Transformation of Botrytis cinerea by direct hyphal blasting or by wound-mediated transformation of sclerotia

<p>Abstract</p> <p>Background</p> <p><it>Botrytis cinerea </it>is a haploid necrotrophic ascomycete which is responsible for 'grey mold' disease in more than 200 plant species. Broad molecular research has been conducted on this pathogen in recent years, resulting in the sequencing of two strains, which has generated a wealth of information toward developing additional tools for molecular transcriptome, proteome and secretome investigations. Nonetheless, transformation protocols have remained a significant bottleneck for this pathogen, hindering functional analysis research in many labs.</p> <p>Results</p> <p>In this study, we tested three different transformation methods for <it>B. cinerea</it>: electroporation, air-pressure-mediated and sclerotium-mediated transformation. We demonstrate successful transformation with three different DNA constructs using both air-pressure- and sclerotium-mediated transformation.</p> <p>Conclusions</p> <p>These transformation methods, which are fast, simple and reproducible, can expedite functional gene analysis of <it>B. cinerea</it>.</p

Postharvest losses of fruit and vegetables during retail and in consumers’ homes: Quantifications, causes, and means of prevention

The issue of food loss and waste (FLW) reduction has recently achieved much public attention as part of worldwide efforts to combat global hunger and improve food security. Studies conducted by various international and national organizations led by the FAO indicated that about one third of all food produced on the planet and about a half of all fruit and vegetables (F&V) are lost and not consumed. FLW occurs during five key stages of the food supply chain: agricultural production, postharvest handling and storage, processing, distribution, and consumption. Large portions of FLW in developed countries occur during retail and consumption, and are largely related to logistic management operations and consumer behaviors. In light of the great importance of FLW reduction, the United Nations set up in September 2015 an ambitious goal to halve per capita global food waste by 2030, and this decision was adapted by the US Federal Government, the EU Parliament, and many other countries. This first Adel Kader review article is dedicated to the subject of F&V losses during retail and consumption, and contains the following chapters: 1) Introduction of the problem of global food losses; 2) Quantifications of F&V losses during retail and consumption in the UK, US and other countries; 3) Causes and consumer decisions related to F&V wastage; 4) Emerging new technologies for prevention of F&V losses, including advances in logistics and cold chain management, retail packaging and technological innovations; 5) Other means to reduce F&V losses, including consumer awareness campaigns, advertisement of home storage instructions and policy and legislative measures. Due to the great importance of reducing F&V losses, we encourage postharvest researchers to become more engaged with logistics and food supply-chain operations, and to conduct multidisciplinary research incorporating consumer behavior studies into postharvest research

Postharvest ethanol and potassium sorbate treatments of table grapes to control gray mold

Germination of Botrytis cinerea spores on potato dextrose agar after a 30 s immersion in 10 or 20% ethanol was 87 and 56%, respectively, compared to 99% among untreated controls. After similar immersion in 0.5 or 1.0% potassium sorbate, 84 and 68% of the spores germinated, respectively. Addition of 0.5 and 1.0% potassium sorbate to 10 and 20% ethanol solution significantly increased the inhibition of spore germination. The germination of spores after 30 s immersion in 20% ethanol plus 0.5% potassium sorbate was 9.7%. The incidence of gray mold, caused by B. cinerea, on detached berries of 'Flame Seedless' grapes immersed for 30 s in water, 10 and 20% ethanol, and 0.5 or 1.0% potassium sorbate was 55.2, 42.1, 31.0, 37.7, or 24.4%, respectively. Addition of 0.5 and 1.0% potassium sorbate to 10 and 20% ethanol reduced decay to 10% or less and was more effective than either alone. After 30 days of storage at 1 degrees C, the combination of 20% ethanol either with 0.5 or 1.0% potassium sorbate was equal in efficacy to commercial SO2 generator pads in reducing the incidence of gray mold on 'Thompson Seedless' grapes. None of the combinations of ethanol and potassium sorbate injured the berries

Disinfecting agents for controlling fruit and vegetable diseases after harvest

Disinfection of fresh fruit and vegetables after harvest is an essential first step of postharvest handling. The minimal requirement from disinfection procedures is to maintain commodities and facilities free of fungal postharvest pathogens and bacterial human pathogens and thus improve food safety. Disinfection of postharvest pathogens that accumulate on the fruit surface before and during harvest is a direct benefit and in particular cases it can by itself prevent decay after storage. The current review includes historical, chemical, and regulatory background on some of the major disinfectants available for usage today. These include chlorine, chlorine dioxide, ozone, ethanol, hydrogen peroxide, organic acids, and electrolyzed water. Some of the disinfectants described in this review are in wide usage for many years and some are considered ‘alternative’ and are at initial levels of usage. Information is given on experimental reports, practical application, phytotoxicity, residues, advantages, disadvantages and mode of action of the compounds and technologies. Special emphasis is given to vapor and gas phase applications due to their unexploited potential and to some complementary technologies that have been reported in recent years. The conclusion from the many details in this review is that disinfection is an important tool to ensure management of postharvest decay of fresh produce. In some cases, disinfection is a precondition to successful implementation of major postharvest technologies and in particular cases it can become the major technology. An important aspect arising from this review is also that some of the bad reputation of chemical disinfectants is unjustified because they leave no or non-toxic levels of residues and their environmental impact is minor in view of their potential benefits

Cytokinin but not gibberellin application had major impact on the phenylpropanoid pathway in grape.

Cytokinin and gibberellic acid (GA) are growth regulators used to increase berry size in seedless grapes and it is of interest to understand their effects on the phenylpropanoid pathway and on ripening processes. GA3 and synthetic cytokinin forchlorfenuron (N-(2-chloro-4-pyridyl)-N'-phenylurea, CPPU) and their combination were applied to 6 mm diameter fruitlets of 'Sable Seedless', and berries were sampled 51 and 70 days (d) following application. All treatments increased berry size and delayed sugar accumulation and acid degradation with a stronger effect of CPPU. CPPU, but not GA, reduced berry color and the levels of anthocyanins. While CPPU reduced the levels of anthocyanins by more than 50%, the combined treatment of GA+CPPU reduced the levels by about 25% at 51 d. CPPU treatment had minor effects on flavonols content but increased the levels of monomeric flavan-3-ols by more than two-fold. Phloroglucinol analysis using HPLC showed that proanthocyanidin content was significantly increased by CPPU, whereas mean degree of polymerization was reduced from 26 to 19. Volatile analysis by GC-MS showed changes in composition with CPPU or GA treatment with potential impact on flavor. RNA-seq analysis showed that GA had a minor overall effect on the transcriptome whereas CPPU had pronounced effects on gene expression at both 51 and 70 d. Comparing the control and CPPU at similar Brix of ca. 19.7°, a reduced expression of stilbene synthases (STSs) including their regulators MYB14 and MYB15, and other phenylpropanoid-related genes was observed in CPPU-treated grapes. Overall, our study shows that CPPU had a major influence on the phenylpropanoid pathway and affected multiple ripening-related processes

Large-Scale, High-Throughput Phenotyping of the Postharvest Storage Performance of &lsquo;Rustenburg&rsquo; Navel Oranges and the Development of Shelf-Life Prediction Models

We conducted a large-scale, high-throughput phenotyping analysis of the effects of various pre-harvest and postharvest features on the quality of &lsquo;Rustenburg&rsquo; navel oranges, in order to develop shelf-life prediction models to enable the use of the First Expired, First Out logistics strategy. The examined pre-harvest features included harvest time and yield, and the examined postharvest features included storage temperature, relative humidity during storage and duration of storage. All together, we evaluated 12,000 oranges (~4 tons) from six different orchards and conducted 170,576 measurements of 14 quality parameters. Storage time was found to be the most important feature affecting fruit quality, followed by storage temperature, harvest time, yield and humidity. The examined features significantly affected (p &lt; 0.001) fruit weight loss, firmness, decay, color, peel damage, chilling injury, internal dryness, acidity, vitamin C and ethanol levels, and flavor and acceptance scores. Four regression models were evaluated for their ability to predict fruit quality based on pre-harvest and postharvest features. Extreme gradient boosting (XGBoost) combined with a duplication approach was found to be the most effective approach. It allowed for the prediction of fruit-acceptance scores among the full data set, with a root mean square error (RMSE) of 0.217 and an R2 of 0.891

Cytokinin but not gibberellin application had major impact on the phenylpropanoid pathway in grape.

Cytokinin and gibberellic acid (GA) are growth regulators used to increase berry size in seedless grapes and it is of interest to understand their effects on the phenylpropanoid pathway and on ripening processes. GA3 and synthetic cytokinin forchlorfenuron (N-(2-chloro-4-pyridyl)-N'-phenylurea, CPPU) and their combination were applied to 6 mm diameter fruitlets of 'Sable Seedless', and berries were sampled 51 and 70 days (d) following application. All treatments increased berry size and delayed sugar accumulation and acid degradation with a stronger effect of CPPU. CPPU, but not GA, reduced berry color and the levels of anthocyanins. While CPPU reduced the levels of anthocyanins by more than 50%, the combined treatment of GA+CPPU reduced the levels by about 25% at 51 d. CPPU treatment had minor effects on flavonols content but increased the levels of monomeric flavan-3-ols by more than two-fold. Phloroglucinol analysis using HPLC showed that proanthocyanidin content was significantly increased by CPPU, whereas mean degree of polymerization was reduced from 26 to 19. Volatile analysis by GC-MS showed changes in composition with CPPU or GA treatment with potential impact on flavor. RNA-seq analysis showed that GA had a minor overall effect on the transcriptome whereas CPPU had pronounced effects on gene expression at both 51 and 70 d. Comparing the control and CPPU at similar Brix of ca. 19.7°, a reduced expression of stilbene synthases (STSs) including their regulators MYB14 and MYB15, and other phenylpropanoid-related genes was observed in CPPU-treated grapes. Overall, our study shows that CPPU had a major influence on the phenylpropanoid pathway and affected multiple ripening-related processes

The Role of Aquaporins in pH-Dependent Germination of Rhizopus delemar Spores.

Rhizopus delemar and associated species attack a wide range of fruit and vegetables after harvest. Host nutrients and acidic pH are required for optimal germination of R. delemar, and we studied how this process is triggered. Glucose induced spore swelling in an acidic environment, expressed by an up to 3-fold increase in spore diameter, whereas spore diameter was smaller in a neutral environment. When suspended in an acidic environment, the spores started to float, indicating a change in their density. Treatment of the spores with HgCl2, an aquaporin blocker, prevented floating and inhibited spore swelling and germ-tube emergence, indicating the importance of water uptake at the early stages of germination. Two putative candidate aquaporin-encoding genes-RdAQP1 and RdAQP2-were identified in the R. delemar genome. Both presented the conserved NPA motif and six-transmembrane domain topology. Expressing RdAQP1 and RdAQP2 in Arabidopsis protoplasts increased the cells' osmotic water permeability coefficient (Pf) compared to controls, indicating their role as water channels. A decrease in R. delemar aquaporin activity with increasing external pH suggested pH regulation of these proteins. Substitution of two histidine (His) residues, positioned on two loops facing the outer side of the cell, with alanine eliminated the pH sensing resulting in similar Pf values under acidic and basic conditions. Since hydration is critical for spore switching from the resting to activate state, we suggest that pH regulation of the aquaporins can regulate the initial phase of R. delemar spore germination, followed by germ-tube elongation and host-tissue infection