Penicillium Funiculosum Research Articles

Evaluation and isolation of anti-cancer compounds from the endophytic fungus Penicillium funiculosum isolated from Persicaria salicifolia (Brouss. Ex Willd.) seeds growing in Egypt

Background The endophytic fungus Penicillium is an important source of natural bioactive products. Persicaria salicifolia (Brouss. ex Willd) (family Polygonaceae) is a widely distributed plant on the Nile River and was reported to have several biological activities, such as antioxidative, antibacterial, and anti-inflammatory effects. Objective This study aimed to explore the potential activities (cytotoxic, anti-oxidant, and anti-microbial) of the endophytic fungus Penicillium funiculosum isolated from Persicaria salicifolia seeds growing in Egypt. Materials and methods The endophytic fungus Penicillium funiculosum was isolated from the seeds of the Persicaria salicifolia plant. The fungi were grown on Basmati rice as a solid media for the enhancement of pure fungi production. The collected fungi were extracted with ethyl acetate and fractionated using n-hexane and methanol. All fractions were examined for their cytotoxic, antioxidant, and antimicrobial activities. The secondary metabolites were isolated from the active fractions through column chromatography, and the isolated compounds were identified by spectroscopic technique. Molecular docking analysis was applied to the isolated compounds. Results and conclusion Ethyl acetate extract (Pf-2) of P. funiculosum was proved to have a highly potent antioxidant (IC50 37.5±0.70 μg/ml) and cytotoxic effects. It was affected on hepatic cancer (HepG-2), human colon carcinoma (HCT-116), lung carcinoma (A-549), and human breast cancer (MCF-7) cells with IC50 values (μg/ml) 4.26±0.2, 6.66±0.9, 9.36±0.3, and 9.41±0.7, respectively. Pf 2 was subjected to fractionation, resulting in four fractions (Pf 2–1 to Pf 2–4). The most potent cytotoxic fraction, Pf 2–2, was further fractionated into six sub-fractions: Pf 2–2 A to Pf 2–2 F. Pf 2–2 A possessed the most potent cytotoxic activity. The ethyl acetate extract (Pf 2) also had antimicrobial activity against gram-positive Bacillus subtilis and gram-negative bacteria (E. coli). Four compounds were isolated from Pf 2–2 A and identified by spectroscopic methods: NMR (1H and 13C) and Mass as (1) (9E, 11E, 13Z, 15Z)-tetracosa-9, 11, 13, 15-tetraenoic acid, (2) 3-(1Z, 3E-hexa-1, 3-dienyl)-4b-methyl-tetradecahydrophenanthrene, (3) mannitol, (4) d-Cerebroside A-glucose. It is the first time to report the isolation of compound (2) from the genus Penicillium and compounds (1, 3, and 4) from the species P. funiculosum. Compound 2 was the most potent cytotoxic one. Molecular docking was studied for polar compounds 1, 3, and 4 using the COX-2 enzyme, which indicated that compound 4 was the most potent anti-inflammatory one.

Novel insights into dimethylsulfoniopropionate cleavage by deep subseafloor fungi

Dimethylsulfoniopropionate (DMSP), a key organic sulfur compound in marine and subseafloor sediments, is degraded by phytoplankton and bacteria, resulting in the release of the climate-active volatile gas dimethylsulfide (DMS). However, it remains unclear if dominant eukaryotic fungi in subseafloor sediments possess specific abilities and metabolic mechanisms for DMSP degradation and DMS formation. Our study provides the first evidence that fungi from coal-bearing sediments ∼2 km below the seafloor, such as Aspergillus spp., Chaetomium globosum, Cladosporium sphaerospermum, and Penicillium funiculosum, can degrade DMSP and produce DMS. In Aspergillus sydowii 29R-4-F02, which exhibited the highest DMSP-dependent DMS production rate (16.95 pmol/μg protein/min), two DMSP lyase genes, dddP and dddW, were identified. Remarkably, the dddW gene, previously observed only in bacteria, was found to be crucial for fungal DMSP cleavage. These findings not only extend the list of fungi capable of degrading DMSP, but also enhance our understanding of DMSP lyase diversity and the role of fungi in DMSP decomposition in subseafloor sedimentary ecosystems.

Economically Imperative Ananas comosus Diseases, Status, and Its Control Measures Documented in Producing Countries

Ananas comosus, commonly known as pineapple, is a fruit with a large potential market as a commodity and commercial fruit. Numerous pests and diseases affect pineapple, directly or indirectly, by lowering the quality and quantity. The fungal causative agents, namely Fusarium ananatum and Thalaromyces stolii (previously named Penicillium funiculosum), cause fruitlet core rot (FCR) and fusariosis by Fusarium guttiforme. Bacteria heart rot (BHR) is an infection by Erwinia chrysanthemi, newly known as Dickeya zeae. Nevertheless, the mealybug wilt of pineapple (MWP) is another pineapple treat to susceptible pineapple varieties caused by pineapple mealybug wilt-associated viruses (PMWaVs). Other diseases include destruction caused by pathogenic nematodes. This review discusses the status of these diseases and the control measures that greatly affect the economy of pineapple-producing countries due to the economic significance of these crops. Growers need up-to-date information on the identity of the diseases that affect pineapple crops in the various countries that produce them to effectively manage the diseases in the field.

Marine cyanobacterial biomass is an efficient feedstock for fungal bioprocesses

BackgroundMarine cyanobacteria offer many sustainability advantages, such as the ability to fix atmospheric CO2, very fast growth and no dependence on freshwater for culture. Cyanobacterial biomass is a rich source of sugars and proteins, two essential nutrients for culturing any heterotroph. However, no previous study has evaluated their application as a feedstock for fungal bioprocesses.ResultsIn this work, we cultured the marine cyanobacterium Synechococcus sp. PCC 7002 in a 3-L externally illuminated bioreactor with working volume of 2 L with a biomass productivity of ~ 0.8 g L−1 day−1. Hydrolysis of the biomass with acids released proteins and hydrolyzed glycogen while hydrolysis of the biomass with base released only proteins but did not hydrolyze glycogen. Among the different acids tested, treatment with HNO3 led to the highest release of proteins and glucose. Cyanobacterial biomass hydrolysate (CBH) prepared in HNO3 was used as a medium to produce cellulase enzyme by the Penicillium funiculosum OAO3 strain while CBH prepared in HCl and treated with charcoal was used as a medium for citric acid by Aspergillus tubingensis. Approximately 50% higher titers of both products were obtained compared to traditional media.ConclusionsThese results show that the hydrolysate of marine cyanobacteria is an effective source of nutrients/proteins for fungal bioprocesses.

Engineering of glycoside hydrolase family 7 cellobiohydrolases directed by natural diversity screening

Protein engineering and screening of processive fungal cellobiohydrolases (CBHs) remain challenging due to limited expression hosts, synergy-dependency, and recalcitrant substrates. In particular, GH7 CBHs are a family critically important for the bioeconomy as well as a methodology for improving these and other difficult to engineer enzymes. Here, we target the discovery of highly active natural GH7 CBHs and engineering of variants with improved activity. Using experimentally assayed activities of genome mined CBHs, we applied sequence and structural alignments to top performers to identify key point mutations linked to improved activity. From ∼1500 known GH7 sequences, an evolutionarily diverse subset of 57 GH7 CBH genes was expressed in Trichoderma reesei and screened using a multiplexed activity screening assay. Ten catalytically enhanced natural variants were identified, produced, purified, and tested for efficacy using industrially-relevant conditions and substrates. Three key amino acids in CBHs with performance comparable or superior to Penicillium funiculosum Cel7A were identified and combinatorically engineered into P. funiculosum cel7a, expressed in T. reesei, and assayed on lignocellulosic biomass. The top performer generated using this combined approach of natural diversity genome mining, experimental assays, and computational modeling produced a 41% increase in conversion extent over native P. funiculosum Cel7A, a 55% increase over the current industrial standard T. reesei Cel7A, and 10% improvement over Aspergillus oryzae Cel7C, the best natural GH7 CBH previously identified in our laboratory.

Calcium signaling positively regulates cellulase translation and secretion in a Clr-2-overexpressing, catabolically derepressed strain of Penicillium funiculosum

BackgroundLow-cost cellulase production is vital to sustainable second-generation biorefineries. The catabolically derepressed strain of Penicillium funiculosum NCIM1228 (PfMig188 or ∆Mig1) secretes a superior set of cellulolytic enzymes, that are most suitable for 2G biorefineries. At a 3% (w/w) load, the ∆Mig1 secretome can release > 80% of fermentable sugars from lignocellulose at a 15% (w/v) biomass load, irrespective of the type of biomass and pretreatment. The robustness of the secretome can be further increased by improving the cellulase production capacity of the fungal strain.ResultsWe began by identifying the transcription factor responsible for cellulase production in NCIM1228. An advanced RNA-seq screen identified three genes, clr-2, ctf1a and ctf1b; the genes were cloned under their native promoters and transformed into NCIM1228. Of the three, clr-2 overexpression led to twofold higher cellulase production than the parent strain and was thus identified as the transcriptional activator of cellulase in NCIM1228. Next, we overexpressed clr-2 in ∆Mig1 and expected an exponential increase in cellulolytic attributes accredited to the reinforced activation mechanisms, conjoint with diminished negative regulation. Although clr-2 overexpression increased the transcript levels of cellulase genes in ∆Mig1, there was no increase in cellulase yield. Even a further increase in the transcript levels of clr-2 via a stronger promoter was ineffective. However, when the CaCO3 concentration was increased to 5 g/l in the growth medium, we achieved a 1.5-fold higher activity of 6.4 FPU/ml in the ∆Mig1 strain with clr-2 overexpression. Enthused by the calcium effect, a transcriptomic screen for genes encoding Ca2+-activated kinase identified ssp1, whose overexpression could further increase cellulase yield to ~ 7.5 FPU/ml. Investigation of the mechanism revealed that calcium signaling exclusively enhances the translation and secretion of cellulase in Penicillium funiculosum.ConclusionsOur study identifies for the first time that cellulose activates two discrete signaling events to govern cellulase transcription and posttranscriptional processes (translation, processing and secretion) in P. funiculosum NCIM1228. Whereas Clr-2, the transcriptional activator of cellulase, governs transcription, calcium signaling specifically activates cellulase translation and secretion.

Bioprocess Optimization of Penicillium funiculosum NCIM 1228 for Improved Production and Hydrolytic Efficiency of Cellulases on Sugarcane Bagasse

Bioprocess Optimization of Penicillium funiculosum NCIM 1228 for Improved Production and Hydrolytic Efficiency of Cellulases on Sugarcane Bagasse

Insulation Materials Susceptibility to Biological Degradation Agents: Molds and Subterranean Termites

Insulation materials are fundamental for decreasing energy losses and guaranteeing thermal and acoustic comfort in buildings, which may significantly contribute to decreasing the energy consumption related with poor thermal building conditions. These insulation materials should have a low susceptibility to biological degradation agents to decrease the risks of degradation of other construction materials, as well as decrease possible health risks related with the development of noxious biological degradation agents regarding indoor air quality, for example, or decrease possible structural risks posed by those agents. The present study aimed at evaluating the susceptibility of several insulation materials to mold growth and subterranean termites’ attack. Insulation materials, including expanded polystyrene (EPS), mineral wool (MW), and expanded cork agglomerate (ICB), were tested against mold development, using maritime pine as a control. Three types of inoculations were made: (1) natural indoor inoculation; (2) artificial inoculation using Aspergillus niger and Penicillium funiculosum; and (3) artificial inoculation using Aureobasidium pullulans. The susceptibility of the insulation materials referred to, plus wood/glass fiber (WGF), was evaluated for two subterranean termite species: Reticulitermes grassei and Reticulitermes flavipes. The expanded cork agglomerate showed a higher susceptibility to molds than the other insulation materials tested. The remaining materials revealed a good performance, showing no growth or traces of growth of molds. All the materials tested showed susceptibility to subterranean termites, with both species being able to cross them to obtain access to the wood. However, wood/glass fiber showed a negative effect, which translated into lower survival rates and attack degrees of the wood. Some tested materials showed a good resistance to the development of biological degradation agents, namely an organic material (coconut fiber), a composite of organic and inorganic materials (WGF), and an inorganic material (EPS). These results indicate that it is possible to pursue the development of innovative and effective insulation materials with a low susceptibility to biological degradation agents, regardless of their organic or inorganic origin.

Systematic identification of CAZymes and transcription factors in the hypercellulolytic fungus Penicillium funiculosum NCIM1228 involved in lignocellulosic biomass degradation

BackgroundPenicillium funiculosum NCIM1228 is a filamentous fungus that was identified in our laboratory to have high cellulolytic activity. Analysis of its secretome suggested that it responds to different carbon substrates by secreting specific enzymes capable of digesting those substrates. This phenomenon indicated the presence of a regulatory system guiding the expression of these hydrolyzing enzymes. Since transcription factors (TFs) are the key players in regulating the expression of enzymes, this study aimed first to identify the complete repertoire of Carbohydrate Active Enzymes (CAZymes) and TFs coded in its genome. The regulation of CAZymes was then analysed by studying the expression pattern of these CAZymes and TFs in different carbon substrates—Avicel (cellulosic substrate), wheat bran (WB; hemicellulosic substrate), Avicel + wheat bran, pre-treated wheat straw (a potential substrate for lignocellulosic ethanol), and glucose (control).ResultsThe P. funiculosum NCIM1228 genome was sequenced, and 10,739 genes were identified in its genome. These genes included a total of 298 CAZymes and 451 TF coding genes. A distinct expression pattern of the CAZymes was observed in different carbon substrates tested. Core cellulose hydrolyzing enzymes were highly expressed in the presence of Avicel, while pre-treated wheat straw and Avicel + wheat bran induced a mixture of CAZymes because of their heterogeneous nature. Wheat bran mainly induced hemicellulases, and the least number of CAZymes were expressed in glucose. TFs also exhibited distinct expression patterns in each of the carbon substrates. Though most of these TFs have not been functionally characterized before, homologs of NosA, Fcr1, and ATF21, which have been known to be involved in fruiting body development, protein secretion and stress response, were identified.ConclusionsOverall, the P. funiculosum NCIM1228 genome was sequenced, and the CAZymes and TFs present in its genome were annotated. The expression of the CAZymes and TFs in response to various polymeric sugars present in the lignocellulosic biomass was identified. This work thus provides a comprehensive mapping of transcription factors (TFs) involved in regulating the production of biomass hydrolyzing enzymes.

Profiling of the β-glucosidases identified in the genome of Penicillium funiculosum: insights from genomics, transcriptomics, proteomics, and homology-modeling studies.

The enzymatic conversion of lignocellulosic biomass to bioethanol depends on efficient enzyme systems with β-glucosidase as one of the key components. In this study, we performed in-depth profiling of the various β-glucosidases present in the genome of the hypercellulolytic fungus Penicillium funiculosum using genomics, transcriptomics, proteomics, and molecular dynamics simulation approaches. Of the eight β-glucosidase genes identified in the P. funiculosum genome, three were predicted to be extracellular based on signal peptide prediction and abundance in the secretome. Among the three secreted β-glucosidases, two belonged to the GH3 family and one belonged to the GH1 family. Homology models of these proteins predicted a deep and narrow active site for the GH3 β-glucosidases (PfBgl3A and PfBgl3B) and a shallow open active site for the GH1 β-glucosidase (PfBgl1A). The enzymatic assays indicated that P. funiculosum-secreted proteins showed high β-glucosidase activities with prominent bands on the 4-methylumbelliferyl β-D-glucopyranoside zymogram. To understand the contributory effects of each of the three secreted β-glucosidases (PfBgls), the corresponding gene was deleted separately, and the effect of the deletion on the β-glucosidase activity of the secretome was examined. Although not the most abundant, PfBgl3A was found to be one of the most important β-glucosidases, as evidenced by a 42% reduction in β-glucosidase activity in the ΔPfBgl3A strain. Our results advance the understanding of the genetic and biochemical nature of all β-glucosidases produced by P. funiculosum and pave the way to design a superior biocatalyst for the hydrolysis of lignocellulosic biomass. IMPORTANCE Commercially available cellulases are primarily produced from Trichoderma reesei. However, external supplementation of the cellulase cocktail from this host with exogenous β-glucosidase is often required to achieve the desired optimal saccharification of cellulosic feedstocks. This challenge has led to the exploration of other cellulase-producing strains. The nonmodel hypercellulolytic fungus Penicillium funiculosum has been studied in recent times and identified as a promising source of industrial cellulases mainly due to its ability to produce a balanced concoction of cellulolytic enzymes, including β-glucosidases. Various genetic interventions targeted at strain improvement for cellulase production have been performed; however, the β-glucosidases of this strain have remained largely understudied. This study, therefore, reports profiling of all eight β-glucosidases of P. funiculosum via molecular and computational approaches. The results of this study provide useful insights that will establish the background for future engineering strategies to transform this fungus into an industrial workhorse.

Phenolic profile, in vitro antimicrobial and in vivo diuretic effects of endemic wild thyme Thymus comosus Heuff ex. Griseb. (Lamiaceae) from Romania.

Thymus comosus Heuff ex. Griseb. (Lamiaceae) is a wild thyme species endemic for Romanian Carpathian areas, frequently collected as substitute for collective herbal product Serpylli herba, cited as antibacterial and diuretic remedy in traditional medicine. The present study aimed to evaluate the in vivo diuretic effect and in vitro antimicrobial properties of three herbal preparations (infusion-TCI, tincture-TCT and an hydroethanolic extract prepared through an optimized ultrasound-assisted method-OpTC) obtained from the aerial parts of T. comosus Heuff ex. Griseb, also evaluating their comprehensive phenolic profile. In vivo diuretic effect was tested using Wistar rats treated orally with each herbal preparation (125 and 250mg/kg dispersed in 25ml/kg isotonic saline solution) and quantified based on cumulative urine output (ml), diuretic action and diuretic activity. Additionally, sodium and potassium excretion were monitored using a potentiometric method with selective electrodes. In vitro antibacterial and antifungal activities were assessed using p-iodonitrotetrazolium chloride assay against six bacterial strains and six fungal strains by monitoring minimum inhibitory concentration (MICs), minimum bactericidal concentrations (MBCs) and minimum fungicidal concentrations (MFCs). Finally, phenolic profile of the aforementioned herbal extracts was evaluated using an ultra-high-pressure liquid chromatography (UHPLC) coupled with high-resolution mass spectrometry (HRMS) method to check the impact of the different preparations on the most abundant and significant compounds. All the extracts exerted a mild diuretic action, TCT and OpTC inducing the most intense diuretic effect. Both herbal preparations produced a statistically significant, dose-dependent and gradual increase of the urine output, the effect being more intense at 24h (6.63-7.13 ml/24h). Potentiometric evaluation of urine samples collected from treated rats revealed a clear and mild natriuretic and kaliuretic effect after the administration. In terms of antimicrobial activity, E. coli (MIC-0.38mg/ml), B. cereus (MIC-0.75mg/ml)), Penicillium funiculosum and P. verrucosum var. cyclopium (MIC-0.19mg/ml) showed the greater sensitivity to the tested extracts, respectively. UHPLC-HRMS screening showed that the bioactive potential of T. comosus herbal preparations was likely related to the higher amounts of phenolic acids (including rosmarinic acid), flavonoids (mainly flavones and derivatives) and other phenolics (such as different isomers of salvianolic acids) in their composition. The obtained results support the ethnopharmacological evidence regarding the mild diuretic and antibacterial potentials of the endemic wild thyme T. comosus, this study being the first one that assessed the aforementioned bioactivities for this species.

Solvent System-Guided Extraction of Centaurium spicatum (L.) Fritch Provides Optimized Conditions for the Biological and Chemical Characteristics of the Herbal Extracts.

Spiked centaury (Centaurium spicatum) is a well-known medicinal plant from the Mediterranean region with various bioactivities, but there are no studies addressing the use of different solvent systems to improve its pharmacological potential. Nine extraction procedures were adapted to study the effects of solvent composition on the content of bioactive compounds in C. spicatum extracts and on corresponding bioactivities. Targeted metabolomics was performed to obtain information on the chemical composition of extracts. Ethanol-water-based extraction procedures were the most efficient in isolating polyphenols, while less polar butanol extract contained the highest amount of iridoids. Antioxidant potential analysis revealed stronger activity in extracts with higher polyphenol content. Bacillus cereus and Staphylococus aureus were designated as the most sensitive bacterial strains to the activity of extracts, while among the micromycetes tested, Penicillium funiculosum was the most susceptible strain. Butanol extract showed antivirulence potential on Candida albicans morphological transition from yeast to hyphal form, and selected extracts were effective against biofilm formation in two Candida species. All the extracts tested in this study showed no cytotoxic activity to immortalize human skin keratinocyte cell line (HaCaT), whereas extracts obtained by ethanol-water extraction stand out for their potent wound healing effects. Moreover, the influence of the extraction solvent system on various bioactivities of C. spicatum is reported herein for the first time. Overall, the results presented in this study promote the use of C. spicatum as a source of natural products with potential antioxidant, wound healing, and antimicrobial applications that are potentially safe for human use.

Integration of agricultural residues as biomass source to saccharification bioprocess and for the production of cellulases from filamentous fungi.

The production of second-generation bioethanol has several challenges, among them finding cheap and efficient enzymes for a sustainable process. In this work, we analyzed two native fungi, Cladosporium cladosporioides and Penicillium funiculosum, as a source of cellulolytic enzyme production, and corn stover, wheat bran, chickpeas, and bean straw as a carbon source in two fermentation systems: submerged and solid fermentation. Corn stover was selected for cellulase production in both fermentation systems, because we found the highest enzymatic activities when carboxymethyl cellulase activity (CMCase) was assessed using CMC as substrate. C. cladosporioides showed the highest CMCase activity (1.6U/mL), while P. funiculosum had the highest filter paper activity (Fpase) (0.39U/mL). The ß-glucosidase activities produced by both fungi were similar in submerged fermentation using corn stover as substrate. Through in-gel zymography, three polypeptides with cellulolytic activities were identified in each fungus: with molecular weights of ~ 38, 45 and 70kDa in C. cladosporioides and ~ 21, 63 and 100kDa in P. funiculosum. The best results for saccharification (10.11g/L of reducing sugars) of diluted acid pretreated corn stover were obtained after 36h of the hydrolytic process at pH 5 and 50°C using the enzyme extract of P. funiculosum. This is the first report of cellulase identification in C. cladosporioides and the saccharification of corn stover using enzymes of this fungus. Enzymatic extracts of C. cladosporioides and P. funiculosum obtained from low-cost lignocellulosic biomass have great potential for use in the production of second-generation bioethanol.

Application of textile (PAN-based) activated carbon fibers decorated with silver nanoparticles in water treatment

Introduction: Water treatment deficit and poor health, hygiene and sanitation infrastructure can contribute to disease transmission by dissemination of contaminants and microorganisms. As an alternative, carbon-based materials coated with antimicrobial molecules have been proposed for water treatment, but few supporting data are available so far. Hence, this study investigates the potential use of PAN-based activated carbon fibers (ACF) decorated with silver nanoparticles in water treatment.Methods: Silver nanoparticles were incorporated into the material using a cheap and electroless method. Field emission scanning electron microscopy (FEGSEM), Raman spectroscopy and X-ray photoelectron spectroscopy (XPS) characterized the whole material. The textile was mounted on a water filter prototype and had its capacity to remove bacterial (Escherichia coli) and fungal (Candida albicans, Aspergillus niger and Penicillium funiculosum) cells evaluated. Composition and toxicity of the filtered water were determined.Results: Water filtered by Ag@ACF for 2 and 24 h contained 0.254 mg/L and 0.964 mg/L Ag, respectively. Ag@ACF filtering successfully removed E. coli, C. albicans, and A. niger from the suspensions, but not P. funiculosum. Treated water was non-toxic for Vero cells and Drosophila melanogaster, but toxic for Raphidocelis subcapitata. Ag@ACF showed efficient microbial elimination when applied in water treatment. Silver nanoparticles released in aqueous medium may be responsible for R. subcapitata toxicity. Future studies should be conducted to reduce silver nanoparticles release from the carbon fiber.

Antibiotic Producing Fungi in Sewage: Inhibitory Effect on 4 Bacterial Test Strains, and Different Fungal Types

Fungi were isolated from raw sewage and sewage that had trickled down soil columns from a waste water treatment plant in Karlsruhe, Germany, using the laboratory techniques.Fusarium sporotrichioides Sherb, Penicillium funiculosum, and Trichoderma harzianum Rafai were named as isolates from raw sewage. P. notatum Westling, P. meleagrinum Biourge, Aspergillus flavus, Link ex Gray, A. repens, A. fumigatus Fresenius, and A. fischeri Wehmer were among the fungi found in the effluent of the soil columns that were isolated under absolutely anaerobic conditions. Fusarium poae (Peck) Wollenw. and Penicillium chrysogenum Thom. were isolated when samples were cultured in anaerobic jars with nitrate. The coloration, smell, and other fungal traits, such as conidial and conidiophore size, etc., were used to identify the organism. The fungi's antibiotic properties against bacteria were investigated. A little amount of the fungal mycelium was plate-plated on new Nutrient agar and Sabouraud agar after isolation. After two days of incubation, bacteria were cross-streaked toward the fungal colonies on the plates. On the plates, six strains of E. coli, Gram-negative Pseudomonas species, and aerobic Gram-positive Enterococci species were streaked in the direction of the fungi. After that, the plates were incubated in an aerobic environment. Similar to this, five anaerobic Gram-positive Bifidobacterium species strains that were isolated from sewage were cross streaked on fungal plates and then further cultured under anaerobic conditions. When compared to A. flavus and A. fumigatus, P. chrysogenum and A. repens were more effective at inhibiting Enterococci. Only P. chrysogenum, A. fumigatus, and A. repens inhibited Pseudomonas sp. A. fumigatus and P. chrysogenum var meleagrinum only little inhibited E. coli, but P. chrysogenum and A. flavus very successfully did so.Pre-growing P. chrysogenum, P. notatum, and P. meleagrinum on plates for two days in an aerobic environment was followed by cross-stripping with test strains of Bifidobacterium and incubation under strictly anaerobic conditions. Both P. chrysogenum and P. notatum had the greatest inhibitory effects on bifidobacterium.

Functioning of Mitochondria in Transgenic Potato Tubers with the gox Gene for Glucose Oxidase from Penicillium funiculosum during Different Storage Periods

Functioning of Mitochondria in Transgenic Potato Tubers with the gox Gene for Glucose Oxidase from Penicillium funiculosum during Different Storage Periods

Design of Nanohydroxyapatite/Pectin Composite from Opuntia Ficus-Indica Cladodes for the Management of Microbial Infections.

Hydroxyapatite (HAp) attracts interest as a biomaterial for use in bone substitution or allografts. In the current work, biomaterial nanocomposites based on HAp and pectin were synthesized by using the double decomposition method, which involved using pectin extracted from fresh cladodes of the prickly pear, Opuntia ficus-indica. The crystallinity, purity, and several analytical techniques like Fourier transform infrared spectroscopy, X-ray diffraction, and scanning electron microscopy were used to understand the surface’s shape. The results revealed that the produced HAp/pectin nanoparticles are pure, spherical, and amorphous. The spectroscopic data indicated a substantial interaction between HAp and pectin, specifically between Ca (II) and pectin hydroxyl and carboxyl groups. The presence of pectin showed a noticeable influence on the prepared nanocomposite texture and porosity. We further assess the antibacterial and antifungal activity of the developed nanocomposite against a number of pathogenic bacteria and fungi, evaluated by the well diffusion method. In the absence of pectin, the XRD analysis revealed that the HAp nanoparticles had 10.93% crystallinity. When the pectin concentration reached 10 wt.%, it was reduced to approximately 7.29%. All synthesized nanocomposites demonstrated strong antimicrobial activity against both Gram-positive (Staphylococcus aureus and Bacillus cereus) and Gram-negative (Escherichia coli and Pseudomonas aeruginosa) bacteria in addition to various fungi (e.g., Aspergillus fumigatus, Penicillium funiculosum, and Trichoderma viride). This study endorses the HAp/Pectin nanocomposite as an efficient antimicrobial material for biomedical advanced applications.

Characterization of a GH5 endoxylanase from Penicillium funiculosum and its synergism with GH16 endo-1,3(4)-glucanase in saccharification of sugarcane bagasse

The production of second-generation fuels from lignocellulosic residues such as sugarcane bagasse (SCB) requires the synergistic interaction of key cellulose-degrading enzymes and accessory proteins for their complete deconstruction to useful monomeric sugars. Here, we recombinantly expressed and characterized unknown GH5 xylanase from P. funiculosum (PfXyn5) in Pichia pastoris, which was earlier found in our study to be highly implicated in SCB saccharification. The PfXyn5 has a molecular mass of ~ 55 kDa and showed broad activity against a range of substrates like xylan, xyloglucan, laminarin and p-nitrophenyl-β-d-xylopyranoside, with the highest specific activity of 0.7 U/mg against xylan at pH 4.5 and 50 °C. Analysis of the degradation products of xylan and SCB by PfXyn5 showed significant production of xylooligosaccharides (XOS) with a degree of polymerization (DP) ranging from two (DP2) to six (DP6), thus, suggesting that the PfXyn5 is an endo-acting enzyme. The enzyme synergistically improved the saccharification of SCB when combined with the crude cellulase cocktail of P. funiculosum with a degree of synergism up to 1.32. The PfXyn5 was further expressed individually and simultaneously with a notable GH16 endoglucanase (PfEgl16) in a catabolite-derepressed strain of P. funiculosum, PfMig188, and the saccharification efficiency of the secretomes from the resulting transformants were investigated on SCB. The secretome of PfMig188 overexpressing Xyn5 or Egl16 increased the saccharification of SCB by 9% or 7%, respectively, over the secretome of PfMig188, while the secretome of dual transformant increased SCB saccharification by ~ 15% at the same minimal protein concentration.

Recombinant Family 1 Carbohydrate-Binding Modules Derived From Fungal Cellulase Enhance Enzymatic Degradation of Lignocellulose as Novel Effective Accessory Protein.

Fungal cellulases usually contain a family 1 carbohydrate-binding module (CBM1), and its role was considered to recognize the substrate specifically. This study testified that the CBM1s derived from cellobiohydrolase I of Trichoderma reesei, Penicillium oxalicum, and Penicillium funiculosum could be used as an effective accessory protein in cellulase cocktails to enhance the saccharification of lignocellulose, and its enhancement effect was significantly superior to some reported accessory proteins, such as bovine serum albumin (BSA). The promoting effects of the CBM1s were related to not only the CBM1 sources and protein dosages, but also the substrate characteristics and solid consistency during enzymatic hydrolysis. The adsorption capacity of the CBM1s, the adsorption kinetic of TrCBM from T. reesei and cellobiohydrolase, endoglucanase, and β-glucosidase from P. oxalicum, and the effect of adding TrCBM on enzyme activities of free cellulases in the hydrolysis system were investigated, and the binding conformations and affinities of CBM1s to cellulose and lignin were predicted by molecular docking. It was speculated that the higher affinity of the CBM1s to lignin than cellulases could potentially enable the CBM1s to displace cellulase adsorbed on lignin or to preferentially adsorb onto lignin to avoid ineffective adsorption of cellulase onto lignin, which enhanced cellulase system efficiency during enzymatic hydrolysis of lignocellulose.

Influence of the high infectious load of the causal agent of bacterial ring rot of potatoes on variations in hydrogen peroxide and cAMP levels in transgenic potato plants <i>in vitro</i> with increased contents of endogenous hydrogen peroxide

Effects of the infectious load of the causal agent Clavibacter michiganensis sps. Sepedonicus (Cms) of ring rot of potatoes on hydrogen peroxide and cAMP levels in potato plants in vitro were studied on three transgenic lines of the Scarb cultivar: line L17.2 – plants transformed with the native gene of glucose oxidase gox from the highly active fungal strain Penicillium funiculosum 46.1; line M7.3 – plants transformed with the modified gene gox-mod; and line Pb14.10 – plants transformed with a vector without a target gene. In addition, non-transgenic plants of the same cultivar (medium resistant against Cms), Lugovskaya (resistant against Cms) and Lukyanovsky (susceptible to Cms) were examined. In the plants infected with Cms (0.2х10 8 cells/ml), the dynamics of cAMP and Н2О2 indicated a balanced system signal “root–stem”. The Cms titre (2х10 8 cells/ml) caused a systemic increase in the concentration of Н 2 О 2 in the organs of almost all plant cultivars; however, a systemic increase in the level of cAMP occurred only in the Lugovskaya cultivar. In all the studied transgenic lines, cAMP levels remained at the control level, accompanied by both local and extensive necrosis. In transgenic potato lines, the observed decrease in protective responses (both local and extensive necrosis) is likely to be associated with an extremely high baseline level of endogenous hydrogen peroxide and an imbalance in cAMP levels. The introduction of the gox gene into potato plants was found to increase their resistance against high Cms titres.