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In two field experiments, performed in 2020 and 2021, potato Nicola plants were sprayed once with three (Exp. 1) or two (Exp. 2) doses of Zorvec Vinabel (oxathiapiprolin+ zoxamide = ZZ), Zorvec Encantia (oxathiapiprolin+ famoxadone = ZF), Zorvec Endavia (oxathiapiprolin+ benthiavalicarb = ZE), Infinito (= INF) or Mefenoxam (= MFX) and thereafter inoculated with genotype 23A1 or 36A2 of Phytophthora infestans. Disease development was recorded at periodic intervals for a month. In both experiments, Zorvec mixtures were significantly more effective in suppressing the disease than INF or MFX. They delayed the onset of the disease and its progress, regardless the genotype used for inoculation. Among the three Zorvec mixtures, ZZ was least effective and ZE most effective. Sensitivity monitoring assays revealed zero mutants of P. infestans resistant to oxathiapiprolin. The data confirmed good efficacy of Zorvec mixtures, especially ZE, in field-grown potato crops as evident by the very effective control of late blight for one month.

Six wild accessions of Cucumis sativum were evaluated for resistance against each of the 23 isolates of the downy mildew oomycete Pseudoperonospora cubensis. The isolates originated from Israel, Europe, USA, and Asia. C. sativum PI 197088 (India) and PI 330628 (Pakistan) exhibited the highest level of resistance against multiple isolates of P. cubensis. Resistance was manifested as reduced lesion number, lesion size, sporangiophores and sporangia per lesion and enhanced encasement of haustoria with callose and intensive accumulation of lignin in lesions of both Plant Introductions (PIs) compared to the susceptible C. sativum SMR-18. In the field, much smaller AUDPC (Area Under Disease Progress Curve) values were recorded in PI 197088 or PI 330628 as compared to SMR-18. Each PI was crossed with SMR-18 and offspring progeny plants were exposed to inoculation with each of several isolates of P. cubensis in growth chambers and the field during six growing seasons. F1 plants showed partial resistance. F2 plants showed multiple phenotypes ranging from highly susceptible (S) to highly resistant (R, no symptoms) including moderately resistant (MR) phenotypes. The segregation ratio between phenotypes in growth chambers ranged from 3:1 to 1:15, depending on the isolate used for inoculation, suggesting that the number of genes, dominant, partially dominant, or recessive are responsible for resistance. In the field, the segregation ratio of 1:15, 1:14:1, or 1:9:6 was observed. F2 progeny plants of the cross between the two resistant PI&rsquo, s were resistant, except for a few plants that were partially susceptible, suggesting that some of the resistance genes in PI 197088 and PI 330328 are not allelic.

Late blight caused by the oomycete pathogen Phytophthora infestans is a devastating disease of potato and tomato worldwide, including Israel. The population structure of this pathogen was monitored in potato and tomato fields in Israel during a 36-year period of 1983-2019. Isolates of the pathogen were tested for sensitivity to phenylamide fungicides, mating type, race structure, and genotype. The phenotypic and genotypic structure of the population from potato have changed greatly from one year to another, from one season to the next, within a season and within a single field. Major changes also occurred in the population collected from tomato crops. The mechanisms driving these multiple changes and the heterogeneous nature of the population in Israel are shown to derive from multiple migration events of the pathogen via seed tubers from Europe and from fitness-driven selection processes.

The efficacy of four oxathiapiprolin (OXPT)-based, novel fungicidal mixtures against downy mildew in cucumber caused by Pseudoperonospora cubensis was examined in growth chambers. OXPT+chlorothalonil (CHT) mixed at weight ratio of 1 + 66.7; OXPT+ azoxystrobin (AZ) 1 + 10.3; OXPT+mandipropamid (MPD) 1 + 8.3; and OXPT+mefenoxam (MFX) 1 + 3, were compared with each other and with individual components. Mixtures performed better than all fungicides alone except for oxathiapiprolin. Of the four mixtures, OXPT+MFX outperformed the other treatments with the highest preventive, curative, translaminar, root treatment and seed treatment efficacies. Deployment in the field of such mixtures with reduced doses of oxathiapiprolin may lower the selection pressure imposed on P. cubensis and delay the buildup of subpopulations resistant to oxathiapiprolin.

Basil downy mildew (BDM) caused by the oomycete Peronospora belbahrii is a destructive disease of sweet basil (Ocimum basilicum) worldwide. It originated in Uganda in the 1930s and recently spread to Europe, the Middle East, Americas, and the Far East. Seed transmission may be responsible for its quick global spread. The pathogen attacks leaf blades, producing chlorotic lesions with ample dark asexual spores on the lower leaf surface. Oospores may form in the mesophyll of infected leaves. The asexual spores germinate on a wet leaf surface within 2 h and penetrate into the epidermis within 4 h. Spore germination and infection occur at a wide range of temperatures from 5 to 28.5°C. Infection intensity depends on the length of dew period, leaf temperature, and inoculum dose. The duration of latent period (from infection to sporulation) extends from 5 to 10 days, depending on temperature and light regime. The shortest is 5 days at 25°C under continuous light. Sporulation requires high humidity but not free leaf wetness. Sporulation occurs at 10 to 26°C. At the optimum temperature of 18°C, the process of sporulation requires 7.5 h at relative humidity ≥ 85%, with 3 h for sporophores emergence from stomata and 4.5 h for spore formation. Sporophores can emerge under light or darkness, but spore formation occurs in the dark only. Limited data are available on spore dispersal. Spores dispersed from sporulating plants contaminate healthy plants within 2 h of exposure. Settled spores may survive on leaf surface of healthy plants for prolonged periods, depending on temperature. Seed transmission of the disease occurs in Europe, but not in Israel or the United States. P. belbahrii in Israel also attacks species belonging to Rosemarinus, Nepeta, Agastache, Micromeria, and Salvia but not Plectranthus (coleus). A Peronospora species that infects coleus does not infect sweet basil. Control of BDM includes chemical, physical, and genetic means. The fungicide mefenoxam was highly effective in controlling the disease but resistant populations were quickly selected for in Israel and Europe rendering it ineffective. A new compound oxathiapiprolin (OSBP inhibitor) is highly effective. Nocturnal illumination of basil crops controls the disease by preventing sporulation. Daytime solar heating suppressed the disease effectively by reducing spore and mycelium viability. The most effective physical means is fanning. Nocturnal fanning prevents or limits dew deposition on leaf surfaces, and as a result, infection and sporulation diminish and epidemics are prevented. Genetic resistance occurs in wild basil and its transfer to sweet basil is under way.

Plenaris (oxathiapiprolin) applied to sunflower seedlings was highly effective in controlling downy mildew incited by the oomycete Plasmopara halstedii. In vitro assays revealed strong suppression of zoospore release and cystospore germination of P.halstedii by Plenaris. Bion (acibenzolar-S-methyl) and Apron (mefenoxam) were partially effective when used singly, but performed synergistically when mixed with Plenaris. Seed treatment (coating) with Plenaris provided dose-dependent control of the disease whereas Bion and Apron provided partial or poor control. However, seeds treated with mixtures containing reduced rates of Plenaris and full rates of Bion and/or Apron provided complete control of the disease due to the synergistic interaction between these components. Such mixtures should be used for seed treatment in the field to minimize selection pressure imposed on the pathogen.

Rubin, A. E., Gotlieb, D., Gisi, U., and Cohen, Y. 2008. Mutagenesis of Phytophthora infestans for resistance against carboxylic acid amide and phenylamide fungicides. Plant Dis. 92:675-683. The carboxylic acid amide (CAA) fungicides mandipropamid, dimethomorph, iprovalicarb, and the phenylamide fungicide mefenoxam (MFX, the active enantiomer of metalaxyl) are antioomycete fungicides effective against downy mildews and late blight. Resistance against MFX was reported in nature in several oomycetes including Phytophthora infestans and Plasmopara viticola, whereas resistance against CAAs was reported in P. viticola but not in P. infestans. In this study the mutability of P. infestans for resistance against CAAs and MFX (as a control) was explored under laboratory conditions. UV light or chemical mutagens (e.g., ethyl methan sulfonate [EMS]) were applied to sporangia, and the emergence of mutants resistant to CAAs or MFX, or with altered mating type, was followed. Many mutants resistant to CAAs developed at generation 0 after mutagenesis, but all showed erratic, instable resistance in planta, diminishing after 1 to 8 asexual infection cycles, and failed to grow on CAA-amended medium. In contrast, 19 mutants resistant to MFX were obtained: 6 with UV irradiation (in isolates 28 or 96) and 13 with EMS (in isolates 408, 409, and 410). In three experiments, a shift in mating type, from A1 to A2, was detected. To elucidate whether or not resistance to CAAs is recessive and therefore might emerge only after sexual recombination, A1 and A2 mutants were crossed and the F1 and F2 progeny isolates were tested for resistance. Offspring isolates segregated for resistance to MFX, with resistant isolates maintaining stable resistance in vitro and in planta, whereas all progeny isolates failed to show stable resistance to CAAs in planta or in vitro. The data suggest that P. infestans could be artificially mutated for resistance against MFX, but not against CAAs.

Four A1 field isolates and one A2 field isolate of Phytophthora infestans were crossed to produce oospores in tomato leaves. The oospores were extracted and mixed with perlite and water, and healthy tomato leaves were used as bait for oospore-progeny infection. Twenty-nine lesions were obtained from the four crosses and 283 single-sporangium isolates were recovered and tested on four tomato differential lines carrying different major genes (Ph-0, Ph-1, Ph-2, and 3707) for late blight resistance. The pathogenic fitness (number of sporangia per unit leaf area) of parental and progeny isolates was strongly dependent on the host genotype; it decreased in the order Ph-0 > Ph-1 > Ph-2 > 3707. The A2 parent had a higher pathogenic fitness than the A1 parents on Ph-0 and Ph-1 but similar, lower fitness on Ph-2. Different levels of pathogenic fitness were observed across all isolates on Ph-0, although Ph-0 lacks resistance genes. Pathogenic fitness on one tomato genotype was not related to the pathogenic fitness on the other tomato genotypes. Some isolates exhibited reduced pathogenic fitness relative to the respective A1 parent, whereas others demonstrated a higher pathogenic fitness compared with the A2 parent. The tomato genotype Solanum pimpinellifolium L3707/5 was resistant to all five parental isolates of P. infestans. However, 37 of the 283 progeny isolates from 11 different lesions had compatible reactions with this line, producing up to 31 × 103 sporangia/cm2. Overall, reduced fitness was more frequent among the progeny isolates than increased fitness.

Pseudoperonospora cubensis is a foliar pathogen of cucurbits. In cucumber, it produces chlorotic, angular lesions with dark sporangia on their under-surface. The distribution of pathotypes and mating types of P. cubensis were investigated in seven provinces in China. Twenty-nine isolates were obtained from Guangdong, Anhui, Hubei, Sichuan, Shandong, Beijing, and Harbin, one to six isolates from each province. They were collected from cucumber during summer 2011, except those from Harbin, which were collected from cucumber in summer 2012. Isolates were tested for pathogenicity and mating type. Pathogenicity was tested by inoculation of detached leaves of: cucumber (Cucumis sativum cvs. Bet-Alpha, SMR18), melon (Cucumis melo reticulatus cv. Ananas-Yokneam), pumpkin (Cucurbita maxima cv. Tripoli), squash (Cucurbita pepo, cv. Beruti), butternut gourd (Cucurbita moschata cv. Waltham), watermelon (Citrullus lanatus, cv. Malali), and sponge gourd (Luffa cylindrica, cultivar unknown). Mating type was determined by oospore production in melon leaf discs inoculated with mixed sporangia of a test isolate and A1 or A2 tester isolates, as recently described (2). The results showed that 22 isolates belonged to pathotype 3, sporulating on cucumber and melon; five isolates belonged to pathotype 6, sporulating on cucumber, melon, pumpkin, squash, and butternut gourd; one isolate belonged to pathotype 5, sporulating on the above five species and watermelon (1), and one isolate was capable of sporulating on the mentioned five species and sponge gourd, herein called pathotype 7. Of the 29 isolates tested, 22 belonged to the A1 mating type, six to the A2 mating type, and one did not produce oospores with either testers. Of the six A2 isolates, two originated from Sichuan, one from Beijing, and three from Harbin. All 22 isolates belonging to pathotype 3 were A1, whereas the other six isolates (four pathotype 6, one pathotype 5, and one pathotype 7) were A2. Crosses made between Chinese isolates belonging to opposite mating types resulted in the formation of abundant oospores in detached melon or cucumber leaves. The results prove that the A2 mating type of P. cubensis occurs in China. This explains the abundant occurrence of oospores of P. cubensis in nature in China (4). The data corroborate with Runge et al. (3), who suggested that the recent changes in the population structure of P. cubensis around the world resulted from the migration of a new genotype of P. cubensis from the Far East to Europe and the U.S.A. In Israel, a new pathotype attacking both Cucumis and Cucurbita appeared in 2002 (1), and A2 mating type first appeared in 2010 (2). References: (1) Y. Cohen et al. Phytoparasitica 31:458, 2003. (2) Y. Cohen and A. E. Rubin. Eur. J. Plant Pathol. 132:577, 2012. (3) F. Runge et al. Eur. J. Plant Pathol. 129:135, 2011. (4) Y. J. Zhang et al. J. Phytopathol. 160:469, 2012.

Downy mildew in basil was first reported from Uganda in 1933 (4). In 2004, it was reported from Italy (3) and, thereafter, from other countries around the world. In Israel, the disease was first observed in November 2011 in two greenhouses located in the northern part of the Jordan Valley. Within a month, second and third outbreaks of the disease occurred simultaneously near the southwest and southeast borders of Israel, 250 km from the initial disease outbreak. By the summer of 2012, the disease had appeared throughout the country, causing major economic damage. The causal agent, identified as Peronospora belbahrii (see below), produced chlorotic lesions on leaf blades with sporangia developing on the lower leaf surfaces. Lesions gradually turn necrotic, and infected leaves abscised. Sporangia were dark purple, oval, 30.4 ± 2.9 μm long × 21.4 ± 1.7 μm wide. Sporangiophores emerged from stomatal openings in a saturated atmosphere, were hyaline, 400 to 600 μm long, dichotomously branched, with three to five branches per sporangiophore, and bore a single sporangium on each branchlet tip. Oospores, seldom seen, were brown, round, and 46.2 ± 2.8 μm in diameter. Sporangia germinated directly, each producing a single germ tube that penetrated the periclinal wall of epidermal cells. PCR assays using sporangia and infected leaves as the template, and specific BAZ primers (1), produced a 134-bp band typical of P. belbahrii (1,2). Twenty isolates, collected from 12 locations in Israel from December 2011 to September 2012, were all sensitive to mefenoxam as the isolates did not cause symptoms on 15-leaf, potted basil plants (cv. Peri, Volcani Center, Israel) that were sprayed with 10 μg mefenoxam/ml (Ridomil Gold 48%, Syngenta, Basel, Switzerland) prior to inoculation. However, one isolate collected in early October 2012 from a severely infected plant in a greenhouse at Rehov in Bet-Shaan Valley, in which the plants had been treated with mefenoxam, was resistant to mefenoxan, showing abundant sporulation on leaves of potted basil plants that had been sprayed with 1,000 μg of mefenoxam/ml prior to inoculation. To our knowledge, this is the first report of the occurrence of downy mildew in basil in Israel. This is also the first global report of resistance to mefenoxam in P. belbahrii. References: (1) L. Belbahri et al. Mycol. Res. 109:1276, 2005. (2) R. Djalali et al. Mycol. Progress 11:961, 2012. (3) A. Garibaldi et al., Plant Dis. 89:683, 2004. (4) C. G. Hansford. Rev. Appl. Mycol. 12:421, 1933.

Root treatment with oxathiapiprolin, benthiavalicarb or their mixture Zorvec-Endavia [ZE (3+7, w/w)] was shown to provide prolonged systemic protection against foliar oomycete pathogens attacking cucumber, tomato and basil. Here we report that these fungicides can effectively protect potato plants against late blight when applied to the soil in which such potato plants are grown. In two field experiments, performed in 2019 and 2020, potato plants grown in 64 L containers were treated with a soil drench of oxathiapiprolin, benthiavalicarb or ZE at 12.5, 25 or 50 mg ai/five plants in a container. Artificial inoculations with Phytophthora infestans revealed that such treated plants were protected against late blight in a dose-dependent manner all along the season. Interestingly, oxathiapiprolin persisted in the treated soil for at least 139 days, providing systemic protection against late blight to the following potato crops grown in that treated soils. Potato plants grown in loess soil in the field were either sprayed or drenched with ZE. Plants treated via the soil were significantly better protected against late blight compared to the plants treated by a spray. The data demonstrate a new strategy for season-long protection of potato against late blight by a single soil application of ZE. The systemic nature of oxathiapiprolin and benthiavalicarb composing ZE assures the translocation to the foliage of two fungicides with different modes of action. This shall minimize the risk of developing resistance against either fungicide in the treated crops.

Oxathiapiprolin is a new fungicide with extremely high efficacy against oomycete plant pathogens. Solo components oxathiapiprolin (OXPT), chlorothalonil (CHT), azoxystrobin (AZ), mandipropamid (MPD), and mefenoxam (MFX) were compared with each other and with four oxathiapiprolin pre-packed fungicidal mixtures, OXPT+CHT 1+66.7, OXPT+AZ 1+10.3, OXPT+MPD 1+8.3, and OXPT+MFX 1+3 (weight active ingredient ratio), for control efficacy of late blight induced by MFX-insensitive Phytophthora infestans strains in tomato in growth chambers and the field. Mixtures performed better than all partner fungicides alone, except OXPT. Of the four mixtures, OXPT+MFX outperformed, with the highest preventive, curative, translaminar, and systemic efficacies. In the field, OXPT+MFX was superior to other fungicides in controlling late blight epidemics induced by MFX-insensitive isolates. Its deployment in the field will combat the dominating MFX-insensitive isolates, reduce the selection pressure imposed on P. infestans and delay the buildup of subpopulations resistant to oxathiapiprolin.

The oomycete Pseudoperonospora cubensis is a leaf pathogen causing severe damage to members of the Cucurbitaceae, especially cucumber and melon. It propagates clonally by sporangia. Oospores of P. cubensis were previously observed in nature but their formation in the laboratory was never reported nor their germination or infection. Here we report on the sexual reproduction of P. cubensis under controlled conditions in the laboratory. When field isolates were inoculated singly onto detached leaves of cucurbits in growth chambers no oospores were produced. However, when pairs of selected isolates were mixed and inoculated onto detached leaves, oospores were formed in the mesophyll within 6–11 days, suggesting that P. cubensis is heterothallic, having two opposite mating types, A1 and A2. Isolates belonging to pathotype 3 were all A1 whereas isolates belonging to the new pathotype 6 were either A1 or A2. Oospores were spherical, ~40 μm in diameter, hyaline to red-brown in color. Oospores were produced regularly, in large numbers, in Cucumis sativum and Cucumis melo, very seldom and in very small numbers in Cucurbita pepo, Cucurbita maxima and Citrullus lanatus, and not in Cucurbita moschata. Oospores were formed at 12.5–21°C but not at 25°C. Under moisture-saturated atmosphere oospores were also produced in leaves of intact plants. Oospores inoculated onto detached leaves in growth chambers produced F1 downy mildew lesions at 6–21 days after inoculation, many in Cucumis sativum, Cucumis melo and Cucurbita moschata, very few in Cucurbita pepo or Citrullus lanatus, and none in Cucurbita maxima. This report shows that P. cubensis is heterothallic, having A1 and A2 mating types which can cross and enable sexual reproduction in cucurbits. A preliminary report on part of the results has been published earlier.

DL-3-amino-butyric acid (BABA) induces local and systemic resistance against disease in numerous plant species. In a recent study we showed that preventive application of BABA to lettuce (Lactuca sativa) plants induced resistance against downy mildew caused by the oomycete Bremia lactucae by callose encasement of the primary infection structures of the pathogen. Now we show that post-infection application of BABA to the foliage or the roots, even at progressive stages of disease development, is highly protective against B. lactucae. Resistance induced by BABA is manifested in multiple microscopic forms, depending on the time of its application. When applied at 1 day post inoculation (dpi) BABA induced HR in penetrated epidermal cells; at 2 dpi it caused massive encasement with callose of the primary haustoria; and, at 3 or 4 dpi it enhanced the accumulation of H2O2 in the developing mycelia runners and altered their colour to red. The pronounced change in the colour of the mycelium was visually apparent to the naked eye. In all cases the pathogen failed to sporulate on the treated plants. This is the first indication that an immunizing compound may be protective at advanced stages of disease development.

Wild type (WT) field isolates of Bremia lactucae failed to germinate in vitro or infect lettuce leaves in the presence of CAA (carboxylic acid amide) fungicides. Minimal inhibitory concentrations (MIC) for mandipropamid, dimethomorph, benthiavalicarb and iprovalicarb were 0.005, 0.5, 0.5 and 5 μg ml−1, respectively. Mutagenesis experiments showed that spores exposed to EMS (ethyl methane sulphonate) or UV irradiation (254 nm) could infect lettuce leaves in the presence of up to 100 μg ml−1 CAA. The proportion of infected leaves relative to the number of spores inoculated (infection frequency) was inversely related to the concentration of CAA used, ranging between 0 and 160 per 1 × 106 spores. Resistant mutants (RM) lost their resistance within 1–14 reproduction cycles on CAA-treated plants. Crosses were made between RMxWT isolates and RMxRM isolates with an attempt to obtain stable homozygous resistant off-springs. Such crosses yielded few resistant but unstable progeny isolates. Mutagenic treatments given to hybrid isolates also failed to produce stable resistance. Previous gene sequencing data showed that stable resistance to CAAs is based on a single SNP in the cellulose synthase 3 (CesA3) gene of Plasmopara viticola. Therefore, we sequenced a 582 bp DNA fragment of Ces3A of WT, RM and hybrid isolates of B.lactucae. No mutation in this gene fragment was found. We conclude that mutagenic agents like EMS or UV may induce resistance to CAA in Bremia lactucae but this resistance is not stable and not linked to mutations in CesA3 gene.

BABA induced local and systemic resistance in lettuce (Lactuca sativa) against the Oomycete Bremia lactucae. Structure-activity analysis showed no induced resistance by related amino-butanoic acids or β-alanine. The R-enantiomer of BABA induced resistance whereas the S-enantiomer did not, suggesting binding to a specific receptor. Other compounds known to be involved in SAR signaling, including abscisic acid, methyl-jasmonate, ethylene, sodium-salicylate and Bion® (BTH) did not induce resistance. Systemic translocation of 14C-BABA and systemic protection against downy mildew were tightly correlated. BABA did not affect spore germination, appressorium formation, or penetration of B. lactucae into the host. Epifluorescence and confocal microscopy revealed that BABA induced rapid encasement with callose of the primary infection structures of the pathogen, thus preventing it from further developing intercellular hyphae and haustoria. Invaded host cells treated with BABA did not accumulate phenolics, callose or lignin, or express HR. In contrast, cells of genetically-resistant cultivars accumulated phenolics, callose and lignin and exhibited HR within one day after inoculation. The callose synthesis inhibitor DDG did not inhibit callose encasement nor compromised the resistance induced by BABA. PR-proteins accumulated too late to be responsible for the induced resistance. DAB staining indicated that BABA induced a rapid accumulation of H2O2 in the penetrated epidermal host cells. Whether H2O2 stops the pathogen directly or via another metabolic route is not known.

Four carboxylic acid amide (CAA) fungicides, mandipropamid (MPD), dimethomorph (DMM), benthiavalicarb (BENT) and iprovalicarb (IPRO) were examined for their effects on various developmental stages of Bremia lactucae, the causal agent of downy mildew in lettuce, in vitro and in planta. Spore germination in vitro or on leaf surfaces was inhibited by all CAA fungicides (technical or formulated). MPD was more effective in suppressing germination than DMM or BENT, whereas IPRO was least effective. CAA induced no disruption of F-actin microfilament organisation in germinating spores of B. lactucae. CAA applied to germinating spores in vitro prevented further extension of the germ tubes. When applied to germinated spores on the leaf surface they prevented penetration. Preventive application of CAA to intact plants inhibited infection. MPD was more effective in suppressing infection than DMM or BENT, whereas IPRO was least effective. Curative application was effective at ≤3 h post-inoculation (hpi) but not at ≥18 hpi. CAA (except IPRO) applied to upper leaf surfaces inhibited spore germination on the lower surface and hence reduced infection. CAA suppressed sporulation of B. lactucae on floating leaf discs and when sprayed onto infected plants two days before onset of sporulation. BENT and DMM were more effective in suppressing sporulation than MPD or IPRO. Epidemics of downy mildew in shade-house grown lettuce were suppressed by CAA. A single spray applied to five-leaf plants before transplanting controlled the disease for 50 days. The results suggest that CAA are effective inhibitors of spore germination and therefore should be used as preventive agents against downy mildew of lettuce caused by B. lactucae.

Pseudoperonospora cubensis, an obligate biotrophic oomycete causing devastating foliar disease in species of the Cucurbitaceae family, was never reported in seeds or transmitted by seeds. We now show that P. cubensis occurs in fruits and seeds of downy mildew-infected plants but not in fruits or seeds of healthy plants. About 6.7% of the fruits collected during 2012–2014 have developed downy mildew when homogenized and inoculated onto detached leaves and 0.9% of the seeds collected developed downy mildew when grown to the seedling stage. This is the first report showing that P. cubensis has become seed-transmitted in cucurbits. Species-specific PCR assays showed that P. cubensis occurs in ovaries, fruit seed cavity and seed embryos of cucurbits. We propose that international trade of fruits or seeds of cucurbits might be associated with the recent global change in the population structure of P. cubensis.

Peronospora belbahrii is a biotrophic oomycete attacking sweet basil. It propagates asexually by producing spores on dichotomously branched sporophores emerging from leaf stomata. Sporulation occurs when infected plants are incubated for at least 7.5h in the dark in moisture-saturated atmosphere at 10-27°C. Exposure to light suppresses spore formation but allows sporophores to emerge from stomata. Incandescent or CW fluorescent light of 3.5 or 6 µmoles.m(2).s(-1) respectively, caused 100% inhibition of spore formation on lower leaf surface even when only the upper leaf surface was exposed to light. The inhibitory effect of light failed to translocate from an illuminated part of a leaf to a shaded part of the same leaf. Inhibition of sporulation by light was temperature-dependent. Light was fully inhibitory at 15-27°C but not at 10°C, suggesting that enzyme(s) activity and/or photoreceptor protein re-arrangement induced by light occur at ≥15°C. DCMU or paraquat could not abolish light inhibition, indicating that photosystem I and photosystem II are not involved. Narrow band led illumination showed that red light (λmax 625 nm) was most inhibitory and blue light (λmax 440 nm) was least inhibitory, suggesting that inhibition in P. belbahrii, unlike other oomycetes, operates via a red light photoreceptor. Nocturnal illumination of basil in the field (4-10 µmoles.m(2).s(-1) from 7pm to 7am) suppressed sporulation of P. belbahrii and reduced epidemics of downy mildew, thus reducing the need for fungicide applications. This is the first report on red light inhibition of sporulation in oomycetes and on the practical application of light for disease control in the field.

The A2 mating type of Pseudoperonospora cubensis was first discovered in Israel in May 2010 on butternut gourd (Cucurbita moschata) (1). We monitored the occurrence of the A2 mating type of P. cubensis in isolates collected during May 2010 through September 2012 from downy mildew-infected cucurbit crops growing along the coastal plain of Israel. Mating type was determined by oospore production in melon leaf discs co-inoculated with sporangia of a test isolate mixed with sporangia of A1 or A2 tester isolates (2). The A1 and A2 tester isolates were maintained at 14°C (14 h light/day) by repeated inoculation of detached leaves of cucumber and pumpkin, respectively. The 29 isolates that were collected from cucumber (Cucumis sativum) were all A1. Of the 33 isolates collected from pumpkin (Cucurbita maxima), squash (C. pepo), or butternut gourd (C. moschata), 88% were A2 and 12% were A1. The host preference of mating type in P. cubensis was monitored at Bar-Ilan University farm during April to July 2012, among about 800 plants of eight cucurbit species (~100 plants per species) that were grown side-by-side in three adjacent net-houses (two 6 × 50 m and one 6 × 100 m) and exposed to natural infection. Downy mildew developed on cucumber, melon, pumpkin, squash, and butternut gourd, but not on watermelon, sponge gourd (Luffa cylindrica), or Momordica balsamina. Three-hundred and three isolates of P. cubensis were collected and tested for mating type: 123 from cucumber, 53 from melon, 30 from pumpkin, 48 from butternut gourd, and 41 from squash. The cucumber isolates expressed A1, A2, and A1A2 at a ratio of 94.3%, 3.3%, and 2.4%, respectively; the melon isolates 58.5%, 26.4%, and 15.1%; the pumpkin isolates 0%, 96.7%, and 3.3%; the butternut isolate 7.3%, 87.3%, and 5.5%; and the squash isolates 2.4%, 97.6%, and 0%, respectively. A1A2 isolates produce oospores when crossed with either A1 or A2 tester isolates. This is the first evidence suggesting a preference of A1 isolates to Cucumis spp. and of A2 isolates to Cucurbita spp. similar preference was recently observed among Chinese isolates of this pathogen (unpublished data). The mechanism(s) controlling this preference is not known. Classical genetics is currently employed to P. cubensis in order to understand if it derives from true linkage. The practical implication for downy mildew management is that growing cucumber/melon in close proximity to pumpkin/squash/butternut gourd should be avoided as it may enhance oospore production in nature. Oospores in soil were recently shown to serve as a primary source of downy mildew infection in cucumber (3). References: (1) Y. Cohen, A. E. Rubin, and M.Galperin. Plant Dis. 95:874, 2011; (2) Y. Cohen and A. E. Rubin. Eur. J. Plant Pathol. 132:577, 2012; (3) Y. J. Zhang et al. J. Phytopathol. 160:469, 2012.

The oomycete Pseudoperonospora cubensis attacks members of the Cucurbitaceae, causing severe foliage damage especially to cucumber and melon. Recently, new pathotypes of this oomycete appeared in Israel (2) and Italy (1) and highly aggressive isolates appeared in the United States (3). Since oospores of P. cubensis were rarely seen and sexual propagation by oospores was never reported (4), it is assumed that it propagates clonally by sporangia. Here we report on sexual reproduction of P. cubensis under controlled conditions in the laboratory. We found that field isolates belonging to the old pathotype 3 or to the new pathotype 6 (2) inoculated singly onto detached leaves of cucurbits in growth chambers at 15 or 20°C produced no oospores, even after prolonged incubation periods. However, when sporangia of some paired field isolates were mixed together at a 1:1 ratio, similarly inoculated onto detached leaves, and incubated at 15 or 20°C, numerous oospores (up to ~300/cm2) were formed in the mesophyll within 6 to 11 days, depending on the isolates pair, the host inoculated, and temperature. Oospores were also formed at 12.5°C but not at 25°C. Oospores developed in intact plants when kept at 15 or 20°C under a humidity-saturated atmosphere during disease development. Oospores were round, light brown to brown with an average diameter of ~40 μm. Oospores were produced in Cucumis sativum (cvs. Nadiojni and Dalila) and Cucumis melo (cvs. Ananas-Yokneam and Ein-Dor) but not in Cucurbita pepo (cv. Arlika, Beiruti), C. moschata (cv. Dalorit), or C. maxima (cv. Tripoli). To verify that oospores are infective, cucumber or melon leaves containing oospores were homogenized in water. The homogenate was twice brought to dryness at 25 to 30°C in petri dishes to differentially kill the vegetative structures of the pathogen (sporangia, cystospores, zoospores, and mycelia), resuspended in water, and inoculated onto detached leaves of various cucurbits in growth chambers at 15 or 20°C. Downy mildew lesions carrying sporangia appeared within 7 to 20 days in leaves of Cucumis sativum, Cucumis melo, and C. moschata but not in C. pepo or C. maxima. The recombinant origin of the F1 offspring isolates was confirmed by mefenoxam sensitivity tests, random amplified polymorphic DNA, and simple sequence repeat analyses. F1 progeny isolates of some crosses lost pathogenicity to C. moschata or C. maxima, toward which one of their parents was pathogenic, while others gained pathogenicity to Luffa cylindrica or Citrullus lanatus toward which neither parent was pathogenic. Data confirmed that isolates of P. cubensis can mate to produce oospores, especially under constant humidity conditions; such oospores are infective to cucurbits and F1 progeny isolates show altered sensitivity to fungicides or altered host range relative to their parents. To our knowledge, this is the first report of oospore formation by P. cubensis in the laboratory and on their pathogenicity to cucurbits. Reasons for the parallel appearance of new pathotypes of P. cubensis in Israel in 2002 (2) and Italy in 2003 (1) and the reemergence of highly aggressive isolates of the pathogen in the United States in 2004 (3) are not known. They may be related to oospore production and sexual recombination in P. cubensis. References: (1) C. Cappelli et al. Plant Dis. 87:449, 2003. (2) Y. Cohen et al. Phytoparasitica 31:458, 2003. (3) G. J. Holmes et al. Am. Veg. Grower. February, 14-15, 2006. (4) A. Lebeda and Y. Cohen. Eur. J. Plant Pathol.129:157, 2011.