Volatile Organic Compounds of Wickerhamomyces anomalus Prevent Postharvest Black Spot Disease in Tomato.

Postharvest diseases, such as black spots caused by Alternaria alternata, have caused huge economic losses to the tomato industry and seriously restricted its development. In recent years, biological control has become a new method to control postharvest diseases of fruits and vegetables. Our research group screened W. anomalus, a yeast demonstrating a promising control effect on a postharvest black spot disease of tomatoes, and explored its physiological mechanism of prevention and control. Therefore, this study investigated the prevention and control effect of metabolites of W. anomalus on tomato black spot disease and the inhibition effect of main components on A. alternata. A GC-MS analysis found that isoamyl acetate was the main component of W. anomalus that played an inhibitory role. The results showed that isoamyl acetate could inhibit the growth of A. alternata and had a certain control effect on postharvest black spots in tomatoes. Our findings suggest that isoamyl acetate could be a promising alternative to fungicides for controlling postharvest black spots in tomatoes.

Advances in the Multifaceted Functions of Cys2/His2-Type Zinc Finger Proteins in Distinctive Plant Characters.

Recent research findings established the cruciality of Cys2/His2-type Zinc Finger Proteins (C2H2-ZFPs) in plant growth and their relevance in coping with various stressors. Nevertheless, the complex structure of the C2H2-ZFPs network and the molecular mechanisms of response to stress in adversity have received considerable attention and now require more in-depth examination. This paper reviews the structural characteristics, classification, and recent functional research advances of C2H2-ZFPs. In addition, it systematically introduces the roles of these proteins across diverse facets of plant biology, encompassing growth and development, responses to biotic and abiotic stresses, and laying the foundation for future functional studies of C2H2-ZFPs.

Deciphering the mechanism of E3 ubiquitin ligases in plant responses to abiotic and biotic stresses and perspectives on PROTACs for crop resistance.

With global climate change, it is essential to find strategies to make crops more resistant to different stresses and guarantee food security worldwide. E3 ubiquitin ligases are critical regulatory elements that are gaining importance due to their role in selecting proteins for degradation in the ubiquitin-proteasome proteolysis pathway. The role of E3 Ub ligases has been demonstrated in numerous cellular processes in plants responding to biotic and abiotic stresses. E3 Ub ligases are considered a class of proteins that are difficult to control by conventional inhibitors, as they lack a standard active site with pocket, and their biological activity is mainly due to protein-protein interactions with transient conformational changes. Proteolysis-targeted chimeras (PROTACs) are a new class of heterobifunctional molecules that have emerged in recent years as relevant alternatives for incurable human diseases like cancer because they can target recalcitrant proteins for destruction. PROTACs interact with the ubiquitin-proteasome system, principally the E3 Ub ligase in the cell, and facilitate proteasome turnover of the proteins of interest. PROTAC strategies harness the essential functions of E3 Ub ligases for proteasomal degradation of proteins involved in dysfunction. This review examines critical advances in E3 Ub ligase research in plant responses to biotic and abiotic stresses. It highlights how PROTACs can be applied to target proteins involved in plant stress response to mitigate pathogenic agents and environmental adversities.

Revisiting the current and emerging concepts of postharvest fresh fruit and vegetable pathology for next-generation antifungal technologies.

Fungal infections of fresh fruits and vegetables (FFVs) can lead to safety problems, including consumer poisoning by mycotoxins. Various strategies exist to control fungal infections of FFVs, but their effectiveness and sustainability are limited. Recently, new concepts based on the microbiome and pathobiome have emerged and offer a more holistic perspective for advancing postharvest pathogen control techniques. Understanding the role of the microbiome in FFV infections is essential for developing sustainable control strategies. This review examines current and emerging approaches to postharvest pathology. It reviews what is known about the initiation and development of infections in FFVs. As a promising concept, the pathobiome offers new insights into the basic mechanisms of microbial infections in FFVs. The underlying mechanisms uncovered by the pathobiome are being used to develop more relevant global antifungal strategies. This review will also focus on new technologies developed to target the microbiome and members of the pathobiome to control infections in FFVs and improve safety by limiting mycotoxin contamination. Specifically, this review stresses emerging technologies related to FFVs that are relevant for modifying the interaction between FFVs and the microbiome and include the use of microbial consortia, the use of genomic technology to manipulate host and microbial community genes, and the use of databases, deep learning, and artificial intelligence to identify pathobiome markers. Other approaches include programming the behavior of FFVs using synthetic biology, modifying the microbiome using sRNA technology, phages, quorum sensing, and quorum quenching strategies. Rapid adoption and commercialization of these technologies are recommended to further improve the overall safety of FFVs.

Yarrowia lipolytica increased the activities of disease defense related enzymes and anti-fungal compounds in asparagus (Asparagus officinalis).

Fungal diseases pose significant threats to the production of asparagus, resulting in economic losses and decreased crop quality. The potential of the yeast Yarrowia lipolytica as a biocontrol agent against Fusarium proliferatum, a common pathogen of asparagus, was investigated in this study. The effects of Y. lipolytica treatment on decay incidence, disease index, and activities of major disease defense-related enzymes were investigated. In addition, we examined the levels of antifungal compounds such as total phenols, flavonoids, and lignin in asparagus plants exposed to Y. lipolytica. The results showed that Y. lipolytica treatment significantly reduced decay incidence and disease index caused by F. proliferatum when compared to the control group. Furthermore, Y. lipolytica-treated plants showed increased activity of disease defense-related enzymes, indicating that defense responses were activated. The activities of all evaluated enzymes were significantly higher in Y. lipolytica-treated asparagus, indicating an improved ability to combat fungal pathogens. Furthermore, Y. lipolytica treatment increased the content of antifungal compounds such as total phenols, flavonoids, and lignin, which are known to possess antimicrobial properties. These findings highlight the potential of Y. lipolytica as a biocontrol agent for fungal diseases in asparagus crops. The ability of Y. lipolytica to reduce disease incidence, boost disease defense-related enzymes, and increase antifungal compound content provides valuable insights into its efficacy as a natural and sustainable approach to disease management. However, further investigations are needed to optimize application methods and determine its efficacy under field conditions.

A transcriptome profile of Wickerhamomyces anomalus incubated with chitosan revealed dynamic changes in gene expression and metabolic pathways.

Previous studies have shown that Wickerhamomyces anomalus can control postharvest diseases of fruits and incubation of the yeast with chitosan can improve its efficiency. In this study, transcriptome study was conducted to determine molecular mechanisms involved in the yeast-chitosan interaction. The bioinformatics analysis of the RNA-seq data confirmed that incubating W. anomalus with 1 % chitosan for 24 h significantly altered the expression of differential genes involved in yeast metabolic and cellular activities. Genes involved in ethyl acetate production, reactive oxygen species regulation, cell wall reinforcement, stress resistance, and signalling were all significantly up-regulated. Pathways which have significant role in the yeast growth and reproduction, energy production, cellular homeostasis, signal transduction, catalytic, and antioxidant activities were significantly enriched. In general, incubation of the yeast with chitosan genes metabolic pathways which are important for the yeast survival, adaptation, and reproduction. Molecular studies are important in providing fundamental theoretical foundation for the practical application of antagonistic yeasts for future uses. As a result, this research will be an input for use of the antagonistic yeast as microbial or biochemical pesticides instead of synthetic chemicals which have both health and environmental effects.

Genome wide and comprehensive analysis of the cytochrome P450 (CYPs) gene family in Pyrus bretschneideri: Expression patterns during Sporidiobolus pararoseus Y16 enhanced with ascorbic acid (VC) treatment.

Cytochrome P450s (CYPs) constitute the largest group of enzymes in plants and are involved in a variety of processes related to growth and protection. However, the CYP gene superfamily in pear (Pyrus bretschneideri) and their characteristics is unclear. Through a comprehensive genome-wide analysis, this article identified a total of 74 CYP genes in the P. bretschneideri genome, which were categorized into fourteen families. Motif analysis reveals that most of the ten motifs predicted were with the p450 conserved domain. The majority of the CYP genes have exon arrangements. Furthermore, promoter analysis unveiled a multitude of cis-acting elements associated with diverse responsiveness including hormones, light responsive, anoxic specific inducibility and anaerobic induction. Analysis of the transcriptome data reveal that about 80% of the pear CYPs genes were upregulated and they were positively correlated with the antioxidant's parameters such as total flavonoids and total phenol content as well as ABTS and DPPH radicals. RT-qPCR analysis confirmed that the CYP genes could be regulated in pear. Collectively, our results reveal comprehensive insights into the CYP superfamily in pear and make a valuable contribution to the ongoing process of functional validation.

Biotechnological and Biocontrol Approaches for Mitigating Postharvest Diseases Caused by Fungal Pathogens and Their Mycotoxins in Fruits: A Review.

Postharvest diseases caused by fungal pathogens are significant contributors to the postharvest losses of fruits. Moreover, some fungal pathogens produce mycotoxins, which further compromise the safety and quality of fruits. In this review, the potential of biotechnological and biocontrol approaches for mitigating postharvest diseases and mycotoxins in fruits is explored. The review begins by discussing the impact of postharvest diseases on fruit quality and postharvest losses. Next, it provides an overview of major postharvest diseases caused by fungal pathogens. Subsequently, it delves into the role of biotechnological approaches in controlling these diseases. The review also explored the application of biocontrol agents, such as antagonistic yeasts, bacteria, and fungi, which can suppress pathogen growth. Furthermore, future trends and challenges in these two approaches are discussed in detail. Overall, this review can provide insights into promising biotechnological and biocontrol strategies for managing postharvest diseases and mycotoxins in fruits.

A First Expression, Purification and Characterization of Endo-ß-1,3-Glucanase from Penicillium expansum.

ß-1,3-glucanase plays an important role in the biodegradation, reconstruction, and development of ß-1,3-glucan. An endo-ß-1,3-glucanase which was encoded by PeBgl1 was expressed, purified and characterized from Penicillium expansum for the first time. The PeBgl1 gene was amplified and transformed into the competent cells of E. coli Rosetta strain with the help of the pET-30a cloning vector. The recombinant protein PeBgl1 was expressed successfully at the induction conditions of 0.8 mmol/L IPTG at 16 °C for 16 h and then was purified by nickel ion affinity chromatography. The optimum reaction temperature of PeBgl1 was 55 °C and it had maximal activity at pH 6.0 according to the enzymatic analysis. Na2HPO4-NaH2PO4 buffer (pH 6.0) and NaCl have inhibitory and enhancing effects on the enzyme activities, respectively. SDS, TritonX-100 and some metal ions (Mg2+, Ca2+, Ba2+, Cu2+, and Zn2+) have an inhibitory effect on the enzyme activity. The results showed that PeBgl1 protein has good enzyme activity at 50-60 °C and at pH 5.0-9.0, and it is not a metal dependent enzyme, which makes it robust for storage and transportation, ultimately holding great promise in green biotechnology and biorefining.

TMT-Based Proteomic Analysis of Hannaella sinensis-Induced Apple Resistance-Related Proteins.

Studies on the molecular mechanism of antagonistic yeasts to control apple postharvest diseases are not comprehensive enough. Our preliminary investigations screened the biocontrol effect of Hannaella sinensis, an antagonistic yeast, and discovered its control efficacy on apple blue mold decay. However, the molecular mechanism of H. sinensis-induced resistance in apple has not been studied. In this study, proteins from apple treated with H. sinensis and sterile saline were analyzed using TMT proteomics technology. It was found that H. sinensis treatment induced the expressions of apple resistance-related proteins. Among the proteins in H. sinensis-induced apple, proteins related to plant defense mechanisms, such as reactive oxygen species scavenging, improvement of plant resistance and synthesis of resistant substances, improvement of plant disease resistance, the degradation of the pathogen cell wall, cell signaling, antibacterial activity, transport of defense-related substances, and protein processing, were differentially regulated. The results of this study revealed the underlying molecular mechanisms of H. sinensis-induced apple resistance at the protein level; the results also provided a theoretical basis for the commercial application of H. sinensis.

Molecular Response of Meyerozyma guilliermondii to Patulin: Transcriptomic-Based Analysis.

Patulin (PAT), mainly produced by Penicillium expansum, is a potential threat to health. In recent years, PAT removal using antagonistic yeasts has become a hot research topic. Meyerozyma guilliermondii, isolated by our group, produced antagonistic effects against the postharvest diseases of pears and could degrade PAT in vivo or in vitro. However, the molecular responses of M. guilliermondii over PAT exposure and its detoxification enzymes are not apparent. In this study, transcriptomics is used to unveil the molecular responses of M. guilliermondii on PAT exposure and the enzymes involved in PAT degradation. The functional enrichment of differentially expressed genes indicated that the molecular response mainly includes the up-regulated expression of genes related to resistance and drug-resistance, intracellular transport, growth and reproduction, transcription, DNA damage repair, antioxidant stress to avoid cell damage, and PAT detoxification genes such as short-chain dehydrogenase/reductases. This study elucidates the possible molecular responses and PAT detoxification mechanism of M. guilliermondii, which could be helpful to further accelerate the commercial application of antagonistic yeast toward mycotoxin decontamination.

Sustainable and efficient method utilizing N-acetyl-L-cysteine for complete and enhanced ochratoxin A clearance by antagonistic yeast.

With the increasing global climate change, ochratoxin A (OTA) pollution in food and environment has become a serious and potential risk element threatening food safety and human health. Biodegradation of mycotoxin is an eco-friendly and efficient control strategy. Still, research works are warranted to develop low-cost, efficient, and sustainable approaches to enhance the mycotoxin degradation efficiency of microorganisms. In this study, the activities of N-acetyl-L-cysteine (NAC) against OTA toxicity were evidenced, and its positive effects on the OTA degradation efficiency of antagonistic yeast, Cryptococcus podzolicus Y3 were verified. Co-culturing C. podzolicus Y3 with 10 mM NAC improved 100% and 92.6% OTA degradation rate into ochratoxin α (OTα) at 1 d and 2 d. The excellent promotion role of NAC on OTA degradation was observed even at low temperatures and alkaline conditions. C. podzolicus Y3 treated with OTA or OTA+NAC promoted reduced glutathione (GSH) accumulation. GSS and GSR genes were highly expressed after OTA and OTA+NAC treatment, contributing to GSH accumulation. In the early stages of NAC treatment, yeast viability and cell membrane were reduced, but the antioxidant property of NAC prevented lipid peroxidation. Our finding provides a sustainable and efficient new strategy to improve mycotoxin degradation by antagonistic yeasts, which could be applied to mycotoxin clearance.

Overexpression of the SDR gene improves the ability of Meyerozyma guilliermondii to degrade patulin in pears and juices.

The intracellular enzymes of antagonistic yeast are effective in controlling patulin (PAT) contamination. However, countless enzymes that have been revealed remain functionally uncharacterized. The study built on previous transcriptomic data obtained by our research group to amplify and express a gene encoding a short-chain dehydrogenase/reductase (SDR) in Meyerozyma guilliermondii. Overexpression of SDR increased the tolerance of M. guilliermondii to PAT and the ability to degrade PAT of the intracellular enzymes. Furthermore, MgSDR-overexpressed M. guilliermondii showed higher PAT degradation in juices (apple and peach) and controlled the blue mold of pears at 20 °C and 4 °C while significantly reduced the content of PAT and the biomass of Penicillium expansum in decayed tissues than wild-type M. guilliermondii. This study provides theoretical references for the subsequent heterologous expression, formulation, and application of the SDR protein from M. guilliermondii and contributes to elucidating the PAT degradation mechanism of antagonistic yeasts.

Ultrastructural observation and transcriptome analysis provide insights into mechanisms of Penicillium expansum invading apple wounds.

Penicillium expansum is a pathogen causing enormous postharvest losses of fruits, especially apples. In this study, we first investigated the morphological changes of P. expansum within apple wounds during infectious process by microscopic observation. We found that conidia swelled and secreted potential hydrophobin in 4 h, germinated in 8 h, and finally formed conidiophores in 36 h, a critical control time point to prevent the second contamination of spores. We then compared the transcript accumulation of P. expansum in apple tissues and liquid culture at 12 h. In total, 3168 and 1318 up-regulated and down-regulated genes were identified. Among them, genes regarding the biosynthesis of substances such as ergosterol, organic acid, cell wall degrading enzymes, and patulin were induced in expression. Pathways were activated, including autophagy, the mitogen-activated protein kinase, and pectin degradation. Our findings provide insights into the lifestyle and the mechanisms of P. expansum invading apple fruits.

Recent advances in Penicillium expansum infection mechanisms and current methods in controlling P. expansum in postharvest apples.

One of the most significant challenges associated with postharvest apple deterioration is the blue mold caused by Penicillium expansum, which leads to considerable economic losses to apple production industries. Apple fruits are susceptible to mold infection owing to their high nutrient and water content, and current physical control methods can delay but cannot completely inhibit P. expansum growth. Biological control methods present promising alternatives; however, they are not always cost effective and have application restrictions. P. expansum infection not only enhances disease pathogenicity, but also inhibits the expression of host-related defense genes. The implementation of new ways to investigate and control P. expansum are expected with the advent of omics technology. Advances in these techniques, together with molecular biology approaches such as targeted gene deletion and whole genome sequencing, will lead to a better understanding of the P. expansum infectious machinery. Here, we review the progress of research on the blue mold disease caused by P. expansum in apples, including physiological and molecular infection mechanisms, as well as various methods to control this common plant pathogen.

Changes of the microbial community in kiwifruit during storage after postharvest application of Wickerhamomyces anomalus.

High-throughput sequencing techniques can provide important information for understanding the interaction between exogenous microbial agents and fruit microbial communities, and explain how it controls postharvest fungal diseases. In this study, we found that Wickerhamomyces anomalus could control the postharvest disease of kiwifruit. Meanwhile, high-throughput sequencing technology results showed that the composition and structure changes of the fungal community in microbial flora were significantly greater than those of bacteria after W. anomalus treated. W. anomalus could colonize inside the fruit and regulate the community composition of bacteria to reduce the abundance of pathogens and eventually maintain the healthy state of the fruit. The dominant genus in the microbiota of kiwifruit after application of W. anomalus showed an increased ability to interact. Some fungi or bacteria are positively associated with yeast in the epiphytic and endophytic sample communities, guiding the synthesis of compound biocontrol strains for kiwifruit postharvest diseases.

Unveiling ochratoxin a controlling and biodetoxification molecular mechanisms: Opportunities to secure foodstuffs from OTA contamination.

Anarchic growth of ochratoxin A (OTA) producing fungi during crop production, prolonged storage, and processing results in OTA contamination in foodstuffs. OTA in food exacerbates the risk of health and economic problems for consumers and farmers worldwide. Although the toxic effects of OTA on human health have not been well established, comprehensive preventive and remedial measures will be essential to eliminate OTA from foodstuffs. Strict regulations, controlling OTA at pre- or post-harvest stage, and decontamination of OTA have been adopted to prevent human and animal OTA exposure. Biological control of OTA and bio-decontamination are the most promising strategies due to their safety, specificity and nutritional value. This review addresses the current understanding of OTA biodegradation mechanisms and recent developments in OTA control and bio-decontamination strategies. Additionally, this review analyses the strength and weaknesses of different OTA control methods and the contemporary approaches to enhance the efficiency of biocontrol agents. Overall, this review will support the implementation of new strategies to effectively control OTA in food sectors. Further studies on efficacy-related issues, production issues and cost-effectiveness of OTA biocontrol are to be carried out to improve the knowledge, develop improved delivery technologies and safeguard the durability of OTA biocontrol approaches.

Study on the biocontrol effect and physiological mechanism of Hannaella sinensis on the blue mold decay of apples.

Blue mold decay is a major postharvest disease of apples, causing considerable losses to the apple industry. In the early stage of this research, an antagonistic yeast, Hannaella sinensis, with a good control effect on the blue mold of apples, was selected. On this basis, the main purpose of this work was to study the biocontrol effect of H. sinensis on the blue mold of apples and the mechanisms involved. The results showed that H. sinensis could effectively control the blue mold decay of apples, reduce the rot rate and diameter, and the antagonistic effect strengthened with the increase of H. sinensis concentration (1 × 108 cells/mL). Further in vitro experiments proved that H. sinensis could significantly inhibit the spore germination and germ tube length of P. expansum. In addition, stable colonization of H. sinensis on apple wounds and surfaces confirmed the environmental adaptability and the ability to compete with other microbiota for nutrition and space. Moreover, H. sinensis induced the activities of resistance-related enzymes such as polyphenol oxidase (PPO), peroxidase (POD), ascorbate peroxidase (APX), superoxide dismutase (SOD), and phenylalanine ammonia-lyase (PAL) in apples and the content of the coding genes corresponding to these enzymes was also higher than that of the control group. Our results indicate that H. sinensis treatment could induce the disease resistance of apples. In summary, H. sinensis served as a promising antagonistic yeast for the prevention and treatment of postharvest blue mold decay of apples.

The necrosis-inducing protein (NIP) gene contributes to Penicillium expansum virulence during postharvest pear infection.

Penicillium expansum is the causative fungus of blue mold decay in postharvest pears resulting in substantial economic losses. Investigating P. expansum-pear fruit interactions is necessary to help develop P. expansum control strategies for effective and safe pear production. Investigating the P. expansum gene expression alterations and essential gene functions during the infection process is indispensable. Based on our results, the necrosis-inducing protein (NIP) gene was closely associated with genes related to plant cell wall degrading enzymes (CWDEs) and involved in P. expansum virulence. The NIP has high homology with other already-known fungal NIPs. To evidence the role of NIP in P. expansum virulence, NIP mutant (including knockout (ΔNIP) and complementation mutant (cNIP)) P. expansum were generated. Despite the NIP deletion did not affect the basic morphology and structure of P. expansum, it slowed down the fungal growth and hyphal production, thus reducing P. expansum's sporulation and patulin (PAT) accumulation. Furthermore, the deletion of NIP reduced the pathogenicity of P. expansum in pear. The complementation of NIP (cNIP) restored the growth, conidia production, PAT accumulation, and virulence of ΔNIP to the level of wild-type P. expansum. In addition, PAT can cause decay and aggravate the disease severity of wild-type P. expansum and ΔNIP on pears. Our results confirmed NIP plays a crucial role in P. expansum's growth, hyphal production, and pathogenicity in pears.

Efficacy of the Yeast Wickerhamomyces anomalus in Biocontrol of Gray Mold Decay of Tomatoes and Study of the Mechanisms Involved.

Gray mold decay is a widespread postharvest disease in tomato that results from infection by the pathogen Botrytis cinerea, leading to huge economic losses. The objective of this study was to select the most effective antagonistic yeast to control tomato gray mold from six potential biocontrol agents and to investigate the possible control mechanism. The results showed that the yeast Wickerhamomyces anomalus was the most effective in inhibiting B. cinerea among the six strains both in vivo and in vitro on tomato, with a colony diameter of 11 mm, a decay diameter of 20 mm, and the lowest decay incidence (53%)-values significantly smaller and lower than the values recorded for the control group and the other yeasts. The efficacy of the control depended on the increase in yeast concentration, and the decay incidence and lesion diameter were reduced to 31%, 28% and 7 mm, 6 mm, respectively, when treated with 1 × 108 and 1 × 109 cells/mL W. anomalus. In addition, W. anomalus was able to rapidly colonize and stably multiply in tomato, occupying the space to control pathogen infection. W. anomalus was also able to motivate the defense mechanism of tomato with stimulation of defense-related enzymes PPO, POD, APX, and SOD and promotion of the content of total phenols and flavonoid compounds. All these results suggest that W. anomalus exhibited exceptional ability to control gray mold in tomato.