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Penicillium expansum is a necrotrophic pathogen, which actively kills host cells and obtains nutrients from dead cells to achieve infection. However, few reports have elucidated the differential levels of carbon and nitrogen sources over increasing distances of the leading edge in fungal colonized fruit tissues during colonization. Our results showed that the highest consumption of sucrose and fructose, as well as the accumulation of glucose, were found in the decayed region of P. expansum-colonized 'Delicious' apple fruit compared with the healthy region at the leading edge and the healthy region 6 mm away from the leading edge. As nitrogen sources, the contents of methionine, glutamate, leucine, valine, isoleucine and serine were the lowest in the decayed region compared with the healthy regions during colonization. In addition, the titratable acidity, oxalic acid, citric acid, succinic acid and malic acid showed the highest accumulation in the decayed region compared with the healthy regions. P. expansum colonization induced the accumulation of saturated fatty acids in the decayed region, while the level of unsaturated fatty acids was the lowest. These changes were not observed in the healthy regions. These results indicated that P. expansum kills cells in advance of its colonization in order to obtain the nutrients of the apple tissue from the distal leading tissue of the colonized apple. It is understood that more carbon and nitrogen sources are required for fungal colonization, and a stronger defense response against colonization occurred in the fruit, causing the transit of nutrients from the distal tissue to the infected sites. [ABSTRACT FROM AUTHOR]

Two new compounds, including one meroterpenoid (1) and a polyketide derivative (2), along with six compounds (3 − 8), were isolated from the rice solid fermentation of Penicillium expansum GY618. The structures of these new compounds were elucidated through interpretations of spectroscopic data and highresolution electrospray ionization mass spectrometry. The absolute configurations of all new compounds were unambiguously established through the optical rotation or calculation of the ECD spectrum. All isolated compounds were evaluated for their antibacterial and antitumor activities. Compound 6 exhibited weak antitumor activity. [Display omitted] • One new meroterpenoid and a new polyketide derivative were isolated. • The absolute configuration of all new compounds was unambiguously established. • Compound 6 showed weak antitumor activity. [ABSTRACT FROM AUTHOR]

Apples (Malus domestica) are one of the most consumed fruits globally. It is a relevant crop in Argentina and Spain, and one of the main fruits for export and industrialization in these countries. Quality control of apples, fundamentally in the postharvest stage, is critical to prevent fungal diseases. The blue mould, caused by Penicillium expansum , is responsible for great economic losses due to the deterioration of the fruit and mycotoxin production. Many studies have characterized this pathogen; however, little is known about the differences between populations from distant geographical origins. The objective of the present study was to characterize two P. expansum populations, from Argentina and Spain, through morphological, metabolomic and molecular approaches, and to evaluate the existence of differences related to their geographical source. A total of 103 isolates, 53 from Argentina and 50 from Spain were studied. Their morphological features were consistent with the species description. The secondary metabolite profiles revealed low chemical diversity. All 103 isolates shared the production of 13 compounds, namely andrastins, aurantioclavine, chaetoglobosins, communesins, expansolides, roquefortine C and patulin. Penostatins and citrinin were produced by 102 and 101 isolates, respectively. A region of the β-tubulin gene was selected to analyse the diversity of the P. expansum isolates. No substantial differences were observed between isolates of different geographical origins through morphology, patulin accumulation, secondary metabolite profiles and phylogenetic analysis. However, the analysis of polymorphisms revealed 29 haplotypes with a relative separation between isolates of both populations; 13 haplotypes contained Argentinean isolates, while Spanish isolates were separated into 16 haplotypes. The diversity indices of Shannon (H'= 2.075; H'= 2.402) and Simpson (Si D = 0.850; Si D = 0.895) for isolates from Argentina and Spain, respectively, indicated that the diversity of P. expansum is greater in Spain than in Argentina. This distribution could be explained both by the existence of haplotype exchange between both countries, with the ancestral haplotypes originating in Spain, and the subsequent adaptation to the environmental conditions or apples varieties grown in each region. • Penicillium expansum populations from Argentinean and Spanish apples were compared. • P. expansum populations showed low secondary metabolite diversity. • Phylogenetic analyses did not allow establishing evolutionary relationships. • Relative separation was revealed by analysis of polymorphisms in the β-tub fragment. • A significant geographical genetic structure was found in the populations. [ABSTRACT FROM AUTHOR]

Penicillium expansum (P. expansum), a widespread fungal pathogen, causes serious economic loss and public health concerns. The aim of this research is to investigate the antifungal effect of Kluyveromyces marxianus YG-4 (K. marxianus YG-4) against P. expansum and possible mechanism. The results showed that competition for nutrients and space, as well as the release of volatile organic compounds (VOCs), are the antifungal mechanisms. Citronellol may be the antifungal component of K. marxianus YG-4 VOCs based on GC-MS analysis. Further experiments had shown that citronellol inhibited the growth of P. expansum LPH9 by damaging the cell structure, disrupting the redox system, reducing antioxidant enzyme activity, and causing oxidative damage. K. marxianus YG-4, K. marxianus YG-4 VOCs and citronellol can effectively inhibit the spore germination of P. expansum on apples. The above results indicated that K. marxianus YG-4 had strong biocontrol activity and can be used as an excellent candidate strain for fruit preservation., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Penicillium expansum is the major fungus that causes blue mold and produces patulin, threatening human health. Due to health and environmental pollution concerns, chitosan (CS) has attracted more and more attention as a safer alternative to synthetic fungicides for the control of blue mold. In the present study, four different benzimidazole groups were introduced onto CS by the acylation reaction to obtain benzimidazole-based chitosan derivatives (R1b-R4b). After being well-characterized with Fourier transform infrared spectroscopy (FTIR), ultraviolet-visible spectroscopy (UV-vis spectra), and nuclear magnetic resonance (NMR), their antifungal activities against P. expansum were screened. Results showed that the inhibitory effects of chitosan derivatives against the pathogen were significantly correlated with chitosan derivatives' concentration and their structures. R4b was shown as optimum with good solubility and antifungal activity with a minimum inhibitory concentration (MIC) value of 0.5 mg/mL and a minimum fungicidal concentration (MFC) value of 2.0 mg/mL. The remarkable antifungal efficiency of R4b against P. expansum was further demonstrated in terms of spore germination, mycelial growth, patulin production, and the preliminary antifungal mechanism. R4b exhibited significant inhibition of patulin production, while its antifungal mechanism was revealed by destroying cell membrane integrity and inducing membrane depolarization. Furthermore, R4b treatment could significantly reduce the incidence of blue mold rot in apple fruit, and the MTT assay showed the nontoxicity of R4b against Raw 264.7, HBZY-1, and Caco-2 cells. Altogether, these results indicate that it is promising to be used as a fruit preservative in the future.

Antioxidant and antifungal activities of three essential oils (EOs): Melaleuca ericifolia Smith (rosalina; REO), Melaleuca quinquenervia (niaouli; NEO) and Abies alba (fir; FEO) were determined. The antioxidant capacity was investigated by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, and the disc diffusion method was applied to evaluate their antifungal efficacy (in four concentrations 62.5 μL/L, 125 μL/L, 250 μL/L, 500 μL/L used) against Penicillium (P.) expansum microscopic filamentous fungus isolated from bread samples. From the findings it can be clearly evident that antioxidant activities of the EOs were very weak with values ranging from 25.81 ± 7.8 TEAC (6.2 ± 1.4%; FEO) to 162.0 ± 2.1 TEAC (15.9 ± 0.4%; NEO). Regarding antifungal properties, our results revealed that the effects of the EOs on P. expansum growth inhibition were dose-dependent, and they were proportionally increased with increasing EOs concentrations. Detected inhibition zones ranged from 0.00 ± 0.00 mm (for all EOs at 62.5 μL/L) to 11.67 ± 1.15 mm (for REO at 500 μL/L). In conclusion, all analyzed EOs possess promising in vitro antifungal activity (despite their weak antioxidant capacity) suggesting their use as a promising natural preservative in the food industry.

Penicillium expansum is a main producer of patulin that causes severe postharvest decay and food safety issues in the fruit industry. Development, pathogenicity, and patulin production of P. expansum are strongly influenced by the PacC-pH signaling pathway. Global transcription factor PacC regulates various fungal biological processes through a complicated molecular network. In the present study, three Ena family genes (PeEnas), PeEnaA, PeEnaB, and PeEnaC, as important downstream targets of PePacC, were identified in P. expansum. Deletion of PeEnaA, PeEnaB, and PeEnaC showed little effect on mycelial growth under alkaline or high salinity conditions, but double and triple deletion of these genes impaired the virulence of P. expansum on apple fruit. Notably, patulin biosynthesis of P. expansum was distinctly inhibited in the deletion mutants of PeEnas. PeEnas regulated expressions of the patulin gene cluster, AP1, CreA, Sge1, and Hog1 at the transcriptional level and played roles in maintaining membrane potential. Overexpression of PeEnaC in ΔPePacC restored the patulin production defect of ΔPePacC. Our results indicated that, as downstream targets of PePacC, the PeEna family proteins play a crucial role in patulin biosynthesis in P. expansum.

Penicillium digitatum and Penicillium expansum are plant pathogenic fungi that cause the green and blue mold diseases, respectively, leading to serious postharvest economic losses worldwide. Moreover, P. expansum can produce mycotoxins, which are hazardous compounds to human and animal health. The development of tools that allow multiple and precise genetic manipulation of these species is crucial for the functional characterization of their genes. In this sense, CRISPR/Cas9 represents an excellent opportunity for genome editing due to its efficiency, accuracy and versatility. In this study, we developed protoplast generation and transformation protocols and applied them to implement the CRISPR/Cas9 technology in both species for the first time. For this, we used a self-replicative, recyclable AMA1-based plasmid which allows unlimited number of genomic modifications without the limitation of integrative selection markers. As test case, we successfully targeted the wetA gene, which encodes a regulator of conidiophore development. Finally, CRISPR/Cas9-derived ΔwetA strains were analyzed. Mutants showed reduced axenic growth, differential pathogenicity and altered conidiogenesis and germination. Additionally, P. digitatum and P. expansum ΔwetA mutants showed distinct sensitivity to fungal antifungal proteins (AFPs), which are small, cationic, cysteine-rich proteins that have become interesting antifungals to be applied in agriculture, medicine and in the food industry. With this work, we demonstrate the feasibility of the CRISPR/Cas9 system, expanding the repertoire of genetic engineering tools available for these two important postharvest pathogens and open up the possibility to adapt them to other economically relevant phytopathogenic fungi, for which toolkits for genetic modifications are often limited. [ABSTRACT FROM AUTHOR]

The active regulation of extracellular pH is critical for the virulence of fungal pathogens. Penicillium expansum is the causal agent of green-blue mold on stored pome fruits and during its infection process acidifies the host tissues by secreting organic acids. P. expansum is also the main producer of patulin (PAT), a mycotoxin found in pome fruit-based products and that represents a serious health hazard for its potential carcinogenicity. While it is known that PAT biosynthesis in P. expansum is regulated by nutritional factors such as carbon and nitrogen and by the pH, the mechanistic effects of biocontrol on PAT production by P. expansum are not known. In this work, we assessed how optimal and suboptimal concentrations of the biocontrol agent (BCA) Papiliotrema terrestris LS28 affect both extracellular pH and PAT biosynthesis in P. expansum. In wounded apples, the optimal and suboptimal concentrations of the BCA provided almost complete and partial protection from P. expansum infection, respectively, and reduced PAT contamination in both cases. However, the suboptimal concentration of the BCA increased the specific mycotoxigenic activity by P. expansum. In vitro, the rate of PAT biosynthesis was strictly related to the extracellular pH, with the highest amount of PAT detected in the pH range 4-7, whereas only traces were detectable at pH 3. Moreover, both in vitro and in apple wounds the BCA counteracted the extracellular P. expansum-driven acidification maintaining extracellular pH around 4, which is within the pH range that is optimal for PAT biosynthesis. Conversely, in the absence of LS28 the pathogen-driven acidification led to rapidly achieving acidic pH values (<3) that lie outside of the optimal pH range for PAT biosynthesis. Taken together, these results suggest that pH modulation by LS28 is important to counteract the host tissue acidification and, therefore, the virulence of P. expansum. On the other hand, the buffering of P. expansum-driven acidification provided by the BCA increases the specific rate of PAT biosynthesis through the extension of the time interval at which the pH value lies within the optimal range for PAT biosynthesis. Nevertheless, the antagonistic effect provided by the BCA greatly reduced the total amount of PAT. [ABSTRACT FROM AUTHOR]

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Blue mold caused by Penicillium expansum is one of the most common apple diseases, and it is becoming a serious threat in apple production. The strain Bacillus amyloliquefaciens Ba168 showed high levels of antimicrobial activity in our previous study. To analyze the antimicrobial protein of Ba168, a high-resolution LC-MS/MS proteomic analysis was performed. A total of 1155 proteins were identified from 5233 unique peptides. A total of 16 potential antimicrobial-activity-related proteins were identified; 10 of these proteins have direct antimicrobial effects, while 6 of these proteins are associated with the formation of antimicrobial substances. Then, an antifungal protein of Ba168 was isolated and purified by the sequential chromatography of DEAE Bio-sep FF anion exchange and Sephadex G-75. The single protein, named BP8-2, showed antifungal activity towards Penicillium expansum. The peptide mass fingerprinting of the protein band of BP8-2 had a high similarity with the amino acid sequences of flagellin protein. The results showed that BP8-2 significantly inhibited the growth of P. expansum and slowed the spread of apple blue mold. The results indicated that flagellin is one of the important antimicrobial substances from Ba168. [ABSTRACT FROM AUTHOR]

Blue mold is one of the most serious postharvest diseases in apples caused by Penicillium expansum. The purpose of this study is to determine the effect of ε-poly-L-lysine (ε-PL) on the pathogenicity of P. expansum and explore the potential mechanism from the perspective of organic acid. The study investigates the effect of ε-PL treatment on the growth and acid production of P. expansum in vitro and in vivo. When the concentration of ε-PL was 50 mg/L, the growth of P. expansum was inhibited and the decrease in pH value was delayed in the medium. For example, on the third day of culture, P. expansum reduced the pH of the medium from 6.1 to 4.15, and ε-PL inhibited the decrease in the pH value at most 34.4%. When the concentration reached 1000 or 2000 mg/L, the infection of P. expansum in fruits was effectively inhibited. During the growth and infection of P. expansum, gluconic acid is one of the main factors leading to the pH value falling in the local environment. After ε-PL treatment, the accumulation of gluconic acid decreased, the activity of glucose oxidase was suppressed, and then the decline in the local environmental pH slowed down. In addition, after ε-PL treatment, the activities of cell-wall-degrading enzymes, such as cellulase (CL) and polygalacturonase (PG), in the different areas of the P. expansum–apple interaction were also affected by pH change. The results show that ε-PL inhibited the pathogenicity of P. expansum by affecting the accumulation of gluconic acid and slowing the decline in pH in fruit tissues, so as to affect the pathogenicity of P. expansum. This is the first time that the mechanism of ε-PL interfering with the pathogenicity of P. expansum from the perspective of organic acids is clarified. [ABSTRACT FROM AUTHOR]

The aim of this work is the application of pectin coatings containing Cryptococcus laurentii as a method of biocontrol of Penicillium expansum for postharvest protection of apples. For this purpose, the yeast was incorporated into a pectin matrix, and its viability and biocontrol activity in vitro and in vivo against P. expansum was evaluated over time. In addition, the influence of the sterilization process on coating thickness was studied. Results showed that pectin coating with C. laurentii enhanced mycelial growth inhibition in vitro studies, while no significant differences were observed in disease incidence and severity reduction in vivo studies. The sterilization process reduced the viscosity of the pectin solution, resulting in coating thicknesses ranging from 0.5 to 1 μm. As a general evaluation, in vitro and in vivo , biocontrol assays were useful in demonstrating better postharvest protection of the yeast at 7 °C concerning 25 °C. [Display omitted] • Pectin coatings maintain Cryptococcus laurentii viability over 8 weeks. • Penicillium expansum is one of the most damaging postharvest fungi for apples. • In vitro assays revealed mycelial growth inhibition of up to 80 %. • In vivo assays showed 45 % of severity reduction and 27 % disease incidence at 7 °C. • Temperature is a determinant in the efficacy of yeast against blue mold. [ABSTRACT FROM AUTHOR]

The active regulation of extracellular pH is critical for the virulence of fungal pathogens. Penicillium expansum is the causal agent of green-blue mold on stored pome fruits and during its infection process acidifies the host tissues by secreting organic acids. P. expansum is also the main producer of patulin (PAT), a mycotoxin found in pome fruit-based products and that represents a serious health hazard for its potential carcinogenicity. While it is known that PAT biosynthesis in P. expansum is regulated by nutritional factors such as carbon and nitrogen and by the pH, the mechanistic effects of biocontrol on PAT production by P. expansum are not known. In this work, we assessed how optimal and suboptimal concentrations of the biocontrol agent (BCA) Papiliotrema terrestris LS28 affect both extracellular pH and PAT biosynthesis in P. expansum. In wounded apples, the optimal and suboptimal concentrations of the BCA provided almost complete and partial protection from P. expansum infection, respectively, and reduced PAT contamination in both cases. However, the suboptimal concentration of the BCA increased the specific mycotoxigenic activity by P. expansum. In vitro, the rate of PAT biosynthesis was strictly related to the extracellular pH, with the highest amount of PAT detected in the pH range 4–7, whereas only traces were detectable at pH 3. Moreover, both in vitro and in apple wounds the BCA counteracted the extracellular P. expansum-driven acidification maintaining extracellular pH around 4, which is within the pH range that is optimal for PAT biosynthesis. Conversely, in the absence of LS28 the pathogen-driven acidification led to rapidly achieving acidic pH values (

erg4 is a key gene for ergosterol biosynthesis in filamentous fungi, but its function in Penicillium expansum remains unknown. Our results showed that P. expansum contains three erg4 genes, including erg4A, erg4B and erg4C. The expression levels of the three genes showed differences in the wild-type (WT) strain, and the expression level of erg4B was the highest, followed by erg4C. Deletion of erg4A, erg4B or erg4C in the WT strain revealed functional redundancy between them. Compared to the WT strain, erg4A, erg4B or erg4C knockout mutants reduced ergosterol levels, with erg4B deletion having the greatest effect. Furthermore, deletion of the three genes reduced sporulation of the strain, and Δerg4B and Δerg4C mutants showed defective spore morphology. In addition, Δerg4B and Δerg4C mutants were found to be more sensitive to cell wall integrity and oxidative stress. However, deletion of erg4A, erg4B or erg4C had no significant effect on colony diameter, spore germination rate, conidiophore structure of P. expansum or pathogenicity to apple fruit. Taken together, erg4A, erg4B and erg4C have redundant functions and are all involved in ergosterol synthesis and sporulation in P. expansum. In addition, erg4B and erg4C contribute to spore morphogenesis, cell wall integrity and response to oxidative stress in P. expansum.

Penicillium expansum is one of the primary pathogens causing postharvest blue mold of pears, which leads to the spoiling of pulp and produces patulin, endangering food security. Mitogen-activated protein kinase (MAPK) signaling pathway regulates the growth and development, cell wall integrity, and pathogenicity of fungi. Mitogen-activated protein kinase kinase (MKK) in the MAPK signaling pathway has been reflected to be associated with fungal cell wall integrity. Nevertheless, the pivotal function of MKK in the process of P. expansum infection remained unexplored. In this study, MKK was knocked out and repaired by the Agrobacterium -mediated homologous recombination technology to explore the function of MKK in P. expansum infection process. It has been found that the deletion of MKK induces morphological changes in the mutant strain and delayed mycelium and spore growth. Further studies showed that the mutant strain was susceptible to cell wall interference compounds and showed a significant decrease in decay diameters and virulence during the infection of the pears. RT-qPCR analysis showed that the deletion of MKK affected the relative expression levels of its upstream and downstream genes. Our results suggest that MKK is essential in the growth and development, cell wall integrity, pathogenicity, and toxigenic capacity of P. expansum. • MAP kinase MKK negatively affects the growth of Penicillium expansum. • The knockout of MKK reduces the ability of P. expansum to infect the pears. • The knockout of MKK diminishes the toxigenic capacity of P. expansum infecting pears. [ABSTRACT FROM AUTHOR]

[Display omitted] • Penicillium expansum has the smallest mitogenome among other Penicillium mitogenomes reported. • Mitochondrial genomes of all Penicillium species retained the core protein-coding genes and had fewer intronic regions compared to other ascomycetes. • Among Penicillium species, cob exhibited the highest genetic distance while atp 8 exhibited the lowest genetic distance. Penicillium expansum is an important phytopathogenic fungus that causes blue mold disease. In this study, the novel mitochondrial genome of P. expansum was sequenced, assembled, annotated, and compared with the previously published Penicillium mitogenomes. P. expansum mitogenome is composed of circular DNA molecules with a genome size of 25,496 bp. It encodes 16 protein-encoding genes (PCGs), two rRNA genes, and 25 tRNA genes. Comparative analysis with six other Penicillium species revealed that gene length, GC content, AT skew, and GC skew were variable among the core protein-coding genes. The Penicillium species' gene synteny analysis identified several gene rearrangements. Among the core 15 PCGs, atp8 had the lowest K2P genetic distance, which shows that this gene is highly conserved. The Ka/Ks value of most PCGs was less than 1, which shows that these genes have undergone purifying selection. Phylogenetic analysis based on 14 concatenated core mitochondrial genes revealed that P. expansum shares a close relationship with P. solitum. This study served as a first report on the complete mitochondrial genome of P. expansum and its comparative analysis that will contribute to population genetics and rapid evolutionary studies among Penicillium species. [ABSTRACT FROM AUTHOR]

Fruit decay and toxin contamination caused by pathogenic fungi have attracted public attention. As the causal agent of blue mold, Penicillium expansum has caused serious losses to various fruit. Additionally, P. expansum produces patulin, a carcinogenic toxin that can contaminate fruit and fruit products. Wuyiencin has been widely used to control fungal diseases. However, the inhibitory effects of wuyiencin on P. expansum and its mechanisms of action are still unclear. In this study, the effects of wuyiencin on the growth, spore germination, virulence and patulin production of P. expansum were investigated. Possible mechanisms were examined and verified by transcriptomic analysis, RT-qPCR, FDA/PI staining and cell leakage detection. Wuyiencin inhibited the growth, spore germination, virulence and patulin production of P. expansum. A large number of genes encoding membrane proteins were up-regulated and expressed. However, genes encoding membrane-bound organelles were down-regulated under wuyiencin treatment. The integrity of the cell membrane was destroyed. In addition, the expression of some genes related to patulin synthesis and translation/protein processing was altered. These results suggested that wuyiencin effectively controlled the growth, spore germination, virulence, and patulin production of P. expansum by destroying cell membrane integrity and altering the expression of patulin synthesis-related genes and translation/protein processing related genes. • Wuyiencin inhibited the growth, virulence and patulin production of P. expansum. • Wuyiencin could disrupt the cell membrane integrity of P. expansum. • Patulin synthesis-related genes expression in P. expansum were altered by wuyiencin. • Wuyiencin altered the expression of translation/protein processing-related genes. [ABSTRACT FROM AUTHOR]

Apple blue mold is a significant postharvest disease caused by Penicillium expansum. pH modification in colonized tissues leads to the production of organic substances, the modulation of enzymes, and then increases fungal pathogenicity. This study evaluated Penicillium expansum-inoculated apple fruits’ quality responding to pH treatments ranging from 2.5 to 8.5 and analyzed the reactive oxygen species (ROS) metabolism modulation in inoculated apple fruits at the same pH. The results showed that the fruit quality of the firmness, total soluble solids, and titratable acid displayed a quick loss at pHs 5.0 and 7.0, compared with 2.5 and 8.5. Similarly, higher disease incidence was observed at pHs 5.0 and 7.0. Apple fruits infected with P. expansum at pHs 2.5 and 8.5 had less content of O2•−, H2O2, and malondialdehyde (MDA); lower enzymatic activity of NADPH oxidase (NOX); and greater cell membrane integrity than those at pHs 5.0 and 7.0. The analysis of the antioxidant enzymatic activities showed upregulation of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), and glutathione reductase (GR) at pHs 2.5 and 8.5 compared with those at pHs 5.0 and 7.0. Similar trends were shown in ascorbic acid and glutathione. These results support the hypothesis that inoculated apple fruits at pHs 2.5 and 8.5 improve resistance to P. expansum by modulating ROS metabolism, compared with pHs 5.0 and 7.0.

Aims: This research sought to improve the ability of biocontrol yeast to suppress postharvest fungal disease and explore possible mechanisms of action. Methods and Results: The addition of 2% sodium glutamate (SG), which is edible and recognized as safe, enhances the inhibitory effect of Rhodosporidium paludigenum Fell & Tallman on Penicillium expansum in vivo and in vitro. Rhodosporidium paludigenum cells grown in medium with a final concentration of 2% SG, displayed viability under a variety of stress conditions, including sodium chloride (NaCl), calcofluor white (CFW), Congo red (CR) and sodium dodecyl sulphate (SDS). Activity and relative gene expression levels of antioxidant‐related enzymes in R. paludigenum, including peroxisomal catalase (CAT), thioredoxin reductase (TrxR), glutathione peroxidase (GSH‐PX), glutathione reductase (GR) and superoxide dismutase (SOD) were altered in the presence of SG. Levels of reactive oxygen species (ROS) increased in cells grown in the presence of SG as well as the content of several amino acids. Conclusions: In the presence of 2% SG R. paludigenum inhibited P. expansum and exhibited tolerance to a number of stressful conditions which may involve the upregulation of antioxidant enzymes and amino acids. Significance and Impact of the Study: The ability of culture conditions to enhance the fungal suppressive abilities of yeast has the potential to enhance the management of postharvest disease in fruit. [ABSTRACT FROM AUTHOR]

The present work aimed to study the essential oil (EO) variation of Dracocephalum moldavica L. under different phenological stages. The EO yield was highest in the flowering stage, followed by floral budding, fruit set, and vegetative stages. Twenty-three components in EOs were identified by GC-MS and quantified using GC-FID. Geranyl acetate (31–50 %), geranial (16–27 %), geraniol (12–13.4 %) and neral (10.67–17.30 %) were four main constituents. Antifungal assays of EO through mycelial growth inhibition percentages (MGI %) with four concentrations (0, 250, 500, and 750 µl/l) showed concentration-dependent responses. EOs showed the high potential of antifungal activity with ID50 values of (89–145), (27–88) and (183–339) µl/l and ID90 values of (667–1293), (509–1296), and (389–866) µl/l for Aspergillus niger, Penicillium expansum, and Trichoderma harzianum respectively. EO of D. moldavica showed considerable promise as a natural bio-preservative and bio-antifungal agent. [ABSTRACT FROM AUTHOR]

The effectiveness of induced resistance chemicals including chitosan, methyl jasmonate, salicylic acid, silicon and thiabendazole for comparison and some resistance parameters including phenylalanine ammonia-lyase (PAL) activity, scopoletin, and scoparone levels and lignin amount were determined in controlling gray and blue mold of apple caused by Botrytis cinerea and Penicillium expansum. Silicon reduced the incidence decay by 80-96 % for B. cinerea. Chitosan reduced the incidence decay by 92-100 % for P expansum. PAL levels were increased in the highest doses of chitosan applications for P expansum and chitosan and silicon applications for B. cinerea. It was also determined that all chitosan doses promote scopoletin and scoparone activity in the fruit for both pathogens. Lignin content of inoculated fruit was increased around the inoculated tissues for both pathogens. Silicon and chitosan provided increased lignin amount against pathogens at increasing concentrations. In conclusion, applications of chitosan, methyl jasmonate, salicylic acid, and silicon were effective to control gray and blue mold decay of apple fruit caused by B. cinerea and P expansum. PAL activity and scopoletin were increased. Furthermore, scoparone was detected in chitosan and silicon applications. [ABSTRACT FROM AUTHOR]

Antioxidant and antifungal activities of three essential oils (EOs): Melaleuca ericifolia Smith (rosalina; REO), Melaleuca quinquenervia (niaouli; NEO) and Abies alba (fir; FEO) were determined. The antioxidant capacity was investigated by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, and the disc diffusion method was applied to evaluate their antifungal efficacy (in four concentrations 62.5 µL/L, 125 µL/L, 250 µL/L, 500 µL/L used) against Penicillium (P.) expansum microscopic filamentous fungus isolated from bread samples. From the findings it can be clearly evident that antioxidant activities of the EOs were very weak with values ranging from 25.81 ± 7.8 TEAC (6.2 ± 1.4%; FEO) to 162.0 ± 2.1 TEAC (15.9 ± 0.4%; NEO). Regarding antifungal properties, our results revealed that the effects of the EOs on P. expansum growth inhibition were dose-dependent, and they were proportionally increased with increasing EOs concentrations. Detected inhibition zones ranged from 0.00 ± 0.00 mm (for all EOs at 62.5 µL/L) to 11.67 ± 1.15 mm (for REO at 500 µL/L). In conclusion, all analyzed EOs possess promising in vitro antifungal activity (despite their weak antioxidant capacity) suggesting their use as a promising natural preservative in the food industry. [ABSTRACT FROM AUTHOR]

Penicillium expansum is the main cause of apple rot. Besides, it can also produce mycotoxin patulin (PAT). Therefore, the search for substances that can inhibit the activity and toxigenicity of P. expansum has become a hot research topic. This study investigates the inhibitory effects of fengycin on patulin production in P. expansum. P. expansum was cultured under different environments with different concentrations of fengycin. The patulin content produced per unit weight of P. expansum mycelium was detected and determined by high pressure liquid chromatography (HPLC). Synergy brands (SYBR) GreenI Real-time PCR was used to detect the expression levels of 6-methylsalicylic acid synthase (6-MSAS) and isoepoxydon dehydrogenase (IDH), which were the key genes of producing patulin of P. expansum mycelium, in the conditions treated by fengycin and untreated. After fengycin treatments, not only the patulin content in every unit weight of P. expansum mycelium but also the expression level of 6-MSAS decreased significantly. The expression level of 6-MSAS of treatment was 0.11 folds of control. However, the expression level of IDH treated by fengycin decreased slightly. Fengycin could inhibit the P. expansum from producing patulin by downregulating the expression of key synthetic genes 6-MSAS. [ABSTRACT FROM AUTHOR]

Summary: Although several studies have shown the ability of l‐glutamate (glutamate) to mitigate the stress imposed by pathogens, the underlying mechanism is still in its infancy. To gain further knowledge, this study focussed on the effect of glutamate on primary nitrogen and carbon metabolisms during apple‐Penicillium expansum interaction. The obtained result showed that glutamate could effectively restrict blue mould rot development in apples, but had no direct impact on fungal growth in vitro. The application of glutamate increased the level of nitrogen in apples, resulting in a disrupted balance of carbon and nitrogen. Consistently, the key enzymes glutamine synthetase (GS) and glutamate synthase (GOGAT) involved in the GS/GOGAT cycle, and the deaminating activity of glutamate dehydrogenase (GDH), forming 2‐oxoglutarate and ammonium from glutamate, were promptly stimulated by glutamate. Interestingly, glutamate led to considerable consumption of the tricarboxylic acid (TCA) cycle intermediates, such as isocitric acid and citric acid, accompanied by the enhancement of malate dehydrogenase and succinate dehydrogenase activities. Collectively, exogenous application of glutamate might confer blue mould resistance in apples, at least in part, by redirecting host's primary nitrogen and carbon metabolisms, such as the activation of the GS/GOGAT cycle, deaminating activity of GDH and the TCA cycle. [ABSTRACT FROM AUTHOR]

To provide fruit throughout the whole year, maintain quality and reduce spoilage, apples and pears are stored at low temperature. However, the development of rots, caused mainly by Penicillium expansum, cannot be avoided. To prevent fruit losses, biological control has been proposed as a potential alternative. In this work, 16 psychrotrophic, non-pectinolytic Debaryomyces hansenii strains were evaluated in a bioassay for their potential biocontrol against P. expansum rots in apples and pears. Isolates with different degrees of biocontrol effectiveness were further investigated in vitro to elucidate mechanisms of antagonism that may have contributed to biocontrol. No correlation between any of the studied mechanisms and biocontrol activity could be established. One of the isolates, designated F9D, was selected due to its ability to reduce rot incidence in more than 95% in apples and 85% in pears. This strain could be a good candidate for the development of a yeast-based formulation to protect both types of fruit. An ISSR-PCR method was developed for typing the selected strain. This molecular marker could be a useful tool to follow the fate of the strain applied on fruit.

Penicillium expansum is the causal agent of blue mold decay on apple fruits and is also known to be the major producer of patulin, a mycotoxin that represents serious hazard to human health. Several mechanisms have been suggested to explain the pathogenesis of P. expansum in host plants. Secreted effector proteins are vital for the pathogenicity of many fungal pathogens through manipulating their hosts for efficient colonization. In this study, we performed a RNA-Seq analysis followed by computational prediction of effector proteins from P. expansum during infection of the host apple fruits, and a total of 212 and 268 candidate effector protein genes were identified at 6 and 9 h after inoculation (hai), respectively. One of the candidate effector protein genes was identified as a concanavalin A-like lectin/glucanase (Peclg), which was dramatically induced during the pathogen–host interaction. Targeted knockout of Peclg resulted in significant reduction in conidial production and germination relative to the wild type. Further studies showed that in addition to salt stress, the mutant was much more sensitive to SDS and Congo red, suggesting a defect in cell wall integrity. Pathogenicity assays revealed that the ΔPeclg mutant showed significant decrease in virulence and infectious growth on apple fruits. All these results suggest that Peclg is required for fungal growth, stress response, and the virulence of P. expansum.

Blue mold, caused by Penicillium expansum, is the most destructive fungal disease of apples and causes great losses during the post-harvest storage of the fruit. Although some apple cultivars are resistant to P. expansum, there has been little information on the molecular mechanism of resistance. In this study, differential proteomic analysis was performed on apple samples infected and uninfected with P. expansum. Parallel reaction monitoring (PRM) technology was used to target and verify the expression of candidate proteins. The label-free technique identified 343 differentially expressed proteins, which were mainly associated with defense responses, metal ion binding, stress responses, and oxidative phosphorylation. The differential expression of enzymes related to reactive oxygen species (ROS) synthesis and scavenging, the activation of defense-related metabolic pathways, and the further production of pathogenesis-related proteins (PR proteins) during P. expansum infection in apples, and direct resistance to pathogen invasion were determined. This study reveals the mechanisms of apple response at the proteomic level with 9 h of P. expansum infection.

Nowadays, increase fruit losses are being reported due to the development of fungal postharvest diseases. In an attempt to reduce the use of synthetic fungicides, a turn towards natural products such as essential oils (EOs) and natural compounds has been made. The objective of this study was to investigate the effects of eucalyptus (Euc), rosemary (Ros) EO, their mixture (50:50 v/v) and their common main component (i.e., eucalyptol) on the quality parameters, fruit response and inhibition of blue rot (Penicillium expansum) in apple and pear fruits during their shelf life. The results of the present study revealed that fungal colony growth decreased in vitro with exposure at eucalyptus EO (Euc-300 μL/L), rosemary EO (Ros-300 μL/L) and their mixture (Euc + Ros 100 and 300 μL/L). The exposure at Ros-100 μL/L stimulated spore production, whilst Euc + Ros (100 and 300 μL/L) and eucalyptol (100 and 300 μL/L) decreased spore germination. Moreover, the in vivo applied treatments resulted in decreased lesion growth of P. expansum in apple and pear fruits. Respiration rate increased with the application of Euc + Ros at 300 μL/L and eucalyptus EO (Euc-100 μL/L and Euc-300 μL/L) for both assessed fruits. On the other hand, no significant differences were reported on apples and pears total soluble solids and acidity values. The application of Euc + Ros-300 μL/L in apples increased hydrogen peroxide (H2O2) levels, whilst Euc-100 and Euc-300 μL/L increased lipid peroxidation levels. Regarding pear fruits, exposure to Euc-100 μL/L and Ros-100 μL/L resulted in increased H2O2 whereas, Euc-100 μL/L, Ros- (100 and 300 μL/L) and eucalyptol (100 and 300 μL/L) also increased lipid peroxidation. The findings of this study indicate that the investigated natural products can be explored for the preservation of fresh apples and pears, as alternative natural fungicides with consideration of the fresh produce quality attributes. [ABSTRACT FROM AUTHOR]

Penicillium expansum is a necrotrophic wound fungal pathogen that secrets virulence factors to kill host cells including cell wall degrading enzymes (CWDEs), proteases, and mycotoxins such as patulin. During the interaction between P. expansum and its fruit host, these virulence factors are strictly modulated by intrinsic regulators and extrinsic environmental factors. In recent years, there has been a rapid increase in research on the molecular mechanisms of pathogenicity in P. expansum; however, less is known regarding the bacteria–fungal communication in the fruit environment that may affect pathogenicity. Many bacterial species use quorum-sensing (QS), a population density-dependent regulatory mechanism, to modulate the secretion of quorum-sensing signaling molecules (QSMs) as a method to control pathogenicity. N-acyl homoserine lactones (AHLs) are Gram-negative QSMs. Therefore, QS is considered an antivirulence target, and enzymes degrading these QSMs, named quorum-quenching enzymes, have potential antimicrobial properties. Here, we demonstrate that a bacterial AHL lactonase can also efficiently degrade a fungal mycotoxin. The mycotoxin is a lactone, patulin secreted by fungi such as P. expansum. The bacterial lactonase hydrolyzed patulin at high catalytic efficiency, with a kcat value of 0.724 ± 0.077 s−1 and KM value of 116 ± 33.98 μM. The calculated specific activity (kcat/KM) showed a value of 6.21 × 10³ s−1M−1. While the incubation of P. expansum spores with the purified lactonase did not inhibit spore germination, it inhibited colonization by the pathogen in apples. Furthermore, adding the purified enzyme to P. expansum culture before infecting apples resulted in reduced expression of genes involved in patulin biosynthesis and fungal cell wall biosynthesis. Some AHL-secreting bacteria also express AHL lactonase. Here, phylogenetic and structural analysis was used to identify putative lactonase in P. expansum. Furthermore, following recombinant expression and purification of the newly identified fungal enzyme, its activity with patulin was verified. These results indicate a possible role for patulin and lactonases in inter-kingdom communication between fungi and bacteria involved in fungal colonization and antagonism and suggest that QQ lactonases can be used as potential antifungal post-harvest treatment. [ABSTRACT FROM AUTHOR]

Penicillium expansum is one of the most common and destructive post-harvest fungal pathogens that can cause blue mold rot and produce mycotoxins in fruit, leading to significant postharvest loss and food safety concerns. Arginine methylation by protein arginine methyltransferases (PRMTs) modulates various cellular processes in many eukaryotes. However, the functions of PRMTs are largely unknown in post-harvest fungal pathogens. To explore their roles in P. expansum, we identified four PRMTs (PeRmtA, PeRmtB, PeRmtC, and PeRmt2). The single deletion of PeRmtA, PeRmtB, or PeRmt2 had minor or no impact on the P. expansum phenotype while deletion of PeRmtC resulted in decreased conidiation, delayed conidial germination, impaired pathogenicity and pigment biosynthesis, and altered tolerance to environmental stresses. Further research showed that PeRmtC could regulate two core regulatory genes, PeBrlA and PeAbaA, in conidiation, a series of backbone genes in secondary metabolism, and affect the symmetric ω-NG, N’G-dimethylarginine (sDMA) modification of proteins with molecular weights of primarily 16–17 kDa. Collectively, this work functionally characterized four PRMTs in P. expansum and showed the important roles of PeRmtC in the development, pathogenicity, and secondary metabolism of P. expansum. [ABSTRACT FROM AUTHOR]

In this study, green and eco-friendly biosynthesis of selenium nanoparticles (Se-NPs) were performed using Penicillium expansum ATTC 36200 for multiple biomedical applications. Mycosynthesized Se-NPs were completely characterized using UV, FT-IR, XRD, SEM, and TEM techniques. Se-NPs biosynthesized by P. expansum was characterized as a spherical shape with average size 4 to 12.7 nm. Moreover, Se-NPs were evaluated for multiple biomedical applications as antimicrobial, antioxidant, and anticancer activities and hemocompatibility. Results illustrated that Se-NPs have potential antimicrobial activity against Gram-positive (Bacillus subtilis ATCC6051 and Staphylococcus aureus ATCC23235), Gram-negative bacteria (Escherichia coli ATCC8739and Pseudomonas aeruginosa ATCC9027), fungi (Candida albicans ATCC90028, Aspergillus niger RCMB 02724 and Aspergillus fumigatus RCMB 02568), and antioxidant activity. Additionally, Se-NPs exhibited anticancer activity against PC3 cell line; IC50 was 99.25 μg/mL. Meanwhile, they showed non-hemolytic activity on human RBCs at concentration up to 250 μg/mL. In conclusion, biosynthetic Se-NPs by P. expansum are promising for many safe-use biomedical applications. [ABSTRACT FROM AUTHOR]

Blue mold of apple is caused by several different Penicillium species, among which P. expansum and P. solitum are the most frequently isolated. P. expansum is the most aggressive species, and P. solitum is very weak when infecting apple fruit during storage. In this study, we report complete genomic analyses of three different Penicillium species: P. expansum R21 and P. crustosum NJ1, isolated from stored apple fruit; and P. maximae 113, isolated in 2013 from a flooded home in New Jersey, USA, in the aftermath of Hurricane Sandy. Patulin and citrinin gene cluster analyses explained the lack of patulin production in NJ1 compared to R21 and lack of citrinin production in all three strains. A Drosophila bioassay demonstrated that volatiles emitted by P. solitum SA and P. polonicum RS1 were more toxic than those from P. expansum and P. crustosum strains (R27, R11, R21, G10, and R19). The toxicity was hypothesized to be related to production of eight-carbon oxylipins. Putative lipoxygenase genes were identified in P. expansum and P. maximae strains, but not in P. crustosum. Our data will provide a better understanding of Penicillium spp. complex secondary metabolic capabilities, especially concerning the genetic bases of mycotoxins and toxic VOCs. [ABSTRACT FROM AUTHOR]

As a multifunctional signaling molecule, hydrogen sulfide (H2S) has been reported to induce plant responses to a variety of abiotic stresses. However, there are no reports on H2S treatment inducing resistance in apples against Penicillium expansum , a biotic factor, and its possible mechanism of action. In this study, fumigating apples with 5 mM sodium hydrosulfide (NaHS), the exogenous donor of H2S, for 12 h reduced the diameter of lesions in fruit colonized by P. expansum. NaHS treatment markedly promoted the synthesis of endogenous H2S, hydrogen peroxide (H2O2), and nitrogen oxide (NO). In vivo NaHS treatment enhanced the activities of phenylalanine ammonia-lyase, cinnamate 4-hydroxylase, p -coumarate:coenzyme A ligase isoenzymes, caffeoyl-CoA-O-methyltransferase, caffeic acid-O-methyltransferase, ferulic acid-5-hydroxylase, cinnamyl-CoA reductase, and cinnamyl-alcohol dehydrogenase. The treatment also facilitated the production of specific phenolic acids, such as cinnamic acid, p -coumaric acid, caffeic acid, ferulic acid, and sinapic acid; total phenolic compounds; p- coumaryl alcohol; coniferyl alcohol; sinapyl alcohol; and lignin. NaHS treatment induced resistance against P. expansum in apples through H2O2- and NO-mediated activation of phenylpropanoid metabolism. [ABSTRACT FROM AUTHOR]

As a multifunctional signaling molecule, hydrogen sulfide (H2S) has been reported to induce plant responses to a variety of abiotic stresses. However, there are no reports on H2S treatment inducing resistance in apples against Penicillium expansum, a biotic factor, and its possible mechanism of action. In this study, fumigating apples with 5 mM sodium hydrosulfide (NaHS), the exogenous donor of H2S, for 12 h reduced the diameter of lesions in fruit colonized by P. expansum. NaHS treatment markedly promoted the synthesis of endogenous H2S, hydrogen peroxide (H2O2), and nitrogen oxide (NO). In vivo NaHS treatment enhanced the activities of phenylalanine ammonia-lyase, cinnamate 4-hydroxylase, p-coumarate:coenzyme A ligase isoenzymes, caffeoyl-CoA-O-methyltransferase, caffeic acid-O-methyltransferase, ferulic acid-5-hydroxylase, cinnamyl-CoA reductase, and cinnamyl-alcohol dehydrogenase. The treatment also facilitated the production of specific phenolic acids, such as cinnamic acid, p-coumaric acid, caffeic acid, ferulic acid, and sinapic acid; total phenolic compounds; p-coumaryl alcohol; coniferyl alcohol; sinapyl alcohol; and lignin. NaHS treatment induced resistance against P. expansum in apples through H2O2- and NO-mediated activation of phenylpropanoid metabolism.

Transcription factors play a key role in Penicillium expansum infection process. Although the crucial characteristics of some transcription factors of pathogenic fungi have been found, many transcription factors involved in P. expansum infections have not been explored and studied. This study aimed to screen the transcription factors of P. expansum involved in postharvest pear infections by ATAC-seq analysis and to analyze the differentially expressed peak-related genes by GO enrichment and KEGG pathway analysis. Our results found the up-regulation of differentially expressed peak-related genes involved in the MAPK signaling pathway, pentose phosphate pathway, starch and sucrose metabolism, and pentose and glucuronate interconversions. Our study especially confirmed the differential regulation of transcription factors MCM1, Ste12 and gene WSC in the MAPK signaling pathway and PG1, RPE1 in the pentose and glucuronate interconversions pathway. These transcription factors and related genes might play an essential role in pear fruit infection by P. expansum. RT-qPCR validation of twelve expressed peak-related genes in P. expansum showed that the expression levels of these twelve genes were compatible with the ATAC-Seq. Our findings might shed some light on the regulatory molecular networks consisting of transcription factors that engaged in P. expansum invasion and infection of pear fruits.

Salicylic acid (SA) is a natural inducer of disease resistance in fruit, but its application in the food industry is limited due to low water solubility. Here, SA was encapsulated in β-cyclodextrin (β-CD) via the host–guest inclusion complexation method, and the efficacy of SA microcapsules (SAM) against blue mold caused by Penicillium expansum in postharvest apple fruit was elucidated. It was observed that SAM was the most effective in inhibiting the mycelial growth of P. expansum in vitro. SAM was also superior to SA for control of blue mold under in vivo conditions. Enzyme activity analysis revealed that both SA and SAM enhanced the activities of superoxide dismutase (SOD) and phenylalanine ammonia lyase (PAL) in apple fruit, whereas SAM led to higher SOD activities than SA. Total phenolic contents in the SAM group were higher than those in the SA group at the early stage of storage. SAM also improved fruit quality by retarding firmness loss and maintaining higher total soluble solids (TSS) contents. These findings indicate that microcapsules can serve as a promising formulation to load SA for increasing P. expansum inhibition activity and improving quality attributes in apple fruit.

Penicillium expansum is a necrotrophic pathogen, which actively kills host cells and obtains nutrients from dead cells to achieve infection. However, few reports have elucidated the differential levels of carbon and nitrogen sources over increasing distances of the leading edge in fungal colonized fruit tissues during colonization. Our results showed that the highest consumption of sucrose and fructose, as well as the accumulation of glucose, were found in the decayed region of P. expansum-colonized ‘Delicious’ apple fruit compared with the healthy region at the leading edge and the healthy region 6 mm away from the leading edge. As nitrogen sources, the contents of methionine, glutamate, leucine, valine, isoleucine and serine were the lowest in the decayed region compared with the healthy regions during colonization. In addition, the titratable acidity, oxalic acid, citric acid, succinic acid and malic acid showed the highest accumulation in the decayed region compared with the healthy regions. P. expansum colonization induced the accumulation of saturated fatty acids in the decayed region, while the level of unsaturated fatty acids was the lowest. These changes were not observed in the healthy regions. These results indicated that P. expansum kills cells in advance of its colonization in order to obtain the nutrients of the apple tissue from the distal leading tissue of the colonized apple. It is understood that more carbon and nitrogen sources are required for fungal colonization, and a stronger defense response against colonization occurred in the fruit, causing the transit of nutrients from the distal tissue to the infected sites.

The occurrence of reactive oxygen species (ROS) during the colonization of necrotrophic pathogens attacking fruit is critical during the attack, but its importance in Penicillium expansum remains unclear. This study aimed to determine the regulatory effects of NADPH oxidase (Nox) genes on the growth and pathogenicity of P. expansum in apple fruits. Deletion mutants of ΔPeNoxA, ΔPeNoxR, and ΔPeRacA genes were constructed to determine the contribution to the colonization process. The ΔPeRacA strain had a significant effect on the reduction of growth and pathogenicity, the ΔPeNoxA strain negatively regulated the growth and development of P. expansum and did not show any significant effect on the pathogenicity, and the ΔPeNoxR strain showed no effect on the growth or pathogenicity of P. expansum in the apple fruits. However, analysis of the content of O2– and H2O2 in the mycelium of all the Nox mutants showed a significant reduction, confirming the functionality of Nox mutations. Growth under stress conditions in the presence of Congo red, sodium lauryl sulfate, and H2O2 showed a negative effect on the radial growth of ΔPeNoxA, but a positive effect on radial growth reduction by ΔPeNoxR and ΔPeRacA mutants was shown. Interestingly, the host antioxidant activity levels of superoxide dismutase (SOD) andcatalase (CAT) in the fruits after inoculation with ΔPeNoxA, ΔPeNoxR, and ΔPeRacA mutants declined, suggesting reduced ROS accumulation in the colonized region. These results suggest that PeNoxA, PeNoxR, and PeRacA differentially regulate the growth and pathogenicity of P. expansum by producing ROS, and that PeRacA showed the strongest regulatory effect.

To study the mechanism by which Pichia anomala induced with chitosan (1% w/v) controls blue mold disease in table grapes caused by Penicillium expansum, this study evaluated alterations in three yeast enzymatic activities. The changes in the five primary disease defense-related enzymes and two non-enzyme activities of table grapes were assayed. The results of the study showed that chitosan (1% w/v) significantly increased the yeast β-1,3-glucanase, catalase (CAT), and malondialdehyde (MDA) activities. Furthermore, P. anomala alone or induced with chitosan (1% w/v) significantly increased the table grapes enzymatic activities of Polyphenol oxidase (PPO), phenylalanine (PAL), peroxidase (POD), and catalase (CAT) compared to the control. The RT-qPCR results also confirmed that the genes of these major disease defense enzymes were up-regulated when the table grapes were treated with P. anomala. The highest results were recorded when the fruit was treated by yeast induced with chitosan (1% w/v). The phenolic compounds, in addition to their nutritional value, can also increase the antimicrobial properties of table grapes. The current experiment determined that the total phenol and flavonoid contents of table grapes showed the highest results for fruits treated by P. anomala induced with chitosan compared with the control. Generally, the increment of these fruit enzymatic and non-enzymatic activities shows improved table grape defense against the pathogenic fungus. The induction of the yeast with chitosan also increases its bio-control efficacy against the pathogen. This study will enable future detailed investigation in the yeast pathogen control mechanisms and the use of yeasts as bio-pesticides.

The occurrence of reactive oxygen species (ROS) during the colonization of necrotrophic pathogens attacking fruit is critical during the attack, but its importance in Penicillium expansum remains unclear. This study aimed to determine the regulatory effects of NADPH oxidase (Nox) genes on the growth and pathogenicity of P. expansum in apple fruits. Deletion mutants of Δ PeNoxA , Δ PeNoxR , and Δ PeRacA genes were constructed to determine the contribution to the colonization process. The Δ PeRacA strain had a significant effect on the reduction of growth and pathogenicity, the Δ PeNoxA strain negatively regulated the growth and development of P. expansum and did not show any significant effect on the pathogenicity, and the Δ PeNoxR strain showed no effect on the growth or pathogenicity of P. expansum in the apple fruits. However, analysis of the content of O2– and H2O2 in the mycelium of all the Nox mutants showed a significant reduction, confirming the functionality of Nox mutations. Growth under stress conditions in the presence of Congo red, sodium lauryl sulfate, and H2O2 showed a negative effect on the radial growth of Δ PeNoxA , but a positive effect on radial growth reduction by Δ PeNoxR and Δ PeRacA mutants was shown. Interestingly, the host antioxidant activity levels of superoxide dismutase (SOD) andcatalase (CAT) in the fruits after inoculation with Δ PeNoxA , Δ PeNoxR , and Δ PeRacA mutants declined, suggesting reduced ROS accumulation in the colonized region. These results suggest that PeNoxA , PeNoxR , and PeRacA differentially regulate the growth and pathogenicity of P. expansum by producing ROS, and that PeRacA showed the strongest regulatory effect. [ABSTRACT FROM AUTHOR]

Blue mold decay is the one of most important postharvest disease of apples caused by the fungus, Penicillium expansum. This study aimed to investigate the biocontrol efficacy of the yeast, Wickerhamomyces anomalus, on postharvest blue mold decay of apples and the relative defense mechanisms. The results indicated that W. anomalus could significantly reduce the blue mold decay of apples, and the maximum inhibition was obtained when the concentration of W. anomalus was 1 × 108 cells ml−1. Furthermore, W. anomalus significantly reduced the fruit decay under ambient conditions, without generating any change in fruit quality. In vitro experiments showed that W. anomalus greatly inhibited the spore germination and germ tube elongation of P. expansum. Besides, its ease of adaptation, stable growth and potential colonization of in apple wounds or surfaces indicated that W. anomalus could compete with P. expansum for nutrients and space, leading to considerable inhibition blue mold decay. W. anomalus significantly induced the activities of polyphenol oxidase (PPO), peroxidase (POD), catalase (CAT), phenylalanine ammonia-lyase (PAL), and ascorbate peroxidase (APX) in apples. Moreover, W. anomalus increased the contents of flavonoid and total phenols. All these results suggested that W. anomalus has potential biocontrol efficacy to control the postharvest blue mold decay of apples [ABSTRACT FROM AUTHOR]

Botrytis cinerea and Penicillium expansum are the most important pathogens of apples during the postharvest phase. In the present study, the effect of ozone (O3) and carbon dioxide (CO2) gases combined with application of the yeast isolates Candida membranifaciens A2, Saccharomyces cerevisiae 69, Pichia guilliermondii M47 and A6, was investigated to control blue and gray molds of apples in vitro and in vivo conditions. In vitro, O3 and CO2 treatments exerted respectively fungistatic and fungicidal activity on fungal spores. In dual culture tests, the combination of antagonistic yeasts with CO2 was the most efficient way to control the growth of both pathogens. In vitro, O3 completely inhibited the growth of yeast isolates, whereas CO2 did not influence the yeast growth. Inoculation of apples with C. membranifaciens A2, P. guilliermondii A6 or S. cerevisiae PTCC 69 in combination with CO2 completely suppressed blue or gray mold symptoms for up to 62 days of storage, both at 20 °C and 4 °C. Yeasts combined with O3 (10 ppm) were poorly effective in controlling disease development on fruit. However, CO2 or O3 treatments used alone resulted in a decrease of apple quality after 62 days of storage at 4 °C; necrotic spots were observed for fruits treated with O3. [ABSTRACT FROM AUTHOR]