Antifungal Activity of Lactobacillus plantarum ZZUA493 and Its Application to Extend the Shelf Life of Chinese Steamed Buns

Lactic acid bacteria (LAB) can produce many kinds of antifungal substances, which have been widely proven to have antifungal activity. In this study, 359 strains of LAB were screened for antifungal activity against Aspergillus niger (A. niger) using the 96-well microtiter plate method, and three showed strong activity. Of these, ZZUA493 showed a broad-spectrum antifungal ability against A. niger, Aspergillus oryzae, Trichoderma longibrachiatum, Aspergillus flavus and Fusarium graminearum. ZZUA493 was identified as Lactobacillus plantarum. Protease treatment, the removal of hydrogen peroxide with catalase and heat treatment had no effect on the antifungal activity of the cell-free supernatant (CFS) of ZZUA493; organic acids produced by ZZUA493 appeared to have an important role in fungal growth inhibition. The contents of lactic acid, acetic acid and phenyllactic acid in the CFS tended to be stable at 48 h, and amounted to 28.5, 15.5 and 0.075 mg/mL, respectively. In addition, adding ZZUA493, as an ingredient during their preparation, prolonged the shelf life of Chinese steamed buns. Overall, ZZUA493 appears to have good potential as a fungal inhibitor for food preservation.

Trichoderma citrinoviride: Anti-Fungal Biosurfactants Production Characteristics

The mechanism of direct impact of Trichoderma fungi on other organisms is a multilayer process. The level of limiting the growth of other microorganisms is determined by the strain and often by the environment. Confirmation of the presence of extracellular biosurfactants in certain strains of Trichoderma considered as biocontrol agents was regarded as a crucial topic complementing the characterization of their interactive mechanisms. Selected strains of T. citrinoviride were cultured in media stimulating biosurfactant biosynthesis, optionally supplemented with lytic enzyme inducers. Results confirmed the anti-fungal properties of surface-active compounds in the tested culture fluids. Preparations that displayed high fungal growth inhibition presented marginal enzymatic activities of both chitinases and laminarinases, implying the inhibitory role of biosurfactants. Fractions from the foam of the culture fluid of the C1 strain, cultured on Saunders medium, and HL strain on MGP medium, without an additional carbon source, exhibited the most prominent ability to inhibit the growth of phytopathogens. Filamentous fungi capable of producing fungicidal compounds, including surfactants, may find applications in protecting the plants against agri-food pathogenic molds.

Potential use of lemongrass essential oil as fungicide against Aspergillus brasiliensis and as post-harvest protectant of wheat

Fungi are among the main responsible for damage and loss in stored grains, its control has been done through synthetic substances, which are harmful to man and the environment. Brazil, one of the leading countries in agriculture, has optimal environmental conditions for the development of mycotoxigenic fungi. Most of the synthetic chemicals used as preservatives have often been realized to be toxic to humans and also cause adverse environmental effects. Thus, it is necessary to search for alternative methods of controlling. In this study the aimed was to evaluate the efficacy of lemongrass essential oil in the control of the Aspergillus brasiliensis. In vitro and serial microdilution tests were carried out at different concentrations of essential oil and citral, which corresponds to 72% of the total oil composition. Inhibition of fungal growth on contaminated wheat grain was evaluated. The in vitro test results showed that the essential oil has fungicidal potential at concentrations from 0.6 μL mL-1, the minimum inhibitory concentration was determined at 0.8 μL mL-1. The tests with citral showed fungal control at concentrations from 0.6 μL mL-1 onwards. For wheat grain, fungal growth inhibition was obtained at the concentration of 1.6 μL mL-1. The essential oil of Cymbopogon flexuosus showed fungicidal activity against the fungus Aspergillus brasiliensis.

Antifungal Siderophore Conjugates for Theranostic Applications in Invasive Pulmonary Aspergillosis Using Low-Molecular TAFC Scaffolds

Invasive pulmonary aspergillosis (IPA) is a life-threatening form of fungal infection, primarily in immunocompromised patients and associated with significant mortality. Diagnostic procedures are often invasive and/or time consuming and existing antifungals can be constrained by dose-limiting toxicity and drug interaction. In this study, we modified triacetylfusarinine C (TAFC), the main siderophore produced by the opportunistic pathogen Aspergillus fumigatus (A. fumigatus), with antifungal molecules to perform antifungal susceptibility tests and molecular imaging. A variation of small organic molecules (eflornithine, fludioxonil, thiomersal, fluoroorotic acid (FOA), cyanine 5 (Cy5) with antifungal activity were coupled to diacetylfusarinine C (DAFC), resulting in a “Trojan horse” to deliver antifungal compounds specifically into A. fumigatus hyphae by the major facilitator transporter MirB. Radioactive labeling with gallium-68 allowed us to perform in vitro characterization (distribution coefficient, stability, uptake assay) as well as biodistribution experiments and PET/CT imaging in an IPA rat infection model. Compounds chelated with stable gallium were used for antifungal susceptibility tests. [Ga]DAFC-fludioxonil, -FOA, and -Cy5 revealed a MirB-dependent active uptake with fungal growth inhibition at 16 µg/mL after 24 h. Visualization of an A. fumigatus infection in lungs of a rat was possible with gallium-68-labeled compounds using PET/CT. Heterogeneous biodistribution patterns revealed the immense influence of the antifungal moiety conjugated to DAFC. Overall, novel antifungal siderophore conjugates with promising fungal growth inhibition and the possibility to perform PET imaging combine both therapeutic and diagnostic potential in a theranostic compound for IPA caused by A. fumigatus.

Effect of Deterpenated Origanum majorana L. Essential Oil on the Physicochemical and Biological Properties of Chitosan/β-Chitin Nanofibers Nanocomposite Films

Herein, the effect of three deterpenated fractions from Origanum majorana L. essential oil on the physicochemical, mechanical and biological properties of chitosan/β-chitin nanofibers-based nanocomposite films were investigated. In general, the incorporation of Origanum majorana L. original essential oil or its deterpenated fractions increases the opacity of the nanocomposite films and gives them a yellowish color. The water solubility decreases from 58% for chitosan/β-chitin nanofibers nanocomposite film to around 32% for the nanocomposite films modified with original essential oil or its deterpenated fractions. Regarding the thermal stability, no major changes were observed, and the mechanical properties decreased. Interestingly, data show differences on the biological properties of the materials depending on the incorporated deterpenated fraction of Origanum majorana L. essential oil. The nanocomposite films prepared with the deterpenated fractions with a high concentration of oxygenated terpene derivatives show the best antifungal activity against Aspergillus niger, with fungal growth inhibition of around 85.90%. Nonetheless, the only nanocomposite film that does not present cytotoxicity on the viability of L929 fibroblast cells after 48 and 72 h is the one prepared with the fraction presenting the higher terpenic hydrocarbon content (87.92%). These results suggest that the composition of the deterpenated fraction plays an important role in determining the biological properties of the nanocomposite films.

Antifungal siderophore conjugates for theranostic applications in invasive pulmonary aspergillosis using low molecular TAFC scaffolds

Invasive pulmonary aspergillosis (IPA) is a life-threatening form of fungal infection, primarily in immunocompromised patients and associated with a significant mortality. Diagnostic procedures are often invasive and/or time consuming and existing antifungals can be constrained by dose limiting toxicity and drug interaction. In this study, we modified triacetylfusarinine C (TAFC), the main siderophore produced by the opportunistic pathogen Aspergillus fumigatus, with antifungal molecules to perform antifungal susceptibility tests and molecular imaging. Methods: A variation of small organic molecules (eflornithine, fludioxonil, thiomersal, fluoroorotic acid (FOA), cyanine 5 (Cy5)) with antifungal activity were coupled to TAFC, resulting in a Trojan horse to deliver antifungal compounds specifically into Aspergillus fumigatus hyphae by the major facilitator transporter MirB. Radioactive labelling with gallium-68 allowed to perform in vitro characterization (LogD, stability, uptake assay) as well as biodistribution experiments and PET/CT imaging in an IPA rat infection model. Compounds labelled with stable gallium were used for antifungal susceptibility tests. Results: [Ga]DAFC-fludioxonil, -FOA and Cy5 revealed a MirB dependent active uptake with fungal growth inhibition at 16 μg/mL after 24 h. Visualization of an Aspergillus fumigatus infection in lungs of a rat was possible with gallium-68 labelled compounds using PET/CT. Heterogeneous biodistribution patterns revealed the immense influence of the antifungal moiety conjugated to DAFC. Conclusion: Overall, novel antifungal siderophore conjugates with promising fungal growth inhibition and the possibility to perform PET-imaging, combine both therapeutic and diagnostic potential in a theranostic compound for IPA caused by Aspergillus fumigatus.

Exploration of Novel TOSMIC Tethered Imidazo[1,2-a]pyridine Compounds for the Development of Potential Antifungal Drug Candidate.

New candidates of imidazo [1, 2-a] pyridine were designed by combining 2-amino pyridine, TOSMIC isocyanide and various assorted aldehydes were synthesized to explore their antioxidant and antifungal potential. The design of these derivatives was based on utilizing the antifungal potential of azoles and TOSMIC moiety. These derivatives were synthesized by adopting multi-component reaction methodology, as it serves as a rapid and efficient tool to target structurally diverse heterocyclic compounds in quantitative yield. The resulting imidazo [1, 2-a] pyridine derivatives were structurally verified by 1HNMR and 13CNMR. The compounds were analyzed for their antioxidant and fluorescent properties and it was observed that compound 15 depicted good fluorescence and antioxidant potential. Additionally, the compounds were evaluated for their antifungal potential against both Aspergillus fumigatus 3007 & Candida albicans 3018 and it was observed that all the compounds had moderate to significant antifungal effect. Confocal images depicted the porous nature of compound treated fungal cell membranes leading to fungal growth inhibition. Molecular docking was performed to establish the interaction with the target enzyme (Lanosterol 14 alpha demethylase) which also corroborated with the in vitro results. In silico tools were employed to determine drug-likeliness of the compounds along with determination of ADME properties.

Potentiated inhibition of Trichoderma virens and other environmental fungi by new biocide combinations

Fungi cause diverse, serious socio-economic problems, including biodeterioration of valuable products and materials that spawns a biocides industry worth ~$11 billion globally. To help combat environmental fungi that commonly colonise material products, this study tested the hypothesis that combination of an approved fungicide with diverse agents approved by the FDA (Food and Drug Administration) could reveal potent combinatorial activities with promise for fungicidal applications. The strategy to use approved compounds lowers potential development risks for any effective combinations. A high-throughput assay of 1280 FDA-approved compounds was conducted to find those that potentiate the effect of iodopropynyl-butyl-carbamate (IPBC) on the growth of Trichoderma virens; IPBC is one of the two most widely used Biocidal Products Regulations–approved fungicides. From this library, 34 compounds in combination with IPBC strongly inhibited fungal growth. Low-cost compounds that gave the most effective growth inhibition were tested against other environmental fungi that are standard biomarkers for resistance of synthetic materials to fungal colonisation. Trifluoperazine (TFZ) in combination with IPBC enhanced growth inhibition of three of the five test fungi. The antifungal hexetidine (HEX) potentiated IPBC action against two of the test organisms. Testable hypotheses on the mechanisms of these combinatorial actions are discussed. Neither IPBC + TFZ nor IPBC + HEX exhibited a combinatorial effect against mammalian cells. These combinations retained strong fungal growth inhibition properties after incorporation to a polymer matrix (alginate) with potential for fungicide delivery. The study reveals the potential of such approved compounds for novel combinatorial applications in the control of fungal environmental opportunists. Key points • Search with an approved fungicide to find new fungicidal synergies in drug libraries. • New combinations inhibit growth of key environmental fungi on different matrices. • The approach enables a more rapid response to demand for new biocides.

Fabrication and Characterization of Chitosan/Poly(Lactic-Co-glycolic Acid) Core-Shell Nanoparticles by Coaxial Electrospray Technology for Dual Delivery of Natamycin and Clotrimazole

Natamycin (NAT) is the drug of choice for the treatment of fungal keratitis (FK). However, its inherent shortcomings, such as poor solubility, high dosing frequency, and long treatment cycle, need to be urgently addressed by designing a new delivery to widen its clinical utility. Growing research has confirmed that clotrimazole (CLZ) plays a significant role in fungal growth inhibition. Hence, coaxial electrospray (CO-ES) technology is used herein to prepare nano-systems with an average hydrodynamic particle size of 309-406 nm for the co-delivery of NAT and CLZ in chitosan (CTS) and poly(lactic-co-glycolic acid) (PLGA). The resulting NAT/CLZ@CTS/PLGA formulations were characterized by a transmission electron microscope (TEM) and in vitro release test. The results show that the formulations had obvious core-shell structures, uniform particle distribution, and also can sustain the release of drugs over 36 h. Furthermore, in vitro hemolysis, in vivo corneal irritation test, local allergenic test, and antifungal activity analyses are performed to evaluate the safety and efficiency of the formulations. Thus, good biosafety along with a significant anti-candidiasis effect are found in the NAT/CLZ@CTS/PLGA nanoparticles (NPs). Taken together, the results suggest that this design may provide a promising drug delivery system and a new option for the treatment of FK.