We studied the diversity and biocontrol potential of 100 fungal endophytes isolated from Espeletia spp., endemic plant species from the Paramo in the Andean mountain range. Our sample was genotypically highly diverse at all ITS similarity levels. The antagonistic properties of these isolates were tested against common crop pathogens in Colombia, including Pectobacterium carotovorum, Ralstonia solanacearum, Pseudomonas syringae, Xanthomonas campestris, Rhizoctonia solani, Botrytis cinerea, Fusarium oxysporum, and Phytophthora infestans. All endophytic isolates were able to significantly inhibit the growth of at least one of the plant pathogens tested (P \ 0.05). Three main types of endophyte/pathogen interactions were observed.However, only those endophytes that produced an evident inhibition halo were further studied using their crude extracts to confirm that the inhibitory effect was due to the production of endophytic bioactive metabolites. From these experiments, nine promising isolates were selected for co-inoculation tests with R. solani in tomato plants. The isolates identified as Aureobasidium pullulans and Paraconiothyrium sporulosum not only protected the plants against this pathogen but also allowed them to exhibit similar growth and development as the uninoculated control. This work explores new alternatives for disease management without the application of chemical pesticides.
Powdery mildew, caused by Erysiphe necator, is the most common and destructive disease of grapes (Vitis spp.) worldwide. In Michigan, it is primarily controlled with fungicides, including strobilurins (quinone outside inhibitors [QoIs]). Within the United States, resistance to this class of fungicides has been reported in E. necator populations in some east coast states. Among 12 E. necator isolates collected from five Michigan vineyards in 2008, one carried the G143A single-nucleotide mutation responsible for QoI resistance. This isolate was confirmed to be resistant in a conidium germination assay on water agar amended with trifloxystrobin at 0.001, 0.01, 0.1, 1, 10, or 100 μg/ml and salicylhydroxamic acid (100 mg/liter). The mutant isolate was able to germinate on media amended with 100 μg/ml trifloxystrobin, whereas a representative wild-type isolate did not germinate at concentrations higher than 0.1 μg/ml. In 2009, 172 isolates were collected from a total of 21 vineyards (juice and wine grapes): three vineyards with no fungicide application history (baseline sites), six research vineyards, and 12 commercial vineyards. QoI resistance was defined as the effective concentration that inhibited 50% of conidial germination (EC50) > 1 μg/ml. Isolates from baseline sites had EC50 values mostly below 0.01 μg/ml, while isolates that were highly resistant to trifloxystrobin (EC50 > 100 μg/ml) occurred in five research and three commercial wine grape vineyards at frequencies of 40 to 100% and 25 to 75% of the isolates, respectively. The G143A mutation was detected in every isolate with an EC50 > 1 μg/ml. These results suggest that fungicide resistance may play a role in suboptimal control of powdery mildew observed in some Michigan vineyards and emphasizes the need for continued fungicide resistance management.
Field isolates of Alternaria solani, which causes early blight of potato in Idaho, USA were evaluated in vitro for their sensitivity towards the succinate dehydrogenase inhibitor (SDHI) fungicides boscalid, fluopyram and penthiopyrad. A total of 20 isolates were collected from foliar-infected tissue in 2009, 26 in 2010 and 49 in 2011. Fungicide sensitivity was tested using the spiral-gradient end point dilution method. The frequency of boscalid-resistant isolates (>50% relative growth when using a spiral dilution gradient starting at 507 mg L À1 ) drastically increased over the duration of this study (15% in 2009, 62% in 2010 and 80% in 2011). Increasing resistance to fluopyram and penthiopyrad was observed. However, cross-resistance was only observed between boscalid and penthiopyrad. The target site of this fungicide class is the succinate dehydrogenase (SDH) enzyme complex, which is vital for fungal respiration. Sequence analysis of the SDH complex revealed mutations in the subunits B and D that were correlated with the emergence of boscalid resistance in potato fields in Idaho. In particular, H277R and H133R were identified in SDH subunits B and D, respectively. The presence of restriction sites in the gene sequences allowed the development of a rapid PCR-RFLP method to assess boscalid sensitivity in A. solani populations.
European hazelnut (Corylus avellana L.) is an emerging crop for export, mainly in southern Chile. Stem cankers and dieback of twigs on sixyear-old European hazelnut cultivar Barcelona were observed during the 2012 growing season on plantations in Panguipulli (39° 38' 37.12" S and 72° 20' 10.87" W), Region de Los Rios, Chile. The incidence has been variable according to the place of plantation; it was estimated at approximately 15%. Cankers were characterized by brownish-gray and brown to reddish discoloration of the vascular stem system. Hazelnut plants between 1 and 3 years old developed stem basal canker, especially at conditions of high humidity and overpopulation of weeds; at critical conditions, the affected plants generally die. Small pieces of cankered stems, selected from 10 European hazelnuts, were surface sterilized in 0.5% sodium hypochlorite for 2 min and rinsed twice in sterile distilled water prior to incubation in a humid chamber for 7 days (25 ± 2°C) to stimulate production of reproductive bodies. Black sub-epidermal perithecia with unitunicate, cylindrical-clavate, 8-spored asci {n = 20) were obtained. Ascospores were septated, hyaline, multigutulate, and slightly constricted at the septum, the average measurements were (n = 20) 13.4 ± 0.6 |am x 3.9 ± 0.2 (im. The ascospores were transferred to potato dextrose agar (PDA) and incubated for 6 days at 25°C in the dark, then hyphal tips were transferred to fresh PDA and obtained a mycelia with white, cottony, and sparse colonies. Pycnidia and smooth, unicellular, hyaline, and biguttulate alpha conidia of 6.1 to 7.2 |am x 2.8 to 3.1 |jm (n = 40) were observed. Beta conidia were not observed in culture media. Mature pycnidia were also detected on hazelnut shells remaining on the soil from the previous season. The identification of the species (isolate IMI-501237) was confirmed at CABI, United Kingdom, using an internal transcribed spacer (ITS), rDNA, BLASTn analysis of the 524-bp fragment, and showed 100% identity with Diaporthe australafricana Crous & J.M. van Niekerk (accessions KC343039, KC343038). These molecular and morphological characteristics were similar that reported from Vitis vinifera (2) and Chilean blueberry (3). The sequence obtained was deposited in GenBank (Accession No. JX3162I8.I). A pathogenicity test was conducted with isolate IMl-501237 on four 1-year-old plants from the hazelnut cultivar Barcelona. Plants were maintained in individual bags in greenhouse conditions (14/10 h dark/light, 20°C; 70% relative humidity). Prior to inoculation, plant tissues were surface disinfected with 2% sodium hypochlorite and rinsed with sterile distilled water. Each plant was inoculated at fresh wound sites on three stems and three vegetative buds on twigs. The inoculum consisted of an agar plug with mycelia (5 mm) from the edge of an actively growing colony cultured on PDA for 6 days. Each inoculation was covered with moistened cotton and sealed with Parafilm; a control plant was inoculated in the same way with agar only. After 30 days, necrotic le...
Mummy berry is one of the most important fungal diseases of cultivated blueberry (Vaccinium spp.) worldwide causing yield losses of up to 70 to 85% and entire lot rejections. This disease is caused by the ascomycete fungus Monilinia vaccinii-corymbosi and is controlled primarily using cultural practices and the prophylactic use of fungicides early in the growing season. This pathogen has multiple phases and depending on the life cycle varies in its difficulty to diagnose. This diagnostic guide provides details about the current taxonomy, host, geographic range, symptoms and signs, as well as effective techniques to aid pathogen identification, evaluation, isolation, and storage for M. vaccinii-corymbosi.
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