Polygalacturonase Enzyme Research Articles

In-silico study of E169G and F242K double mutations in leucine-rich repeats (LRR) polygalacturonase inhibiting protein (PGIP) of Gossypium barbadense and associated defense mechanism against plant pathogens

BackgroundPolygalacturonase inhibiting proteins (PGIPs) play a pivotal role in plant defense against plant pathogens by inhibiting polygalacturonase (PG), an enzyme produced by pathogens to degrade plant cell wall pectin. PGIPs, also known as leucine-rich repeat pathogenesis-related (PR) proteins, activate the host’s defense response upon interaction with PG, thereby reinforcing the host defense against plant pathogens attacks. In Egyptian or extra-long staple cotton (Gossypium barbadense), the interaction between PGIP and PG is one of the crucial steps in the defense mechanism against major pathogens such as Xanthomonas citri pv. malvacearum and Alternaria macrospora, which are responsible for bacterial leaf blight and leaf spot diseases, respectively.ResultsTo unravel the molecular mechanisms underlying these PR proteins, we conducted a comprehensive study involving molecular modeling, protein-protein docking, site-specific double mutation (E169G and F242K), and molecular dynamics simulations. Both wild-type and mutated cotton PGIPs were examined in the interaction with the PG enzyme of a bacterial and fungal pathogen. Our findings revealed that changes in conformations of double-mutated residues in the active site of PGIP lead to the inhibition of PG binding. The molecular dynamics simulation studies provide insights into the dynamic behaviour and stability of the PGIP-PG complexes, shedding light on the intricate details of the inhibitory and exhibitory mechanism against the major fungal and bacterial pathogens of G. barbadense, respectively.ConclusionsThe findings of this study not only enhance our understanding of the molecular interactions between PGs of Xanthomonas citri pv. malvacearum and Alternaria macrospora and PGIP of G. barbadense but also present a potential strategy for developing the disease-resistant cotton varieties. By variations in the binding affinities of PGs through specific mutations in PGIP, this research offers promising avenues for the development of enhanced resistance to cotton plants against bacterial leaf blight and leaf spot diseases.

Quorum Quenching Lactonase Alters Virulence of Pectobacterium atrosepticum and Reduces Maceration in Potatoes.

Pectobacterium atrosepticum is a soft rot phytopathogenic bacterium mainly infecting potatoes. The virulence of P. atrosepticum is controlled by quorum sensing (QS), a communication mechanism which enables bacteria to coordinate their behavior in a population density-dependent manner. Inhibiting QS has gained interest as a sustainable alternative to conventional treatments to control pathogens in agriculture. Here, we investigate the potential of a robust lactonase to inhibit P. atrosepticum virulence in vitro, combining phenotypic and proteomic studies. We report that exogenous lactonase treatment reduced the secretion of pectate lyase, cellulase, and polygalacturonase enzymes, leading to decreased virulence on potato tubers. Major changes in the P. atrosepticum proteome revealed that lactonase affects the abundance of proteins with various functions, including virulence. Taken as a whole, these results suggest that lactonase-mediated disruption of QS in P. atrosepticum is a promising strategy to limit P. atrosepticum infections.

Postharvest application of sodium nitroprusside improves the quality of ‘Umran’ ber fruit under cold storage conditions by regulating enzyme activities

ABSTRACT Ber fruit is highly perishable, with a storage life of only 3–5 days under ambient conditions, which makes it difficult to transport the fruit even to domestic markets. An experiment was conducted to study the effect of postharvest treatment with sodium nitroprusside (50, 100 and 150 µM) on the storage life, quality attributes, bioactive composition and antioxidant enzyme contents of ‘Umran’ ber fruit. The treated fruit were stored for 28 days to investigate the effects of sodium nitroprusside after 7 days of cold storage at 7.5°C and 90–95% RH. It was found that fruit treated with 100 μM sodium nitroprusside experienced lower fruit weight loss and spoilage, maintained fruit firmness, and had a higher sensory quality, phenolic content, carotenoid content, soluble solids content, sugar content, ascorbic acid and antioxidant activity than the control. Furthermore, the activities of the cell wall degrading enzymes polygalacturonase, polyphenol oxidase, pectin methyl esterase and catalase were lower, while the activities of superoxide dismutase and peroxidase enzymes were higher in the treated fruit. Therefore, treatment with 100 μM sodium nitroprusside shows high potential for improving the storage life and maintaining the quality attributes of ber fruit for up to 21 days under cold storage.

Isolation and characterization of polygalacturonase producing thermophilic Aspergillus niger isolated from decayed tomato fruits

This study aimed to isolate a fungal strain capable of producing acidophilic and thermostable polygalacturonase. In this study, the fungal isolate was isolated from decaying tomatoes. Based on the colony characteristics, microscopic and morphological observations, the isolated fungal pathogen has been identified as Aspergillus niger. The isolated fungus was used in solid-state fermentation to produce an acidic polygalacturonase enzyme. The enzyme was then purified using ammonium sulphate precipitation and column chromatography, and its activity was assayed by measuring the releasing sugar group from citrus pectin using a 3, 5-dinitrosalicylic acid (DNSA) reagent assay. The crude extract obtained from solid-state fermentation had an activity of 94.6 U/mL. Ammonium sulphate precipitation increased the enzyme's specific activity from 6.89 U/mg to 12.42 U/mg. Sephadex G-200 was used to purify the enzyme 3.58 times, and its specific activity was determined to be 24.66 U/mg. The Sephacryl S-100 column achieved a final fold purification of 9.93 times and a specific activity of 68.41 U/mg. The purified enzyme performed best when polygalacturonic acid was used as a substrate. The enzyme's optimum temperature and pH were 55°C and 5, respectively. CaCl2 was found to be the best chelating ion for the enzyme. This enzyme is recommended for use in a variety of industrial applications as the enzyme was found to be stable at acidic pH and high temperature.

Genome-wide analysis of the PG gene family in Penicillium expansum: Expression during the infection stage in pear fruits (Pyrus bretschneideri)

Penicillium expansum is a necrotrophic plant pathogen that has a broad range of fruit hosts and is responsible for blue mold rot. This rot has a significant impact on the fruit industry, causing economic losses during storage, transport, and sale. Polygalacturonase (PGs) enzymes are essential for pathogenic fungi to break down pectin, as they hydrolyze the polygalacturonic acid chain along the oxygen bridge. This enzyme has been identified in decayed tissue and is linked to multiple soft rot diseases. This study encompasses a thorough examination of the PGs gene family within the genome of P. expansum, utilizing a genome-wide analysis approach. As a result, a total of seven PG genes were successfully found. These genes were phylogenetically divided into four distinct clades. The gene structural characteristics of each group exhibited a fundamental level of conservation. Furthermore, the gene architectures and motif compositions exhibited robust evidence in favor of the credibility of the phylogenetic relationship. Motif analysis reveals that most of the motifs predicted had Glycosyl hydrolases family 28 conserved domains (PF00295). Analysis of RNA-seq data and the PG genes identified by RT-qPCR were mostly upregulated. Hence, comprehending the characteristics of PePGs holds significant scientific importance and will offer comprehensive insights into the PG superfamily in P. expansum and will make a valuable contribution to the ongoing validation of functional genes.

Polygalacturonase inhibiting protein enhances cell wall strength of strawberry fruit for resistance to Botrytis cinerea infection

Polygalacturonase inhibitor protein (PGIP) plays an active role in resisting pathogens from degrading cell walls. Strawberry fruit quality is strongly affected by Botrytis cinerea (B. cinerea), so PGIP genes from heterologous species were cloned and analyzed to identify genes having good B.cinerea infection resistance. In this study, a PGIP gene from grape fruit was isolated and heterologously transiently expressed in strawberry fruit. The results revealed that VvPGIP (PGIPO) overexpression in strawberry fruit led to B. cinerea infection resistance, but RNA interference of VvPGIP (PGIPI) made fruit susceptible to B. cinerea infection. qPCR indicated that expression levels of FaPGIP was increased after transient expression of VvPGIP in PGIPO fruit for 48 h as compared to the control and that polygalacturonase enzyme (PG) was significantly down-regulated, whereas PGIPI fruit had the opposite effect, suggesting that VvPGIP may activate the FaPGIP expression levels, thereby reducing the disease incidence and index of strawberry fruits and improving the disease resistance of the fruits. VvPGIP also inhibited the decrease of total pectin and cellulose content. Transcriptome and metabolome results showed that PGIP is widely involved in photosynthesis, signal transduction, and secondary metabolism of fruits. In addition, many enzyme activities and defense reaction were also significantly enriched. Using Y2H and BiFC technology, we screened and verified the interaction protein of VvPGIP with Endo-1, 3-Beta-Glucanase (VvGLU) and pathogen-related protein 4 (VvPR-4) on the cell membrane. Taken together, PGIP as a key protein in plant defense response functions a key role in regulating the strength of cell walls and can inhibit the degradation of plant cell walls by pathogens.

Overexpression of the Rubus idaeus Polygalacturonases Gene RiPG2 Accelerates Fruit Softening in Solanum lycopersicum

The high susceptibility of raspberries to softening restricts the development of the raspberry industry. The primary causes of fruit softening are the breakdown of components linked to the cell wall and the destruction of the cell wall structure itself. Polygalacturonase (PG), a key enzyme that catalyzes pectin degradation, plays a critical role in fruit softening. However, there are currently limited studies on the mechanism of PG genes in raspberry fruit softening. In this study, a PG gene, RiPG2, was isolated from raspberry (Rubus idaeus L.). ‘Polka’ fruits and tomato plants overexpressing RiPG2 were obtained by Agrobacterium tumefaciens-mediated leaf disc transformation to elucidate the role of RiPG2 in fruit softening. The total length of the RiPG2 gene is 1185 bp, and the gene encodes a total of 394 amino acids. The GFP fusion protein was expressed at the chloroplast under laser confocal microscopy, indicating that the RiPG2 protein is localized to the chloroplasts. Phenotypic analysis revealed that the fruit firmness of three strains was considerably less than that of controls, but PG enzyme activity was increased. Overexpression of RiPG2 altered the content of cell wall components, with an increase in water-soluble pectin (WSP) and ion-bound pectin (ISP) but a decrease in protopectin, cellulose, hemicellulose, and covalently bound pectin (CSP). In addition, RiPG2 positively regulated the expression of cell wall metabolism-related genes such as SlEXP1, SlTBG4, SlXTH5, and SlPL. These results suggest that the RiPG2 gene regulates the structure and composition of the cell wall and acts synergistically with other cell wall metabolism-related genes to promote fruit softening. This study provides a new candidate gene for molecular breeding to improve raspberry firmness.

Maturation and antioxidant activity of 'Giombo' and 'Rama Forte' persimmons produced in the Brazilian semiarid.

The objective of this work was characterizing persimmons of the 'Giombo' and 'Rama Forte' cultivars harvested at different ripening stages in the Brazilian semiarid. Fruits were harvested at three ripening stages - green, semi-ripe and ripe - then evaluated for the following characteristics: fruit weight and diameter, skin and pulp color, fruit firmness, pulp pH, soluble solids content, titratable acidity, SSC/TA ratio, total soluble sugars, reducing sugars, astringency index, and the contents of tannin, vitamin C, carotenoid, β-carotene, and total extractable polyphenols. Also, total antioxidant activity by the DPPH and ABTS methods and pectin methylesterase, and polygalacturonase enzyme activities were evaluated. Two experiments were carried out in a completely randomized design, one for each cultivar, with treatments consisting of different stages of maturation, with five replications of three fruits each. Data were submitted to analysis of variance and the differences between the means were compared using the Tukey test at 5% probability. Fruit firmness and soluble solids content did not vary between maturation stages for any of the cultivars. However, the skin color index increased with advancing maturation for both 'Giombo' and 'Rama Forte'. The astringency index, the content of total extractable polyphenols, soluble tannins and the antioxidant capacity were lower in fruits harvested at the ripe stage, for both cultivars. It can be concluded that persimmons of the 'Giombo' and 'Rama Forte' cultivars present better physicochemical quality characteristics when harvested when ripe, with a totally yellow skin.

Combined In Silico and In Vitro Study to Reveal the Structural Insights and Nucleotide-Binding Ability of the Transcriptional Regulator PehR from the Phytopathogen Ralstonia solanacearum.

The transcriptional regulator PehR regulates the synthesis of the extracellular plant cell wall-degrading enzyme polygalacturonase, which is essential in the bacterial wilt of plants caused by one of the most devastating plant phytopathogens, Ralstonia solanacearum. The bacterium has a wide global distribution infecting many different plant species, resulting in massive agricultural and economic losses. Because the PehR molecular structure has not yet been determined and the structural consequences of PehR on ligand binding have not been thoroughly investigated, we have used an in silico approach combined with in vitro experiments for the first time to characterize the PehR regulator from a local isolate (Tezpur, Assam, India) of the phytopathogenic bacterium R. solanacearum F1C1. In this study, an in silico approach was employed to model the 3D structure of the PehR regulator, followed by the binding analysis of different ligands against this regulatory protein. Molecular docking studies suggest that ATP has the highest binding affinity for the PehR regulator. By using molecular dynamics (MD) simulation analysis, involving root-mean-square deviation, root-mean-square fluctuations, hydrogen bonding, radius of gyration, solvent-accessible surface area, and principal component analysis, it was possible to confirm the sudden conformational changes of the PehR regulator caused by the presence of ATP. We used an in vitro approach to further validate the formation of the PehR-ATP complex. In this approach, recombinant DNA technology was used to clone, express, and purify the gene encoding the PehR regulator from R. solanacearum F1C1. Purified PehR was used in ATP-binding experiments using fluorescence spectroscopy and Fourier transform infrared spectroscopy, the outcomes of which showed a potent binding to ATP. The putative PehR-ATP-binding analysis revealed the importance of the amino acids Lys190, Glu191, Arg192, Arg375, and Asp378 for the ATP-binding process, but further study is required to confirm this. It will be simpler to comprehend the catalytic mechanisms of a crucial PehR regulator process in R. solanacearum with the aid of the ATP-binding process hints provided by these structural biology applications.

Validation of Pasteurisation Temperatures for a Tomato-Oil Homogenate (salmorejo) Processed by Radiofrequency or Conventional Continuous Heating.

Salmorejo is a viscous homogenate based on tomato, olive oil and breadcrumbs commercialised as a "fresh-like" pasteurised-chilled purée. Due to its penetration, dielectric heating by radiofrequency (RF) might improve pasteurisation results of conventional heating (CH). The objective was to validate the pasteurisation temperature (70-100 °C, at 5 °C intervals) for salmorejo processed by RF (operating at 27.12 MHz for 9.08 s) or conventional (for 10.9 s) continuous heating. The main heat-induced changes include: orangeness, flavour homogenisation, loss of freshness, thickening, loss of vitamin C and lipid oxidation. Both CH and RF equivalent treatments allowed a strong reduction of total and sporulated mesophilic microorganisms and an adequate inhibition of the pectin methylesterase, peroxidase and, to a lesser extent, polyphenol oxidase but did not inhibit the polygalacturonase enzyme. Pasteurisation at 80 °C provided a good equilibrium in levels of microbiological and enzymatic inhibition and thermal damage to the product. Increasing this temperature does not improve enzyme inactivation levels and salmorejo may become overheated. A "fresh-like" good-quality salmorejo can be obtained using either conventional or radiofrequency pasteurisers.

Anti-corrosive and anti-microbial activity of MTMI on CRCA metal

The present study was intended to demonstrate the synthesis, and characterization of 1-(3-methoxyphenyl)-N-(4H-1,2,4-triazol-4-yl)methan-imine(MTMI) as an inhibitor for the corrosion on CRCA in 1 M HCl. In addition to this, molecular docking, and biological activity was carried out. MTMI formation was confirmed by FTIR and NMR spectroscopic studies. The corrosion inhibition effect was studied by weight loss, potentiodynamic polarization (PDP) measurements, and electrochemical impedance spectroscopy (EIS) techniques. The conditions for optimum inhibition efficiency were established by varying the concentration of the inhibitor and temperature by weight loss method. Kinetic and thermodynamic parameters were calculated and discussed. Results were fitted into various adsorption isotherm models. The adsorption of MTMI on CRCA was confirmed by scanning electron microscopy (SEM). Adsorption obeyed the Langmuir isotherm model and followed physical adsorption. Increasing inhibitor concentration led to a significant reduction in the corrosion rate of mild steel, with an inhibitor efficiency value above 93%. A molecular docking study showed the binding capacity of the protein endo-polygalacturonase (1HG8) and the activity of polygalacturonase enzyme on MTMI was measured.

Changes in Pectic Substances and Cell Wall Degrading Enzymes During Muskmelon Fruit Ripening

Changes in pectic substances and the activities of the cell walldegrading enzymes, pectin esterase (PE) and polygalacturonase (PG) wereevaluated during muskmelon fruits ripening. Respiration rateprogressively increased in a climacteric pattern. Fruit flesh firmnesssteadily declined and rind yellow colour increased during ripening. Totaland insoluble pectins progressively decreased, while the soluble pectinsincreased with fruit ripening. Positive correlation coefficients wereobtained between total and insoluble pectins, and fruit flesh firmness.Total protein and PE activity increased up to the climacteric peak ofrespiration and decreased afterwards. PG activity progressively increasedduring fruit ripening, with high correlation between the increase inenzyme activity and fruit softening in the three muskmelon cultivars. Thismay indicate that the two enzymes polygalacturonase (PG) and pectinesterase (PE) are the main enzymes responsible for fruit softening duringmuskmelon fruit ripening.

Phytic acid is a new substitutable plant-derived antifungal agent for the seedling blight of Pinus sylvestris var. mongolica caused by Fusarium oxysporum

Phytic acid (PA) is a new substitutable plant-derived antifungal agent; however, few reports have been published regarding its antifungal effects on pathogenic fungi. The present study explored the in vitro antifungal activity of PA against four phytopathogenic fungi and found that PA was the most effective at inhibiting the growth of Fusarium oxysporum. This study aimed to investigate the in vivo and in vitro antifungal activities of PA against the seedling blight of Pinus sylvestris var. mongolica caused by F. oxysporum and to determine its possible mechanism of action. The results showed that PA inhibited spore germination and mycelial growth of F. oxysporum in a concentration-dependent manner and exhibited strong inhibition when its concentration exceeded 1000 mg/L. It mainly destroyed the integrity of the cell membrane, increasing its cell membrane permeability, causing the cell contents to spill out, and impairing fungal growth. In addition, the leakage of intercellular electrolytes and soluble proteins indicated that PA used at its EC20 and EC50 increased the membrane permeability of F. oxysporum. The increase in malondialdehyde and hydrogen peroxide content confirmed that PA treatment at its EC20 and EC50 damaged the cell membrane of the pathogen. Scanning electron microscopy revealed that PA affected the morphology of mycelia, causing them to shrivel, distort, and break. Furthermore, PA significantly reduced the activities of the antioxidant-related enzymes superoxide dismutase and catalase, as well as that of the pathogenicity-related enzymes polygalacturonase, pectin lyase, and endoglucanase (EG) in F. oxysporum (P < 0.05). In particular, EG enzyme activity was maximally inhibited in F. oxysporum treated with PA at its EC50. Moreover, PA significantly inhibited the incidence of disease, and growth indices in Pinus sylvestris var. mongolica seedling blight was determined. In summary, PA has a substantial inhibitory effect on F. oxysporum. Therefore, PA could serve as a new substitutable plant-derived antifungal agent for the seedling blight of P. sylvestris var. mongolica caused by F. oxysporum.

Effects of postharvest calcium treatmenttreatment on the firmness of persimmon (Diospyros kaki) fruit based on a decline in WSP

• Calcium treatment could maintain the firmness and color of persimmon fruit. • Calcium treatment delayed the decrease of molecular weight and branching rate of WSP. • Calcium treatment inhibited the increase of the activity of PG, PME and PL. Persimmon fruit is a typical respiratory climacteric fruit. Previous studies have found that water-soluble pectin (WSP) was closely related to fruit firmness, and calcium was involved in pectin bridging, which also affected fruit firmness. In this research, the effect of calcium treatment (2% CaCl 2 , 30 min) on persimmon fruit firmness was studied from the perspective of WSP. Firmness of persimmon fruit treated with calcium increased by 40% and the WSP content decreased by 14% after storage for 15 d Meanwhile, the activity of pectin degrading enzymes polygalacturonase (PG), pectin methylesterase (PME) and pectin lyase (PL) decreased by 39%, 24%, and 11%, respectively. Fruit firmness was negatively correlated with the WSP content and activity of pectin degrading enzymes. Therefore, the lower pectin degrading enzyme activity delayed the decrease of the WSP molecular weight and branching rate, and inhibited the increase of WSP content, thus maintaining the integrity of cell wall structure and higher firmness of persimmon fruit. Additionally, calcium treatment could delay the water migration in the cell wall and vacuole, further maintaining persimmon fruit firmness. The results of this research not only help reveal the mechanism of persimmon fruit softening, but also provide reference for the application of calcium in persimmon fruit preservation.

Transcriptomic and physiological analysis reveals the possible mechanism of ultrasound inhibiting strawberry (Fragaria × ananassa Duch.) postharvest softening.

Ultrasound effectively inhibited strawberry softening but the mechanism was not clear. In this study, physical data including firmness, soluble pectin (SP) contents, pectin esterase (PE), polygalacturonase (PG) activity and transcriptome sequencing data were analyzed to explore the mechanism of strawberry response to ultrasonic treatment. After 24 days storage, the firmness reduction rate and soluble contents (SP) increased rate of the strawberry treated with ultrasound (25 kHz, 0.15 W/cm2) for 3 min decreased 41.70 and 63.12% compared with the control, respectively. While the PG and PE enzyme activities of ultrasound-treated strawberries were significantly lower than control after storage for 18 days. A total of 1,905 diferentially expressed genes (DEGs) were identified between ultrasound-treated and control, with 714 genes upregulated and 1,254 genes downregulated, including 56 genes in reactive oxygen species (ROS), auxin (AUX), ethylene (ETH) and jasmonic acid (JA) signaling pathways. At 0 h, 15 genes including LOX, JMT, ARP, SKP, SAUR, IAA, ARF, and LAX were significantly upregulated compared with the control group, which means reactive oxygen specie, auxin, ethylene and jasmonic acid-mediated signaling pathway respond to ultrasound immediately. ERF109, ERF110, and ACS1_2_6 downregulated before 2 days storage indicated ethylene signaling pathway was inhibited, while after 2 days, 9 genes including ERF027, ERF109, and ERF110 were significantly upregulated indicating that the response of the ethylene signaling pathway was lagging. Therefore, in strawberry ultrasound enhanced ROS scavenging and activated JA biosynthesis, which acts as a signal for delaying the activation of ET signaling pathway, thus suppressing the activity of pectin-degrading enzymes PE and PG, and ultimately inhibiting postharvest softening.

Inhibitory effect and mechanism of action of juniper essential oil on gray mold in cherry tomatoes.

Juniper essential oil (JEO), which is mostly known as an immune system booster and effective detoxifier, has substantial antimicrobial activity. A comparison of the inhibitory effects of three plant essential oils from juniper (Juniperus rigida), cedarwood (Juniperus virginiana), and cypress (Crupressus sempervirens) on four plant pathogenic fungi indicated that JEO was the most effective at inhibiting the growth of gray mold (Botrytis cinerea). Additional studies were subsequently conducted to explore the in vivo and in vitro antifungal activity and possible mechanism of JEO against B. cinerea. The results show that JEO inhibited the germination of spores and mycelial growth of B. cinerea in a concentration-dependent manner and exhibited strong inhibition when its concentration exceeded 10 μL/mL. JEO also significantly inhibited the incidence of disease and diameters of gray mold lesions on cherry tomato fruit (Solanum lycopersicum). After 12 h of treatment with JEO, the extracellular conductivity, and the contents of soluble protein, malondialdehyde, and hydrogen peroxide were 3.1, 1.2, 7.2, and 4.7 folds higher than those of the control group, respectively (P < 0.05), which indicated that JEO can damage membranes. Scanning electron microscopy observations revealed that JEO affected the morphology of mycelia, causing them to shrivel, twist and distort. Furthermore, JEO significantly improved the activities of the antioxidant-related enzymes superoxide dismutase and catalase but reduced the pathogenicity-related enzymes polygalacturonase (PG), pectin lyase and endoglucanase of B. cinerea (P < 0.05). In particular, PG was reduced by 93% after treatment with JEO for 12 h. Moreover, the 18 constituents of JEO were identified by gas chromatography/mass spectrometry (GC-MS) analysis, mainly limonene (15.17%), γ-terpinene (8.3%), β-myrcene (4.56%), terpinen-4-ol (24.26%), linalool (8.73%), α-terpineol (1.03%), o-cymene (8.35%) and other substances with antimicrobial activity. Therefore, JEO can be an effective alternative to prevent and control gray mold on cherry tomato fruit.

Toxicity, microbiological parameters and amino acid composition of the organic feed additive Grinat

Recently, the problem of increasing the productivity of farm animals is one of the most important for veterinary medicine. Since the animals “programmed” for maximum productivity by selection turned out to be excessively exposed to anthropogenic and biological negative factors, the result of this is a decrease in their productivity. Conducted toxicological, microbiological, mycological and chromatographic studies of the organic feed additive ˮGreenat“, which includes humic acids. It is promising to study the possibility of using preparations containing humic substances to increase the productivity of farm animals and poultry, to enhance the general nonspecific resistance of the body. Given that peat is the raw material for the manufacture of the additive, toxicity, mycological and microbiological indicators are extremely important. Toxicological studies (bioassays on rabbits and Colpoda steinii culture) proved the absence of toxic substances, as evidenced by the activity of the colpoda infusoria for 10 minutes and 3 hours of the study, and the absence of hyperemia and inflammation on the skin of the rabbit. The number of mold fungi does not exceed the allowable limits of 1.5x102 CFU/g (according to the norm 5.0x104), qualitative studies revealed and identified by mass spectrometry the mold fungi A. alternata; C. cladosporioides, which belong to saprophytes and get into the organic feed additive "Grinat" from the remains of plants during their decomposition. Microscopic fungi Alternaria spp. participate in the decomposition and mineralization of plant residues due to the polygalacturonase enzyme complex, which allows them to attach themselves and exist in various conditions. Cladosporium spp. found outdoors and indoors, in degraded organic debris, produce secondary metabolites such as antibiotics that are inhibitors of B. subtilis, E. coli, and C. albicans. Microbiologically, Maldi Tof mass spectrometry detected Bacillus megaterium, which belongs to soil bacteria, and Staphylococcus hominis, found as a harmless commensal on the skin of humans and animals. Chromatographically confirmed the presence of essential amino acids: leucine (0.120%), valine (0.089%), threonine (0.064%), phenylalanine (0.039%), isoleucine (0.024%), histidine (0.021), methionine (0.011%), lysine (0.05%), tryptophan (0.007%). Accordingly, conditionally replaceable and non-essential amino acids, which can help to avoid a shortage of feed components in animal diets. Key words: Greenate, Colpoda steinii, general toxicity, amino acids, molds, Maldi Tof, Bacillus megaterium, Staphylococcus hominis, A. alternata, C. cladosporioides.

Deciphering the role of Trichoderma sp. bioactives in combating the wilt causing cell wall degrading enzyme polygalacturonase produced by Fusarium oxysporum: An in-silico approach

The cell wall degrading enzymes polygalacturonase (PG) secreted by Fusarium oxysporum f. sp. radicis-lycopersici (FOL) is testified to trigger Fusarium crown and root rot disease in tomato crops; instigated due to the degradation of the pectin. Trichoderma sp. is documented as a potential biocontrol agent playing a pivotal role in plant health and disease management. An in-silico approach employing homology modelling, molecular docking, molecular dynamics (MD) simulation and MMPBSA was employed to assess the prospective role of bioactives produced by Trichoderma sp. in combating the PG2 enzyme. The studies revealed that amongst the wide range of bioactives screened, Trichodermamide B produced by T. harzianum and Viridin, Virone, and Trichosetin produced by T. virens emerged as the potential inhibitors of the PG2. Docking results revealed that the complexes possessed most stable energy for Trichodermamide B (−8.1 kcal/mol) followed by Viridin (−7.7 kcal/mol), Virone (−7.1 kcal/mol), and Trichosetin (−7 kcal/mol), respectively. Interaction studies of FOL with T. virens and T. harzianum reported an inhibition of 83.33% and 75.87%, respectively. The structural rigidity and stability of the docked complex was confirmed through MD simulations evaluated across multiple descriptors from the simulation trajectories. Further, MMPBSA analysis validated the results that binding of the enzyme to the screened ligands was spontaneous. The study unravels new insights on the versatile potential of Trichoderma sp. Bioactives as a prospective agent for the inhibition of cell-wall degrading enzymes secreted by phytopathogens. The proposed study can be implemented for design of bioformulations that serve the role of biopesticide, promising a sustainable alternate to chemical-based products.

Relationship of Polygalacturonase Enzyme Activity between Brown Rot Isolates and their Virulence on Potato Cultivars

Relationship of Polygalacturonase Enzyme Activity between Brown Rot Isolates and their Virulence on Potato Cultivars

Effects of procyanidin treatment on the ripening and softening of banana fruit during storage

Cell wall degradation accompanied with softening is very common in harvested fruit. To develop a facile postharvest approach, which can be used at ambient temperature, for suppressing softening and maintaining quality of banana fruit, influences of 1% procyanidin (PA) solution treatment on pulp firmness, total soluble solids (TSS) in pulp, peel hue value, chlorophyll content of peel, fruit ethylene production, fruit respiration rate, malondialdehyde (MDA) concentration in peel, peel ion leakage, activities and expression levels of enzymes related with fruit softening during storage at 23 ± 2°C were investigated. Results indicated that PA treatment effectively inhibited ripening-associated processes, as compared to the control banana fruit, PA-treated fruit exhibited higher firmness, hue value and chlorophyll content, lower TSS content, ethylene production, respiration rate, MDA concentration, and ion leakage. Besides, lower activities of cell wall-degrading enzymes polygalacturonase (PG), pectin methyl esterase (PME), pectate lyase (PL), and cellulase (CX) and expression levels of PG, PME, PL were also observed in PA-treated fruit. These results would provide theoretical basis to help solve fruit quality decline of banana fruit during storage in further research.