Evaluating predictors of apple scab with receiver operating characteristic curve analysis

ABSTRACT Apple scab (Venturia inaequalis) is a perennial threat to apple production in temperate climates throughout the world. In the eastern United States, apple scab is managed almost exclusively through the regular application of fungicides. Management of the primary phase of disease is focused on preventing infection by ascospores. Management of secondary cycles of infection is largely dependent on how well primary infections were controlled. In this study, we used receiver operating characteristic curve analysis to evaluate how well mid-season assessments of the incidence of apple scab on cluster leaves, clusters (i.e., the whorl of cluster leaves), or immature fruit can serve as predictors of apple scab on harvested fruit (harvest scab) and whether these mid-season assessments of scab could be used reliably to manage scab under various damage thresholds. Results showed that assessment of scab on immature fruit was superior at predicting harvest scab than were assessments made on clusters or cluster leaves at all damage thresholds evaluated. A management action threshold of 7% scab incidence on immature fruit was identified by Youden's index as the optimal action threshold to prevent harvest scab incidence from exceeding 5%. Action thresholds could be higher or lower than 7% when economic assumptions were factored in to the decision process. The utility of such a predictor is discussed.     

梨叶叶龄与梨黑星病菌侵染发病的关系

田间试验结果表明，鸭梨叶片在生长发育期抗病性差，易感染黑星病；发育成熟后，抗病性逐渐增强。自然条件下，受叶片抗病性和孢子着落量的综合影响，鸭梨成熟叶片很少感染黑星病。鸭梨叶片从展叶到发育成熟，在梨树生长发育前期（盛花期后60d内）约需40d，后期（盛花期60d后）约需30d。正处在生长发育期的鸭梨叶片受侵染，潜育期为18－25d，发病后，病斑大，产孢期短；成熟叶片受侵染，潜育期多在40d以上，发病后，病斑小，产孢期长。梨树生长前期梨黑星病的流行主要由于大部分叶片正处在生长发育期抗病性较差；后期果实发病则主要源于成熟叶片内长期潜育和长期产孢的黑星病菌。     

Evidence for the Predisposition of Fungicide-Resistant Isolates of Venturia inaequalis to a Preferential Selection for Resistance to Other Fungicides

In the United States, populations of the apple scab pathogen Venturia inaequalis have progressed through three consecutive rounds of fungicide resistance development, first to dodine, then to the benzimidazoles, and most recently to the sterol demethylation inhibitors (DMIs). Analysis of extensive monitoring data have to date provided no indication of detectable cross-resistance or partial cross-resistance of V. inaequalis populations to the three unrelated classes of fungicides prior to the selection of resistant subpopulations. However, in this study, resistance to both benomyl and DMIs developed to significantly higher frequencies within the previously established dodine-resistant population than in the population sensitive to dodine. Accelerated selection of phenotypes double resistant to dodine and the DMI fenarimol was apparent over the course of distinct seasons of apple scab management with either dodine or fenarimol. The data provide evidence for an accelerated speed of resistance development among phenotypes of V. inaequalis already resistant to an unrelated fungicide. This finding represents a departure from the previous model, which assumed entirely independent rounds of resistance developments. The data indicate that phenotypes of V. inaequalis might not only be selected for the trait of fungicide resistance but also for traits allowing a more flexible response to changes in the environment where they compete.     

Quantitative Trait Loci (QTL) Analysis Reveals Both Broad-Spectrum and Isolate-Specific QTL for Scab Resistance in an Apple Progeny Challenged with Eight Isolates of Venturia inaequalis

ABSTRACT The major scab resistance gene Vf, extensively used in apple breeding programs, was recently overcome by the new races 6 and 7 of the fungal pathogen Venturia inaequalis. New, more durable, scab resistance genes are needed in apple breeding programs. F(1) progeny derived from the cross between partially resistant apple cv. Discovery and apple hybrid 'TN10-8' were inoculated in the greenhouse with eight isolates of V. inaequalis, including isolates able to overcome Vf. One major resistance gene, Vg, and seven quantitative trait loci (QTL) were identified for resistance to these isolates. Three QTL on linkage group (LG)12, LG13, and LG15 were clearly isolate-specific. Another QTL on LG5 was detected with two isolates. Three QTL on LG1, LG2, and LG17 were identified with most isolates tested, but not with every isolate. The QTL on LG2 displayed alleles conferring different specificities. This QTL co-localized with the major scab resistance genes Vr and Vh8, whereas the QTL on LG1 colocalized with Vf. These results contribute to a better understanding of the genetic basis of the V. inaequalis-Malus x domestica interaction.     

Effect of Fall Application of Fungal Antagonists on Spring Ascospore Production of the Apple Scab Pathogen, Venturia inaequalis

ABSTRACT The influences of Microsphaeropsis sp., M. arundinis, Ophiostoma sp., Diplodia sp., and Trichoderma sp., all antagonists of Venturia inaequalis, on ascospore production were evaluated under natural conditions and compared with urea and Athelia bombacina, a known antagonist. In the autumn, the fungi were applied to leaf disks artificially inoculated with V. inaequalis and to scabbed apple (Malus domestica) leaves incubated under controlled and natural conditions. In addition, large-scale trials were conducted with Microsphaeropsis sp. applied either as a foliar postharvest spray or as a ground application at 90% leaf fall. All fungal isolates, except Ophiostoma sp., were recovered from the leaf material that overwintered in the orchard. All treatments, except those with Ophiostoma sp., resulted in a significant reduction in V. inaequalis ascospore production on the leaf disks incubated under controlled conditions or in the orchard. In 1997, leaves with apple scab lesions treated with urea or Microsphaeropsis sp. produced significantly fewer ascospores of V. inaequalis than did nontreated leaves, with a reduction of 73.0 and 76.3%, respectively. In 1998, leaves treated with Microsphaeropsis sp., urea, Trichoderma sp., A. bombacina, and M. arundinis reduced ascospore production by 84.3, 96.6, 75.2, 96.6, and 52.2%, respectively. Based on all tests combined, the most efficient isolate was Microsphaeropsis sp. Postharvest applications of Microsphaeropsis sp. reduced the total amount of airborne ascospores trapped by 70.7 and 79.8% as compared with the nontreated plots in 1997 and 1998, respectively. Microsphaeropsis sp. provided a significant and consistent reduction in ascospore production in all tests.     

Evidence of Two Formae Speciales in Venturia inaequalis, Responsible for Apple and Pyracantha Scab

ABSTRACT Genetic relationships, mating crosses, and host specificity of Venturia inaequalis isolates from Malus spp. and of Spilocaea pyracanthae isolates from Pyracantha spp. were evaluated. Sequence analysis of the complete internal transcribed spacer (ITS) region (ITS1-5.8S to ITS2) revealed a total similarity between these two putative species. ITS restriction fragment length polymorphism carried out with five restriction enzymes on a collection of 28 isolates confirmed a lack of diversity in this region between and within these two populations. Additional isolates from three related species (V. pirina, V. nashicola, and S. eriobotryae) were divided into two distinct monophyletic groups in a phylogenetic tree using ITS sequence comparison. These groups were related to their anamorph (i.e., Spilocaea or Fusicladium). When inoculated on their host of origin, fields isolates caused typical symptoms of scab disease, and a host specificity was demonstrated by cross pathogenicity of isolates from Malus x domestica and Pyracantha spp. Mating on dried leaves in vitro between one isolate of each putative species led to production of numerous perithecia. Ninety-six sporulating monoascosporic progenies were isolated from this cross. Based on these genetic and pathogenic data, we proposed that pathogens responsible for scab on Malus spp. and Pyracantha spp. are considered as two formae speciales belonging to V. inaequalis.     

A new scab‐like disease on apple caused by the formerly saprotrophic fungus Venturia asperata

Atypical scab‐like symptoms were reported for the first time in 2007 in the south of France on fruits of apple cultivars carrying the Rvi6 (=Vf) major resistance gene to Venturia inaequalis. With microscopic observations, nucleotide sequence data and pathological tests, it was shown that the causal agent was Venturia asperata. Scanning electron microscopy was used to compare its infection process and conidiogenesis to those of Venturia inaequalis on apple and Venturia pirina on pear. Venturia asperata produced fewer hyphae and fewer spores than the two other Venturia species, and resulted in weaker symptoms. This fungal species was previously described as a saprotroph on apple leaf litter. This is the first report of damage on apple fruits caused by V. asperata. Changes in host and cultural practices may have created a new context favourable for the emergence of this pathogen. It was also detected on symptomless leaves and on overwintered leaves on the ground. Pseudothecia developed on overwintered leaves and released ascospores over a 2‐month period from the end of March until the end of May, suggesting that the fungus is able to survive from season to season. However, it is not yet known if this new disease will establish over coming years and become an emergent disease.     

Accumulation of Pathogenesis-Related Proteins in the Apoplast of a Susceptible Cultivar of Apple (Malus domestica cv. Elstar) after Infection by Venturia inaequalis and Constitutive Expression of PR Genes in the Resistant Cultivar Remo

Leaves of apple (Malus domestica cv. Elstar) were infected with a cloned isolate of the apple scab Venturia inaequalis. The intercellular washing fluid (IWF) of these plants was collected and the variation in the composition of proteins in the IWF was analysed by SDS-PAGE and two-dimensional gel electrophoresis during and after the infection with V. inaequalis, the causal agent of apple scab. The subsequent analysis of induced proteins by electron spray ionization quadrupole time of flight mass spectroscopy revealed the presence of -1,3-glucanase, chitinase, thaumatin-like protein and a cysteine-like protease in M. domestica leaves infected by V. inaequalis. These results were confirmed by immunoblotting with antibodies against some of these proteins. Moreover, a non-specific lipid transfer protein was identified in uninfected leaves: the amount declined to a non-detectable level within the first week after infection by V. inaequalis. The analysis of the IWF of M. domestica cv. Remo, bearing resistances to apple scab, powdery mildew and fire blight, showed a protein pattern comparable to that of the IWF from V. inaequalis infected leaves from cultivar Elstar indicating the constitutive production at least of some of the pathogenesis-related proteins in the resistant cultivar.     

Effectiveness of lime sulphur and other inorganic fungicides against pear scab as affected by rainfall and timing application

Journal of Plant Diseases and Protection - Pear scab, caused by Venturia pirina, is the most significant pear disease, causing economic losses in many pear production areas. In organic pear...     

抗黑星病芽变鸭梨组织结构、生理生化与抗病性关系研究

对一株抗黑星病芽变‘鸭梨’的叶片组织结构和生理生化特性进行了研究,发现其叶片蜡质含量高、栅栏组织双层出现时间早、厚度大、海绵组织结构致密、气孔数目少,推断这些特性决定黑星病菌分生孢子不易侵入叶片内部;叶绿素含量高为植株抗病性提供了生理基础,可溶性糖含量低有可能抑制黑星病营养菌丝在细胞间的生长和扩展。通过研究该抗黑星病芽变‘鸭梨’的抗病机理,为‘鸭梨’抗黑星病育种提供了理论依据。     

Effect of Timing of Application of the Biological Control Agent Microsphaeropsis ochracea on the Production and Ejection Pattern of Ascospores by Venturia inaequalis

ABSTRACT Field and in vitro trials were conducted to establish the influence of the biological control agent Microsphaeropsis ochracea on the ejection pattern of ascospores by Venturia inaequalis and on apple scab development, and to establish the best timing of application. The ejection pattern of ascospores was similar on leaves sprayed with M. ochracea and on untreated leaves. Fall application of M. ochracea combined with a delayed-fungicide program was evaluated in orchards with intermediate and high scab risk. For both orchards, it was possible to delay the first three and two infection periods in 1998 and 1999, respectively, without causing significant increase or unacceptable leaf and fruit scab incidence. To evaluate the best timing of application, sterile leaf disks were inoculated with V. inaequalis and then with M. ochracea 0, 2, 4, 6, 8, 10, 12, 14, and 16 weeks later. After incubation under optimal conditions for pseudothecia development, the number of ascospores was counted. Similarly, M. ochracea was sprayed on scabbed leaves on seven occasions from August to November 1999 and 2000. Leaves were overwintered on the orchard floor and ascospore production was evaluated the following spring. Ascospore production was reduced by 97 to 100% on leaf disks inoculated with M. ochracea less than 6 weeks after inoculation with V. inaequalis, but ascospore production increased with increasing period of time when M. ochracea was applied 8 to 16 weeks after the inoculation with V. inaequalis. In the orchard, the greatest reduction in production of ascospores (94 to 96% in 2000 and 99% in 2001) occurred on leaves sprayed with M. ochracea in August. The production of ascospores was reduced by 61 to 84% in 2000 and 93% in 2001 on leaves sprayed with M. ochracea in September, reduced by 64 to 86% in 2000 and 74 to 89% in 2001 on leaves sprayed in October, and reduced by 54 and 67% in 2000 and 2001, respectively, on leaves sprayed in November. It was concluded that M. ochracea should be applied in August or September and that ascospore maturation models and delayed-fungicide program could be used in orchards treated with this biological control agent.     

Modelling the effects of wetness duration and fruit maturity on infection of apple fruits of Cox's Orange Pippin and two clones of Gala by Venturia inaequalis

The effects were investigated of fruit maturity and duration of wetness on infection of apple fruits by Venturia inaequalis , and subsequent scab development. Incubation rate (inverse of median incubation period) increased linearly with increasing temperature (5–20°C) on detached 5-week-old fruits of cv. Royal Gala. Fruits were highly susceptible in the early stages of development, but became increasingly resistant as they matured. Inoculation of attached 12-week-old and detached near-mature fruits did not result in any lesions, while inoculation of attached 4-, 5-, 7- and 9-week-old fruits resulted in various levels of infection. Fruits of cv. Mondial Gala were more susceptible than those of cv. Cox's Orange Pippin. On cv. Mondial Gala, a wet period of 9 h resulted in ≈ 90% infection of 4-week-old fruits, but only 9% infection of 9-week-old fruits. Numbers of scab lesions on an apple generally followed a Neyman type A rather than a Poisson distribution, indicating a certain degree of aggregation of lesions on a fruit. A two-parameter generalization of the Poisson model described the observed incidence–density relationship well. A longer duration of wetness was required to result in a similar level of scab infection on old fruits to that on young fruits. On cv. Mondial Gala, wet periods of 9 and 32 h were required for ≈ 90% incidence of fruit scab on 4- and 7-week-old fruits, respectively. A mathematical model was developed to relate the incidence of fruit scab to duration of wetness and fruit maturity. The potential use of these results in practical disease management is discussed.     

Ultrastructural study on acibenzolar-S-methyl-induced scab resistance in epidermal pectin layers of Japanese pear leaves

The infection behavior of Japanese pear scab pathogen Venturia nashicola race 1 was studied ultrastructurally in acibenzolar-S-methyl (ASM)-pretreated susceptible Japanese pear (cv. Kousui) leaves to determine the mechanism of ASM-induced scab resistance. On ASM-pretreated leaf surfaces, the infection behavior (conidial germination and appressorial formation) was similar to that on distilled water (DW)-pretreated leaves prior to cuticle penetration by the pathogen. However, after penetration, differentiated behavior was found in epidermal pectin layers and middle lamellae of the ASM-pretreated leaves. Subcuticular hyphae in epidermal pectin layers and middle lamellae of ASM-pretreated pear leaves were observed at lower frequency than in DW-treated leaves. The results indicated that fungal growth was suppressed in ASM-pretreated pear leaves. In the pectin layers of ASM- and DW-pretreated leaves, some hyphae showed morphological modifications, which were used as criteria to judge collapse of hyphal cells, including plasmolysis, necrotic cytoplasm, and cell wall destruction. More hyphae had collapsed in ASM-pretreated leaves than in DW-treated ones. In addition, the cell walls of collapsed hyphae broke into numerous fibrous and amorphous pieces, suggesting that ASM-induced scab resistance might be associated with cell-wall-degrading enzymes from pear plants. In addition, results from morphometrical analysis suggested that the activity or production of pectin-degrading enzyme from hyphae were inhibited by ASM application when compared with DW treatment.     

Mapping Quantitative Field Resistance Against Apple Scab in a 'Fiesta' x 'Discovery' Progeny

ABSTRACT Breeding of resistant apple cultivars (Malus x domestica) as a disease management strategy relies on the knowledge and understanding of the underlying genetics. The availability of molecular markers and genetic linkage maps enables the detection and the analysis of major resistance genes as well as of quantitative trait loci (QTL) contributing to the resistance of a genotype. Such a genetic linkage map was constructed, based on a segregating population of the cross between apple cvs. Fiesta (syn. Red Pippin) and Discovery. The progeny was observed for 3 years at three different sites in Switzerland and field resistance against apple scab (Venturia inaequalis) was assessed. Only a weak correlation was detected between leaf scab and fruit scab. A QTL analysis was performed, based on the genetic linkage map consisting of 804 molecular markers and covering all 17 chromosomes of apple. With the maximum likelihood-based interval mapping method, eight genomic regions were identified, six conferring resistance against leaf scab and two conferring fruit scab resistance. Although cv. Discovery showed a much stronger resistance against scab in the field, most QTL identified were attributed to the more susceptible parent 'Fiesta'. This indicated a high degree of homozygosity at the scab resistance loci in 'Discovery', preventing their detection in the progeny due to the lack of segregation.     

GUIDELINE FOR THE EFFICACY EVALUATION OF FUNGICIDES

This guideline combines EPPO Guidelines nos 5 and 42 for the efficacy evaluation of fungicides against Venturia inaequalis and V. pirina -protectant applications and curative treatments, respectively.     

亚洲梨黑星病菌致病性及其寄主抗病机制的研究进展

梨黑星病是亚洲梨的主要病害之一。该病是由纳雪黑星病菌(Venturia nashicola)感染所致。V.nashicola主要寄生在亚洲梨叶片表皮细胞壁的果胶质层中。该菌的感染可能主要与分泌的细胞外分泌物质、角质分解酶、过氧化氢和果胶质分解酶有关。而亚洲梨对V.nashicola的抗性可能主要与多聚半乳糖醛酸酶抑制蛋白、多种病程相关蛋白、富亮氨酸重复类受体蛋白激酶等有关。另外,不具直接杀菌能力的系统抗性诱导剂acibenzolar-S-methyl(ASM)在大田试验中对梨黑星病菌有较好控制效果。这与ASM诱导的植物防御反应,包括多聚半乳糖醛酸酶抑制蛋白和几丁质酶等有关。     

Further studies of the inheritance of benomyl resistance in Venturia pirina isolated from pear orchards in Israel

Fourteen single-spore cultures of benomyl-resistant Venturia pirina were isolated from pear scab lesions at four sites in Israel. According to the ability of the isolates to germinate and grow at varying benomyl concentrations, four levels of resistance were determined in vitro : three isolates with low resistance (LR) grew at 0.5 but not at 5 μg/ml benomyl: five moderately resistant (MR) isolates grew at 5 but not at 50 μg/ml benomyl: five highly resistant (HR) isolates grew at 50 μg/ml but their hyphae were curled: and one isolate with very high resistance (VHR) grew unaffected at 50 μg/ml benomyl. The difference between the HR and the VHR phenotypes was clearly shown on medium amended with N -(3.5-dichlorophenyl) carbamate (MDPC): only the VHR isolate showed negative cross-resistance to 1 μ g/ml MDPC. whereas HR isolates grew unaffected. Crosses between resistant isolates and sensitive wild types, as well as between different resistant isolates, showed that the various levels of resistance are conferred by four allelic mutations that constitute a polymorphic series at a single locus.     

In vitro evaluation of fungal isolates for their ability to influence leaf rheology,production of pseudothecia,and ascospores of Venturia inaequalis

As part of an ongoing research project on biological control of apple scab, this study presents a novel approach for the in vitro selection of potential antagonists of the saprophytic phase of Venturia inaequalis. A collection of forty-two fungal isolates were tested for their in vitro ability to degrade apple leaf tissue, inhibit pseudothecia, and ascospore production. The inhibition of ascospore production cannot always be linked reliably with leaf degradation or the evaluation of pseudothecia production. Consequently, ascospore production was retained as the most useful screening parameter. Six isolates proved to significantly reduce the ascospore production of Venturia inaequalis. Two were as effective as Athelia bombacina, a previously reported antagonist of pseudothecia formation and inhibited over 98% of the ascospore production. These new organisms are now available for future field tests. Future selections from a large collection of fungal and bacterial saprophytes can now be based on a reliable and simple in vitro screening method.     

Localized Melanization of Appressoria Is Required for Pathogenicity of Venturia inaequalis

ABSTRACT During formation of appressoria produced from conidia and ascospores of Venturia inaequalis, a dark brown ring structure was detected at the base of appressoria. This melanized appressorial ring structure (MARS) was attached to the leaf surface like a sealing ring and formed the fungus-plant interface; it is believed to be required for pathogen penetration of the cuticle. Neither germ tubes nor infection structures beneath the cuticle were found to be visibly melanized. MARS were formed not only on apple leaves but also on leaves of nonhost plants and artificial surfaces differing in hydrophobicity; the formation of appressoria and MARS was confined to hard surfaces. The melanin nature of the ring was confirmed by using melanin biosynthesis inhibitors. Applications prior to inoculation largely inhibited the melanization and reduced infection rate by 45 to 80%; curative applications were not effective. Transmission electron microscopy verified a localized melanization of the cell wall around the penetration pore, and melanin was incorporated into all layers of the fungal cell wall. Appressoria without MARS were not able to infect the plant, suggesting that this structure can be considered to be a pathogenicity factor in V. inaequalis.     

Effects of cultivar mixtures on scab control in apple orchards

The effects of two mixtures of resistant and susceptible apple cultivars on the development of scab caused by Venturia inequalis were observed in an experimental orchard over four years, initially for two years without fungicides against scab, and subsequently for two years with a moderate fungicide schedule. The row-by-row and within-row mixtures included a susceptible cultivar and a resistant cultivar in equal proportions. Without fungicides, the results showed a significant reduction of disease incidence over both years (7·3 to 21·3%), and severity in the second year (35·4%) in the within-row mixtures, compared to the monoculture of the susceptible cultivar. The best results were obtained when the within-row mixture was associated with moderate fungicide treatments; in this case the reduction in disease incidence reached 75·1% on leaves and 69·7% on fruits during the growth phase. The characteristics of the Venturia inaequalis / Malus  ×  domestica pathosystem and the results obtained in this experiment suggest a moderate but not negligible ability of cultivar mixtures for reducing epidemics of the disease.