Broad-spectrum Antifungal Activity Research Articles

Screening for Antifungal Indigenous Lactobacilli Strains Isolated from Local Fermented Milk for Developing Bioprotective Fermentates and Coatings Based on Acid Whey Protein Concentrate for Fresh Cheese Quality Maintenance.

The demand for healthy foods without artificial food additives is constantly increasing. Hence, natural food preservation methods using bioprotective cultures could be an alternative to chemical preservatives. Thus, the main purpose of this work was to screen the indigenous lactobacilli isolated from fermented cow milk for their safety and antifungal activity to select the safe strain with the strongest fungicidal properties for the development of bioprotective acid whey protein concentrate (AWPC) based fermentates and their coatings intended for fresh cheese quality maintenance. Therefore, 12 lactobacilli strains were isolated and identified from raw fermented cow milk as protective cultures. The safety of the stains was determined by applying antibiotic susceptibility, haemolytic and enzymatic evaluation. Only one strain, Lacticaseibacillus paracasei A11, met all safety requirements and demonstrated a broad spectrum of antifungal activity in vitro. The strain was cultivated in AWPC for 48 h and grew well (biomass yield 8 log10 cfu mL-1). L. paracasei A11 AWPC fermentate was used as a vehicle for protective culture in the development of pectin-AWPC-based edible coating. Both the fermentate and coating were tested for their antimicrobial properties on fresh acid-curd cheese. Coating with L. paracasei A11 strain reduced yeast and mould counts by 1.0-1.5 log10 cfu mL-1 (p ≤ 0.001) during cheese storage (14 days), simultaneously preserving its flavour and prolonging the shelf life for six days.

Synthesis, Antifungal Activity, 3D-QSAR and Controlled Release on Hydrotalcite Study of Longifolene-Derived Diphenyl Ether Carboxylic Acid Compounds

Twenty-two novel longifolene-derived diphenyl ether-carboxylic acid compounds 7a-7v were synthesized from renewable biomass resources longifolene, and their structures were confirmed by FT-IR, 1H NMR, 13C NMR, and HRMS. The preliminary evaluation of in vitro antifungal activity displayed that compound 7b presented inhibition rates of 85.9%, 82.7%, 82.7%, and 81.4% against Alternaria solani, Cercospora arachidicola, Rhizoctonia solani, and Physalospora piricola, respectively, and compound 7l possessed inhibition rates of 80.7%, 80.4%, and 80.3% against R. solani, C. arachidicola, P. piricola, respectively, exhibiting excellent and broad-spectrum antifungal activities. Besides, compounds 7f and 7a showed significant antifungal activities with inhibition rates of 81.2% and 80.7% against A.solani, respectively. Meanwhile, a reasonable and effective 3D-QSAR mode (r2 = 0.996, q2 = 0.572) has been established by the CoMFA method. Furthermore, the drug-loading complexes 7b/MgAl-LDH were prepared and characterized. Their pH-responsive controlled-release behavior was investigated as well. As a result, complex 7b/MgAl-LDH-2 exhibited excellent controlled-releasing performance in the water/ethanol (10:1, v:v) and under a pH of 5.7.

Discovery of N-Phenylpropiolamide as a Novel Succinate Dehydrogenase Inhibitor Scaffold with Broad-Spectrum Antifungal Activity on Phytopathogenic Fungi.

Based on the structural features of both succinate dehydrogenase inhibitors (SDHIs) and targeted covalent inhibitors, a series of N-phenylpropiolamides containing a Michael acceptor moiety were designed to find new antifungal compounds. Nineteen compounds showed potent inhibition activity in vitro on nine species of plant pathogenic fungi. Compounds 9 and 13 showed higher activity on most of the fungi than the standard drug azoxystrobin. Compound 13 could completely inhibit Physalospora piricola infection on apples at 200 μg/mL concentration over 7 days and showed high safety to seed germination and seedling growth of plants at ≤100 μg/mL concentration. The action mechanism showed that 13 is an SDH inhibitor with a median inhibitory concentration, IC50, value of 0.55 μg/mL, comparable with that of the positive drug boscalid. Molecular docking studies revealed that 13 can bind well to the ubiquinone-binding region of SDH by hydrogen bonds and undergoes π-alkyl interaction and π-cation interaction. At the cellular level, 1 as the parent compound could destruct the mycelial structure of P. piricola and partly dissolve the cell wall and/or membrane. Structure-activity relationship analysis showed that the acetenyl group should be a structure determinant for the activity, and the substitution pattern of the phenyl ring can significantly impact the activity. Thus, N-phenylpropiolamide emerged as a novel and promising lead scaffold for the development of new SDHIs for plant protection.

Functional Characterization of a Bacillus-Derived Novel Broad-Spectrum Antifungal Lipopeptide Variant against Candida tropicalis and Candida auris and Unravelling Its Mode of Action.

Limited treatment options, recalcitrance, and resistance to existing therapeutics encourage the discovery of novel antifungal leads for alternative therapeutics. Antifungal lipopeptides have emerged as potential candidates for developing new and alternative antifungal therapies. In our previous studies, we isolated and identified the lipopeptide variant AF4 and purified it to homogeneity via chromatography from the cell-free supernatant of Bacillus subtilis. AF4 was found to have broad-spectrum antifungal activity against more than 110 fungal isolates. In this study, we found that clinical isolates of Candida tropicalis and Candida auris exposed to AF4 exhibited low MICs of 4 to 8 mg/L. Time-kill assays indicated the in vitro pharmacodynamic potential of AF4. Biocompatibility assays demonstrated ~75% cell viability at 8 mg/L of AF4, indicating the lipopeptide's minimally cytotoxic nature. In lipopeptide-treated C. tropicalis and C. auris cells, scanning electron microscopy revealed damage to the cell surface, while confocal microscopy with acridine orange(AO)/propidium iodide (PI) and FUN-1 indicated permeabilization of the cell membrane, and DNA damage upon DAPI (4',6-diamidino-2-phenylindole) staining. These observations were corroborated using flow cytometry (FC) in which propidium iodide, 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA), and rhodamine 123 (Rh123) staining of cells treated with AF4 revealed loss of membrane integrity, increased reactive oxygen species (ROS) production, and mitochondrial membrane dysfunction, respectively. Membrane perturbation was also observed in the 1,6-diphenyl-1,3,5-hexatriene (DPH) fluorescence study and the interaction with ergosterol was observed by an ergosterol binding assay. Decreased membrane dipole potential also indicated the probable binding of lipopeptide to the cell membrane. Collectively, these findings describe the mode of action of AF4 against fungal isolates by membrane disruption and ROS generation, demonstrating its antifungal potency. IMPORTANCE C. tropicalis is a major concern for candidiasis in India and C. auris has emerged as a resistant yeast causing difficult-to-treat infections. Currently, amphotericin B (AMB) and 5-flucytosine (5-FC) are the main therapeutics for systemic fungal infections; however, the nephrotoxicity of AMB and resistance to 5-FC is a serious concern. Antifungal lead molecules with low adverse effects are the need of the hour. In this study, we briefly describe the antifungal potential of the AF4 lipopeptide and its mode of action using microscopy, flow cytometry, and fluorescence-based assays. Our investigation reveals the basic mode of action of the investigated lipopeptide. This lipopeptide with broad-spectrum antifungal potency is apparently membrane-active, and there is a smaller chance that organisms exposed to such a compound will develop drug resistance. It could potentially act as a lead molecule for the development of an alternative antifungal agent to combat candidiasis.

Combination of Cetylpyridinium Chloride and Chlorhexidine Acetate: A Promising Candidate for Rapid Killing of Gram-Positive/Gram-Negative Bacteria and Fungi.

Combined use of the present antimicrobial drugs has been proved to be an alternative approach for antimicrobial agents' development since the co-existed of the drugs working in different mechanism have been demonstrated potentially enhance their antimicrobial activity. In this work, antibacterial and antifungal activity of the cetylpyridinium chloride (CPC)/chlorhexidine acetate (CHA) combination was evaluated for the first time, while a universal concentration for the rapid killing of gram-positive/gram-negative bacteria and fungi was also proposed. The minimum inhibitory concentrations (MIC) of CPC and CHA used alone or in combination were first measured, showing that the combined treatment decreased the MIC against tested gram-positive/gram-negative bacteria and fungi to 1/8-1/2. Growth curve assays demonstrated CPC and CHA had dynamic combined effects against the tested microorganisms at the concentration equal to MIC. Besides, combined use of these two drugs could also enhance their biocidal activity, which was illustrated by fluorescence microscopy and SEM images, as well as soluble protein measurement. More importantly, in vitro acute eye and skin irritation tests showed short-term contact with CPC/CHA combination would not cause any damage to mammalian mucosa and skin. In a word, CPC/CHA combination exhibited broad-spectrum antibacterial and antifungal activity against tested gram-positive/gram-negative bacteria and fungi while without any acute irritation to mammalian mucosa and skin, providing a new perspective on the selection of personal disinfectants.

Use of synthetic peptides in the prevention of oral diseases

Among the protective factors of mixed saliva, antimicrobial peptides have received much attention. They have broad-spectrum antibacterial and antifungal activities. Synthetic peptides are found in hygiene products, such as mouthwashes, sprays, and toothpastes. According to their manufacturers, the use of these products helps prevent oral bacterial infections. Objective. To study the effect of toothpastes and mouth spray, which contain synthetic peptide complexes, on the physicochemical parameters of mixed saliva in order to substantiate their preventive antibacterial effect. Material and methods. Mixed saliva from patients was examined before and after the use of the synthetic peptide-containing spray and toothpaste. The following salivary physicochemical parameters (pH, surface tension, and base buffer capacity) were measured. Results. The peptide-containing spray and toothpaste caused changes in the oral fluid pH within the normal range, which assisted in maintaining oral homeostasis and in effectively exerting the functions of mixed saliva. When the spray or toothpaste was used, the base buffer capacity was slightly reduced, which suggested that the synthetic peptides were able to neutralize oral fluid acids and thereby to enhance the anti-inflammatory and antibacterial functions of mixed saliva. The surface tension of saliva was increased with the peptide-containing spray and that was decreased with the peptide-containing toothpaste. This effect indicated the active ability of the synthetic peptides to be absorbed into blood and to have a direct effect on cell metabolism at the site of lesion (when using the spray), as well as the ability of the complexes to integrate into the colloidal system of saliva, which enhanced its reparative and protective properties (when using the toothpaste). Conclusion. The study shows an increase in the antibacterial effect of mixed saliva when using oral hygiene products containing peptides, which contributes to the prevention of risk factors for inflammatory diseases of the oral mucosa and periodontium.

Synthesis and Development of N-2,5-Dimethylphenylthioureido Acid Derivatives as Scaffolds for New Antimicrobial Candidates Targeting Multidrug-Resistant Gram-Positive Pathogens

The growing antimicrobial resistance to last-line antimicrobials among Gram-positive pathogens remains a major healthcare emergency worldwide. Therefore, the search for new small molecules targeting multidrug-resistant pathogens remains of great importance. In this paper, we report the synthesis and in vitro antimicrobial activity characterisation of novel thiazole derivatives using representative Gram-negative and Gram-positive strains, including tedizolid/linezolid-resistant S. aureus, as well as emerging fungal pathogens. The 4-substituted thiazoles 3h, and 3j with naphthoquinone-fused thiazole derivative 7 with excellent activity against methicillin and tedizolid/linezolid-resistant S. aureus. Moreover, compounds 3h, 3j and 7 showed favourable activity against vancomycin-resistant E. faecium. Compounds 9f and 14f showed broad-spectrum antifungal activity against drug-resistant Candida strains, while ester 8f showed good activity against Candida auris which was greater than fluconazole. Collectively, these data demonstrate that N-2,5-dimethylphenylthioureido acid derivatives could be further explored as novel scaffolds for the development of antimicrobial candidates targeting Gram-positive bacteria and drug-resistant pathogenic fungi.

Novel quinolin-2(1H)-one analogues as potential fungicides targeting succinate dehydrogenase: design, synthesis, inhibitory evaluation and molecular modeling.

Succinate dehydrogenase is an important target of fungicides. Succinate dehydrogenase inhibitors (SDHIs) have widely been used to combat destructive plant pathogenic fungi because they possess efficient and broad-spectrum antifungal activities and as well as unique mode of action. The research and development of novel SDHIs have been ongoing. Thirty-six novel quinolin-2(1H)-one derivatives were designed, synthesized and characterized. The single crystal structure of compound 3c was determined through the X-ray diffraction of single crystals. The bioassay results displayed that most compounds had good antifungal activities at 16 μg mL-1 against Rhizoctonia cerealis, Erysiphe graminis, Botrytis cinerea, Penicillium italicum and Phytophthora infestans. Compounds 6o, 6p and 6r had better antifungal activities than the commercialized fungicide pyraziflumid against Botrytis cinerea. Their half maximal effective concentration (EC50 ) values were 0.398, 0.513, 0.205 and 0.706 μg mL-1 , respectively. Moreover, the inhibiting activities of the bioactive compounds were tested against succinate dehydrogenase. The results indicated that they possessed outstanding activities. Compounds 6o, 6p and 6r also exhibited better inhibiting activities than pyraziflumid against succinate dehydrogenase. Their half maximal inhibitory concentration (IC50 ) values were 0.450, 0.672, 0.232 and 0.858 μg mL-1 , respectively. The results of molecular dynamic (MD) simulations indicated that compound 6r displayed stronger affinity to succinate dehydrogenase than pyraziflumid. The results of the present study displayed that quinolin-2(1H)-one derivative could be one scaffold of potential SDHIs and will provide some valuable information for the research and development of new SDHIs. © 2022 Society of Chemical Industry.

Multi-arm ε-polylysines exhibit broad-spectrum antifungal activities against Candida species.

Invasive fungal infections pose a crucial threat to public health and are an under-recognized component of antimicrobial resistance, which is an emerging crisis worldwide. Here we designed and synthesized a panel of multi-arm ε-polylysines (ε-mPLs, nR-Km) with a precise number of n = 3-6 arms of ε-oligo(L-lysine)s and a precise arm length of m = 3-7 ε-lysine residues. ε-mPLs have good biocompatibility and exhibited broad-spectrum antifungal activities towards Aspergillus, Mucorales and Candida species, and their antifungal activities increased with residue arm length. Among these ε-mPLs, 3R-K7 showed high antifungal activity against C. albicans with a MIC value of as low as 24 μg mL-1 (only 1/16th that of ε-PL) and also exhibited similar antifungal activity towards the clinically isolated multi-drug resistant (MDR) C. albicans strain. Furthermore, 3R-K7 could inhibit the formation of C. albicans biofilms and kill the cells within mature C. albicans biofilms. Mechanistic studies proved that 3R-K7 killed fungal cells by entering the cells to generate reactive oxygen species (ROS) and induce cell apoptosis. An in vivo study showed that 3R-K7 significantly increased the survival rate of mice in a systemic murine candidiasis model, demonstrating that ε-mPL has great potential as a new antifungal agent.

The elucidation of the multimodal action of the investigational anti-Candida lipopeptide (AF4) lead from Bacillus subtilis.

Background: Candida species are the main etiological agents for candidiasis, and Candida albicans are the most common infectious species. Candida species' growing resistance to conventional therapies necessitates more research into novel antifungal agents. Antifungal peptides isolated from microorganisms have potential applications as novel therapeutics. AF4 a Bacillus-derived lipopeptide demonstrating broad-spectrum antifungal activity has been investigated for its ability to cause cell death in Candida species via membrane damage and oxidative stress. Methods: Using biophysical techniques, the secondary structure of the AF4 lipopeptide was identified. Scanning electron microscopy and confocal microscopy with fluorescent dyes were performed to visualise the effect of the lipopeptide. The membrane disruption and permeabilization were assessed using the 1,6-diphenyl hexatriene (DPH) fluorescence assay and flow cytometric (FC) assessment of propidium iodide (PI) uptake, respectively. The reactive oxygen species levels were estimated using the FC assessment. The induction of apoptosis and DNA damage were studied using Annexin V-FITC/PI and DAPI. Results: Bacillus-derived antifungal variant AF4 was found to have structural features typical of lipopeptides. Microscopy imaging revealed that AF4 damages the surface of treated cells and results in membrane permeabilization, facilitating the uptake of the fluorescent dyes. A loss of membrane integrity was observed in cells treated with AF4 due to a decrease in DPH fluorescence and a dose-dependent increase in PI uptake. Cell damage was also determined from the log reduction of viable cells treated with AF4. AF4 treatment also caused elevated ROS levels, induced phosphatidylserine externalisation, late-stage apoptosis, and alterations to nuclear morphology revealed by DAPI fluorescence. Conclusion: Collectively, the mode of action studies revealed that AF4 acts primarily on the cell membrane of C. albicans and has the potential to act as an antifungal drug candidate.

Broad-spectrum antifungal activity of Phyllanthus niruri leaves tested against Candida species

Introduction: The emergence of resistant pathogenic microorganisms against conventional antimicrobials has become a global concern. To combat new and re-emerging infectious illnesses, new antimicrobial agents with different chemical structures and novel modes of action are required. Therefore, this study evaluated ethanolic extracts of Phyllanthus niruri for their antimicrobial activities against Candida species isolated from different clinical samples. Materials and Methods: This involved the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method for the identification of Candida species. The ethanolic extraction of P. niruri leaves was examined. The agar well diffusion method was used to determine the antifungal activity of the leaf extracts against standard American Type Culture Collection strains as well as clinical isolates of Candida species. Results: Using PCR-RFLP, Candida tropicalis was found to be the most prevalent species of Candida, followed by Candida albicans, Candida glabrata, Candida krusei and Candida parapsilosis. The ethanol extract of P. nirui leaves showed good activity against all the clinical and standard strains of Candida which were comparable to the standard drug fluconazole. The activity of P. niruri against C. albicans was highest followed by C. parapsilosis, C. tropicalis, C. glabrata and C. krusei at a concentration of 100 mg/ml. Conclusion: The findings of this study support the use of P. niruri plant to treat Candida infections, particularly fluconazole-resistant Candida species.

Design, Catalyst-Free Synthesis of New Novel α-Trifluoromethylated Tertiary Alcohols Bearing Coumarins as Potential Antifungal Agents

A new method for the synthesis of α-trifluoromethylated tertiary alcohols bearing coumarins is described. The reaction of 3-(trifluoroacetyl)coumarin and pyrrole provided the target compounds with high yields under catalyst-free, mild conditions. The crystal structure of compound 3fa was investigated by X-ray diffraction analysis. The biological activities, such as in vitro antifungal activity of the α-trifluoromethylated tertiary alcohols against Fusarium graminearum, Fusarium oxysporum, Fusarium moniliforme, Rhizoctonia solani Kuhn, and Phytophthora parasitica var nicotianae, were investigated. The bioassay results indicated that compounds 3ad, 3gd, and 3hd showed broad-spectrum antifungal activity in vitro. Compound 3cd exhibited excellent fungicidal activity against Rhizoctonia solani Kuhn, with an EC50 value of 10.9 μg/mL, which was comparable to that of commercial fungicidal triadimefon (EC50 = 6.1 μg/mL). Furthermore, molecular docking study suggested that 3cd had high binding affinities with 1W9U, like argifin.

Semisynthetic Amides of Polyene Antibiotic Natamycin.

Natamycin is a macrolide polyene antibiotic, characterized by a potent broad spectrum antifungal activity and low toxicity. However, it is not used for the treatment of systemic mycoses due to its low bioavailability and low solubility in aqueous solutions. In order to create new semisynthetic antifungal agents for treatment of mycoses, a series of water-soluble amides of natamycin were synthesized. Antifungal activities of natamycin derivatives were investigated against Candida spp., including a panel of Candida auris clinical isolates and filamentous fungi. Toxicity for mammalian cells was assayed by monitoring antiproliferative activity against human postnatal fibroblasts (HPF) and human embryonic kidney cells (HEK293). By comparing leakage of contents from ergosterol versus cholesterol containing vesicles, a ratio that characterizes the efficacy and safety of natamycin and its derivatives was determined (EI, efficiency index). Ability of all tested semisynthetic natamycines to prevent proliferation of the yeast Candida spp. cells was comparable or even slightly higher to those of parent antibiotic. Interestingly, amide 8 was more potent than natamycin (1) against all tested C. auris strains (MIC values 2 μg/mL vs 8 μg/mL, respectively). Among 7 derivatives, amide 10 with long lipophilic side chains showed the highest EI and strong antifungal activity in vitro but was more toxic against HPF. In vivo experiments with amide 8 showed in vivo efficacy on a mouse candidemia model with a larger LD50/ED50 ratio in comparison to amphotericin B.

Production of iturin A by Bacillus velezensis ND and its biological control characteristics.

Bacillus subtilis, as a biocontrol bacterium, possess a variety of biological functions and the capacity to control plant pathogens. Iturin A is a biosurfactant with broad-spectrum antifungal activity produced by fermentation of B. subtilis. In this study, the dynamic parameters of solid-state fermentation (SSF) and submerged fermentation (SMF) of Bacillus velezensis ND were compared, and a method for producing iturin A with a yield of 12.46 g/kg utilizing SSF was proposed. It has significant advantages over SMF and has the highest yield of all previously reported studies. B. velezensis ND also contains protease activity, cellulase activity, iron-carrying activity, the ability to synthesis 3-indoleacetic acid (IAA), fixation nitrogen, and degrade phosphorus. In cotton pot experiments, it can effectively increase cotton growth and minimize Verticillium wilt. This strain's superior fermentation efficiency, biological function, and biocontrol ability are sufficient to demonstrate its promise for the development and use of biocontrol agents.

Discovery of Novel Orally Bioavailable Triazoles with Potent and Broad-Spectrum Antifungal Activity In Vitro and In Vivo.

In our continuing efforts to discover novel triazoles with improved antifungal activity in vitro and in vivo, a series of 41 novel compounds containing 1,2,3-triazole side chains were designed and synthesized via a click reaction based on our previous work. Most of the compounds showed moderate to excellent broad-spectrum antifungal activity in vitro. Among them, the most promising compound 9A16 displayed excellent antifungal and anti-drug-resistant fungal ability (MIC80 = 0.0156-8 μg/mL). In addition, compound 9A16 showed powerful in vivo efficacy on mice systematically infected with Candida albicans SC5314, Cryptococcus neoformans H99, fluconazole-resistant C. albicans 100, and Aspergillus fumigatus 7544. Moreover, compared to fluconazole, compound 9A16 showed better in vitro anti-biofilm activity and was more difficult to induce drug resistance in a 1 month induction of resistance assay in C. albicans. With favorable pharmacokinetics, an acceptable safety profile, and high potency in vitro and in vivo, compound 9A16 is currently under preclinical investigation.

Isolation, identification, and biocontrol mechanisms of endophytic Burkholderia vietnamiensis C12 from Ficus tikoua Bur against Rhizoctonia solani

Rice sheath blight is a devastating fungal disease caused by Rhizoctonia solani. Biocontrol utilizing endogenous microorganisms has been considered a safe and sustainable strategy. To obtain a new type of antagonistic bacteria with a high biocontrol effect on rice sheath blight, over 50 endophytic bacteria were isolated from the root of the medicinal plant Ficus tikoua Bur and screened for their antifungal activity by dual culture assay. Among them, strain C12 showed the strongest inhibitory activity against R. solani (inhibition rate 94.78%), and exhibited a broad-spectrum antifungal activity. According to the phylogenetic tree based on 16S rDNA and recA genes, C12 was identified as Burkholderia vietnamiensis, belonging to the Burkholderia cepacia complex, and safe for plants and humans. Through the study on B. vietnamiensis C12, the strain could secrete cell wall degrading enzymes (protease and chitinase), produce IAA, and siderophores, fix nitrogen, and solubilize phosphorus. The bacterial culture filtrate (although treated at 121 °C) of strain C12 and its ethyl acetate extract significantly inhibited the growth of R solani and resulted in distorted hyphae. Moreover, when the extract was 50 μg/mL, 100 μg/mL, 200 μg/mL, and 500 μg/mL, the diameter of mycelium was decreased by 50.44%, 56.43%, 67.27%, and 74.80% respectively, the number of sclerotia was decreased by 85.45%, 90.42%, 96.83%, and 99.22% respectively. Multiple antifungal substances in extracts of C12 were detected through LC-MS. Strain C12 gave a 70.98% reduction in lesion length in a detached inoculated leaf assay. In greenhouse and field tests, strain C12 significantly reduced the disease index of rice sheath blight, and the control efficiency reached 68.75% and 66.52%, respectively. Furthermore, C12 enhanced the resistance of rice to R. solani by increasing the activities of defense enzymes SOD, PAL, POD, CAT, and PPO. The strain also exhibited plant-promoting abilities on rice. The results indicated that B. vietnamiensis C12 had great biocontrol potential and could be used as a candidate strain for green control of rice sheath blight.

Antifungal Mechanism of Metabolites from Newly Isolated Streptomyces sp. Y1-14 against Banana Fusarium Wilt Disease Using Metabolomics.

Banana Fusarium wilt caused by Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4) is one of the most destructive banana diseases in the world, which limits the development of the banana industry. Compared with traditional physical and chemical practices, biological control becomes a promising safe and efficient strategy. In this study, strain Y1-14 with strong antagonistic activity against Foc TR4 was isolated from the rhizosphere soil of a banana plantation, where no disease symptom was detected for more than ten years. The strain was identified as Streptomyces according to the morphological, physiological, and biochemical characteristics and the phylogenetic tree of 16S rRNA. Streptomyces sp. Y1-14 also showed a broad-spectrum antifungal activity against the selected 12 plant pathogenic fungi. Its extracts inhibited the growth and spore germination of Foc TR4 by destroying the integrity of the cell membrane and the ultrastructure of mycelia. Twenty-three compounds were identified by gas chromatography-mass spectrometry (GC-MS). The antifungal mechanism was investigated further by metabolomic analysis. Strain Y1-14 extracts significantly affect the carbohydrate metabolism pathway of Foc TR4 by disrupting energy metabolism.

The Activity of Polyhomoarginine against Acanthamoeba castellanii.

Arginine-rich peptides can have broad-spectrum anti-bacterial and anti-fungal activities. Polyhomoarginine consists of highly cationic residues which can act on the negatively charged microbial cell membranes. Acanthamoeba is a free-living protozoan known to cause a rare corneal infection which is difficult to diagnose and treat. This study evaluated the activity of the polyhomoarginines against Acanthamoeba castellanii. Acanthamoeba amoebicidal, amoebistatic, encystation and excystment assays were performed using protocols described in the literature. The activity of polyhomoarginines (PHAs) of different lengths (10 to 400 residues) was measured against the trophozoites and cysts of Acanthamoeba castellanii ATCC30868 in concentrations ranging from 0.93 μM to 15 μM. Data were represented as mean ± SE and analysed using one-way ANOVA. Overall, PHAs demonstrated good anti-acanthamoeba activity against both trophozoites and cysts. PHA 30 reduced the number of viable trophozoites by 99%, inhibited the formation of cysts by 96% and the emergence of trophozoites from cysts by 67% at 3.75 μM. PHA 10 was similarly active, but at a slightly higher concentration of 15 μM, reducing the numbers of viable trophozoites by 98%, inhibiting cyst formation by 84% and preventing the emergence of trophozoites from cysts by 99%. At their greatest anti-amoeba concentrations, PHA 10 gave only 8% haemolysis at 15 μM while PHA 30 gave <40 % haemolysis at 3.75 μM. Polyhomoarginine 10 showed excellent anti-amoebic activity against both forms of Acanthamoeba castellanii and was non-toxic at its most active concentrations. This implies that polyhomoarginines can be developed into a potential therapeutic agent for Acanthamoeba keratitis. However, there is a need to carry out further pre-clinical and then in vivo experiments in the AK animal model.

Liposomal amphotericin B-the present.

Most invasive fungal infections are opportunistic in nature but the epidemiology is constantly changing, with new risk groups being identified. Neutropenia is a classical risk factor for fungal infections, while critically ill patients in the ICU are now increasingly at risk of yeast and mould infections. Factors to be considered when choosing antifungal treatment include the emergence of rarer fungal pathogens, the risk of resistance to azoles and echinocandins and the possibility of drug-drug interactions. Liposomal amphotericin B has retained its place in the therapeutic armamentarium based on its clinical profile: a broad spectrum of antifungal activity with a low risk of resistance, predictable pharmacokinetics with a rapid accumulation at the infection site (including biofilms), a low potential for drug-drug interactions and a low risk of acute and chronic treatment-limiting toxicities versus other formulations of amphotericin B. It is a suitable choice for the first-line empirical or pre-emptive treatment of suspected fungal infections in neutropenic haematology patients and is an excellent alternative for patients with documented fungal disease who can no longer tolerate or continue their first-line azole or echinocandin therapy, both in the haematology setting and in the ICU. Moreover, it is the first-line drug of choice for the treatment of invasive mucormycosis. Finally, liposomal amphotericin B is one of the few antifungal agents approved for use in children of all ages over 1 month and is included in paediatric-specific guidelines for the management of fungal disease.

Improved Production of Fengycin in Bacillus subtilis by Integrated Strain Engineering Strategy.

Fengycin is a lipopeptide with broad-spectrum antifungal activity. However, its low yield limits its commercial application. Therefore, we iteratively edited multiple target genes associated with fengycin synthesis by combinatorial metabolic engineering. The ability of Bacillus subtilis 168 to manufacture lipopeptides was restored, and the fengycin titer was 1.81 mg/L. Fengycin production was further increased to 174.63 mg/L after knocking out pathways associated with surfactin and bacillaene synthesis and replacing the native promoter (PppsA) with the Pveg promoter. Subsequently, fengycin levels were elevated to 258.52 mg/L by upregulating the expression of relevant genes involved in the fatty acid pathway. After blocking spore and biofilm formation, fengycin production reached 302.51 mg/L. Finally, fengycin production was increased to approximately 885.37 mg/L after adding threonine in the optimized culture medium, which was 488-fold higher compared with that of the initial strain. Integrated strain engineering provides a strategy to construct a system for improving fengycin production.