However, at the end of the trials, the efficacy of all the products was not affected by the different, tested environmental conditions. Sulphur consistently provided the highest disease control (75%–85% efficacy). Among the resistant inducers that were tested, calcium oxide was the most effective, in terms of powdery mildew control under all the conditions tested in phytotrons, reducing disease severity from 46% to 61%. Foliar applications of phosphite (14%–28% efficacy), Ampelomyces quisqualis (12%–23% efficacy) and potassium silicate (13%–24% efficacy) only slightly reduced the disease severity for all the tested temperature regimes and CO₂ concentrations, compared to the untreated control. The results obtained under our experimental conditions show that a possible increase in CO₂ concentration and temperature, which is expected for the next few years, should not influence the efficacy of the tested resistance inducers or of sulphur against powdery mildew on zucchini. Moreover, the suppressive effect of calcium oxide is in light of its possible use in greenhouses for zucchini powdery mildew control under 400–450 ppm of CO₂ and under enriched condition of 800–850 ppm of CO₂.     

The curative activity of non-ionic surface-active agents against some powdery mildew diseases

Eight non-ionic surfactants of the general formula RO(C₂H₄O)_nH were examined as fungicides for the control of various powdery mildew diseases. Those based on the condensation products of cetyl and oleyl alcohol with ethylene oxide were effective only as curative fungicides, having very little protective potential. This activity was greatest in products with n= 6–10. The most effective derivative, having eight ethylene oxide units, was particularly active in greenhouse tests against Sphaerotheca fuliginea on cucumber, S. pannosa on roses, and Podosphaera leucotricha on apples at concentrations which were not damaging to these crops. Although this compound caused damage to apples in the field it was effective against rose mildew and apple mildew, and its purely curative action gave a level of control equal to the standard dinitro compounds, which are capable of acting as either protectant or curative fungicides. This suggests that the dinitro-compounds also probably act mainly in a curative fashion against these diseases in the field.