Enhancing biological control of postharvest green mold in lemons: Synergistic efficacy of native yeasts with diverse mechanisms of action.

Argentina is among the most important lemon fruit producers in the world. Penicillium digitatum is the primary lemon fungal phytopathogen, causing green mold during the postharvest. Several alternatives to the use of synthetic fungicides have been developed, being the use of biocontrol yeasts one of the most promising. Although many of the reports are based on the use of a single yeast species, it has been shown that the combination of agents with different mechanisms of action can increase control efficiency through synergistic effects. The combined use of native yeasts with different mechanisms of action had not been studied as a biological control strategy in lemons. In this work, the mechanisms of action of native yeasts (Clavispora lusitaniae AgL21, Clavispora lusitaniae AgL2 and Clavispora lusitaniae AcL2) with biocontrol activity against P. digitatum were evaluated. Isolate AgL21 was selected for its ability to form biofilm, colonize lemon wounds, and inhibit fungal spore germination. The compatibility of C. lusitaniae AgL21 with two killer yeasts of the species Kazachstania exigua (AcL4 and AcL8) was evaluated. In vivo assays were then carried out with the yeasts applied individually or mixed in equal cell concentrations. AgL21 alone was able to control green mold with 87.5% efficiency, while individual killer yeasts were significantly less efficient (43.3% and 38.3%, respectively). Inhibitory effects were increased when C. lusitaniae AgL21 and K. exigua strains were jointly applied. The most efficient treatment was the combination of AgL21 and AcL4, reaching 100% efficiency in wound protection. The combination of AgL21 with AcL8 was as well promising, with an efficiency of 97.5%. The combined application of native yeasts showed a synergistic effect considering that the multiple mechanisms of action involved could hinder the development of green mold in lemon more efficiently than using single yeasts. Therefore, this work demonstrates that the integration of native yeasts with diverse modes of action can provide new insights to formulate effective microbial consortia. This could lead to the development of tailor-made biofungicides, allowing control of postharvest fungal diseases in lemons while remaining competitive with traditionally used synthetic chemicals.

Bio-oxidation of progesterone by Penicillium oxalicum CBMAI 1185 and evaluation of the cytotoxic activity.

We report the biotransformation of progesterone 1 by whole cells of Brazilian marine-derived fungi. A preliminary screening with 12 fungi revealed that the strains Penicillium oxalicum CBMAI 1996, Mucor racemous CBMAI 847, Cladosporium sp. CBMAI 1237, Penicillium oxalicum CBMAI 1185 and Aspergillus sydowii CBMAI 935 were efficient in the biotransformation of progesterone 1 in the first days of the reaction, with conversion values ranging from 75 % to 99 %. The fungus P. oxalicum CBMAI 1185 was employed in the reactions in quintuplicate to purify and characterize the main biotransformation products of progesterone 1. The compounds testololactone 1a, 12ß-hydroxyandrostenedione 1b and 1ß-hydroxyandrostenedione 1c were isolated and characterized by NMR, MS, [α]D and MP. In addition, the chromatographic yield of compound 1a was determined by HPLC-PDA in the screening experiments. In this study, we show a biotransformation pathway of progesterone 1, suggesting the presence of several enzymes such as Baeyer-Villiger monooxygenases, dehydrogenases and cytochrome P450 monooxygenases in the fungus P. oxalicum CBMAI 1185. In summary, the results obtained in this study contribute to the synthetic area and have environmental importance, since the marine-derived fungi can be employed in the biodegradation of steroids present in wastewater and the environment. The cytotoxic results demonstrate that the biodegradation products were inactive against the cell lines, in contrast to progesterone.

The colonization of Penicillium oxalicum SL2 on rice root surface increased Pb interception capacity of iron plaque and decreased Pb uptake by roots.

The exploration of microbial resources to reduce Pb accumulation in rice attracted great attention. In this study, we found Penicillium oxalicum SL2, a Pb-tolerant strain with good capability of dissolving phosphorus and stabilizing Pb in soil, was able to colonize on the root surface of rice seedlings without additional carbon sources, and promoted the secretion of metabolites related to amino acid metabolism, organic acid metabolism, signal transduction and other pathways in rhizosphere exudates, in which the secretion of oxalate increased by 47.7 %. However, P. oxalicum SL2 increased Fe(II) proportion and Fe availability on the root surface, resulting in iron plaque content decrease. Moreover, by converting root surface Pb from Pb-Fe state to PbC2O4 and Pb-P compounds, P. oxalicum SL2 increased Pb intercept capacity of iron plaque by 118.0 %. Furthermore, P. oxalicum SL2 regulated element distribution on the root surface, and reduced the relative content of Pb on the maturation zone of root tip, which was conducive to reducing Pb uptake by apoplastic pathway and the risk of Pb accumulation in root system. Our findings further revealed the interaction between P. oxalicum SL2 and rice root, providing a theoretical basis for the development and application of microbial agents in Pb-contaminated farmland.

Antioxidant compounds produced by endolichenic fungus Penicillium sp.-strain 1322P isolated from Pyxine subcinerea.

Endolichenic fungi are expecting for new bioresources of pharmacological compounds. However, the number of investigations targeting antioxidant compounds produced by endolichenic fungi remains limited. To discover new antioxidant compounds, we analyzed the antioxidant activity of the methanol extracts derived from isolated lichen mycobionts or endolichenic fungi induced from Pyxine subcinerea. We performed this analysis using the oxygen radical absorbance capacity (ORAC) method. As a result, we isolated from an endolichenic fungus identified as Penicillium sp.-stain 1322P in Pyxine subcinerea. This fungus produced a red pigment, and its chemical structure was determined to be sclerotioramine based on the analytical data obtained from NMR, LC-MS/MS, and HPLC-PDA. Sclerotioramine exhibited high antioxidant activity, and the ORAC values (mean ± SD) of sclerotioramine and sclerotiorin were 11.4 ± 0.36 and 4.86 ± 0.70 mmol TE per gram of the respective pure compound. Thus, the antioxidant activity of sclerotioramine was greater than twice that of sclerotiorin. This work represents the first report that the antioxidant activity of sclerotioramine is higher than that of the sclerotiorin.

Species diversity of xerophilic Aspergillus and Penicillium in marine surface waters revealed by isolation using osmophilic medium.

The species diversity of xerophilic and halophilic fungi distributed in marine surface water was studied at four local sites located in two geographically distant regions in Japan. At each site, 5-10 samples were collected and isolated using an osmophilic medium. Species identification was conducted based on nucleotide sequence of calmodulin or ß -tubulin and morphological characteristics for Aspergillus, Penicillium, and Talaromyces, and on the sequences of rRNA internal transcribed spacer for the other taxa. Overall, 231 strains were isolated from all sites and classified into 85 species belonged to 12 orders and 33 genera. The isolates that showed better mycelial growth than the control（no NaCl added) in the halotolerance test were defined as halophilic fungi, and only 22 species（10 Aspergillus species and 12 Penicillium species) were halophilic. Comparison of the halophilic fungal flora of the two regions revealed that four species common to both regions were isolated for Aspergillus, but no such species were isolated for Penicillium. Given that 15 halophilic species（10 Aspergillus and 5 Penicillium species) are known to be xerophilic species distributed in indoor environments, it can be inferred that indoor xerophilic species are likely to be widely distributed in marine surface water.

Transcriptomic investigation of the interaction between a biocontrol yeast, Papiliotrema terrestris strain PT22AV, and the postharvest fungal pathogen Penicillium expansum on apple.

Biocontrol strategies offer a promising alternative to control plant pathogens achieving food safety and security. In this study we apply a RNAseq analysis during interaction between the biocontrol agent (BCA) Papiliotrema terrestris, the pathogen Penicillium expansum, and the host Malus domestica. Analysis of the BCA finds overall 802 upregulated DEGs (differentially expressed genes) when grown in apple tissue, with the majority being involved in nutrients uptake and oxidative stress response. This suggests that these processes are crucial for the BCA to colonize the fruit wounds and outcompete the pathogen. As to P. expansum analysis, 1017 DEGs are upregulated when grown in apple tissue, with the most represented GO categories being transcription, oxidation reduction process, and transmembrane transport. Analysis of the host M. domestica finds a higher number of DEGs in response to the pathogen compared to the BCA, with overexpression of genes involved in host defense signaling pathways in the presence of both of them, and a prevalence of pattern-triggered immunity (PTI) and effector-triggered immunity (ETI) only during interaction with P. expansum. This analysis contributes to advance the knowledge on the molecular mechanisms that underlie biocontrol activity and the tritrophic interaction of the BCA with the pathogen and the host.

Screening of core microorganisms in healthy and diseased peaches and effect evaluation of biocontrol bacteria (Burkholderia sp.).

Biological antagonists serve as the most important green alternatives to chemical fungicides, a class of microorganism that inhibits the growth of pathogenic fungi to reduce fruit incidence. In this paper, healthy and diseased peach fruit was selected for amplicon sequencing of the epiphytic microbiota on their surface to obtain a comprehensive understanding. Community structure, diversity and LefSe analysis were performed to screen Acetobacter, Muribaculaceae and Burkholderia as the core bacteria, Mycosphaerella, Penicillium and Alternaria as the core fungi, they showed significant differences and were highly enriched. Two strains fungi (Penicillium K3 and N1) and one strain antagonistic bacteria (Burkholderia J2) were isolated. The in intro test results indicated the bacterial suspension, fermentation broth and volatile organic compounds of antagonistic bacteria J2 were able to significantly inhibit pathogen growth. In vivo experiments, peach was stored at 28 °C for 6 days after different treatments, and samples were taken every day. It was found that Burkholderia J2 enhanced peach resistance by increasing the activities of antioxidant-related enzymes such as SOD, POD, PAL, PPO, GR, MDHAR, and DHAR. The results improved that Burkholderia J2 has great biocontrol potential and could be used as a candidate strain for green control of blue mold.

Dietary potential of the symbiotic fungus Penicillium herquei for the larvae of a nonsocial fungus-cultivating weevil Euops chinensis.

Many insect taxa cultivate fungi for food. Compared to well-known fungus cultivation in social insects, our knowledge on fungus cultivation in nonsocial insects is still limited. Here, we studied the nutritional potentials of the fungal cultivar, Penicillium herquei, for the larvae of its nonsocial insect farmer, Euops chinensis, a specialist on Japanese knotweed Reynoutria japonica. Overall, fungal hyphae and leaf rolls contained significantly higher carbon (C), stable isotopes of C (δ13C), and nitrogen (δ15N) but significantly lower C/N ratios compared to unrolled leaves, whereas insect bodies contained significantly higher N contents but lower C and C/N ratios compared to other types of samples. The MixSIAR model indicated that fungal hyphae contributed a larger proportion (0.626-0.797) to the diet of E. chinensis larvae than leaf materials. The levels of ergosterol, six essential amino acids, seven nonessential amino acids, and three B vitamins tested in fungal hyphae and/or leaf rolls were significantly higher than in unrolled leaves and/or larvae. The P. herquei genome contains the complete set of genes required for the biosynthesis of ergosterol, the essential amino acids valine and threonine, nine nonessential amino acids, and vitamins B2 and B3, whereas some genes associated with five essential and one nonessential amino acid were lost in the P. herquei genome. These suggest that P. herquei is capable of providing the E. chinensis larvae food with ergosterol, amino acids, and B vitamins. P. herquei appears to be able to synthesize or concentrate these nutrients considering that they were specifically concentrated in fungal hyphae. IMPORTANCE: The cultivation of fungi for food has occurred across divergent insect lineages such as social ants, termites, and ambrosia beetles, as well as some seldom-reported solitary insects. Although the fungal cultivars of these insects have been studied for decades, the dietary potential of fungal cultivars for their hosts (especially for those nonsocial insects) is largely unknown. Our research on the mutualistic system Euops chinensis-Penicillium herquei represents an example of the diverse nutritional potentials of the fungal cultivar P. herquei in the diet of the larvae of its solitary host, E. chinensis. These results demonstrate that P. herquei has the potential to synthesize or concentrate ergosterol, amino acids, and B vitamins and benefits the larvae of E. chinensis. Our findings would shed light on poorly understood fungal cultivation mutualisms in nonsocial insects and underscore the nutritional importance of fungal cultivars in fungal cultivation mutualisms.

Penicillium oxalicum SL2-enhanced nanoscale zero-valent iron effectively reduces Cr(VI) and shifts soil microbiota.

Most current researches focus solely on reducing soil chromium availability. It is difficult to reduce soil Cr(VI) concentration below 5.0 mg kg-1 using single remediation technology. This study introduced a sustainable soil Cr(VI) reduction and stabilization system, Penicillium oxalicum SL2-nanoscale zero-valent iron (nZVI), and investigated its effect on Cr(VI) reduction efficiency and microbial ecology. Results showed that P. oxalicum SL2-nZVI effectively reduced soil total Cr(VI) concentration from 187.1 to 3.4 mg kg-1 within 180 d, and remained relatively stable at 360 d. The growth curve of P. oxalicum SL2 and microbial community results indicated that γ-ray irradiation shortened the adaptation time of P. oxalicum SL2 and facilitated its colonization in soil. P. oxalicum SL2 colonization activated nZVI and its derivatives, and increased soil iron bioavailability. After restoration, the negative effect of Cr(VI) on soil microorganisms was markedly alleviated. Cr(VI), Fe(II), bioavailable Cr/Fe, Eh, EC and urease (SUE) were the key environmental factors of soil microbiota. Notably, Penicillium significantly stimulated the growth of urease-positive bacteria, Arthrobacter, Pseudarthrobacter, and Microvirga, synergistically reducing soil chromium availability. The combination of P. oxalicum SL2 and nZVI is expected to form a green, economical and long-lasting Cr(VI) reduction stabilization strategy.

Immunosuppressive effects of the mycotoxin patulin in macrophages.

Patulin (PAT) is a fungi-derived secondary metabolite produced by numerous fungal species, especially within Aspergillus, Byssochlamys, and Penicillium genera, amongst which P. expansum is the foremost producer. Similar to other fungi-derived metabolites, PAT has been shown to have diverse biological features. Initially, PAT was used as an effective antimicrobial agent against Gram-negative and Gram-positive bacteria. Then, PAT has been shown to possess immunosuppressive properties encompassing humoral and cellular immune response, immune cell function and activation, phagocytosis, nitric oxide and reactive oxygen species production, cytokine release, and nuclear factor-κB and mitogen-activated protein kinases activation. Macrophages are a heterogeneous population of immune cells widely distributed throughout organs and connective tissue. The chief function of macrophages is to engulf and destroy foreign bodies through phagocytosis; this ability was fundamental to his discovery. However, macrophages play other well-established roles in immunity. Thus, considering the central role of macrophages in the immune response, we review the immunosuppressive effects of PAT in macrophages and provide the possible mechanisms of action.

The proteome of Penicillium expansum during infection of postharvest apple is revealed using Label-Free and Parallel Reaction Monitoring(PRM)Techniques.

Penicillium expansum is the main pathogen in the postharvest storage of apples. Penicilliosis caused by P. expansum infection not only seriously affects the appearance and quality of fruits, but also the secondary metabolite Patulin (PAT) can cause harm to human health. Until now, little attention has been paid to the molecular mechanism of P. expansum infecting apples. Studying its molecular mechanism can help us better prevent and control apple postharvest blue mold. In this present investigation, we will use Label-Free technology to perform proteomic sequencing on apple samples at key time points of P. expansum infection, explore and screen key proteins and metabolic pathways during infection, and use Parallel Reaction Monitoring (PRM) technology to thoroughly validate proteomic data. The infection of P. expansum activates the MAPK signaling pathway, plant-pathogen interaction metabolic pathway and phenylpropanoid biosynthesis pathway of apple, participates in the regulation of ROS generation and oxidative stress process, promotes the synthesis of lignin and flavonoids, and the synthesis of Pathogenesis-Related Protein helps apple directly defend against P. expansum infection. This study provides the foundation for relevant postharvest control strategies, paving the way for further exploration of the proteome of pathogens infecting fruit and vegetables. SIGNIFICANCE: Proteins are macromolecules essential to the life of organisms, as they participate in the function and structure of cells. Proteomics technology is currently one of the important means to study the the response mechanism of pathogenic bacteria to plant infection, which can reveal the essence of physiological and pathological processes and help to clarify the possible relationship between protein abundance and plant stress. The present study essentially uses recent proteome analysis technology, namely label-free and PRM techniques, and lays the foundations for studying the of the infection response between P. expansum and apples. In particular, it provides a broad perspective on the molecular mechanism of P. expansum in the early stage of apple infection through detailed functional exploration and verification of associated proteins. Thus, it provides a theoretical basis for preventing and treating apple postharvest blue mold.

Production and immobilization of pectinases from Penicillium crustosum in magnetic core-shell nanostructures for juice clarification.

Global market of food enzymes is held by pectinases, mostly sourced from filamentous fungi via submerged fermentation. Given the one-time use nature of enzymes to clarify juices and wines, there is a crucial need to explore alternatives for enzyme immobilization, enabling their reuse in food applications. In this research, an isolated fungal strain (Penicillium crustosum OR889307) was evaluated as a new potential pectinase producer in submerged fermentation. Additionally, the enzyme was immobilized in magnetic core-shell nanostructures for juice clarification. Findings revealed that Penicillium crustosum exhibited enzymatic activities higher than other Penicillium species, and pectinase production was enhanced with lemon peel as a cosubstrate in submerged fermentation. The enzyme production (548.93 U/mL) was optimized by response surface methodology, determining the optimal conditions at 35 °C and pH 6.0. Subsequently, the enzyme was covalently immobilized on synthesized magnetic core-shell nanoparticles. The immobilized enzyme exhibited superior stability at higher temperatures (50 °C) and acidic conditions (pH 4.5). Finally, the immobilized pectinases decreased 30 % the orange juice turbidity and maintained 84 % of the enzymatic activity after five consecutive cycles. In conclusion, Penicillium crustosum is a proven pectinase producer and these enzymes immobilized on functionalized nanoparticles improve the stability and reusability of pectinase for juice clarification.

Genomic and Comparative Transcriptomic Analyses Reveal Key Genes Associated with the Biosynthesis Regulation of Okaramine B in Penicillium daleae NBP-49626.

Restricted production of fungal secondary metabolites hinders the ability to conduct comprehensive research and development of novel biopesticides. Okaramine B from Penicillium demonstrates remarkable insecticidal efficacy; however, its biosynthetic yield is low, and its regulatory mechanism remains unknown. The present study found that the yield difference was influenced by fermentation modes in okaramine-producing strains and performed genomic and comparative transcriptome analysis of P. daleae strain NBP-49626, which exhibits significant features. The NBP-49626 genome is 37.4 Mb, and it encodes 10,131 protein-encoding genes. Up to 5097 differentially expressed genes (DEGs) were identified during the submerged and semi-solid fermentation processes. The oka gene cluster, lacking regulatory and transport genes, displayed distinct transcriptional patterns in response to the fermentation modes and yield of Okaramine B. Although transcription trends of most known global regulatory genes are inconsistent with those of oka, this study identified five potential regulatory genes, including two novel Zn(II)2Cys6 transcription factors, Reg2 and Reg19. A significant correlation was also observed between tryptophan metabolism and Okaramine B yields. In addition, several transporter genes were identified as DEGs. These results were confirmed using real-time quantitative PCR. This study provides comprehensive information regarding the regulatory mechanism of Okaramine B biosynthesis in Penicillium and is critical to the further yield improvement for the development of insecticides.

Penigrines A-E: Five undescribed azepine-indole alkaloids from Penicillium griseofulvum.

Penigrines A-E (1-5), five undescribed azepine-indole alkaloids, were isolated from the fungus Penicillium griseofulvum. Their structures with absolute configurations were determined by NMR, HRESIMS, ECD calculation, and X-ray diffraction experiments. Penigrine C (3) possesses an undescribed 6-oxa-8-azabicyclo[3.2.2]nonane-7,9-dione moiety that fused to an indole core, and penigrines D and E (4 and 5) are a pair of epimers. The plausible biosynthetic pathways of 1-5 are proposed. Penigrine A (1) shows the potential for heart failure treatment.

Complete genome sequence of a novel gammapartitivirus from Penicillium oxalicum RCEF7482.

Penicillium oxalicum, an important biocontrol fungus in China, has been a subject of extensive study due to its role in combating various pathogenic fungi. Despite the prevalence of mycoviruses with double-stranded (ds) RNA genomes in filamentous fungi, there has been no screening of mycoviruses in P. oxalicum. In this report, we describe the identification and characterization of a novel dsRNA virus isolated from P. oxalicum, designated as "Penicillium oxalicum partitivirus 1" (PoPV1). The genome of PoPV1 consists of two dsRNA segments, dsRNA1 (1,770 bp) and dsRNA2 (1,584 bp), each containing a single open reading frame (ORF): ORF1 and ORF2. Comparative analysis revealed that the RdRp and CP amino acid sequences of PoPV1 share the highest identity (89.18% and 73.97%, respectively) with those of Penicillium aurantiogriseum partitivirus 1 (PaPV1). Motif analysis based on RdRp amino acid sequences places PoPV1 in the genus Gammapartitivirus within the family Partitiviridae, with a distinctive motif VI (R/KV/ILGDD). Phylogenetic analysis further established a close relationship of PoPV1 to PaPV1, forming a unique clade among the gammapartitiviruses. Consequently, we propose that Penicillium oxalicum partitivirus 1 represents a new species in the genus Gammapartitivirus. This is the first report of a dsRNA virus in P. oxalicum.

Five previously undescribed citrinin derivatives from the endophytic fungus Penicillium citrinum GZWMJZ-836.

Penicillium citrinum GZWMJZ-836 is an endophytic fungus from Drynaria roosii Nakaike. Five previously undescribed citrinin derivatives (1-5) and six intermediates related to their biosynthesis (6-11) were obtained from the extract of this strain's solid fermentation using multiple column chromatography separations, including high-performance liquid chromatography. The structures of these compounds were determined through comprehensive spectroscopic analyses, primarily using NMR and HRESIMS data. The stereochemistry was mainly confirmed by ECD calculations, and the configurations of C-7' in compounds 4 and 5 were determined using 13C NMR calculations. Compounds 4-5 and 8 showed antibacterial activity against five strains, with minimum inhibitory concentration values ranging from 7.8 to 125 µM. Compounds 4 and 7 exhibited inhibitions against three plant pathogenic fungi, with IC50 values ranging from 66.6 to 152.1 µM. Additionally, a putative biosynthetic pathway for compounds 1-5 derived from citrinin was proposed.

Anti-diabetic and anti-inflammatory indole diterpenes from the marine-derived fungus Penicillium sp. ZYX-Z-143.

Seven new indole-diterpenoids, penpaxilloids A-E (1-5), 7-methoxypaxilline-13-ene (6), and 10-hydroxy-paspaline (7), along with 20 known ones (8-27), were isolated from the marine-derived fungus Penicillium sp. ZYX-Z-143. Among them, compound 1 was a spiro indole-diterpenoid bearing a 2,3,3a,5-tetrahydro-1H-benzo[d]pyrrolo[2,1-b][1,3]oxazin-1-one motif. Compound 2 was characterized by a unique heptacyclic system featuring a rare 3,6,8-trioxabicyclo[3.2.1]octane unit. The structures of the new compounds were established by extensive spectroscopic analyses, NMR calculations coupled with the DP4 + analysis, and ECD calculations. The plausible biogenetic pathway of two unprecedented indole diterpenoids, penpaxilloids A and B (1 and 2), was postulated. Compound 1 acted as a noncompetitive inhibitor against protein tyrosine phosphatase 1B (PTP1B) with IC50 value of 8.60 ± 0.53 µM. Compound 17 showed significant α-glucosidase inhibitory activity with IC50 value of 19.96 ± 0.32 µM. Moreover, compounds 4, 8, and 22 potently suppressed nitric oxide production on lipopolysaccharide-stimulated RAW264.7 macrophages.

Identification of fungi isolated from commercial bumblebee colonies.

Fungi can have important beneficial and detrimental effects on animals, yet our understanding of the diversity and function of most bee-associated fungi is poor. Over 2 million bumblebee colonies are traded globally every year, but the presence and transport of viable fungi within them is unknown. Here, we explored whether any culturable fungi could be isolated from commercial bumblebee nests. We collected samples of various substrates from within 14 bumblebee colonies, including the honey, honey cup wall, egg cup wall, and frass then placed them on agar and recorded any growth. Fungal morphotypes were then subcultured and their ITS region sequenced for identification. Overall, we cultured 11 fungal species from the various nest substrates. These included both pathogenic and non-pathogenic fungi, such as Aspergillus sp., Penicillium sp., and Candida sp. Our results provide the first insights into the diversity of viable fungal communities in commercial bumblebee nests. Further research is needed to determine if these fungi are unique to commercial colonies or prevalent in wild bumblebee nests, and crucially to determine the ecological and evolutionary implications of these fungi in host colonies.

Novel fungal alternative proteins from Penicillium limosum for enhancing structural and functional properties of plant-based meat analogues.

Fungal proteins are excellent novel protein resources due to their high nutritional value and biological activity. In this study, a non-toxic strain of Penicillium limosum with a high biomass yield, protein, and essential amino acid contents, was isolated from wheat Qu (solid-state fermentation starter culture). Pea protein isolate (PPI) and P. limosum mycelial protein powder were extruded to prepare high-moisture meat analogues (HMMA), and their structural and functional properties were evaluated. Compared with 100% PPI, the addition of 5% mycoprotein enhanced the viscosity, gelling properties, chewiness, fibrous degree and in vitro protein digestibility (68.65%) of HMMA. Protein aggregates formed during high temperature extrusion, which increased the oil absorption capacity of HMMA (5% MY substitution). Conversely, their water absorption capacity indices were reduced by 5%. These findings provide a theoretical basis for the functional application of novel fungal alternative proteins.

Ethanolic extract from fruiting bodies of Cordyceps militaris HL8 exhibits cytotoxic activities against cancer cells, skin pathogenic yeasts, and postharvest pathogen Penicillium digitatum.

Cordyceps militaris is a well-known medicinal mushroom in Asian countries. This edible fungus has been widely exploited for traditional medicine and functional food production. C. militaris is a heterothallic fungus that requires both the mating-type loci, MAT1-1 and MAT1-2, for fruiting body formation. However, recent studies also indicated two groups of C. militaris, including monokaryotic strains carrying only MAT1-1 in their genomes and heterokaryotic strains harboring both MAT1-1 and MAT1-2. These strain groups are able to produce fruiting bodies under suitable cultivating conditions. In previous work, we showed that monokaryotic strains are more stable than heterokaryotic strains in fruiting body formation through successive culturing generations. In this study, we report a high cordycepin-producing monokaryotic C. militaris strain (HL8) collected in Vietnam. This strain could form normal fruiting bodies with high biological efficiency and contain a cordycepin content of 14.43 mg/g lyophilized fruiting body biomass. The ethanol extraction of the HL8 fruiting bodies resulted in a crude extract with a cordycepin content of 69.15 mg/g. Assays of cytotoxic activity on six human cancer cell lines showed that the extract inhibited the growth of all these cell lines with the IC50 values of 6.41-11.51 µg/mL. Notably, the extract significantly reduced cell proliferation and promoted apoptosis of breast cancer cells. Furthermore, the extract also exhibited strong antifungal activity against Malassezia skin yeasts and the citrus postharvest pathogen Penicillium digitatum. Our work provides a promising monokaryotic C. militaris strain as a bioresource for medicine, cosmetics, and fruit preservation.