Using plant growth-promoting microorganisms (PGPMs) to improve plant development under in vitro culture conditions.

MAIN CONCLUSION: The use of beneficial microorganisms improves the performance of in vitro - cultured plants through the improvement of plant nutrition, the biological control of microbial pathogens or the production of phytohormones that promote plant growth and development. Plant in vitro culture techniques are highly useful to obtain significant amounts of true-to-type and disease-free plant materials. One of these techniques is clonal micropropagation which consists on the establishment of shoot tip cultures, shoot multiplication, in vitro rooting and acclimatization to ex vitro conditions. However, in some cases, the existence of recalcitrant genotypes, with a compromised multiplication and rooting ability, or the difficulties to overcome the overgrowth of endophytic contaminations might seriously limit its efficiency. In this sense, the establishment of beneficial interactions between plants and plant growth-promoting microorganisms (PGPMs) under in vitro culture conditions might represent a valuable approach to efficiently solve those restrictions. During the last years, significant evidence reporting the use of beneficial microorganisms to improve the yield of in vitro multiplication or rooting as well as their acclimatization to greenhouse or soil conditions have been provided. Most of these positive effects are strongly linked to the ability of these microorganisms to provide in vitro plants with nutrients such as nitrogen or phosphorous, to produce plant growth regulators, to control the growth of pathogens or to mitigate stress conditions. The culture of A. thaliana under aseptic conditions has provided high-quality knowledge on the root development signaling pathways, involving hormones, triggered in the presence of PGPMs. Overall, the present article offers a brief overview of the use of microorganisms to improve in vitro plant performance during the in vitro micropropagation stages, as well as the main mechanisms of plant growth promotion associated with these microorganisms.

Non-volatile signals and redox mechanisms are required for the responses of Arabidopsis roots to Pseudomonas oryzihabitans.

Soil bacteria promote plant growth and protect against environmental stresses, but the mechanisms involved remain poorly characterized, particularly when there is no direct contact between the roots and bacteria. Here, we explored the effects of Pseudomonas oryzihabitans PGP01 on the root system architecture (RSA) in Arabidopsis thaliana seedlings. Significant increases in lateral root (LR) density were observed when seedlings were grown in the presence of P. oryzihabitans, as well as an increased abundance of transcripts associated with altered nutrient transport and phytohormone responses. However, no bacterial transcripts were detected on the root samples by RNAseq analysis, demonstrating that the bacteria do not colonize the roots. Separating the agar containing bacteria from the seedlings prevented the bacteria-induced changes in RSA. Bacteria-induced changes in RSA were absent from mutants defective in ethylene response factor (ERF109), glutathione synthesis (pad2-1, cad2-1, and rax1-1) and in strigolactone synthesis (max3-9 and max4-1) or signalling (max2-3). However, the P. oryzihabitans-induced changes in RSA were similar in the low ascorbate mutants (vtc2-1and vtc2-2) to the wild-type controls. Taken together, these results demonstrate the importance of non-volatile signals and redox mechanisms in the root architecture regulation that occurs following long-distance perception of P. oryzihabitans.

Global analysis of the apple fruit microbiome: are all apples the same?

We present the first worldwide study on the apple (Malus × domestica) fruit microbiome that examines questions regarding the composition and the assembly of microbial communities on and in apple fruit. Results revealed that the composition and structure of the fungal and bacterial communities associated with apple fruit vary and are highly dependent on geographical location. The study also confirmed that the spatial variation in the fungal and bacterial composition of different fruit tissues exists at a global level. Fungal diversity varied significantly in fruit harvested in different geographical locations and suggests a potential link between location and the type and rate of postharvest diseases that develop in each country. The global core microbiome of apple fruit was represented by several beneficial microbial taxa and accounted for a large fraction of the fruit microbial community. The study provides foundational information about the apple fruit microbiome that can be utilized for the development of novel approaches for the management of fruit quality and safety, as well as for reducing losses due to the establishment and proliferation of postharvest pathogens. It also lays the groundwork for studying the complex microbial interactions that occur on apple fruit surfaces.

Rhizosphere microorganisms enhance in vitro root and plantlet development of Pyrus and Prunus rootstocks.

MAIN CONCLUSION: The in vitro application of rhizosphere microorganisms led to a higher rooting percentage in Pyrus Py12 rootstocks and increased plant growth of Pyrus Py170 and Prunus RP-20. The rooting of fruit tree rootstocks is the most challenging step of the in vitro propagation process. The use of rhizosphere microorganisms to promote in vitro rooting and plant growth as an alternative to the addition of chemical hormones to culture media is proposed in the present study. Explants from two Pyrus (Py170 and Py12) rootstocks and the Prunus RP-20 rootstock were inoculated with Pseudomonas oryzihabitans PGP01, Cladosporium ramotenellum PGP02 and Phoma sp. PGP03 following two different methods to determine their effects on in vitro rooting and plantlet growth. The effects of the microorganisms on the growth of fully developed Py170 and RP-20 plantlets were also studied in vitro. All experiments were conducted using vermiculite to simulate a soil system in vitro. When applied to Py12 shoots, which is a hard-to-root plant material, both C. ramotenellum PGP02 and Phoma sp. PGP03 fungi were able to increase the rooting percentage from 56.25% to 100% following auxin indole-3-butyric acid (IBA) treatment. Thus, the presence of these microorganisms clearly improved root development, inducing a higher number of roots and causing shorter roots. Better overall growth and improved stem growth of treated plants was observed when auxin treatment was replaced by co-culture with microorganisms. A root growth-promoting effect was observed on RP-20 plantlets after inoculation with C. ramotenellum PGP02, while P. oryzihabitans PGP01 increased root numbers for both Py170 and RP-20 and increased root growth over stem growth for RP-20. It was also shown that the three microorganisms P. oryzihabitans PGP01, C. ramotenellum PGP02 and Phoma sp. PGP03 were able to naturally produce auxin, including indole-3-acetic acid (IAA), at different levels. Overall, our results demonstrate that the microorganisms P. oryzihabitans PGP01 and C. ramotenellum PGP02 had beneficial effects on in vitro rooting and plantlet growth and could be applied to in vitro tissue culture as a substitute for IBA.

Verifying the biocontrol activity of novel film-forming formulations of Candida sake CPA-1: resilience in relation to environmental factors, rainfall episodes, and control of Botrytis cinerea on different hosts.

BACKGROUND: The efficacy of Candida sake CPA-1 as a biocontrol agent against several diseases has been studied since it was isolated 20 years ago. However, it was only recently that two suitable and effective film-forming formulations based on potato starch and maltodextrins were developed using the fluidized-bed spray-drying system. The present work aimed to confirm the capability of both novel formulations by testing their resilience on grapes at different temperatures (0, 22, and 30 °C), relative humidities (40% and 85%), and simulated rainfall levels. Another objective was to examine the control of Botrytis cinerea in different hosts. RESULTS: The CPA-1 cells from both dried formulations survived better than the liquid formulation on grapes stored at 0 and 22 °C regardless of the relative humidity. After simulated rainfall, potato starch formulation achieved significantly higher populations than maltodextrin formulation, although the highest reduction was -1.6 log N N0 -1 . A positive effect of cell establishment prior to the simulated rainfall was shown, and recovered cells from the potato starch formulation were significantly higher after 72 h of cell establishment. Finally, both formulations reduced the incidence and severity of B. cinerea on pears, apples, and tomatoes. CONCLUSION: The potential of these novel film-forming formulations of C. sake CPA-1 was verified. The resilience of formulated C. sake was better than the commercialized liquid formulation, the adherence of the formulations to the grapes improved after an establishment period prior to rain exposure, and the control of B. cinerea was verified in a wider range of hosts. © 2019 Society of Chemical Industry.

Exploring sources of resistance to brown rot in an interspecific almond × peach population.

BACKGROUND: Monilinia spp. are responsible for brown rot, one of the most significant stone fruit diseases. Planting resistant cultivars seems a promising alternative, although most commercial cultivars are susceptible to brown rot. The aim of this study was to explore resistance to Monilinia fructicola over two seasons in a backcross one interspecific population between almond 'Texas' and peach 'Earlygold' (named T1E). RESULTS: 'Texas' almond was resistant to brown rot inoculation, whereas peach was highly susceptible. Phenotypic data from the T1E population indicated wide differences in response to M. fructicola. Additionally, several non-wounded individuals exhibited resistance to brown rot. Quantitative trait loci (QTLs) were identified in several linkage groups, but only two proximal QTLs in G4 were detected over both seasons and accounted for 11.3-16.2% of the phenotypic variation. CONCLUSION: Analysis of the progeny allowed the identification of resistant genotypes that could serve as a source of resistance in peach breeding programs. The finding of loci associated with brown rot resistance would shed light on implementing a strategy based on marker-assisted selection (MAS) for introgression of this trait into elite peach materials. New peach cultivars resistant to brown rot may contribute to the implementation of more sustainable crop protection strategies. © 2019 Society of Chemical Industry.

Unravelling the contribution of the Penicillium expansum PeSte12 transcription factor to virulence during apple fruit infection.

Blue mould disease caused by Penicillium expansum infection is one of the most important diseases of pome fruit accounting for important economic losses. In the present study, the PeSte12 transcription factor gene was identified, and deletant mutants were produced by gene replacement. Knockout mutants showed a significant decrease of virulence during apple fruit infection. Virulence was affected by the maturity stage of the fruit (immature, mature and over-mature), and disease severity was notably reduced when the apples were stored at 0 °C. The ΔPeSte12 mutants resulted defective in asexual reproduction, producing less conidia, but this characteristic did not correlate with differences in microscopic morphology. In addition, the ΔPeSte12 mutants produced higher quantity of hydrogen peroxide than the wild type strain. Gene expression analysis revealed that PeSte12 was induced over time during apple infection compared to axenic growth, particularly from 2 dpi, reinforcing its role in virulence. Analysis of transcriptional abundance of several genes in ΔPeSte12 mutants showed that in most of the evaluated genes, PeSte12 seemed to act as a negative regulator during axenic growth, as most of them exhibited an increasing expression pattern along the time period evaluated. The highest expression values corresponded to detoxification, ATPase activity, protein folding and basic metabolism. Gene expression analysis during apple infection showed that 3 out of 9 analysed genes were up regulated; thus, PeSte12 seemed to exert a positive control to particular type of aldolase. These results demonstrate the PeSte12 transcription factor could play an important role in P. expansum's virulence and asexual reproduction.

Fluidised-bed spray-drying formulations of Candida sake CPA-1 by adding biodegradable coatings to enhance their survival under stress conditions.

The biocontrol agent Candida sake CPA-1 has demonstrated to be effective against several diseases on fruit. However, for application of CPA-1 under field conditions, it was necessary to mix it with a food coating to improve survival under stress conditions, as well as adherence and distribution on fruit surfaces. The objective of this study was to obtain a more competitive formulation under field conditions to be applied independently of any product. To achieve this purpose, the drying process of CPA-1 by a fluidised-bed spray-drying system together with biodegradable coatings was optimised. This approach is novel for the drying system used and the formulation obtained which was able to form a film or coating on fruit surfaces. Several substances were tested as carriers and binders, and drying temperature was optimised. The addition of protective compounds was also tested to improve survival of CPA-1 during the dehydration process. Product shelf life, biocontrol efficacy on grapes against Botrytis cinerea, and the improvement of C. sake behaviour under stress conditions were tested. The optimal temperature of drying was 55 °C and two formulations that were able to develop a coating on fruit surfaces were obtained. One of the formulations was created by using a combination of native and pregelatinised potato starch; the other formulation was obtained using maltodextrin and by adding skimmed milk and sucrose as protectant compounds. The formulated products reduced the incidence and severity of B. cinerea, and CPA-1 survival rate was increased under stress conditions of temperature and humidity.

Biological Characterization of the Biocontrol Agent Bacillus amyloliquefaciens CPA-8: The Effect of Temperature, pH and Water Activity on Growth, Susceptibility to Antibiotics and Detection of Enterotoxic Genes.

This work focuses on the biological understanding of the biocontrol agent Bacillus amyloliquefaciens CPA-8 in order to accomplish the characterization required in the registration process for the development of a microorganism-based product. The tolerance of CPA-8 to grow under different pH-temperature and water activity (a w)-temperature conditions was widely demonstrated. Regarding the pH results, optimum growth at the evaluated conditions was observed at 37 °C and pH between 7 and 5. On the contrary, the slowest growth was recorded at 20 °C and pH 4.5. Moreover, the type of solute used to reduce a w had a great influence on the minimum a w at which the bacterium was able to grow. The lowest a w values for CPA-8 growth in media modified with glycerol and glucose were 0.950 and 0.960, respectively. Besides, the lowest a w for CPA-8 growth increased when the temperature decreased to 20 °C, at which CPA-8 was not able to grow at less than 0.990 a w, regardless of the type of solute. Antibiotic susceptibility tests were carried out to determine which antibiotic could affect the behavior of the bacteria and revealed that CPA-8 was clearly resistant to hygromycin. Finally, a PCR amplification assay to detect the presence of enterotoxic genes from Bacillus cereus in CPA-8 was also performed. CPA-8 gave negative results for all the genes tested except for nheA gene, which is not enough for the toxicity expression, suggesting that fruit treated with this antagonist will not be a potential vehicle for foodborne illnesses.

Dry formulations of the biocontrol agent Candida sake CPA-1 using fluidised bed drying to control the main postharvest diseases on fruits.

BACKGROUND: The biocontrol agent Candida sake CPA-1 is effective against several diseases. Consequently, the optimisation of a dry formulation of C. sake to improve its shelf life and manipulability is essential for increasing its potential with respect to future commercial applications. The present study aimed to optimise the conditions for making a dry formulation of C. sake using a fluidised bed drying system and then to determine the shelf life of the optimised formulation and its efficacy against Penicillium expansum on apples. RESULTS: The optimal conditions for the drying process were found to be 40 °C for 45 min and the use of potato starch as the carrier significantly enhanced the viability. However, none of the protective compounds tested increased the viability of the dried cells. A temperature of 25 °C for 10 min in phosphate buffer was considered as the optimum condition to recover the dried formulations. The dried formulations should be stored at 4 °C and air-packaged; moreover, shelf life assays indicated good results after 12 months of storage. The formulated products maintained their biocontrol efficacy. CONCLUSION: A fluidised bed drying system is a suitable process for dehydrating C. sake cells; moreover, the C. sake formulation is easy to pack, store and transport, and is a cost-effective process. © 2017 Society of Chemical Industry.

Relevance of the main postharvest handling operations on the development of brown rot disease on stone fruits.

BACKGROUND: Brown rot caused by Monilinia spp. is one of the most important postharvest diseases of stone fruit. The aim of this study was to evaluate the relevance of the main postharvest operations of fruit - hydrocooling, cold room, water dump, sorting and cooling tunnel - in the development of M. laxa on peaches and nectarines artificially infected 48, 24 or 2 h before postharvest operations. RESULTS: Commercial hydrocooling operation reduced incidence to 10% in 'Pp 100' nectarine inoculated 2 and 24 h before this operation; however, in 'Fantasia' nectarine incidence was not reduced, although lesion diameter was decreased in all studied varieties. Hydrocooling operation for 10 min and 40 mg L-1 of sodium hypochlorite reduced brown rot incidence by 50-77% in nectarines inoculated 2 h before operation; however, in peach varieties it was not reduced. Water dump operation showed reduction of incidence on nectarine infected 2 h before immersion for 30 s in clean water at 4 °C and 40 mg L-1 sodium hypochlorite; however, in peach varieties it was not reduced. Cold room, sorting and cooling tunnel operation did not reduce brown rot incidence. CONCLUSION: From all studied handling operations on stone fruit packing houses, hydrocooling is the most relevant in the development of brown rot disease. Duration of the treatment seems to be more important than chlorine concentration. In addition, hydrocooling and water dump were less relevant in peaches than in nectarines. As a general trend, hydrocooling and water dump reduced incidence on fruit with recent infections (2 or 24 h before operation); however, when infections have been established (48 h before operation) diseases were not reduced. © 2017 Society of Chemical Industry.

Pseudomonas synxantha volatile organic compounds: efficacy against Cadophora luteo-olivacea and Botrytis cinerea of kiwifruit.

Volatile organic compounds (VOCs) are responsible for the antagonistic activity exerted by different biological control agents (BCAs). In this study, VOCs produced by Pseudomonas synxantha strain 117-2b were tested against two kiwifruit fungal postharvest pathogens: Cadophora luteo-olivacea and Botrytis cinerea, through in vitro and in vivo assays. In vitro results demonstrated that P. synxantha 117-2b VOCs inhibit mycelial growth of C. luteo-olivacea and B. cinerea by 56% and 42.8% after 14 and 5 days of exposition, respectively. In vivo assay demonstrated significant inhibitory effects. VOCs used as a biofumigant treatment reduced skin-pitting symptoms disease severity by 28.5% and gray mold incidence by 66.6%, with respect to the untreated control. BCA volatiles were analyzed by solid-phase microextraction coupled with gas chromatography-mass spectrometry (SPME-GC/MS), and among the detected compounds, 1-butanol, 3-methyl and 1-nonene resulted as the most produced. Their efficacy as pure synthetic compounds was assayed against mycelial growth of fungal pathogens by different concentrations (0.34, 0.56, and 1.12 µL mL-1 headspace). The effect of the application of VOCs as a biofumigant was also investigated as the expression level of seven defense-related genes of kiwifruit at different exposition times. The results indicated an enhancement of the expression of almost all the genes starting from 3 h of treatment. These results described P. synxantha VOCs characteristics and their potential as a promising method to adopt for protecting kiwifruit from postharvest diseases caused by C. luteo-olivacea and B. cinerea.

Control of foodborne pathogens on fresh-cut fruit by a novel strain of Pseudomonas graminis.

The consumption of fresh-cut fruit has substantially risen over the last few years, leading to an increase in the number of outbreaks associated with fruit. Moreover, consumers are currently demanding wholesome, fresh-like, safe foods without added chemicals. As a response, the aim of this study was to determine if the naturally occurring microorganisms on fruit are "competitive with" or "antagonistic to" potentially encountered pathogens. Of the 97 and 107 isolates tested by co-inoculation with Escherichia coli O157:H7, Salmonella and Listeria innocua on fresh-cut apple and peach, respectively, and stored at 20 °C, seven showed a strong antagonistic capacity (more than 1-log unit reduction). One of the isolates, CPA-7, achieved the best reduction values (from 2.8 to 5.9-log units) and was the only isolate able to inhibit E. coli O157:H7 at refrigeration temperatures on both fruits. Therefore, CPA-7 was selected for further assays. Dose-response assays showed that CPA-7 should be present in at least the same amount as the pathogen to adequately reduce the numbers of the pathogen. From the results obtained in in vitro assays, competition seemed to be CPA-7's mode of action against E. coli O157:H7. The CPA-7 strain was identified as Pseudomonas graminis. Thus, the results support the potential use of CPA-7 as a bioprotective agent against foodborne pathogens in minimally processed fruit.

Deciphering the Effect of Light Wavelengths in Monilinia spp. DHN-Melanin Production and Their Interplay with ROS Metabolism in M. fructicola.

Pathogenic fungi are influenced by many biotic and abiotic factors. Among them, light is a source of information for fungi and also a stress factor that triggers multiple biological responses, including the activation of secondary metabolites, such as the production of melanin pigments. In this study, we analyzed the melanin-like production in in vitro conditions, as well as the expression of all biosynthetic and regulatory genes of the DHN-melanin pathway in the three main Monilinia species upon exposure to light conditions (white, black, blue, red, and far-red wavelengths). On the other hand, we analyzed, for the first time, the metabolism related to ROS in M. fructicola, through the production of hydrogen peroxide (H2O2) and the expression of stress-related genes under different light conditions. In general, the results indicated a clear importance of black light on melanin production and expression in M. laxa and M. fructicola, but not in M. fructigena. Regarding ROS-related metabolism in M. fructicola, blue light highlighted by inhibiting the expression of many antioxidant genes. Overall, it represents a global description of the effect of light on the regulation of two important secondary mechanisms, essential for the adaptation of the fungus to the environment and its survival.

Identification and Biosynthesis of DHN-melanin Related Pigments in the Pathogenic Fungi Monilinia laxa, M. fructicola, and M. fructigena.

Monilinia is the causal agent of brown rot in stone fruit. The three main species that cause this disease are Monilinia laxa, M. fructicola, and M. fructigena, and their infection capacity is influenced by environmental factors (i.e., light, temperature, and humidity). To tolerate stressful environmental conditions, fungi can produce secondary metabolites. Particularly, melanin-like pigments can contribute to survival in unfavorable conditions. In many fungi, this pigment is due to the accumulation of 1,8-dihydroxynaphthalene melanin (DHN). In this study, we have identified for the first time the genes involved in the DHN pathway in the three main Monilinia spp. and we have proved their capacity to synthetize melanin-like pigments, both in synthetic medium and in nectarines at three stages of brown rot development. The expression of all the biosynthetic and regulatory genes of the DHN-melanin pathway has also been determined under both in vitro and in vivo conditions. Finally, we have analyzed the role of three genes involved in fungi survival and detoxification, and we have proved that there exists a close relationship between the synthesis of these pigments and the activation of the SSP1 gene. Overall, these results deeply describe the importance of DHN-melanin in the three main species of Monilinia: M. laxa, M. fructicola, and M. fructigena.

Transcriptional profiling of the terpenoid biosynthesis pathway and in vitro tests reveal putative roles of linalool and farnesal in nectarine resistance against brown rot.

The most devastating fungal disease of peaches and nectarines is brown rot, caused by Monilinia spp. Among the many plant responses against biotic stress, plant terpenoids play essential protective functions, including antioxidant activities and inhibition of pathogen growth. Herein, we aimed to characterize the expression of terpenoid biosynthetic genes in fruit tissues that presented different susceptibility to brown rot. For that, we performed artificial inoculations with Monilinia laxa at two developmental stages (immature and mature fruit) of two nectarine cultivars ('Venus' -mid-early season cultivar - and 'Albared' -late season cultivar-) and in vitro tests of the key compounds observed in the transcriptional results. All fruit were susceptible to M. laxa except for immature 'Venus' nectarines. In response to the pathogen, the mevalonic acid (MVA) pathway of the 'Venus' cultivar was highly induced in both stages rather than the methylerythritol phosphate (MEP) pathway, being the expression of some MEP-related biosynthetic genes [e.g., PROTEIN FARNESYLTRANSFERASE (PpPFT), and 3S-LINALOOL SYNTHASE (PpLIS)] different between stages. In 'Albared', both stages presented similar responses to M. laxa for both pathways. Comparisons between cultivars showed that HYDROXYMETHYLGLUTARYL-CoA REDUCTASE (PpHMGR1) expression levels were common in susceptible tissues. Within all the terpenoid biosynthetic pathway, linalool- and farnesal-related pathways stood out for being upregulated only in resistant tissues, which suggest their role in mediating the resistance to M. laxa. The in vitro antifungal activity of linalool and farnesol (precursor of farnesal) revealed fungicidal and fungistatic activities against M. laxa, respectively, depending on the concentration tested. Understanding the different responses between resistant and susceptible tissues could be further considered for breeding or developing new strategies to control brown rot in stone fruit.

Phenotypic plasticity of Monilinia spp. in response to light wavelengths: From in vitro development to virulence on nectarines.

The development of brown rot in stone fruit caused by the necrotrophic fungus Monilinia spp. is influenced by many abiotic factors, such as temperature, humidity, and light. Specifically, filamentous fungi perceive light as a signal for ecophysiological and adaptive responses. We have explored how specific light wavelengths affect the in vitro development, the regulation of putative development genes and the virulence of the main species of Monilinia (M. laxa, M. fructicola and M. fructigena). After subjecting Monilinia spp. to different light wavelengths (white, black, blue, red, far-red) for 7 days, several differences in their phenotype were observed among light conditions, but also among species. These species of Monilinia exhibited a different phenotypic plasticity in response to light regarding pigmentation, growth, and specially conidiation of colonies. In this sense, we observed that the conidial production was higher in M. laxa than M. fructicola, while M. fructigena showed an inability to produce conidia under the tested conditions. Growth rate among species was significantly lower in M. fructicola under red light wavelength while among light conditions it was increased under far-red light wavelength for M. laxa and under black light for M. fructicola; in contrast, no statistical differences were observed for M. fructigena. Gene expression analysis of 13 genes involved in fungal development of Monilinia spp. revealed a significant difference among the three species of Monilinia, and especially depended on light wavelengths. Among them, a high expression of OPT1, RGS2, RGS3 and SPP1 genes was observed in M. laxa, and LTF1 and STE12 in M. fructicola under black light. In contrast, a high expression of REG1 and C6TF1 genes occurred in both M. fructicola and M. laxa subject to red and far-red light wavelength, respectively. When nectarines were artificially infected with M. laxa and M. fructicola subjected to black light, the virulence was clearly reduced, but not in M. fructigena. Overall, results presented herein demonstrate that light wavelengths are a key abiotic factor for the biology of Monilinia spp., specially modulating its capacity to form conidia, and thus, influencing its spreading and the onset of the disease on nectarines during postharvest.

ROS-scavenging-associated transcriptional and biochemical shifts during nectarine fruit development and ripening.

ROS are known as toxic by-products but also as important signaling molecules playing a key role in fruit development and ripening. To counteract the negative effects of ROS, plants and fruit own multiple ROS-scavenging mechanisms aiming to ensure a balanced ROS homeostasis. In the present study, changes in specific ROS (i.e. H2O2) as well as enzymatic (SOD, CAT, POX, APX) and non-enzymatic (phenylpropanoids, carotenoids and ascorbate) ROS-scavenging systems were investigated along four different stages of nectarine (cv. 'Diamond Ray') fruit development and ripening (39, 70, 94 and 121 DAFB) both at the metabolic (28 individual metabolites or enzymes) and transcriptional level (24 genes). Overall, our results demonstrate a complex ROS-related transcriptome and metabolome reprogramming during fruit development and ripening. At earlier fruit developmental stages an increase on the respiration rate is likely triggering an oxidative burst and resulting in the activation of specific ethylene response factors (ERF1). In turn, ROS-responsive genes or the biosynthesis of specific antioxidant compounds (i.e. phenylpropanoids) were highly expressed or accumulated at earlier fruit developmental stages (39-70 DAFB). Nonetheless, as the fruit develops, the decrease in the fruit respiration rate and the reduction of ERF1 genes leads to lower levels of most non-enzymatic antioxidants and higher accumulation of H2O2. Based on available literature and the observed accumulation dynamics of H2O2, it is anticipated that this compound may not only be a by-product of ROS-scavenging but also a signaling molecule accumulated during the ripening of nectarine fruit.

Environmentally driven transcriptomic and metabolic changes leading to color differences in "Golden Reinders" apples.

Apple is characterized by its high adaptation to diverse growing environments. However, little is still known about how different environments can regulate at the metabolic or molecular level specific apple quality traits such as the yellow fruit peel color. In this study, changes in carotenoids and chlorophylls, antioxidants as well as differences in the transcriptome were investigated by comparing the peel of "Golden Reinders" apples grown at different valley and mountain orchards. Mountain environment favored the development of yellow color, which was not caused by an enhanced accumulation of carotenoids but rather by a decrease in the chlorophyll content. The yellow phenotype was also associated to higher expression of genes related to chloroplast functions and oxidative stress. Time-course analysis over the last stages of apple development and ripening, in fruit from both locations, further revealed that the environment differentially modulated isoprenoids and phenylpropanoid metabolism and pointed out a key role for H2O2 in triggering apple peel degreening. Overall, the results presented herein provide new insights into how different environmental conditions regulate pigment and antioxidant metabolism in apple leading to noticeable differences in the apple peel color.

Light Intensity Alters the Behavior of Monilinia spp. in vitro and the Disease Development on Stone Fruit-Pathogen Interaction.

The development of brown rot caused by the necrotrophic fungi Monilinia spp. in stone fruit under field and postharvest conditions depends, among others, on environmental factors. The effect of temperature and humidity are well studied but there is little information on the role of light in disease development. Herein, we studied the effect of two lighting treatments and a control condition (darkness) on: (i) several growth parameters of two Monilinia spp. (M. laxa and M. fructicola) grown in vitro and (ii) the light effect in their capacity to rot the fruit (nectarines) when exposed to the different lighting treatments. We also assessed the effect of such abiotic factors in the development of the disease on inoculated nectarines during postharvest storage. Evaluations also included testing the effect of fruit bagging on disease development as well as on ethylene production. Under in vitro conditions, lighting treatments altered colony morphology and conidiation of M. laxa but this effect was less acute in M. fructicola. Such light-induced changes under in vitro development also altered the capacity of M. laxa and M. fructicola to infect nectarines, with M. laxa becoming less virulent. The performance of Monilinia spp. exposed to treatments was also determined in vivo by inoculating four bagged or unbagged nectarine cultivars, indicating an impaired disease progression. Incidence and lesion diameter of fruit exposed to the different lighting treatments during postharvest showed that the effect of the light was intrinsic to the nectarine cultivar but also Monilinia spp. dependent. While lighting treatments reduced M. laxa incidence, they enhanced M. fructicola development. Preharvest conditions such as fruit bagging also impaired the ethylene production of inoculated fruit, which was mainly altered by M. laxa and M. fructicola, while the bag and light effects were meaningless. Thus, we provide several indications of how lighting treatments significantly alter Monilinia spp. behavior both in vitro and during the interaction with stone fruit. This study highlights the importance of modulating the lighting environment as a potential strategy to minimize brown rot development on stone fruit and to extent the shelf-life period of fruit in postharvest, market, and consumer's house.