Monilinia Fructicola Research Articles

Identification of Volatile Organic Compounds in Extremophilic Bacteria and Their Effective Use in Biocontrol of Postharvest Fungal Phytopathogens.

Phytopathogenic fungal growth in postharvest fruits and vegetables is responsible for 20–25% of production losses. Volatile organic compounds (VOCs) have been gaining importance in the food industry as a safe and ecofriendly alternative to pesticides for combating these phytopathogenic fungi. In this study, we analysed the ability of some VOCs produced by strains of the genera Bacillus, Peribacillus, Pseudomonas, Psychrobacillus and Staphylococcus to inhibit the growth of Alternaria alternata, Botrytis cinerea, Fusarium oxysporum, Fusarium solani, Monilinia fructicola, Monilinia laxa and Sclerotinia sclerotiorum, in vitro and in vivo. We analysed bacterial VOCs by using GC/MS and 87 volatile compounds were identified, in particular acetoin, acetic acid, 2,3-butanediol, isopentanol, dimethyl disulphide and isopentyl isobutanoate. In vitro growth inhibition assays and in vivo experiments using cherry fruits showed that the best producers of VOCs, Bacillus atrophaeus L193, Bacillus velezensis XT1 and Psychrobacillus vulpis Z8, exhibited the highest antifungal activity against B. cinerea, M. fructicola and M. laxa, which highlights the potential of these strains to control postharvest diseases. Transmission electron microscopy micrographs of bacterial VOC-treated fungi clearly showed antifungal activity which led to an intense degeneration of cellular components of mycelium and cell death.

Early almond harvest as a strategy for sustainable irrigation, pest and disease management

Almonds (Prunus dulcis (Mill.) D. A. Webb) are one of the most economically important agricultural commodities in California. As production in the state continues to increase, growers seek sustainable methods to manage agricultural water use and pests that are affordable and environmentally sustainable. The primary objectives of this research were to test the efficacy of hull split regulated deficit irrigation in combination with early harvest of almonds (Prunus dulcis (Mill.) D. A. Webb) for improved irrigation efficiency and control of navel orangeworm (NOW) (Amyelois transitella (Walker)) and hull rot (HR) (Rhizopus stolonifer (Ehrenb:Fr.) Vuill.; Monilinia fructicola (G. Wint.) Honey). Standard Harvest (SH) timing was compared to Early Harvest (EH) timing at approximately three to four weeks earlier in two California almond orchards using a randomized complete block design at each orchard. Stem water potential (SWP) measurements were used to monitor tree water status and manipulate irrigation to prepare trees for EH treatments. Trees were maintained at -1400 to -1800 kPa SWP for two to three weeks leading up to both EH and SH. There were significantly lower HR strikes in EH treatment trees than in SH treatment trees, and significant reductions in the percentage of kernels infested with NOW when EH was employed. There were no significant kernel yield, marketable yield, or shaker efficacy differences between the two treatments. Although 83.3 and 40.9 mm less water was applied to EH trees than SH trees at the Denair and Woodland sites respectively, the trees from both treatment regimes at both sites returned to the same water status after SH and recovered from the induced moderate water stress by the same date after harvest. With further development of complementary harvest and processing strategies, early harvest of almonds shows promise as a strategy for improved water use efficiency and pest and disease control in almond orchards.

Postharvest Treatments with Sulfur-Containing Food Additives to Control Major Fungal Pathogens of Stone Fruits.

The sulfur-containing salts, classified as food additives, sodium metabisulfite (SMBS), potassium metabisulfite (PMBS), aluminum sulfate (AlS), and aluminum potassium sulfate (AlPS), were evaluated for their activity against Monilinia fructicola, Rhizopus stolonifer, and Geotrichum candidum, the most economically important fungal pathogens causing postharvest disease of stone fruit. In in vitro tests with potato dextrose agar (PDA) Petri dishes amended with different concentrations of the salts (0, 10, 20, 30, 50, and 100 mM), SMBS and PMBS at all concentrations, AlS above 20 mM, and AlPS above 30 mM, completely inhibited the mycelial growth of the three fungi after incubation at 25 °C for up to 10 days. In in vivo primary screenings with artificially inoculated nectarines, aqueous solutions of the four salts reduced the incidence and severity of brown rot (BR) at concentrations of 10 and 50 mM, whereas only AlS and AlPS reduced Rhizopus rot (RR), and none of the salts was effective against sour rot (SR). Solutions at 100 mM were phytotoxic and injured the fruit peel. In small-scale trials, 1 min dip treatments at 20 °C in SMBS or PMBS at 10 mM significantly reduced the incidence and severity of BR after incubation at 20 °C for up to 8 days. Conversely, dips in AlS and AlPS reduced neither BR nor RR. Results highlight the potential of SMBS and PMBS as new nonpolluting tools for the integrated control of BR, but not RR and SR, on stone fruit.

Mushroom extract of Lactarius deliciosus (L.) Sf. Gray as biopesticide: Antifungal activity and toxicological analysis

ABSTRACT Monilinia fructicola (Wint.) Honey is a plant pathogenic fungus that infects stone fruits such as peach, nectarine and plum, which are high demand cultivars found in Brazil. This pathogen may remain latent in the host, showing no apparent signs of disease, and consequently may spread to different countries. The aim of this study was to evaluate the activity of hydroalcoholic extract (HydE) obtained from Lactarius deliciosus (L.) Sf. Gray a mushroom, against M. fructicola phytopathogenic-induced mycelial growth. In addition, the purpose of this study was to examine phytotoxicity attributed to HydE using Brassica oleracea seeds, as well as cytotoxic analysis of this extract on cells of mouse BALB/c monocyte macrophage cell line (J774A.1 cell line) (ATCC TIB-67). The L. deliciosus HydE inhibited fungal growth and reduced phytopathogen mycelial development at a concentration of 1.25 mg/ml. Our results demonstrated that the extract exhibited phytotoxicity as evidenced by (1) interference on germination percentage and rate index, (2) decreased root and initial growth measures, and (3) lower fresh weight of seedlings but no cytotoxicity in Vero cell lines. Data suggest that the use of the L. deliciosus extracts may be beneficial for fungal control without any apparent adverse actions on mouse BALB/c monocyte macrophage cell line (J774A.1 cell line) viability.

Baseline Sensitivities of Major Citrus, Pome, and Stone Fruits Postharvest Pathogens to Natamycin and Estimation of the Resistance Potential in Penicillium digitatum.

Natamycin is a biofungicide that was registered in the United States in 2016 and approved in California in 2017 for postharvest use on citrus and stone fruits. It has been used as a food preservative for many decades, with no resistance ever observed to date. The objective of this study was to determine baseline sensitivities for mycelial growth of 43 to 72 isolates of seven postharvest pathogens to natamycin and the resistance potential of Penicillium digitatum. Mean effective concentrations to inhibit mycelial growth by 50% (EC50 values), as determined by the spiral gradient method, were 0.90 μg/ml for Alternaria alternata, 0.76 μg/ml for Botrytis cinerea, 3.20 μg/ml for Geotrichum citri-aurantii, 0.17 μg/ml for Monilinia fructicola, 1.54 μg/ml for P. digitatum, 1.14 μg/ml for P. expansum, and 0.48 μg/ml for Rhizopus stolonifer. Distributions of EC50 values for each pathogen were unimodal and mostly normal with no outliers detected. Natamycin was also inhibitory to spore germination with values for five of the species similar to those for mycelial growth. Microscopically, natamycin generally arrested spores at the pregermination swelling stage. Mass platings of a conidial mixture of 10 isolates of P. digitatum were inoculated on agar media with 2.5-log radial concentration gradients of natamycin or fludioxonil, and a conidial mixture of 10 isolates of G. citri-aurantii were plated on media amended with natamycin or propiconazole. No resistant isolates were observed for both species to natamycin or for G. citri-aurantii to propiconazole, whereas a resistance frequency of 4.5 × 10-6 to 3.1 × 10-6 was calculated for P. digitatum to fludioxonil. The wide spectrum of activity against different fungal pathogens and a low resistance potential support the registration of natamycin as a postharvest treatment and its integration into an integrated pest management program with other practices including sanitation and rotation of other fungicides with different modes of action.

Genetic diversity assessments of brown rot pathogen Monilinia fructicola based on the six simple sequence repeat loci

Genetic diversity assessments of brown rot pathogen Monilinia fructicola based on the six simple sequence repeat loci

Chemical Composition and Antimicrobial Properties of Mentha × piperita cv. 'Kristinka' Essential Oil.

Several economically important crops, fruits and vegetables are susceptible to infection by pathogenic fungi and/or bacteria postharvest or in field. Recently, plant essential oils (EOs) extracted from different medicinal and officinal plants have had promising antimicrobial effects against phytopathogens. In the present study, the potential microbicide activity of Mentha × piperita cv. ‘Kristinka’ (peppermint) EO and its main constituents have been evaluated against some common phytopathogens. In addition, the cell membrane permeability of the tested fungi and the minimum fungicidal concentrations were measured. The antifungal activity was tested against the following postharvest fungi: Botrytis cinerea, Monilinia fructicola, Penicillium expansum and Aspergillus niger, whereas antibacterial activity was evaluated against Clavibacter michiganensis, Xanthomonas campestris, Pseudomonas savastanoi and P. syringae pv. phaseolicola. The chemical analysis has been carried out using GC-MS and the main components were identified as menthol (70.08%) and menthone (14.49%) followed by limonene (4.32%), menthyl acetate (3.76%) and β-caryophyllene (2.96%). The results show that the tested EO has promising antifungal activity against all tested fungi, whereas they demonstrated only a moderate antibacterial effect against some of the tested bacteria.

In Vitro Antifungal Activity and Toxicity of Dihydrocarvone-Hybrid Derivatives against Monilinia fructicola.

The aim of this study was to synthesize a series of novel and known dihydrocarvone-hybrid derivatives (2–9) and to evaluate mycelial growth activity of hybrid molecules against two strains of Monilinia fructicola, as well as their toxicity. Dihydrocarvone-hybrid derivatives have been synthesized under sonication conditions and characterized by FTIR, NMR, and HRMS. Antifungal efficacy against both strains of M. fructicola was determined by half maximal effective concentration (EC50) and toxicity using the brine shrimp lethality test (BSLT). Among the synthesized compounds, 7 and 8 showed the best activity against both strains of M. fructicola with EC50 values of 148.1 and 145.9 µg/mL for strain 1 and 18.1 and 15.7 µg/mL for strain 2, respectively, compared to BC 1000® (commercial organic fungicide) but lower than Mystic® 520 SC. However, these compounds showed low toxicity values, 910 and 890 µg/mL, respectively, compared to Mystic® 520 SC, which was highly toxic. Based on the results, these hybrid compounds could be considered for the development of more active, less toxic, and environmentally friendly antifungal agents against phytopathogenic fungi.

Antifungal mechanisms of lavender essential oil in the inhibition of rot disease caused by Monilinia fructicola in postharvest flat peaches

Lavender essential oil (LEO), a natural antimicrobial agent, is generally recognized as safe and effective in the inhibition of phytopathogenic fungi. Direct contact and fumigation (in vivo and in vitro) were used to study the fungistatic effect of LEO on Monilinia fructicola. Additionally, the effect on the ultrastructure of cells and the degree of destruction of the cell membrane of M. fructicola were revealed. In addition, the effects of LEO on the expression levels of apoptosis-related genes in M. fructicola cells were detected, and GC-MS was used to analyze the main components of LEO. LEO had a good inhibitory efficacy against M. fructicola in flat peaches, with almost complete growth inhibition at 800 μL/L. These effects were associated with the leakage of cytoplasmic contents, hyphal distortion, and spore disruption. Moreover, the expression of apoptosis RTG1 and RLM1 genes increased with LEO treatment. These results demonstrate that LEO can inhibit M. fructicola by inducing cytoplasmic membrane damage and cell apoptosis in fungi, and that the major ingredients of LEO are monoterpenes and sesquiterpenes, which are presumed to contribute to the inhibitory effects.

Monilinia fructicola on loquat: An old pathogen invading a new host

Monilinia fructicola has been widely reported as the causal agent of brown rot disease on many Rosaceae family fruits worldwide. It has been reported on stone fruits, e.g., peach, plum, cherry, apricot and mume; as well as pome fruits, e.g., apple, pear and hawthorn. Loquat is a member of the Eriobotrya genus in subfamily Maloideae along with apple, pear and hawthorn. So far, loquat has not been reported as the host of any Monilia species. In June 2019, brown rot symptoms were observed on loquat fruits in an orchard in Wuhan, Hubei Province, China. Thirty single spore isolates were obtained and identified as M. fructicola based on morphological characteristics and molecular analysis. This is the first report of loquat brown rot disease caused by Monilia species in the world. Furthermore, upon artificial inoculation, all three Monilia species from peach in China, i.e., M. fructicola, M. mumecola and M. yunnanensis, could cause typical brown rot disease on loquat fruits. At the same time, M. fructicola isolates from loquat showed virulence similar to those isolates from peach when the pathogenicity test was conducted on peach fruits. These results suggested that loquat could be infected by other Monilia species and that isolates from loquat also have potential to damage other Rosaceae family fruits in practice.

Pan-Mitogenomics Approach Discovers Diversity and Dynamism in the Prominent Brown Rot Fungal Pathogens.

Monilinia fructicola and Monilinia laxa species are the most destructive and economically devastating fungal plant pathogens causing brown rot disease on stone and pome fruits worldwide. Mitochondrial genomes (mitogenomes) play critical roles influencing the mechanisms and directions of the evolution of fungal pathogens. The pan-mitogenomics approach predicts core and accessory regions of the mitochondrial genomes and explains the gain or loss of variation within and between species. The present study is a fungal pan-mitogenome of M. fructicola (N = 8) and M. laxa (N = 8) species. The completely sequenced and annotated mitogenomes showed high variability in size within and between the species. The mitogenomes of M. laxa were larger, ranging from 178,351 to 179,780bp, than the mitogenomes of M. fructicola, ranging from 158,607 to 167,838bp. However, size variation within the species showed that M. fructicola isolates were more variable in the size range than M. laxa isolates. All the mitogenomes included conserved mitochondrial genes, as well as variable regions including different mobile introns encoding homing endonucleases or maturase, non-coding introns, and repetitive elements. The linear model analysis supported the hypothesis that the mitogenome size expansion is due to presence of variable (accessory) regions. Gene synteny was mostly conserved among all samples, with the exception for order of the rps3 in the mitogenome of one isolate. The mitogenomes presented AT richness; however, A/T and G/C skew varied among the mitochondrial genes. The purifying selection was detected in almost all the protein-coding genes (PCGs) between the species. However, cytochrome b was the only gene showing a positive selection signal among the total samples. Combined datasets of amino acid sequences of 14 core mitochondrial PCGs and rps3 obtained from this study together with published mitochondrial genome sequences from some other species from Heliotales were used to infer a maximum likelihood (ML) phylogenetic tree. ML tree indicated that both Monilinia species highly diverged from each other as well as some other fungal species from the same order. Mitogenomes harbor much information about the evolution of fungal plant pathogens, which could be useful to predict pathogenic life strategies.

A novel approach for in vitro fungicide screening and the sensitivity of Monilinia populations from peach orchards in Turkey to respiratory inhibitor fungicides

Brown rot caused by Monilinia fructicola and M. laxa can result in considerable losses in peach production worldwide. Respiratory Inhibitors (RIs) have been used to control the disease in Turkey and worldwide. However, resistance against RIs may have been developing in Monilinia populations in Turkey. The sensitivity level of the total of 128 isolates of Monilinia fructicola and M. laxa from peach fruits in Turkey were tested against certain RIs: azoxystrobin, boscalid, and commercially used signum (26,7% boscalid, 6,7% pyraclostrobin). Fungal growth from both mycelia and conidia was evaluated as fungicide response phenotypes. A new approach was developed to define the sensitivity levels of the isolates. The mean IC50 values of each fungicide on representative collection from each species were used as discriminatory doses in view of both phenotypes. The whole collection was scanned at discriminatory doses and the relative growth (RG) of each isolate was obtained. The distribution of RG data sets in the quantile ranges was considered to define sensitivity levels of the isolates against each fungicide. Resistance levels for mycelial and conidial growth assays varied within and between the species against the three RIs. According to the pairwise correlation analyses between responses of the species to the fungicides, the significant correlation indicating a cross-resistance was only found between boscalid and signum on conidial growth assays for M. fructicola. This study represents in vitro fungicide responses of a large population of Monilinia pathogens from peach in Turkey for the first time which will help to reconsider disease control strategies and suggests a fungicide sensitivity definition be used in all in vitro fungicide assays as a common and comparable strategy at the global stage.

Evaluating the biocontrol potential of Canadian strain Bacillus velezensis 1B-23 via its surfactin production at various pHs and temperatures

BackgroundMicroorganisms, including Bacillus species are used to help control plant pathogens, thereby reducing reliance on synthetic pesticides in agriculture. Bacillus velezensis strain 1B-23 has been shown to reduce symptoms of bacterial disease caused by Clavibacter michiganensis subsp. michiganensis in greenhouse-grown tomatoes, with in vitro studies implicating the lipopeptide surfactin as a key antimicrobial. While surfactin is known to be effective against many bacterial pathogens, it is inhibitory to a smaller proportion of fungi which nonetheless cause the majority of crop diseases. In addition, knowledge of optimal conditions for surfactin production in B. velezensis is lacking.ResultsHere, B. velezensis 1B-23 was shown to inhibit in vitro growth of 10 fungal strains including Candida albicans, Cochliobolus carbonum, Cryptococcus neoformans, Cylindrocarpon destructans Fusarium oxysporum, Fusarium solani, Monilinia fructicola, and Rhizoctonia solani, as well as two strains of C. michiganensis michiganensis. Three of the fungal strains (C. carbonum, C. neoformans, and M. fructicola) and the bacterial strains were also inhibited by purified surfactin (surfactin C, or [Leu7] surfactin C15) from B. velezensis 1B-23. Optimal surfactin production occurred in vitro at a relatively low temperature (16 °C) and a slightly acidic pH of 6.0. In addition to surfactin, B. velenzensis also produced macrolactins, cyclic dipeptides and minor amounts of iturins which could be responsible for the bioactivity against fungal strains which were not inhibited by purified surfactin C.ConclusionsOur study indicates that B. velezensis 1B-23 has potential as a biocontrol agent against both bacterial and fungal pathogens, and may be particularly useful in slightly acidic soils of cooler climates.

First report of brown rot of nectarine caused by Monilia yunnanensis in Tibet.

Brown rot caused by Monilinia spp. is one of the most important diseases of stone fruits. To date, three species of Monilinia have been found to occur on Prunus species worldwide: Monilinia fructicola (G. Winter) Honey, Monilinia fructigena (Aderhold & Ruhland) Honey, and Monilinia laxa (Aderhold & Ruhland) Honey (Zhu et al. 2005; Hu et al. 2011a). While M. fructicola is widespread in the Americas, and parts of Europe and Asia (CABI, 2010), M. laxa and M. fructigena are the primary species causing brown rot of peach in Europe (Bryde et al. 1977). In China, a new species Monilia yunnanensis was identified in 2011 (Hu et al. 2011b; Zhao et al. 2013; Yin et al. 2015; Yin et al. 2017). However, the species causing brown rot of nectarine (Prunus persica var. nectarina) in Tibet have not been undertaken. In the summer of 2017-2018, brown rot disease of nectarine was observed in Nyingchi, Tibet, and approximately 30% of nectarines were affected annually. Therefore, the brown rot disease of nectarine is one of the main factors that restrict the yield and quality of nectarine fruit production, and causes severe economic losses in Tibet. Thirty-six nectarine fruit with typical brown rot symptoms were collected from Tibet during the summer of 2017-2018. In order to isolate the causal agent, small pieces of pericarp were disinfected with 75% ethanol for 1 min, and then for 1 min in 1% NaOCl, rinsed in sterile distilled water for three times, dried on sterile paper and placed on potato dextrose agar (PDA). Thirty-six single-spore isolates were obtained and all morphologically similar, and three representative isolates 2-1, 2-16 and 2-31 which were from different period and years in 2017-2018 were characterized phylogenetically and morphologically to identify them to species level. Pathogenicity of each representative isolate was confirmed by inoculating five surface-disinfected mature nectarines with mycelial plugs in the wound of the fruit. Nectarine fruit inoculated with sterile PDA plugs served as the negative control. The inoculated nectarines developed brown lesions after 6 days incubation at 22°C, and the pathogen was successfully re-isolated. There were no symptoms on the control nectarine fruit. The isolates 2-1, 2-16 and 2-31 produced gray-green colonies with even margins and concentric rings of sporogenous mycelium after 3 days incubation, and abundant black-colored stromata on the media after 16 days of incubation at 22°C, resembling those described for M. yunnanensis (Hu et al. 2011b). Conidia were one-celled, hyaline, ellipsoid to lemon shape (9.24 to 15.58 μm), and borne in branched monilioid chains. The average daily growth of mycelium on PDA at 22°C was 11.56 mm. Therefore, the isolates 2-1, 2-16 and 2-31 were preliminarily identified as M. yunnanensis based on the morphological investigations (Hu et al. 2011b). Morphological identification was confirmed by phylogenetic analysis based on sequences of glyceraldehyde-3-phosphate dehydrogenase (G3PDH) and β-tubulin (TUB2) genes of 2-1, 2-16 and 2-31 which were amplified using primers Mon-G3pdhF/Mon-G3pdhR and Mon-TubF1/Mon-TubR1 (Hu et al. 2011b). In both G3PDH and TUB2 phylogenetic trees, the isolates 2-1, 2-16 and 2-31 formed monophyletic clades within a derived clade with the M. yunnanensis isolates. Additionally, the three isolates were more closely related to M. yunnanensis (HQ908782.1 and HQ908783.1) than to other Monilinia species. Based on morphological and molecular identification, the isolates 2-1, 2-16 and 2-31 were identified as M. yunnanensis. Previously, M. yunnanensis has been reported as a new species causing brown rot of peach in China (Hu et al, 2011b). To our knowledge, this is the first report of M. yunnanensis causing nectarine fruit brown rot in Tibet. These findings suggest that M. yunnanensis is spreading on its principal host plants and causing substantial economic losses in the Tibet fruit production.

A Method for the Examination of SDHI Fungicide Resistance Mechanisms in Phytopathogenic Fungi Using a Heterologous Expression System in Sclerotinia sclerotiorum.

Succinate dehydrogenase inhibitors (SDHIs) are a class of broad-spectrum fungicides used for management of diseases caused by phytopathogenic fungi. In many cases, reduced sensitivity to SDHI fungicides has been correlated with point mutations in the SdhB and SdhC target genes that encode components of the succinate dehydrogenase complex. However, the genetic basis of SDHI fungicide resistance mechanisms has been functionally characterized in very few fungi. Sclerotinia sclerotiorum is a fast-growing and SDHI fungicide-sensitive phytopathogenic fungus that can be conveniently transformed. Given the high amino acid sequence similarity and putative structural similarity of SDHI protein target sites between S. sclerotiorum and other common phytopathogenic ascomycete fungi, we developed an in vitro heterologous expression system that used S. sclerotiorum as a reporter strain. With this system, we were able to demonstrate the function of mutant SdhB or SdhC alleles from several ascomycete fungi in conferring resistance to multiple SDHI fungicides. In total, we successfully validated the function of Sdh alleles that had been previously identified in field isolates of Botrytis cinerea, Blumeriella jaapii, and Clarireedia jacksonii (formerly S. homoeocarpa) in conferring resistance to boscalid, fluopyram, or fluxapyroxad and used site-directed mutagenesis to construct and phenotype a mutant allele that is not yet known to exist in Monilinia fructicola populations. We also examined the functions of these alleles in conferring cross-resistance to more recently introduced SDHIs including inpyrfluxam, pydiflumetofen, and pyraziflumid. The approach developed in this study can be widely applied to interrogate SDHI fungicide resistance mechanisms in other phytopathogenic ascomycetes.

Deciphering the Monilinia fructicola Genome to Discover Effector Genes Possibly Involved in Virulence.

Brown rot is the most economically important fungal disease of stone fruits and is primarily caused by Monilinia laxa and Monlinia fructicola. Both species co-occur in European orchards although M. fructicola is considered to cause the most severe yield losses in stone fruit. This study aimed to generate a high-quality genome of M. fructicola and to exploit it to identify genes that may contribute to pathogen virulence. PacBio sequencing technology was used to assemble the genome of M. fructicola. Manual structural curation of gene models, supported by RNA-Seq, and functional annotation of the proteome yielded 10,086 trustworthy gene models. The genome was examined for the presence of genes that encode secreted proteins and more specifically effector proteins. A set of 134 putative effectors was defined. Several effector genes were cloned into Agrobacterium tumefaciens for transient expression in Nicotiana benthamiana plants, and some of them triggered necrotic lesions. Studying effectors and their biological properties will help to better understand the interaction between M. fructicola and its stone fruit host plants.

In vitro effect of lemon essential oil and garlic extract on Monilinia fructicola growth

Brown rot produced by Monilinia fructicola (MON) is the main disease that causes postharvest losses in peaches of Argentina. There is a public concern in reducing the use of synthetic fungicides. The objective of this work was to evaluate the effect of lemon essential oil (LO) and aqueous garlic extract (GE) on MON in vitro mycelial growth inhibition (INH). In a first trial a MON colony disc was placed in the middle of a Petri dish containing PDA 2%, or over the plate lid containing 1 mL of the following treatments: Control, LO 0.5%, LO 1%, LO 3% and Fludioxonil 0.02%. In a second trial, GE was obtained processing fresh garlic (cultivar Colorado) in a blender with tap water; 1 mL of garlic extract was placed in the cover of each plate, the treatments were: Control, GE 2.27%, GE 5.68%, GE 11.35% and Fludioxonil 0.02%, the MON colony was placed over the lid containing the extract. All concentrations of LO assayed and GE 11.35% showed INH > 90%. Following these results LO and GE could be evaluated in future trials as postharvest treatments on peach fruits for Monilinia fructicola control.

Effect of phosphites on the content of total phenols and chlorogenic acid in peach-Monilinia fructicola pathosystem

Monilinia fructicola is a fungal pathogen responsible for fruit brown rot in many species of the Prunus genus. The use of abiotic elicitors as phosphites is being studied for the induction of fruit resistance as an alternative to the use of synthetic fungicides. The aim of this work was to evaluate the effect of phosphites on the content of total phenols, chlorogenic acid and changes in susceptibility to M. fructicola in peach fruits. Peach fruits of the cultivars Flordaking and Elegant Lady were treated with potassium- and calcium-phosphites, and harvested at three development stages: green fruit (GF), pit hardening (PH) and harvest time (HT). After harvesting, half of them were inoculated with a conidia suspension of M. fructicola under laboratory conditions. Total phenols and chlorogenic acid content were assayed on the skin of the fruits. Results showed different behaviors according to the cultivar. In Flordaking, the phosphite treatment allowed an increase in the content of total phenols in the stages of GF and PH, but not for chlorogenic acid. In the presence of inoculum, chlorogenic acid decreased, but not the total phenols. The Elegant Lady did not show differences in the content of total phenols and chlorogenic acid after the phosphite treatments. Both metabolites exhibited a seasonal behavior.

Botanical Control of Citrus Green Mold and Peach Brown Rot on Fruits Assays Using a Persicaria acuminata Phytochemically Characterized Extract.

Persicaria acuminata (Polygonaceae) is a perennial herb that grows in the central area of Argentina and it is commonly used by native populations to heal infected wounds and other conditions related to fungal infections. In this article, we explored the in vitro antifungal activity of its ethyl acetate extract against a panel of three fruit phytopathogenic fungi including: Penicillium digitatum, P. italicum, and Monilinia fructicola. The sesquiterpenes isolated from the extract were also evaluated against these strains, demonstrating that the dialdehyde polygodial was the responsible for this activity. In order to encourage the use of the extract rather than the pure compound, we displayed ex vivo assays using fresh oranges and peaches inoculated with P. digitatum and M. fructicola, respectively, and subsequently treated by immersion with an extract solution of 250 and 62.5 µg/mL, respectively. There were no statistically significant differences between the treatments with commercial fungicides and the extract over the control of both fruit rots. The concentration of the active compound present in the extract used on fruit experiments was determined by Gas Chromatography-Mass Spectroscopy. Finally, cytotoxicity evaluation against Huh7 cells showed that P. acuminata extract was less cytotoxic than the commercial fungicides at the assayed concentrations. After these findings we could conclude that a chemically characterized extract of P. acuminata should be further developed to treat fungal diseases in fruits from an agro-ecological model.

Survey and Genetic Analysis of Demethylation Inhibitor Fungicide Resistance in Monilinia fructicola From Michigan Orchards.

Resistance to sterol demethylation inhibitor (DMI) fungicides in Monilinia fructicola, causal agent of brown rot of stone fruit, has been reported in the southeastern and eastern United States and in Brazil. DMI resistance of some M. fructicola isolates, in particular those recovered from the southeastern United States, is associated with a sequence element termed "Mona" that causes overexpression of the cytochrome demethylase target gene MfCYP51. In this study, we conducted statewide surveys of Michigan stone fruit orchards from 2009 to 2011 and in 2019, and we determined the sensitivity to propiconazole of a total of 813 isolates of M. fructicola. A total of 80.7% of Michigan isolates were characterized as resistant to propiconazole by relative growth assays, but the Mona insert was not uniformly detected and was present in some isolates that were not characterized as DMI resistant. Gene expression assays indicated that elevated expression of MfCYP51 was only weakly correlated with DMI resistance in M. fructicola isolates from Michigan, and there was no obvious correlation between the presence of the Mona element and elevated expression of MfCYP51. However, sequence analysis of MfCYP51 from 25 DMI-resistant isolates did not reveal any point mutations that could be correlated with resistance. Amplification and sequencing upstream of MfCYP51 resulted in detection of DNA insertions in a wide range of isolates typed by DMI phenotype and the presence of Mona or other unique sequences. The function of these unique sequences or their presence upstream of MfCYP51 cannot be correlated to a DMI-resistant genotype at this time. Our results indicate that DMI resistance was established in Michigan populations of M. fructicola by 2009 to 2011, and that relative resistance levels have continued to increase to the point that practical resistance is present in most orchards. In addition, the presence of the Mona insert is not a marker for identifying DMI-resistant isolates of M. fructicola in Michigan.