Broad-spectrum Antifungal Activity Research Articles

Antifungal Activities of Bacillus mojavensis BQ-33 towards the Kiwifruit Black Spot Disease Caused by the Fungal Pathogen Didymella glomerata.

‘Hongyang’ kiwifruit (Actinidia chinensis, cultivar ‘Hongyang’) black spot disease is caused by the fungal pathogen Didymella glomerata, and is a serious disease, causing considerable losses to the kiwifruit industry during growth of the fruit. Hence, we aimed to identify a potential biocontrol agent against D. glomerata. In this study, bacterial isolates from the rhizosphere soil of kiwifruit were tested for their potential antifungal activity against selected fungal pathogens. Based on a phylogenetic tree constructed using sequences of 16S rDNA and the gyrA gene, BQ-33 with the best antifungal activity was identified as Bacillus mojavensis. We evaluated the antagonistic activity and inhibitory mechanism of BQ-33 against D. glomerata. Confrontation experiments showed that both BQ-33 suspension and the sterile supernatant (SS) produced by BQ-33 possessed excellent broad-spectrum antifungal activity. Furthermore, the SS damaged the cell membrane and cell wall of the mycelia, resulting in the leakage of a large quantity of small ions (Na+, K+), soluble proteins and nucleic acids. Chitinase and β-1,3-glucanase activities in SS increased in correlation with incubation time and remained at a high level for several days. An in vivo control efficacy assay indicated that 400 mL L−1 of SS completely inhibited kiwifruit black spot disease caused by D. glomerata. Therefore, BQ-33 is a potential biocontrol agent against kiwifruit black spot and plant diseases caused by other fungal pathogens. To our knowledge, this is the first report of the use of a rhizosphere microorganism as a biocontrol agent against kiwifruit black spot disease caused by D. glomerata.

INHIBITORY EFFECT OF ANTIFUNGAL ACTIVITY OF GROUND NUT OIL AGENT’S FUNGI

Ground nut (Arachis hypogaea) oil is essential component of our food and it can inhibit the spore germination of Aspergillus sp. The development of protective components with natural origin as alternatives to synthetic fungicides is currently in the spotlight. This oil is known for their broad spectrum of antifungal activity against plant pathogen and human both. The essential oils reduce hyphal growth and induce cell lysis and cytoplasmic evacuation in fungi.

Discovery of Niphimycin C from Streptomyces yongxingensis sp. nov. as a Promising Agrochemical Fungicide for Controlling Banana Fusarium Wilt by Destroying the Mitochondrial Structure and Function.

Banana Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc) is the most destructive soil-borne fungal disease. Tropical race 4 (Foc TR4), one of the strains of Foc, can infect many commercial cultivars, which represents a threat to global banana production. Currently, there are hardly any effective chemical fungicides to control the disease. To search for natural product-based fungicides for controlling banana Fusarium wilt, we identified a novel strain Streptomyces yongxingensis sp. nov. (JCM 34965) from a marine soft coral, from which a bioactive compound, niphimycin C, was isolated using an activity-guided method. Niphimycin C exhibited a strong antifungal activity against Foc TR4 with a value of 1.20 μg/mL for EC50 and obviously inhibited the mycelial growth and spore germination of Foc TR4. It caused the functional loss of mitochondria and the disorder of metabolism of Foc TR4 cells. Further study showed that niphimycin C reduced key enzyme activities of the tricarboxylic acid (TCA) cycle and the electron transport chain (ETC). It displayed broad-spectrum antifungal activities against the selected 12 phytopathogenic fungi. In pot experiments, niphimycin C reduced the disease indexes in banana plantlets and inhibited the infection of Foc TR4 in roots. Hence, niphimycin C could be a promising agrochemical fungicide for the management of fungal diseases.

Activity and cell toxicology of fluazinam on Fusarium graminearum

Fusarium graminearum is an important plant pathogen and the causal agent of Fusarium head blight (FHB). At present, the principal method of controlling FHB is through fungicides. Fluazinam is an agent with strong broad-spectrum antifungal activity and has been used to control many diseases. However, there are no reported uses of fluazinam for controlling FHB. This study reports the activity and cell toxicology mechanisms of fluazinam on the filamentous fungus F. graminearum and its effect on fungal growth and development. The activity of fluazinam was tested for 95 wild-type field strains of F. graminearum. The EC50 values (the 50% effective concentration) of fluazinam for inhibition of mycelial growth and spore germination ranged from 0.037 μg/ml to 0.179 μg/ml and from 0.039 μg/ml to 0.506 μg/ml, respectively. The fluazinam sensitivity of these strains varied in 4.9 and 13.0 folds, implying that the target of the fungicide remained unchanged. After treatment with 0.3 μg/ml (≈EC90) fluazinam, the production of conidia was reduced, and the cell wall and cell membrane had shrunked; the cell nucleus and septum morphology, cell membrane permeability, and sexual development were not affected. When treated with 0.1 μg/ml (≈EC50) or 0.3 μg/ml fluazinam, the mycelial respiration and deoxynivalenol (DON) synthesis of F. graminearum were decreased. Confocal images showed that the formation of toxisomes was disturbed after fluazinam treatment, suggesting that fluazinam reduces DON synthesis by inhibiting toxisome formation. Infection of wheat coleoptiles revealed that fluazinam had a strong protective activity against F. graminearum. At 250 μg/ml fluazinam the control efficacy of protective treatments reached 100% and controlled strains resistant to carbendazim. These results contribute to the understanding of the mode of action of fluazinam and its application.

Formulation and Development of a Water-in-Oil Emulsion-Based Luliconazole Cream: In Vitro Characterization and Analytical Method Validation by RP-HPLC.

Luliconazole (LCZ) is a new antifungal agent containing imidazole moiety which revealed broad-spectrum antifungal activity. The aim of this research was to prepare water-in-oil (w/o) emulsion-based cream formulation of LCZ in addition to the development and validation of an analytical method by reverse-phase high-performance liquid chromatography (RP-HPLC). Cetostearyl alcohol (12.14%), light liquid paraffin (5.00%), white soft paraffin (2.75%), and Tween-80 (1.00%) appeared as the optimized concentration to give better consistency to the cream. Moreover, without adding pH adjusting agents the pH of the optimized formulation (F5) was obtained within the range of human skin pH throughout the stability period. The value of particle size, polydispersity index, and zeta potential was 187.90 ± 2.061 nm, 0.124 ± 0.026, and -10.553 ± 1.349 mV, respectively. In this study, an analytical C18 (4.6 mm × 25 cm), 5 μm column was used for chromatographic separation with a mixture of acetonitrile and water in the proportion of 50 : 50 v/v as the mobile phase at a flow rate of 1.0 mL/min. The calibration curve was obtained linear at 296 nm in the concentration range of 0.08–0.12 mg/mL. Furthermore, the limit of detection (LOD) and limit of quantification (LOQ) were 0.0013 and 0.0042 µg/mL, respectively. In addition, the observed results demonstrated that our developed method was linear (R2 = 0.999), precise (%RSD below than 2.0%), and accurate (mean recovery% = 100.18–100.91). The F5 showed no physical changes until 6th month analysis at room temperature and accelerated conditions. Similarly, the assay obtained 101.99% ± 0.27 and 99.89% ± 0.08 at room temperature and accelerated conditions, respectively. Additionally, all validated parameters were obtained within the acceptable limit as well. These findings conclude that both physically and chemically stable w/o cream formulation of LCZ can be formulated and assessed for their stability by applying the authenticated analytical procedure of RP-HPLC.

S3.5d Identification of fungicidal activity of otilonium bromide by the repurposing approach

S3.5 Environmental exposure - risk for human fungal disease, September 21, 2022, 4:45 PM - 6:15 PM ObjectivesThe development of new antifungal drugs will help to expand the limited antifungal arsenal and curb the emergence of drug resistance. To meet the demand for novel antifungal drugs, we performed a high-throughput screen of an FDA-approved compound library (HY-L022, MCE®) and identified a promising lead antifungal compound, otilonium bromide (OTB).MethodsWe determined antifungal activity of each compound from the library and investigated the antifungal spectrum of OTB using broth microdilution assays (Fig. 1a). We tested the fungicidal effect of OTB by the spot assay and disk diffusion assay. The inhibitory effect of OTB on mycelial growth was identified by observing hyphal growth in hyphal-inducing media containing OTB. The antibiofilm activity of OTB was evaluated by XTT method. We further checked the antifungal activity of OTB in vivo by a murine C. albicans infection model, which was constructed by immunosuppression of mice with cyclophosphamide, inhibiting the intestinal microorganisms by levofloxacin (14 mg/ml), and then gavage with C. albicans cells (1 × 108 cells per mouse). A total of 24 C57 mice were randomly divided into three groups: one group was gavaged with saline, one group with 1 mg/kg fluconazole, and one group with 50 mg/kg OTB. In all 2 h after infection, drugs were given once a day for 4 consecutive days, with 15 days of observation. The death of a mouse was assessed daily and statistically analyzed by Log-rank (Mantel-Cox) test. The toxicity of OTB to human cloned colon adenocarcinoma cells was assessed with MTT method.ResultsWe found that OTB is the most promising lead antifungal molecule among FDA-approved drugs (Fig. 1a). OTB has a broad-spectrum antifungal activity with minimum inhibitory concentration for inhibiting the growth of 50% of strains (MIC50) values ranging from 0.39 to 6.25 μm (Fig. 1b). We found that OTB (≥3.5 μm) has fungicidal effect on C. albicans using spot assay (Fig. 1c). We further found that OTB (25 μg/disk) can form a clear inhibition zone, but not fluconazole (25 μg/disk) (Fig. 1d). Besides, we spotted C. albicans cells from the broth microdilution assays onto YPD solid medium and found that cells treated with OTB (≥1.56 μm) could not recover on YPD solid medium, while cells treated with FLC could (Fig. 1e). It is worth noting that OTB has inhibitory effects on filamentation and biofilm formation at 12.5 μm (Fig. 2a, b), which are key virulence factors of pathogenic fungi. Besides, OTB (50 mg/kg) significantly prolonged the median survival time from 5 days (control group) to 15 days and improved the survival rate from 0% (control group) to 62.5% (P = .0078) (Fig. 2c). Furthermore, we found that OTB exhibited relative low toxicity to mammal cells with IC50 28-57 μm (Fig. 2d), which was much higher than MIC50 against pathogenic fungi.ConclusionOur study highlights that OTB has a broad-spectrum antifungal activity with low MIC50 values and high safety in vitro and potent antifungal efficacy in vivo and will be beneficial to the treatment of devastating invasive fungal infections.

P092 Otilonium bromide is a potent antifungal agent against fluconazole- and flucytosine-resistant Cryptococcus neoformans strains

Poster session 1, September 21, 2022, 12:30 PM - 1:30 PMObjectives Cryptococcus neoformans is a worldwide threat causing global pulmonary and systemic infections in humans. However, only three drugs are available for the treatment of cryptococcosis: fluconazole, amphotericin B, and flucytosine. Drug repurposing, the process of using a drug for an indication different from the initial indication, is an emerging approach in the discovery of new antifungal drugs. We've set out to identify repurposable drugs for cryptococossis.MethodsWe screened the United States FDA-approved drugs for antifungal activity against C. albicans lab strain SC5314. We tested the antifungal profile against major human fungal pathogens including C. neoformans. In order to uncover the mechanism of action against C. neoformans, lab strain H99 was used to evolve drug resistant adaptors. Next-generation sequencing technology was used to investigate the genome change of drug resistant adaptors. SNP calling was also performed to identify possible mutations causing drug resistance.ResultsHere we found that otilonium bromide (OTB), which is extensively used to treat patients affected by the irritable bowel syndrome, had broad-spectrum antifungal activity. OTB was active against fluconaozle-resistant and flucotysine-resistant C. neoformans strains. Furthermore, we found resistance to OTB was mostly due to duplication of chromosome 6. Further work will be on the identification of the Candidate gene on chromosome 6 which is required OTB resistance.ConclusionThis study highlights the potential application of OTB as a new antifungal drug against C. neoformans strains susceptible or resistant to commonly used antifungal drugs.

Penicillium digitatum as a Model Fungus for Detecting Antifungal Activity of Botanicals: An Evaluation on Vietnamese Medicinal Plant Extracts.

Medicinal plants play important roles in traditional medicine, and numerous compounds among them have been recognized for their antimicrobial activity. However, little is known about the potential of Vietnamese medicinal plants for antifungal activity. In this study, we examined the antagonistic activity of twelve medicinal plant species collected in Northern Vietnam against Penicillium digitatum, Aspergillus flavus, Aspergillus fumigatus, and Candida albicans. The results showed that the antifungal activities of the crude extracts from Mahonia bealei, Ficus semicordata, and Gnetum montanum were clearly detected with the citrus postharvest pathogen P. digitatum. These extracts could fully inhibit the growth of P. digitatum on the agar medium, and on the infected citrus fruits at concentrations of 300–1000 µg/mL. Meanwhile, the other tested fungi were less sensitive to the antagonistic activity of the plant extracts. In particular, we found that the ethanolic extract of M. bealei displayed a broad-spectrum antifungal activity against all four pathogenic fungi. Analysis of this crude extract by enrichment coupled with high-performance liquid chromatography revealed that berberine and palmatine are major metabolites. Additional inspections indicated berberine as the key compound responsible for the antifungal activity of the M. bealei ethanolic extract. Our study provides a better understanding of the potential of Vietnamese medicinal plant resources for combating fungal pathogens. This work also highlights that the citrus pathogen P. digitatum can be employed as a model fungus for screening the antifungal activity of botanicals.

Identification of Cyclic Dipeptides and a New Compound (6-(5-Hydroxy-6-methylheptyl)-5,6-dihydro-2H-pyran-2-one) Produced by Streptomyces fungicidicus against Alternaria solani.

As an important microbial resource, Actinomycetes, especially Streptomyces, have important application values in medicine and biotechnology. Streptomyces fungicidicus SYH3 was isolated from soil samples in tomato-growing areas and showed good inhibitory effects on Alternaria solani in tomato. To obtain pure active compounds, SYH3 fermentation broth was subjected to XAD-16 macroporous resin and silica gel column chromatography. Combined with the repeated preparation and separation of preparative high-performance liquid chromatography (HPLC), a total of four monomer compounds were obtained after activity tracking. Compound 4 was identified as a new six-membered lactone ring compound named 6-(5-hydroxy-6-methylheptyl)-5,6-dihydro-2H-pyran-2-one by 1D and 2D nuclear magnetic resonance (NMR) data and mass spectrometry (MS). The other three active compounds belong to the cyclodipeptide, and their half maximal inhibitory concentration (IC50) values against A. solani were 43.4, 42.9, and 30.6 μg/mL, respectively. Compound 4 significantly inhibited the spore germination and induced swollen and deformed local hyphae of A. solani with an IC50 value of 24.9 μg/mL. Compound 4 also had broad-spectrum antifungal activity and had a good antifungal effect on the tested plant-pathogenic fungi. The modes of action of new compound (4) still require further investigation, representing a novel and effective anti-fungal agent for future application.

Spiro[benzoxazine-piperidin]-one derivatives as chitin synthase inhibitors and antifungal agents: Design, synthesis and biological evaluation

Four series of spiro[benzoxazine-piperidin]-one derivatives were designed and synthesized. Their inhibition percentages against chitin synthase and antifungal activities were evaluated. Based on the preliminary biological assays, the series of derivatives containing α, β-unsaturated carbonyl fragment which had moderate to excellent CHS inhibitory activity and antifungal activity were further researched. In this series of compounds, eight out of twenty-one compounds had good to excellent inhibitory activity against chitin synthase with an inhibition percentage value above 60% at the concentration of 300 μg/mL. Among them, compounds 9a, 9o, 9s and 9t showed excellent chitin synthase inhibitory activity with IC50 values of 0.14 mM, 0.11 mM, 0.10 mM and 0.16 mM, respectively, which were equal to that of the control drug (polyoxin B). The results of sorbitol protection assays and evaluation of antifungal activity against micafungin-resistant fungi further proved that the target of these synthesized compounds was chitin synthase. The antifungal activity evaluation showed that compounds 9a, 9d, 9h, 9s and 9t had broad-spectrum antifungal activity in vitro and their antifungal activities are equal to those of fluconazole and polyoxin B. The result of combination use showed this series of compounds combined with fluconazole had additive or synergistic effects. In addition, compounds 9a, 9o and 9t showed good antifungal activity against fluconazole-resistant C. albicans and fluconazole-resistant C. neoformans variants. Consequently, the results showed that these compounds were chitin synthase inhibitors and antifungal agents and had significant activity against drug-resistant fungal variants.

N-Terminus Plays a Critical Role for Stabilizing the Filamentous Assembly and the Antifungal Activity of Bg_9562.

ABSTRACTBg_9562, a prophage tail-like protein was earlier shown to be required for bacterial mycophagy by Burkholderia gladioli strain NGJ1. The purified protein exhibited broad-spectrum antifungal activity; however, the structural and mechanistic details vis-à-vis its activity remained elusive. In this study, we have structurally characterized the protein Bg_9562 using negatively stained transmission electron microscopy, molecular modeling and mutagenesis. We find that Bg_9562 shows structural similarity to Gp13, a tail assembly chaperone. The transmission electron microscopy revealed that, Bg_9562 forms long flexible tubular structures. Molecular modeling of the filament like structure divulges that the inter subunit contacts are meditated largely through hydrophobic interactions. Using mutagenesis, we demonstrate that the N-terminal residues of the protein when deleted results in reduced activity and destabilization of filament formation. Overall, structure-function analysis opens up avenues for further utilization of the protein as a potent antifungal molecule.IMPORTANCE Burkholderia gladioli strain NGJ1, isolated from healthy rice seedling, was earlier demonstrated to have mycophagous properties on a broad range of fungi, including Rhizoctonia solani, a causal agent of deadly sheath blight disease of rice. The purified Bg_9562 protein exerts broad-spectrum antifungal activity. The protein also inhibits the growth of laboratory strain of Candida, an opportunistic human pathogen. In this study, we structurally characterize Bg_9562 using a combination of negative staining transmission electron microscopy, molecular modeling, mutagenesis, and functional antifungal assay. We show that the protein assembles into long filament like structures stabilized by N-terminus residues and this region is important for its activity. Our study has implications in utilizing Bg_9562 or its derivatives as antifungal molecule(s) which will provide environmentally friendly control of fungal diseases of plants and animals.

Potential Biological Control of Endophytic Streptomyces sp. 5-4 Against Fusarium Wilt of Banana Caused by Fusarium oxysporum f. sp. cubense Tropical Race 4.

Fusarium wilt of banana caused by Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4) is one of the most disastrous fungal diseases. Biological control is a promising strategy for controlling Fusarium wilt of banana. To explore endophytic actinomycetes as biocontrol resources against Foc TR4, antagonistic strains were isolated from different tissues of medicinal plants. Here, a total of 144 actinomycetes were isolated and belonged to Nonomuraea, Kitasatospora, and Streptomyces. Forty-three isolates exhibited antifungal activities against Foc TR4. The strain labeled with 5-4 isolated from roots of Piper austrosinense had a broad-spectrum antifungal activity by the production of chitinase and β-1,3-glucanase and was identified as Streptomyces hygroscopicus subsp. hygroscopicus 5-4. Furthermore, disease index of banana wilt was significantly reduced by application of strain 5-4 in comparison with application of Foc TR4 alone. Exogenous application of strain 5-4 increased the expression levels of defense genes such as (PAL), peroxidase (POD), pathogenesis-related protein 1 (PR-1), hydrolytic enzymes (β-1,3-glucanase), lysin motif receptor kinase 1 (LYK-1), and mitogen-activated protein kinase 1 (MPK-1). The antifungal mechanism assay demonstrated that extracts of strain 5-4 inhibited spore gemination and hyphal growth of Foc TR4, and caused abnormally swollen, deformity, and rupture of Foc TR4 hypha. Thus, S. hygroscopicus subsp. hygroscopicus 5-4 could be used as a potential biological agent for controlling Fusarium wilt of banana.

A swamp forest Streptomyces sp. strain KF15 with broad spectrum antifungal activity against chilli pathogens exhibits anticancer activity on HeLa cells.

During the investigation, soil actinomycetes were isolated from Kathlekanu swamp forest and the crude ethyl acetate extract from the potent isolate KF15 was analysed with GC-MS and HPTLC to identify bioactive metabolites. The crude extract was examined for in-vitro antifungal activity on pathogens of chilli; MTT cytotoxicity assay was performed against HeLa cell line to determine the anticancer potential. The isolate Streptomyces sp. strain KF15 exhibited antagonistic activity against fungal pathogens by inhibiting growth and altering growth pattern with increased antimicrobial activity in dose-dependent manner. GC-MS revealed many bioactive compounds and HPTLC depicted metabolite fingerprint. The IC50 of 99.85µg/ml indicated the high potential of KF15 extract to prevent proliferation of HeLa cells. Therefore, the findings of this study indicate that the crude extract from Streptomyces sp. strain KF15 contains antifungal and anticancer metabolites; further study on purification could help in controlling many fungal diseases as well as cervical cancer in humans.

Biocontrol potential of Burkholderia sp. BV6 against the rice blast fungus Magnaporthe oryzae.

To investigate the broad-spectrum antifungal activity of Burkholderia sp. BV6, that is isolated from rice roots and its biocontrol potential against rice blast caused by Magnaporthe oryzae. We evaluated the ability to isolate BV6 in the biological control of rice blast disease and investigated its antifungal mechanisms. BV6 strongly inhibited the hyphal growth of M. oryzae Guy11 and other plant pathogenic fungi, and pot experiments showed that BV6 significantly decreases the disease index of rice blast from 47.5 to 24.6. The secreted small-molecule secondary metabolites were regarded as weapons during the antifungal process by inhibiting the germination of M. oryzae conidia and mycelial growth, and thereby prevent the following infection. Liquid chromatography-mass spectrometry analysis of the metabolites from the supernatant of isolate BV6 showed that the antifungal weapons of isolate BV6 are novel, small, molecular hydrophilic compounds that are different from reported antifungal compounds. The isolate BV6 inhibits the M. oryzae infection by the production of small-molecule antifungal compounds. The current study discovers the role of the Burkholderia sp. BV6 in the biocontrol of plant pathogenic fungi. Therefore, isolate BV6 is a potential candidate for developing a microbial formulation for the biocontrol of the most common disease of rice blast.

Antifungal CoAl layered double hydroxide ultrathin nanosheets loaded with oregano essential oil for cereal preservation

Fungal infestation of cereals seriously affects human health and the economy. Protecting cereals from fungal infestation is meaningful. Therefore, safe and economical antifungal agents must be explored. Oregano essential oil (OEO) has broad-spectrum antifungal activity but is unstable and poorly water-dispersible. CoAl-LDH ultrathin nanosheets (CoAl-LDH UNs) are a biosafe carrier with peroxidase-like activity and good water dispersibility. A new nanocomposite was constructed by integrating OEO and CoAl-LDH UNs named OEO/CoAl-LDH UNs. The antifungal activity of OEO was improved for the synergetic effect of OEO and CoAl-LDH UNs. OEO/CoAl-LDH UNs showed excellent antifungal activity against Fusarium moniliforme, Fusarium graminearum and Aspergillus flavus. A polyvinyl alcohol film containing OEO/CoAl-LDH UNs was prepared as a food package and effectively inhibited fungal infestation on cereals. This work can provide some insights for the exploration of other new antifungal materials. The good biosafety of this antifungal material can be effectively applied to food packaging.

New Developments in Pediatric Antifungal Pharmacology.

New Developments in Pediatric Antifungal Pharmacology.

Curvicollide D, a new modified γ-lactone from the culture broth of Albifimbria verrucaria and its antifungal activity against plant pathogenic fungi.

In our screening program for new antifungal active compounds, a new modified γ-lactone curvicollide D (1) together with five known trichothecenes (2-6) were isolated from the culture filtrate of fungus Albifimbria verrucaria based on the in vitro antifungal assay. The chemical structure of new compound 1 was elucidated by NMR and HR-MS spectroscopic analyses, and the relative configurations of 1 were deduced from NOE experiments and coupling constant analysis. Compound 1 exhibited moderate antifungal activities against plant pathogenic fungi Botrytis cinerea, Colletotrichum coccodes, and Magnaporthe oryzae with MIC value in a range of 100-200 µg ml-1. Moreover, trichothecene compounds (2-6) displayed a broad spectrum of antifungal activities with MIC values in a range of 6.3-100 µg ml-1.

Synthesis, Antifungal Activity, Cytotoxicity and QSAR Study of Camphor Derivatives.

Control of fungal phytopathogens affecting crops and woodlands is an important goal in environmental management and the maintenance of food security. This work describes the synthesis of 37 camphor derivatives, of which 27 were new compounds. Their antifungal effects on six fungi were evaluated in vitro. Compounds 3a, 4a and 5k showed strong antifungal activity against Trametes versicolor, with EC50 values of 0.43, 6.80 and 4.86 mg/L, respectively, which were better than that of tricyclazole (EC50 118.20 mg/L) and close to or better than that of carbendazim (EC50 1.20 mg/L). The most potent compound, 3a, exhibited broad-spectrum antifungal activity towards six fungi with EC50 values within the range of 0.43–40.18 mg/L. Scanning electron microscopy demonstrated that compounds 3a, 4a and 5k gave irregular growth and shriveling of the mycelia. In vitro cytotoxicity evaluation revealed that the tested camphor derivatives had mild or no cytotoxicity for LO2 and HEK293T cell lines. Quantitative structure−activity relationship (QSAR) analysis revealed that the number of F atoms, relative molecular weight, the atomic orbital electronic population and total charge on the positively charged surfaces of the molecules of camphor derivatives have effects on antifungal activity. The present study may provide a theoretical basis for a high-value use of camphor and could be helpful for the development of novel potential antifungals.

Design, synthesis, and biological evaluation of novel berberine derivatives against phytopathogenic fungi.

The abuse of chemical fungicides not only leads to toxic residues and resistance in plant pathogenic fungi, but also causes environmental pollution and side effects on in humans and animals. Based on the antifungal activities of berberine, seven different types of berberine derivatives (A1-G1) were synthesized, and their antifungal activities against six plant pathogenic fungi were evaluated (Rhizoctonia solani, Botrytis cinerea, Fusarium graminearum, Phytophthora capsici, Sclerotinia sclerotiorum, and Magnaporthe oryzae). The results for antifungal activities in vitro showed that berberine derivative E1 displayed good antifungal activity against R. solani with a median effective concentration (EC50 ) of 1.77 μg ml-1 , and berberine derivatives F1 and G1 demonstrated broad-spectrum antifungal activities with EC50 values ranging from 4.43 to 42.23 μg ml-1 against six plant pathogenic fungi. Berberine derivatives (E2-E29, F2-F18, and G2-G9) were further synthesized to investigate the structure-activity relationship (SAR), and compound E20 displayed significant antifungal activity against R. solani with an EC50 value of 0.065 μg ml-1 . Preliminary mechanism studies showed that E20 could cause mycelial shrinkage, cell membrane damage, mitochondrial abnormalities and the accumulation of harmful reactive oxygen species, resulting in cell death in R. solani. Moreover, in vivo experimental results showed that the protective effect of E20 was 97.31% at 5μg ml-1 , which was better than that of the positive control thifluzamide (50.13% at 5μg ml-1 ). Berberine derivative E20 merits further development as a new drug candidate with selective and excellent antifungal activity against R. solani. © 2022 Society of Chemical Industry.

Bioactivities evaluation of an endophytic bacterial strain Bacillus velezensis JRX-YG39 inhabiting wild grape

BackgroundBotrytis cinerea can cause serious disease on lots of plant hosts during growth and postharvest storage. Biocontrol is known to be eco-friendly methods to control pathogens. Plant endophytic bacteria are generally considered as beneficial organisms, since they can promote plant growth and enhance plant immune system. Thus, screening biological control agents is very important for sustainable plant protection.ResultsFifty-six endophytic bacteria were obtained from wild grape. Sixteen isolates and their extracts exhibited significant antifungal activity against B. cinerea. Particularly, strain JRX-YG39 with the strongest inhibition ability had a broad-spectrum antifungal activity. Combining 16S rDNA analysis and the phylogenetic results based on gyrA and gyrB genes, JRX-YG39 was assigned as Bacillus velezensis. JRX-YG39 could produce bioactive VOCs and obviously depressed mycelia growth of B. cinerea. It was confirmed that VOCs released by JRX-YG39 could significantly promote growth and induce defense of Arabidopsis thaliana. Thirty-one bioactive secondary metabolites were further identified from JRX-YG39 culture by gas chromatography–mass spectrometry analysis. Dibutyl phthalate, a potential antifungal substance, was the major compound accounting for 78.65%.ConclusionsB. velezensis JRX-YG39 has wide broad-spectrum antagonistic activity and significant plant promotion activity. Hence, B. velezensis JRX-YG39 will provide a valuable constituent of modern agricultural practice as biofertilizers and biocontrol agents.