Production Of Apothecia Research Articles

Loss of nitrogen fixing capacity in a montane lichen is linked to increased nitrogen deposition

AbstractThe circumboreal/circumpolar N2‐fixing lichenStereocaulon vesuvianumis among the most widespread and abundant fruticose species in montane Britain but has lost the capacity to fix N2over large areas of the country.To investigate whether loss of N2‐fixation inS. vesuvianumis linked to increased N deposition, we examined thallus morphology, physiology and chemistry at twelve locations representing an N deposition gradient of 3–40 kg ha−1 year−1. Measurements were made in parallel on a non‐N2‐fixing reference species (Parmelia saxatilis). The presence or absence of cephalodia (N2‐fixing nodules containing the cyanobacteriumStigonemasp) was recorded in over 500 herbarium specimens ofS. vesuvianumdating back to 1820.Cephalodium abundance inS. vesuvianum, and15N concentration inS. vesuvianumandP. saxatilis, were strongly negatively correlated with N deposition and particularly with dry deposited N; cephalodia do not form at total N deposition rates ≥8–9 kg ha−1 year−1. Other morphological oddities inS. vesuvianumat N‐polluted sites include increased apothecium (fungal reproductive structure) production and green algal biofilm development. Biofilm covered thalli without cephalodia lacked nitrogenase activity and cephalodia at sites where they rarely develop had nitrogenase activities typical for this species. The presence or absence of cephalodia in herbarium specimens ofS. vesuvianumsuggest that the present‐day N‐deposition linked gradient in N2‐fixing capacity did not exist in the 19th century and largely developed between 1900–1940.Synthesis.We provide clear evidence that N2‐fixing capacity inS. vesuvianumhas been lost in regions subjected to many decades of enhanced atmospheric N deposition. This loss is consistent with established models of diazotrophy, which identify supply of combined N as an inhibitor of N2‐fixation. Progressive depletion of thallus15N with increasing N deposition is in line with available data indicating that much atmospheric N pollution is15N‐depleted. Rates of nitrogenase activity inS. vesuvianumare low compared to other symbiotic systems and perhaps more likely supplanted by elevated N deposition. We suggest that other ecosystem compartments with low rates of fixation (e.g. soils) might also be susceptible to N pollution and merit investigation.

A Trojan horse approach for white mold biocontrol: Paraconiothyrium endophytes promotes grass growth and inhibits Sclerotinia sclerotiorum

Endophytic Paraconiothyrium strains associated with Brachiaria spp. and Panicum maximum were screened in vitro and in vivo for antagonism against Sclerotinia sclerotiorum and grass growth promotion. Based on multilocus phylogenetic analyses, three Paraconiothyrium estuarinum strains and two Paraconiothyrium cyclothyrioides strains were identified. In vitro, P. estuarinum strain CML 3699 was the most efficient in S. sclerotiorum mycelial growth inhibition. Volatile organic compounds (VOCs) produced by Paraconiothyrium strains reduced the sclerotia number, sclerotia weight and apothecia number in comparison to the control. The VOCs of P. cyclothyrioides strain CML 3698 promoted higher inhibitory effects on apothecium production. Paraconiothyrium estuarinum CML 3695, when inoculated into Brachiaria ruziziensis seeds, promoted increased biomass in comparison to those in non-inoculated seeds. All strains were sensitive to higher glyphosate doses, which reduced their mycelial development. However, when the mycelia of the endophytic fungi were removed and transferred to new PDA medium, the fungi recovered growth. After desiccation of Paraconiothyrium-inoculated B. ruziziensis with glyphosate, Paraconiothyrium strains recovered the ability to colonize the sclerotia of S. sclerotiorum and reduced carpogenic germination via a Trojan-horse-like mode of action. Considering that grass can be deployed by farmers as a physical barrier to S. sclerotiorum ascospore release, the growth promotion and parasitism of sclerotia exerted by Paraconiothyrium strains towards B. ruziziensis reinforce this disease control strategy.

First Report of Sclerotinia sclerotiorum Causing Strawberry Fruit Rot in Florida.

During the 2019-2020 Florida strawberry season (October to April), a strawberry (Fragaria × ananassa) fruit rot was observed in two fields (Plant City and Wimauma, FL) with up to 5% incidence on short-day cultivars SensationTM Florida127, and Florida Brilliance. Symptoms on pink and ripe fruit consisted of softening, discoloration, watery rot with white fuzzy mycelium, and initial sclerotium formation. Diseased tissue was placed on General Isolation (GI) medium (Amiri et al. 2018) and incubated at 25°C under a 12-h photoperiod. A fungus producing spreading cottony white colonies with dark sclerotia near the outer edges of the plates was consistently isolated. One isolate from each cultivar field (20-51 and 20-55) was selected for identification and pathogenicity tests. Apothecial production was induced following the protocol of Li and Rollins (2009), and apothecia (n=30) had an average diameter of 4.5 (3.5 to 7.2) mm. Sclerotia were 3.6 (2.5 to 6.2) mm by 4.5 (3.1 to 5.9) mm (n=30) in size. Dimensions of asci were 130.2 (115.1 to 160.5) μm by 8.5 (6.2 to 13.1) μm (n=30), and those of ascospores were 12.2 (10.8 to 14.6) μm by 6.8 (5.7 to 8.1) μm (n=30). Based on these morphological features, the pathogen was tentatively identified as Sclerotinia sclerotiorum (Lib.) de Bary (Maas 1998). DNA was extracted from the same two isolates using the FastDNA kit (MP Biomedicals, Solon, OH), and the ribosomal internal transcribed spacer (ITS) region was amplified using the primers ITS1 and ITS4 (White et al. 1990). Sequences were deposited in GenBank (accession nos. MT378215 and MT378216). BLASTn searches revealed that isolates 20-51 and 20-55 were 99.62% identical (526 / 528 bp) to S. sclerotiorum CBS 499.50 (MH856725.1). Immature pink fruit of SensationTM 'Florida127' were harvested, surface disinfested in bleach solution (0.08% NaClO) for 90 sec, rinsed twice with deionized water, then placed into styrofoam egg cartons inside clean plastic boxes (30x24x7 cm) containing 150 ml of sterile deionized water to maintain moisture, and kept at 25°C with a 12-h photoperiod. Ascospores and sclerotia were used for inoculation tests with three repetitions in an egg carton containing 12 fruit each per isolate and inoculation method. The experiment was repeated once. Fruit were inoculated by placing 20 μl of a 1 × 106 ascospores/mL suspension or a single sclerotium on the upper half of the fruit. Controls were included, by placing 20 μl of sterile DI water or fruit with no sclerotia. Evaluations were done 6, 10, and 15 days after inoculation (DAI). Control fruit remained healthy, while inoculated fruit developed symptoms of softening and discoloration. For ascospore inoculation, disease incidence was 55 (± 5) and 78% (± 4), for 6 and 15 DAI, respectively, whereas for sclerotia inoculation incidence was 100% 6 DAI. Morphologically identical fungi to the original isolates were re-isolated from the diseased fruit. No other fruit decay fungi were observed. S. sclerotiorum has been previously reported causing strawberry fruit rot in Washington state in the United States, England, Israel, and Scotland (Alcorn 1966; Maas 1998; McLean 1957). It has also been listed in the indices of plant diseases from Florida, North Carolina, and California as causing crown rot (Farr and Rossman 2020). To our knowledge, this is the first report of S. sclerotiorum causing strawberry fruit rot in Florida. The pathogen is an aggressive necrotroph with prolonged survival and affects several vegetable crops grown in Florida (Paret et al. 2018). Because only the strawberry beds are fumigated, sclerotia remaining in the alleys could serve as inoculum sources. Currently, the disease is rare and of minor significance to strawberry production. However, efforts should be implemented to monitor its occurrence and spread.

Spatiotemporal Distribution Pattern of Sclerotinia sclerotiorum Apothecia is Modulated by Canopy Closure and Soil Temperature in an Irrigated Soybean Field.

Identifying the optimal timing for fungicide application is crucial in order to maximize the control of Sclerotinia stem rot (SSR), which is caused by Sclerotinia sclerotiorum. In this study, the impact of canopy closure and soil temperature on apothecia production was investigated to optimize fungicide application timing. Replicated soybean plots with a row spacing of 0.36 and 0.38 or 0.76 m were established in 2015 and 2016 in an irrigated soybean field at Michigan State University's Montcalm Research Center. The number of apothecia and ascospores and the incidence of SSR were monitored two times per week for 10 to 12 weeks. In both row-spacing trials, apothecia were observed earlier in 2016 (before the R1 growth stage) than in 2015 (between R1 and R2). The maximum number of apothecia was 50 times higher with the 0.36-m row spacing than with the 0.76-m row spacing in 2015 but was 2.5 times higher with the 0.76-m row spacing than with the 0.38-m row spacing in 2016, though the overall numbers were much lower in 2016. The apothecia distribution pattern was synchronized with the canopy closure pattern and the soil temperature profile. The peak number of apothecia was observed when canopy closure reached at least 50% and when average soil temperature in the row was between 21.5 and 23.5°C. In 91% of the cases, the presence of apothecia was observed when the percentage of light blocked was 70%, and no apothecia germinated in the absence of light or under full light exposure. During the first 50 days after plant emergence, the rate of canopy closure was higher in 2016 than in 2015, and the first diseased plant was observed earlier in 2016 (R2) than in 2015 (R5). Canopy closure and the distance of the sampling point from the soybean row explained much of the variability in the number of apothecia. These results can partially explain the inconsistent efficacy of fungicide applications based on the soybean growth stage and will be helpful for informing disease models and fine-tuning fungicide application strategies.

First report of Sclerotinia subarctica in France detected with a rapid PCR-based test

White mould can affect the production of a wide range of economically important crops worldwide. The symptoms may be caused by several species, including Sclerotinia subarctica, a species mostly occurring in northern latitudes in sympatry with S. sclerotiorum. Although the two species are morphologically indistinguishable, S. subarctica was reported to have different climatic requirements for mycelial growth and production of apothecia. These differences may affect the precision of white mould risk prediction models that are based on the production of ascospores by S. sclerotiorum. To assess the presence of S. subarctica in France, we adapted a rapid PCR-based test to distinguish S. subarctica from other commonly found species of Sclerotinia. This test was used to characterize a collection of 969 Sclerotinia sp. isolates originating from various plants (bean, canola, carrot, lettuce, melon and witloof chicory), air and soil samples in different regions of France. One single isolate, collected from witloof chicory in northern France, was identified as S. subarctica. When genotyped with five microsatellite markers designed for S. sclerotiorum, this isolate had a haplotypic profile that was clearly distinct from the other isolates. The ITS sequence of this isolate was identical to those of isolates collected in northern Europe and Alaska. Koch’s postulates were verified. When inoculated on witloof chicory, the isolate identified as S. subarctica induced white mould symptoms. This study is the first to report the presence of S. subarctica south of the 51st parallel north and on witloof chicory.

Mycelial growth, pathogenicity, aggressiveness and apothecial development of Sclerotinia sclerotiorum isolates from Brazil and the United States in contrasting temperature regimes

ABSTRACT Fungi can adapt to environmental conditions and produce different physiological responses. The aim of this study was to verify the existence of Sclerotinia sclerotiorum temperature ecotypes in isolates from Brazil and the USA. Ten S. sclerotiorum isolates from tropical and subtropical regions of Brazil and six isolates from the USA were used to measure mycelial growth, pathogenicity and aggressiveness on bean, canola and soybean, as well as apothecial formation at contrasting temperatures. For mycelial growth, regardless of the origin, all isolates grew faster at 20°C, compared to 27°C. For pathogenicity and aggressiveness, disease severity was greater at 20°C than at 30°C considering all isolates. As regards apothecial production, only Brazilian isolates were capable of producing apothecia with no preconditioning. After preconditioning at 4°C during 40 days, isolates from Brazil and the USA produced apothecia. None of the 16 isolates was capable of producing apothecia at 30oC after 40 days. Results indicated no adaptation of S. sclerotiorum isolates from Brazil to grow or colonize leaflets at higher temperatures, compared to isolates from the USA. Only sclerotia from S. sclerotiorum isolates from Brazil were capable of germinating carpogenically without preconditioning.

The susceptibility of Asian, European and North American Fraxinus species to the ash dieback pathogen Hymenoscyphus fraxineus reflects their phylogenetic history

In Europe, common ash (Fraxinus excelsior) is being decimated because of the invasive fungus Hymenoscyphus fraxineus. In its native range in Asia this ascomycete is considered a harmless leaf associate of F. mandshurica and F. chinensis subsp. rhynchophylla. Field observations from Europe suggest that there is species-specific variation in disease susceptibility among European and North American Fraxinus species, but a wider comparison at the genus level has been missing so far. We assessed disease symptoms and pathogen apothecium development in 17 Fraxinus species from Asia, Europe and North America exposed to high infection pressure in a Danish arboretum. We also tested their susceptibility to pathogen infection through controlled stem and leaf inoculations and subsequently assessed the level of pathogen DNA by a qPCR assay. The results suggested the presence of a phylogenetic signal in disease susceptibility where closely related Asian, European and North American species in section Fraxinus had relatively high levels of H. fraxineus DNA in the leaves and supported high production of apothecia. Leaves from some North American species also contained relatively high levels of H. fraxineus DNA, supported moderate production of apothecia and developed lesions—stating the need to avoid introduction of H. fraxineus to North America.

Biocontrol of Sclerotinia sclerotiorum infection of cabbage by Coniothyrium minitans and Trichoderma spp.

Nine fungal isolates [Clonostachys rosea (1), Coniothyrium minitans (1), Trichoderma crassum (1), T. hamatum (4), T. rossicum (1) and T. virens (1)] were tested in two bioassays for their ability to degrade sclerotia and reduce apothecial production and carpogenic infection of cabbage seedlings. C. minitans LU112 reduced apothecial production in both bioassays, with T. virens LU556 significantly reducing apothecial production in the sclerotial parasitism assay. Both isolates also reduced sclerotial viability in this assay to 5% for C. minitans and 22% for T. virens. C. minitans LU112 and T. virens LU556 reduced the infection of cabbage seedlings in the pot bioassay 126 days after sowing but not after 147 days, partly due to ascospore cross-infection between treatments. C. minitans LU112, T. virens LU556 and T. hamatum LU593 as maizemeal-perlite (MP) soil incorporation and transplant potting mix incorporation were evaluated for their ability to control Sclerotinia sclerotiorum disease of cabbage in field experiments. S. sclerotiorum infection of cabbage was reduced by 46–52% and 31–57% by both C. minitans LU112 and T. hamatum LU593 as MP soil incorporations, respectively, in the two field experiments. T. virens LU556 MP soil incorporation and C. minitans LU112 and T. hamatum LU593 transplant potting mix incorporations reduced S. sclerotiorum disease in the first experiment but not in the second experiment. A commercial C. minitans LU112 formulation, C. Mins LU112 WG, also significantly reduced S. sclerotiorum disease by 59%. Soil incorporation of C. minitans and T. hamatum was shown to have potential to control S. sclerotiorum disease in cabbage.

菌核病菌の子のう盤形成能を消失する湛水処理の条件

菌核病菌の子のう盤形成能を消失する湛水処理の条件

Ophiostoma mitovirus 3a, ascorbic acid, glutathione, and photoperiod affect the development of stromata and apothecia by Sclerotinia homoeocarpa

Hypovirulence in Sclerotinia homoeocarpa is associated with infection by Ophiostoma mitovirus 3a (OMV3a). OMV3a is also present in asymptomatic isolates, with growth and virulence comparable to that of virus-free isolates. Hypovirulent isolates have impaired mitochondrial function resulting in increased activity of the alternative oxidase pathway, which is implicated in the reduction of reactive oxygen species in other fungi. In this study, hypovirulent, asymptomatic, and virus-free isolates were grown on potato dextrose agar amended with ascorbic acid or glutathione and were incubated under various photoperiods to determine the importance of reactive oxygen species, light, and OMV3a infection for differentiation of stromata and apothecia by S. homoeocarpa. Hypovirulent isolates did not form stromata or apothecia. Glutathione and darkness reduced stromata size and apothecia production by virulent and asymptomatic isolates. Apothecia formed under several different photoperiods, and ascorbic acid increased apothecia production. Ascospores were not detected in these apothecia. The results suggest that hypovirulence, light, and the superoxide radical are important factors in the formation of stromata and apothecia by S. homoeocarpa isolates. This is the first report of sterile apothecia production by North American isolates of S. homoeocarpa and provides a starting point for attempts to produce fertile apothecia.

In vitro and in vivo antifungal activity of synthetic pure isothiocyanates against Sclerotinia sclerotiorum

Isothiocyanates (ITCs) released by the enzymatic hydrolysis of glucosinolates in the Brassicaceae are potentially useful for controlling fungal pathogens. In vitro activity of pure ITCs against Sclerotinia sclerotiorum (Lib.) de Bary was studied by adding them to glass filters in petri dishes and dissolving them in the growing media. Methyl, allyl and benzyl ITCs were the most fungitoxic of the compounds in bioassays with S. sclerotiorum isolate Ss31. In the volatile phase, mycelial growth was completely inhibited by these three compounds. Aromatic ITCs were less toxic in the petri dishes but were more toxic than aliphatic ITCs when dissolved in the agar. Benzyl ITC exhibited the highest inhibitory effect on sclerotial germination, with an EC(50) value of 75.1 µmol L(-1) . Butyl and benzyl ITCs reduced apothecial production of S. sclerotiorum by 92.5% at the highest concentration. In in vivo assay, only allyl and 2-phenylethyl ITCs reduced disease incidence (by 76.7 and 70% respectively) at low concentrations. Sclerotinia sclerotiorum in the soil might be suppressed by the higher concentrations of allyl and benzyl ITCs released from decomposition of Brassica juncea, B. carinata, B. nigra and Sinapis spp.

Effect of Salinity, Temperature and Carbon Source on the Growth and Development of Sclerotia of Sclerotinia sclerotiorum Isolated from Semi-arid Environment

Studies were conducted to determine the effects of temperature, solute potential and carbon source on the mycelial growth, sclerotia development, and apothecium production of an isolate of Sclerotinia sclerotiorum. Mycelial growth rate was greatest at <TEX>$25^{\circ}C$</TEX> on potato dextrose agar (PDA) medium amended with up to 2% NaCl (<TEX>${\psi}s{\leq}1.91\;MPa$</TEX>) and thereafter, growth rate declined. The least number of sclerotia were produced at <TEX>$20^{\circ}C$</TEX>on both PDA and malt extract agar (MEA) amended with 8% NaCl (<TEX>${\psi}s=6.62\;MPa$</TEX>). With increasing temperature and decreasing solute potential the number and size of sclerotia were significantly reduced. The combined effect of temperature, solute potential and carbon source on sclerotia production were highly significant and had an impact on the development of the rind layer cells of sclerotia. These cells lacked a transparent cell wall which was surrounded by a compact melanized layer, and some of these cells appeared to be devoid of cell contents or were totally vacuolated. The survival of the sclerotia with increase in salinity and temperature appeared to affect melanization and the nature of the rind cells. The observations of this study re-enforces the need for an integrated disease management to control S. sclerotiorum.

Effects of Fluctuating Soil Temperature and Water Potential on Sclerotia Germination and Apothecial Production of Sclerotinia sclerotiorum

The effects of fluctuating soil temperature and water potential on sclerotial germination and apothecial production by Sclerotinia sclerotiorum were investigated in growth chamber experiments. In the temperature experiments, temperature fluctuations of 4, 8, 12, and 16°C around a median of 20°C, and a constant of 20°C, were tested. Daily temperature fluctuations of 8°C resulted in highest levels of sclerotial germination and apothecial production. The earliest appearance of apothecia occurred in the 8°C fluctuation treatment, 24 days after the start of the experiment. Sclerotia in the 12°C fluctuation treatment germinated last; its first sclerotium germinated 44 days after experiment initiation. For the soil water potential experiments, constant saturation (approximately -0.001 MPa) and three levels of soil water potential fluctuation from saturation-"low" (-0.03 to -0.04 MPa), "medium" (-0.06 to -0.07 MPa), and "high" (-0.09 to -0.1 MPa)-were tested. Constant saturation yielded the highest number of germinated sclerotia and apothecia. All soil water potential fluctuations were detrimental to sclerotial germination and apothecial production, with sclerotial germination under fluctuating moisture conditions less than a tenth of that occurring under constant saturation. The first sclerotium in the constant saturation treatment germinated in 35 days; however, 76 days were required in the high soil water potential fluctuation treatment.

Stem rot disease incidence on faba beans in an artificially infested field

Stem rot, a destructive fungal disease caused by Sclerotinia trifoliorum Eriksson, is a serious problem for many important crops including faba beans (Vicia faba L.). Field experiments were carried out for five consecutive years in northern Greece to study the period of infection of S. Trifoliorum and the disease incidence in faba beans grown in a field that was artificially infested by placing various numbers of fungal sclerotia into the soil during the sowing period. Disease severity was very low (< 4%) in all growing seasons, whereas a high percentage of infection (13.3 to 95.0%) was observed under controlled environment. Apothecial production in the field occurred from November to early January. Symptoms of fungal infection on faba bean plants were observed from early February to early April, with the most infection occurring during March. Faba bean plants were infected by mycelium, after myceliogenic germination of sclerotia, at the base of the stem close to the soil surface. Visible, water-soaked lesions appeared before flower initiation. Disease severity was not significantly correlated with monthly rainfall (r = 0.11) nor with mean monthly temperature (r = 0.15) over five growing seasons. The growth stage of plants and the absence of an energy source during ascospore release, as well as the mode of sclerotial germination (myceliogenic), may have been the critical factors responsible for the low disease severity in the field.

Studies on the germination of sclerotia of Sclerotinia sclerotiorum

Sclerotial germination of 33 isolates of Sclerotinia sclerotio-rum, which were collected from different regions and crops, was compared at 10†C in a climate chamber. Germination of S. sclerotiorum, measured as the number of sclerotia producing stipes and apothecia, was the predominant mode that was affected significantly by isolate. Isolates which originated from a warmer area generally germinated more quickly. The culture media affected the viability of sclerotia of isolates M01 and MZ. Sclerotia germinated on different substrates such as standard gardener`s soil, sand and vermiculite, but did not germinate well when the sclerotia had developed on nutrient-rich or nutrient-poor medium. The optimum medium for sclerotial development was wheat grain. Sclerotia produced on wheat grain were well-suited for studies on germination of sclerotia and formation of apothecia. The production of apothecia was optimal at a temperature of 10†C and an illumination of 500 lux, and after an incubation period of 2 to 3 months. One apothecium produced over 1 million ascospores and survived over 15 days in the climate chamber.

Size is not a reliable measure of sexual fecundity in two species of lichenized fungi

Since a number of studies have shown a strong positive correlation between apothecia production and lichen thallus size, it has been suggested that size can serve as an easy measure of lichen sexual fecundity. The reliability of the relationship between size and apothecia production among different environments has not been studied. We measured apothecia development and thallus area in populations of the lichenized fungi Xanthoparmelia cumberlandia and Xanthoparmelia coloradoensis. Our data suggest that while size is a significant predictor of apothecia production, the relationship between size and apothecia production is not consistent among various environments, nor does size adequately explain variation in apothecia production.

The relationship between thallus mass, surface area and apothecium production in Umbilicaria rigida

Abstract: The relationship between thallus mass, surface area and apothecium production in the alpine, saxicolous umbilicate lichen Umbilicaria rigida was investigated. There is a very strong and significant relationship between an increase in thallus mass and an increase in upper surface area. The relationship is strictly linear, adding 2 mm2 to the upper surface area for each 1 mg added to the mass, and does not exhibit allometric change from small to large individuals. There is a moderately strong and significant relationship between the two size parameters (mass, upper surface area) and the number of apothecia produced on the upper surface. The relationship is linear and does not indicate increased allocation to reproduction with increased size. However, the thalli have to reach a certain minimum size to produce apothecia (= 30 mg), indicating the existence of a developmental threshold for reproduction. Thalli up to 660 mg may however be without apothecia. The observations indicate that thallus size is a fairly good predictor of current reproductive status in this lichen.

Control of apothecia of Sclerotinia sclerotiorum by soil amendment with S–H mixture or Perlka ® in bean, canola and wheat fields

Ascospores of Sclerotinia sclerotiorum produced from apothecia are the primary source of inoculum for causing diseases such as white mold of common bean, pod rot of pea, stem blight of canola and head rot of sunflower and safflower in the Canadian prairies. A field study was conducted for 4 years to determine efficacy of control of production of apothecia from carpogenically germinated sclerotia of S. sclerotiorum by soil amendment with Perlka ® (calcium cyanamide) and S–H mixture (a formulated compound). Results of the 4-year experiments showed that amendment of soil with Perlka ® at low (30 g/m 2) or high (60 g/m 2) rate was effective in reducing carpogenic germination of sclerotia and production of apothecia under the canopy of host crops (common bean and canola) and a non-host crop (wheat). In the experiments of 1988, for example, the numbers of apothecia produced in the treatments of Perlka ®-low rate (30 g/m 2), Perlka ®-high rate (60 g/m 2) and untreated control were 42, 46, and 182 apothecia/plot (m 2), respectively, for bean; 89, 42, and 318 apothecia/plot (m 2), respectively, for canola; and 146, 143, and 412 apothecia/plot (m 2), respectively, for wheat. However, soil amendment of S–H mixture at low (30 g/m 2) or high (60 g/m 2) rate was ineffective in reducing carpogenic germination of sclerotia and production of apothecia for all the 4 years of testing in all three crops. The ineffectiveness of S–H mixture and the practicality of Perlka ® for control of Sclerotinia diseases of crops grown under Canadian prairie conditions are discussed.

White Mold of Houndstongue (Cynoglossum officinale) Caused by Sclerotinia sclerotiorum in Canada.

Houndstongue (Cynoglossum officinale L.) is a rangeland weed introduced into Canada from Eurasia, and it can be highly toxic to livestock feeding in pastures (3). During 2004, houndstongue plants near Creston, BC, Canada developed water-soaked lesions with white mycelia and black sclerotia on leaves and crowns. Isolations from diseased leaf tissues and sclerotia on potato dextrose agar (PDA) at 20°C for 5 to 7 days produced fungal colonies with formation of black sclerotia 5 to 10 mm in diameter. A single hyphal tip isolate from houndstongue. Ss-HT-C. was compared with a sunflower isolate of S. sclerotiorum, sun-87 (1), for morphology and pathogenicity. For apothecial production, Ss-HT-C and sun-87 were grown on PDA in petri dishes at 10°C for 10 weeks, and sclerotia produced were harvested, placed on moist vermiculite in petri dishes, and incubated at 20°C under light for 3 weeks. Mature apothecia were excised, stained with acid fuchsin, mounted on slides, and examined for asci and ascospores with a microscope. There were no morphological differences between Ss-HT-C and sun-87, each producing an ascus with eight binucleate, elliptical ascospores, measuring 4 × 10 μm (width × length), supporting the identity of Ss-HT-C as S. sclerotiorum (2,4). For pathogenicity tests of Ss-HT-C and sun-87, mycelial plugs (8 mm in diameter) were removed from the margin of colonies grown on PDA for 5 days at 20°C, and placed on leaves of C. officinale plants that were grown in a greenhouse (20 ± 4°C) to the rosette stage. Inoculated plants were covered with clear plastic bags, kept in the same greenhouse for 3 days, and the diameters of the leaf lesions developed at inoculation sites were measured. The experiment was run twice with 30 plants per isolate and five leaves per plant. Uninoculated plants covered with plastic bags were used as controls. Experiments used a completely randomized design. Results of leaf inoculations showed that Ss-HT-C and sun-87 were pathogenic to hound-stongue. There was no statistical difference between isolates or trials. The frequency of leaves with lesions was 90% for Ss-HT-C and 93% for sun-87. The mean leaf lesion diameters were 32 and 35 mm for Ss-HT-C and sun-87, respectively. Leaves of control plants remained healthy. S. sclerotiorum was reisolated from leaves with lesions, but not from controls. After 14 to 21 days, new sclerotia, 5 to 10 mm in diameter, were formed on leaves of inoculated plants. The plants eventually died. This study confirms that S. sclerotiorum is the causal agent for the disease of hound-stongue in Canada, and to our knowledge, this is the first world record of infection of this weed by S. sclerotiorum.

Effect of soil microorganisms on viability of sclerotia of Ciborinia camelliae, the causal agent of camellia flower blight

Two isolates each of Trichoderma virens and Coniothyrium minitans, four of T. longipile, and one each of T. tomentosum and Clonostachys rosea, all known to be sclerotial parasites of Sclerotinia sclerotiorum, were tested for their antagonism to sclerotia of the closely related pathogen, Ciborinia camelliae. Treatment of sclerotia in sterile sand or potting‐mix assays with isolate suspensions of 106 or 107 spores/ml had little effect on the viability of sclerotia, 12–37 weeks after treatment. In a field trial, spore suspensions of T. virens LU556, C. rosea LU155, and C. minitans LU1 12, sprayed in July, August, and September, at a total of 0.8–2.9 × 1011 spores/m2 onto soil naturally infested with over 600 sclerotia/m2, had no effect on apothecial production or on sclerotial numbers recovered at the end of the trial, 8 weeks after the last mycoparasite application. Parasitism of sclerotia by C. minitans, C. rosea, and T. virens was not observed under scanning electron microscopy examination. Thus, the test isolates were not considered effective parasites of C. camelliae sclerotia. Subsequently, 376 microorganisms isolated from C. camelliae sclerotia baits that had been buried in soil, and 30 microorganisms from decaying C. camelliae sclerotia, were screened in vitro for their ability to parasitise healthy sclerotia of the pathogen. The most effective antagonists were T. virens LU569 and T. viride LU570, which reduced numbers of viable C. camelliae sclerotia after 14 weeks by 53% and 43%, respectively. These isolates warrant further evaluation under field conditions where one can potentially manipulate the soil environment by the addition of soil conditioners to enhance their efficacy in situ.