Optimizing biodegradable plastics: Molecular dynamics insights into starch plasticization with glycerol and oleic acid.

Petroleum-based plastics dominate everyday life, necessitating the exploration of natural polymers as alternatives. Starch, abundant and biodegradable, is a promising raw material. However, understanding the molecular mechanisms underlying starch plasticization has proven challenging. To address this, we employ molecular dynamics simulations, focusing on amylose as a model. Our comprehensive evaluation revealed that chain size affects solubility, temperature influenced diffusivity and elastic properties, and oleic acid expressed potential as an alternative plasticizer. Furthermore, blending glycerol or oleic acid with water suggested the enhancement amylose's elasticity. These findings contribute to the design of sustainable and improved biodegradable plastics.

In Vitro Evaluation of the Antifungal Effect of AgNPs on Fusarium oxysporum f. sp. lycopersici.

The application of nanomaterials in the agri-food industry can lead us to the formulation of new sustainable and effective pesticides for the control of fungi such as Fusarium oxysporum f. sp. lycopersici (Fol). This is a fungal plant pathogen for the tomato plant. In this work, silver nanoparticles (AgNPs) were synthesized by a green methodology from Geranium leaf extract as a reducing agent. The poisoned food technique was used to determine the percentage of inhibition of Fol mycelial growth by the action of AgNPs. They were characterized by transmission electron microscopy (TEM, JEOL JEM-2100, Tokyo, Japan) and ultraviolet-visible spectroscopy (UV-VIS, DU 730 Beckman Coulter, Brea, CA, USA). Five different concentrations of AgNPs (10, 20, 40, 75, and 150 mg/L) were evaluated in vitro in order to determine the minimum inhibitory concentration (MIC) as well as the behavior of their antifungal activity in tomato fruit. Nanoparticles with spherical morphology and average diameters of 38.5 ± 18.5 nm were obtained. The maximum percentage of inhibition on the mycelial growth of Fol was 94.6 ± 0.1%, which was obtained using the AgNPs concentration of 150 mg/L and it was determined that the MIC corresponds to 75 mg/L. On the other hand, in a qualitative way, it was possible to observe an external inhibitory effect in the tomato fruit from the concentration of 10 mg/L. Finally, we can conclude that AgNPs are a viable alternative for alternative formulations applied in the agri-food industry as pesticide solutions.

Role of the nonribosomal peptide synthetase siderophore enzyme (Rfs) of Mucor lusitanicus in controlling the growth of fungal phytopathogens.

Dimorphic species of Mucor, which are cosmopolitan fungi belonging to subphylum Mucoromycotina, are metabolically versatile. Some species of Mucor are sources of biotechnological products, such as biodiesel from Mucor circinelloides and expression of heterologous proteins from Mucor lusitanicus. Furthermore, Mucor lusitanicus has been described as a model for understanding mucormycosis infections. However, little is known regarding the relationship between Mucor lusitanicus and other soil inhabitants. In this study, we investigated the potential use of Mucor lusitanicus as a biocontrol agent against fungal phytopathogens, namely Fusarium oxysporum f. sp. lycopersici, Fusarium solani, and Alternaria solani, which destroy economically important crops. Results showed that aerobic cell-free supernatants of the culture broth (SS) from Mucor lusitanicus inhibited the growth of the fungal phytopathogens in culture, soil, and tomato fruits. The SS obtained from a strain of Mucor lusitanicus carrying the deletion of rfs gene, which encodes an enzyme involved in the synthesis of siderophore rhizoferrin, had a decreased inhibitory effect against the growth of the phytopathogens. Contrarily, this inhibitory effect was more evident with the SS from an rfs-overexpressing strain compared to the wild-type. This study provides a framework for the potential biotechnological use of the molecules secreted from Mucor lusitanicus in the biocontrol of fungal phytopathogens.

Expression of bik cluster and production of bikaverin by Fusarium oxysporum f. sp. lycopersici grown using two alternate nitrogen sources / Expresión de bik cluster y producción de bikaverina por Fusarium oxysporum f. sp. lycopersici cultivados utilizando dos fuentes alternativas de nitrógeno

The genus Fusarium can be utilized to produce a great variety of secondary metabolites under specific culture conditions, including pigments of increasing biotechnological interest, such as bikaverin. Such pigments are important due to the biological properties they possess, including antitumor and antibiotic activities, among others. In Fusarium fujikuroi, bik1–bik6 have been identified as the genes that are responsible for the synthesis of bikaverin. Therefore, in this study, we screened for the presence of bik genes and examined changes in mRNA levels of the bik genes under the influence of NH4NO3 (0.024, 0.048, 0.50, 1.0, and 4.60 g L−1) and NH4Cl (0.50 and 1.0 g L−1) as nitrogen sources for the phytopathogen Fusarium oxysporum f. sp. lycopersici. Our results indicated the presence of at least six bik (bik1–bik6) genes and showed increased mRNA levels for bik4, bik5, and bik6 in conditions where NH4NO3 was used at pH 3.0. The characteristic coloration of bikaverin was obtained in 10 out of 16 culture conditions, except when the fungus was grown with higher concentrations of NH4NO3 (1.0 and 4.60 g L−1). The pigment was chloroform-extracted from the culture conditions of NH4NO3 (0.024, 0.048, and 0.50 g L−1) and NH4Cl (0.50 and 1.0 g L−1) with 3 and 9 days of incubation. Analysis via visible spectroscopy and matrix-assisted laser desorption ionization-time of flight mass spectrometry were used for the identification of bikaverin.(AU)

Expression of bik cluster and production of bikaverin by Fusarium oxysporum f. sp. lycopersici grown using two alternate nitrogen sources.

The genus Fusarium can be utilized to produce a great variety of secondary metabolites under specific culture conditions, including pigments of increasing biotechnological interest, such as bikaverin. Such pigments are important due to the biological properties they possess, including antitumor and antibiotic activities, among others. In Fusarium fujikuroi, bik1-bik6 have been identified as the genes that are responsible for the synthesis of bikaverin. Therefore, in this study, we screened for the presence of bik genes and examined changes in mRNA levels of the bik genes under the influence of NH4NO3 (0.024, 0.048, 0.50, 1.0, and 4.60 g L-1) and NH4Cl (0.50 and 1.0 g L-1) as nitrogen sources for the phytopathogen Fusarium oxysporum f. sp. lycopersici. Our results indicated the presence of at least six bik (bik1-bik6) genes and showed increased mRNA levels for bik4, bik5, and bik6 in conditions where NH4NO3 was used at pH 3.0. The characteristic coloration of bikaverin was obtained in 10 out of 16 culture conditions, except when the fungus was grown with higher concentrations of NH4NO3 (1.0 and 4.60 g L-1). The pigment was chloroform-extracted from the culture conditions of NH4NO3 (0.024, 0.048, and 0.50 g L-1) and NH4Cl (0.50 and 1.0 g L-1) with 3 and 9 days of incubation. Analysis via visible spectroscopy and matrix-assisted laser desorption ionization-time of flight mass spectrometry were used for the identification of bikaverin.

Transcript pattern analysis of Arf-family genes in the phytopathogen Fusarium oxysporum f. sp. lycopersici reveals the role of Arl3 in the virulence.

Fusarium oxysporum f. sp. lycopersici is an important plant pathogen that has been used to understand the virulence mechanisms that soil inhabiting fungi exhibit during the infection process. In F. oxysporum many of the virulence factors are secreted, and the secretion process requires the formation of vesicles. Arf family members, represented by Arf (ADP- Ribosylation Factor), Arl (Arf-like), and Sar (Secretion-associated and Ras-related) proteins, are involved in the vesicle creation process. In this study we identified the Arf family members in F. oxysporum f. sp. lycopersici, which includes seven putative proteins: Arf1, Arf3, Arl1 through Arl3, Arl8B, and Sar1. Quantification of the mRNA levels of each arf encoding gene revealed that the highest expression corresponds to arf1 in all tested conditions. The phylogenetic analysis revealed that no other Arf1 paralogue, such as Arf2 from yeast, is present in F. oxysporum f. sp. lycopersici. The essential function suggested of Arf1 in F. oxysporum f. sp. lycopersici was corroborated experimentally when, after several attempts, it was impossible to obtain a knockout mutant in arf1. Moreover, arl3 mRNA levels increased significantly when plant tissue was added as a sole carbon source, suggesting that the product of these genes could play pivotal roles during plant infection, the corresponding mutant ∆arl3 was less virulent compared to the wild-type strain. These results describe the role of arl3 as a critical regulator of the virulence in F. oxysporum f. sp. lycopersici and stablish a framework for the arf family members to be studied in deeper details in this phytopathogen.

Influence of iron and copper on the activity of laccases in Fusarium oxysporum f. sp. lycopersici.

Fusarium oxysporum f. sp. lycopersici is a phytopathogenic fungus that causes vascular wilt in tomato plants. In this work we analyze the influence of metal salts such as iron and copper sulphate, as well as that of bathophenanthrolinedisulfonic acid (iron chelator) and bathocuproinedisulfonic acid (copper chelator) on the activity of laccases in the intra (IF) and extracellular fractions (EF) of the wild-type and the non-pathogenic mutant strain (rho1::hyg) of F. oxysporum. The results show that laccase activity in the IF fraction of the wild and mutant strain increased with the addition of iron chelator (53.4 and 114.32%; respectively). With copper, it is observed that there is an inhibition of the activity with the addition of CuSO4 for the EF of the wild and mutant strain (reduction of 82 and 62.6%; respectively) and for the IF of the mutant strain (54.8%). With the copper chelator a less laccase activity in the IF of the mutant strain was observed (reduction of 53.9%). The results obtained suggest a different regulation of intracellular laccases in the mutant strain compared with the wild type in presence of CuSO4 and copper chelator which may be due to the mutation in the rho gene.

Influence of iron and copper on the activity of laccases in Fusarium oxysporum f. sp. lycopersici

ABSTRACT Fusarium oxysporum f. sp. lycopersici is a phytopathogenic fungus that causes vascular wilt in tomato plants. In this work we analyze the influence of metal salts such as iron and copper sulphate, as well as that of bathophenanthrolinedisulfonic acid (iron chelator) and bathocuproinedisulfonic acid (copper chelator) on the activity of laccases in the intra (IF) and extracellular fractions (EF) of the wild-type and the non-pathogenic mutant strain (rho1::hyg) of F. oxysporum. The results show that laccase activity in the IF fraction of the wild and mutant strain increased with the addition of iron chelator (53.4 and 114.32%; respectively). With copper, it is observed that there is an inhibition of the activity with the addition of CuSO4 for the EF of the wild and mutant strain (reduction of 82 and 62.6%; respectively) and for the IF of the mutant strain (54.8%). With the copper chelator a less laccase activity in the IF of the mutant strain was observed (reduction of 53.9%). The results obtained suggest a different regulation of intracellular laccases in the mutant strain compared with the wild type in presence of CuSO4 and copper chelator which may be due to the mutation in the rho gene.

Influence of iron and copper on the activity of laccases in Fusarium oxysporum f. sp. lycopersici

ABSTRACT Fusarium oxysporum f. sp. lycopersici is a phytopathogenic fungus that causes vascular wilt in tomato plants. In this work we analyze the influence of metal salts such as iron and copper sulphate, as well as that of bathophenanthrolinedisulfonic acid (iron chelator) and bathocuproinedisulfonic acid (copper chelator) on the activity of laccases in the intra (IF) and extracellular fractions (EF) of the wild-type and the non-pathogenic mutant strain (rho1::hyg) of F. oxysporum. The results show that laccase activity in the IF fraction of the wild and mutant strain increased with the addition of iron chelator (53.4 and 114.32%; respectively). With copper, it is observed that there is an inhibition of the activity with the addition of CuSO4 for the EF of the wild and mutant strain (reduction of 82 and 62.6%; respectively) and for the IF of the mutant strain (54.8%). With the copper chelator a less laccase activity in the IF of the mutant strain was observed (reduction of 53.9%). The results obtained suggest a different regulation of intracellular laccases in the mutant strain compared with the wild type in presence of CuSO4 and copper chelator which may be due to the mutation in the rho gene.

GTPase Rho1 regulates the expression of xyl3 and laccase genes in Fusarium oxysporum.

The Rho1 protein is a GTPase that participates in cell wall biogenesis. We analyzed the transcript levels of laccase genes (lccl, lcc2, lcc3, lcc4, lcc5, and lcc9), and a xylanase gene (xyl3) in Fusarium oxysporum f. sp. lycopersici strain 4287 (wild type) and two mutant strains; rhol::hyg that lacks a functional Rho1, and rho1::hyg + rho1 (G14V) that has a constitutively active Rho1. The transcript levels of lcc2, lcc3, lcc5, and xyl3 differed among the three strains, but those of lcc1 and lcc9 did not. Xylanase activities were higher in rho1::hyg than in both the wild type and rho1::hyg + rho1 (G14V) . Laccase activities were significantly higher in the two mutants than in the wild type. Rho1 thus plays a role in regulating xyl3, lcc2, lcc3, and lcc5 at the transcriptional and/or translational level.

Isolation and expression of enolase gene in Fusarium oxysporum f. sp. lycopersici.

Fusarium oxysporum f. sp. lycopersici is a fungus responsible for the tomato disease known as fusariosis. Enolase, which is the enzyme that catalyzes the reaction of 2-phosphoglycerate to phosphoenolpyruvate, is present during glycolysis. Enolase genes have been isolated from bacteria and fungi, among other organisms. In this research, a large portion of the enolase, eno, gene sequence was isolated from F. oxysporum and compared with those of other microorganisms, revealing a similarity of 51-69 %. We analyzed the copy number of the eno gene and determined that only a single copy is present in F. oxysporum, as in several fungi, such as Candida albicans and Aspergillus oryzae. We also detected the expression of the eno gene by reverse transcription-polymerase chain reaction during in vitro growth under two growth conditions where glucose was used as the carbon source, and we observed the same eno gene expression levels under both growth conditions.

Rho1 and other GTP-binding proteins are associated with vesicles carrying glucose oxidase activity from Fusarium oxysporum f. sp. lycopersici.

In eucaryotic cells, the delivery of a secreted protein to the plasma membrane via vesicles must include transport, recognition, and fusion events. Proteins exposed on the cytoplasmic face of the secretory vesicles play a role in these events; these include the GTP-binding proteins, which are crucial components in this process. Fractions enriched with vesicles carrying glucose oxidase (GOX) activity from Fusarium oxysporum f. sp. lycopersici, a soilborne fungal pathogen causing vascular wilt on tomato plants, were obtained using two successive sucrose gradients, the first a linear-log and the second an isopycnic gradient. In this study, we used the following Fusarium strains: a wild-type and a strain carrying a Δrho1 loss-of-function mutation (presenting dramatically reduced virulence). By ADP-ribosylation with C3 exotoxin, and Western blot analysis with specific antibodies, we identified the small GTPases Rho1, Rho4, Cdc42 and Rab8, and a heterotrimeric Gα protein associated with vesicles carrying GOX activity. This was done for both strains, with the exception of Rho1, which was absent in the mutant strain; in addition, the levels of the Cdc42 protein were observed to be higher in the Δrho1 strain. These data indicate that three Rho proteins, Rho1, Rho4, and Cdc42, are present in secretory vesicles carrying GOX activity in F. oxysporum, and that Rho1 is not essential for the transport and secretion of, at least, cargo proteins carried in secretory vesicles, or Cdc42/Rho4 can fulfill its role in these events.