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The Crop protection system is dominated by the use of synthetic insecticides however incorporation of fungal biocontrol agents (FBA) at lower doses is considered to consolidate the integrated pest management (IPM) program. The success of IPM is delimited and relies on the understanding of how its performance is affected by adverse effects on FBA by agrochemicals deployed for its management as well as of other pests and diseases. In this study, laboratory grade actives of three different insecticides used for the control of Tetranychus urticae were tested at different concentrations to determine their effects on germination, mycelial growth and sporulation of eight entomopathogenic fungal isolates under in vitro conditions. The fungal isolate with the most adverse effects on the biological index was further studied for underlying reasons of antagonism. This paper reports for the first time changes in conidial surface morphology, its germination and penetration capacity on mite cuticle, post insecticide treatment. Bioassays showed that all insecticide actives at their lowest concentration (12.5% MC) were most toxic to Beauveria bassiana P isolate. Ethion and chlorpyriphos were compatible with Hirsutella thompsonii PDBC-1 at 12.5% MC while propargite showed compatibility with B. bassiana MTCC 6097, Metarhizium anisopliae MTCC 4104 and Cordyceps fumosorosea MTCC 4636 at the same concentration. Further, SEM studies showed that post-insecticide treatments, there were structural deformations on the conidial surface with a decrease in its germination and germ tube penetration capacity on the cuticle of T. urticae. Future studies in this area will help in improving IPM along with overcoming insecticide resistance problems.

Berberine hydrochloride (BH), an active component of Coptis chinensis and other plant taxa, has broad antimicrobial activity and may be useful for the treatment of Candida infections. In this study, the mechanisms underlying the inhibitory effect of BH against Candida albicans were evaluated, with a focus on the high-osmolarity glycerol mitogen-activated protein kinase (HOG-MAPK) pathway, which regulates multiple physiological functions. BH (256 and 64 μg ml-1) significantly increased intracellular glycerol and ROS levels in C. albicans, inhibited germ tube and hyphal formation, and increased chitin and β-1,3-glucan exposure on the cell wall. The inhibitory effect of BH was positively correlated with its concentration, and the inhibitory effect of 256 μg ml-1 BH was greater than that of 4 μg ml-1 fluconazole (FLC). Furthermore, RT-PCR analysis showed that 256 and 64 μg ml-1 BH altered the HOG-MAPK pathway in C. albicans. In particular, the upregulation of the core genes, SLN1, SSK2, HOG1, and PBS2 may affect the expression of key downstream factors related to glycerol synthesis and osmotic pressure (GPD1), ROS accumulation (ATP11 and SOD2), germ tube and hyphal formation (HWP1), and cell wall integrity (CHS3 and GSC1). BH affects multiple biological processes in C. albicans; thus, it can be an effective alternative to conventional azole antifungal agents.

This study evaluated the efficacy of the combined application of well-characterized chitosan polymer (degree of acetylation = 10%, degree of polymerization [DPn] = 90, and dispersity [ÐDP] = 2.8) and oligomers (partially acetylated chitosan polymers and oligosaccharides [paCOS]) (DP = 2 to 17) on conidia germination and mycelial growth of Fusarium graminearum, the major causal agent of Fusarium head blight in wheat. The polymer alone showed a higher inhibitory effect than the paCOS mixture alone, with half-maximal inhibitory concentrations of less than 50 µg ml−1 and more than 100 µg ml−1, respectively. Using time-lapse microscopy, we also showed that paCOS did not affect conidia germination at 50 µg ml−1, whereas chitosan polymer at the same concentration led to a delay in germination and in elongation of germ tubes. Scanning electron microscopy was used to observe the chitosan-induced changes in hyphal morphology. Surprisingly, the combination of chitosan polymer and paCOS led to strong synergistic effects in inhibiting conidia germination and fungal growth, as quantified by both the Abbot and Wadley equations. To our knowledge, this is the first report on a synergistic effect of a combination of chitosan polymers and oligomers, also highlighting for the first time the importance of ÐDP when studying structure–function relationships of functional biopolymers such as chitosan. The consequences of this finding for the improvement of chitosan-based antimicrobial or plant protective products are discussed. Given the economic importance of F. graminearum, this study suggests that the combination of chitosan polymer and oligomers can be used to support an efficient, sustainable plant protection strategy. [Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

Candida species are opportunistic pathogens that may cause infections in predisposed persons. This work aimed to detect the antifungal and antibiofilm potentials of Lactobacillus acidophilus and Lactobacillus plantarum supernatants on Candida spp. About 60 % and 80 % of Candida isolates were recovered from saliva samples of 20 patients with fixed orthodontic appliances, and 20 children having dental caries, respectively. The antifungal susceptibility of Candida spp. was investigated using disk diffusion assay. C. albicans strains showed low resistance to fluconazole (15 %) and amphotericin B (10 %). Using the agar well diffusion assay, both L. acidophilus and L. plantarum supernatants inhibited the in vitro growth of all tested Candida spp. with inhibition zone diameters of (11-19 mm) and (7-16 mm), respectively. On observing the effects of L. plantarum and L. acidophilus supernatants on Candida cells’ morphology; the light microscopic examination demonstrated the inhibition in germ tube formation of all tested C. albicans with percentages of 68 % and 53 %, and for C. krusei with inhibition percentages of 69 % and 59 %, respectively. L. acidophilus and L. plantarum strains showed high co-aggregation ability with C. albicans strains with ranges of 42-49 % and 30-35 %, respectively. The antibiofilm activities of the two Lactobacillus supernatants were determined using the tissue culture plate assays. Significant inhibition of biofilms formation by Candida spp. was recorded on treatment with L. plantarum and L. acidophilus supernatants, with reduction percentages of 50-72 % and 74-85 %, respectively.

Bionanotechnology has increasingly gained attention in biomedical fields as antifungal and antibiofilm agents. In this study, biosynthesized silver nanoparticles (bio-AgNPs) using aqueous Eucalyptus camaldulensis leaf extract were successfully performed by a one-step green approach. Spherical-shaped nanoparticles, approximately 8.65 nm, exhibited noncytotoxicity to erythrocytes, HeLa, and HaCaT cells. The synthesized nanoparticles showed strong fungicidal activity ranging from 0.5 to 1 µg/ml. The nanoparticles affected Candida adhesion and invasion into host cells by reduced germ tube formation and hydrolytic enzyme secretion. Inhibitory effects of bio-AgNPs on Candida biofilms were evaluated by the prevention of yeast-to-hyphal transition. A decrease in cell viability within mature biofilm demonstrated the ability of bio-AgNPs to penetrate into the extracellular matrix and destroy yeast cell morphology, leading to cell death. Molecular biology study on biofilms confirmed downregulation in the expression of genes ALS3, HWP1, ECE1, EFG1, TEC1, ZAP1, encoding hyphal growth and biofilm development and PLB2, LIP9, SAP4, involved in hydrolytic enzymes. In addition to candida treatment, the bio-AgNPs could be applied as an antioxidant to protect against oxidative stress-related human diseases. The findings concluded that bio-AgNPs could be used as an antifungal agent for candida treatment, as well as be incorporated in medical devices to prevent biofilm formation.

The increased resistance/tolerance of Candida infections to antimicrobial treatment can be attributed to biofilm-associated cells. A way to overcome this situation is to re-purpose non-anti-fungal drugs that could be active against fungi. We have explored the potential of a small library of eighteen non-antifungal drugs used in different human diseases. Candida albicans was cultured in the presence and absence of different concentrations of these drugs. Subsequently, inhibition of growth, germ tube formation, adhesion, and biofilm development were studied. Out of eighteen drug molecules, six showed a reduction in planktonic and biofilm growth in a dose-dependent manner and three drugs inhibited germ tube formation. This study shows the potential of non-antifungal drugs for the development of new anti-Candida agents.

Inhibition of B. cinerea by alginate (Alg) / cajuput essential oil (Caj) at different concentration (0%, 0.25%, 0.5%, 0.75%) and the composite’s surface properties were investigated. Among the samples, the treated spore with Caj 0.75% showed the lowest germination percentage and germ tube length. The regarded antifungal action was confirmed by observing the changes of membrane integrity stained with propidium iodide under fluorescence microscope. Moreover, the film composite roughness was characterized which Caj affected significantly (P

Aim Candida-associated fungal infections are prevalent in hospitalized and immune-compromised patients. Their biofilm architecture and high rate of antifungal resistance make treatment challenging. Eucalyptol (EPTL), a monoterpene majorly present in the essential oil of eucalyptus is well known for curing respiratory infections. Hence, the present study investigated the anti-biofilm efficacy of EPTL against the laboratory strains and clinical isolates of Candida to delineate its mode of action. Methods The effect of EPTL on the viability, biofilm formation, and mature biofilm of Candida strains was studied. Furthermore, its effect on cell cycle arrest, mitochondrial membrane potential (MMP), ROS generation, germ tube formation, ergosterol content and transcriptional expression of selected genes was also investigated. Results EPTL exhibited anti-biofilm activity against mature and developing biofilm of Candida albicans and Candida glabrata along with their clinical isolates. The biochemical components and enzyme activity were differentially modulated in EPTL-treated biofilm extracellular matrix. EPTL generated ROS and arrested cell cycle at the G1 /S phase in both the species, while altered MMP was recorded in C. glabrata. Transcriptional analysis evidenced for differential gene expression of selected ABC transporters, secreted hydrolytic enzymes, and cell wall biogenesis in C. albicans/C. glabrata upon treating with EPTL. Conclusion The current data on anti-biofilm activity of EPTL establish its candidacy for drug development or as an adjuvant with existing antifungal formulations. Significance and impact of the study Present investigation elucidates the mode of action of Eucalyptol as antifungal agent and would stand as a candidate for management of topical fungal infection.

Background Alteration in the normal composition of gasses in the atmosphere referred to as air pollution can occur as a result of different processes, including emissions from vehicles and power generators. Gasses affect living things directly and indirectly by playing roles in respiration, membrane function, synthesis, and growth. The gasses contained in fumes emitted from vehicles and generators may likely have effect on microorganisms in the environment including microfungi. Two microfungi were selected to study the effect of generator emissions on their growth. Results The gaseous emissions from power generators fueled by gasoline caused reduction in spore germination and germ tube lengths of R. stolonifer and F. oxysporum. For the former, spore germination decreased with length of exposure after an initial increase by ~ 100%. The fungus exhibited a single major germination peak at 10 min and a minor one at 30–35 min exposures. Germ tube length of the fungus also decreased with increased exposure. F. oxysporum too showed reduced spore germination and germ tube length with exposure but the fungus seemed to adjust better to the unfavorable environment created by emitted gasses showing multiple peaks of reduced heights as time progressed, though another rise that could reach a peak appeared at the 45-min maximum exposure for germ tube length. The peaks were however more broad for spore germination experiments indicating more stability in adjustment than observed for germ tube length. Greatest reduction in spore germination was by 25% in R. stolonifer and 71% in F. oxysporum Germ tube length reduction for R. stolonifer was by 24–76%, the greatest occurring at 35-min exposure, while for F. oxysporum it was 5–83%, the greatest occurring at 40 min exposure. Conclusions These observations reveal the toxicity of the gasses emitted to the growth of the two filamentous fungi and the potential harmful effect to other fungi which might be useful in the ecosystem as decomposers and to those that may be pathogenic to higher plants.

The transition of Candida albicans from yeast to hyphae cause the attachment to epithelial cells, forming biofilm and invasion. Therefore, the effect of Pediococcus acidilactici on morphogenesis and pathogenesis of C. albicans were examined. Inhibitory activity of P. acidilactici on Candida species growth was investigated and inhibition of C. albicans biofilm formation was measured by XTT method. Also, expression of seven genes as: Agglutinin-like protein 1 (ALS1,3), hyphal cell-wall protein (HWP1), secreted aspartyl proteinase (SAP4,6), Enhanced filamentous growth 1 (EFG1) and Enhanced activated protein 1 (EAP1) were studied by RT-PCR with different concentrations of P. acidilactici. The experimental activity of this probiotic bacterium was evaluated in an animal model by culture and histopathological methods. P. acidilactici inhibited the growth of Candida species at concentrations of 8-512 µg/mL (approximately 8 × 106 to 5 × 108 CFU/ml). This probiotic bacterium inhibited the germ tube and biofilm formation in a dose-dependent manner. RT-PCR analysis showed a reduction in genes expression. The P. acidilactici reduced the CFUs in mice receiving this probiotic bacterium. Histopathological analyses showed that Candida colonization was diminished in mice following the administration of probiotic. Since the wide range of antifungal activity of this bacterium bacterium, it can be used to manage oral candidiasis.

The aim of this study was to evaluate the phytochemical constituents, antioxidant, antifungal, and anti-virulence activities of traditionally used Mezoneuron benthamianum leaves. Extracts were prepared using acetone and methanol, and the preliminary phytochemical screening was performed. The antioxidant activity was studied using the DPPH method. Anti-Candida albicans activity was established and the effect on the germ tube and phospholipase production, as well as on the host cell adherence was assessed. The extracts showed the presence of anthraquinones, cardiac glycosides, flavonoids, reducing sugars, saponins, steroids, tannins, and terpenoids. Gallic acid and trans-resveratrol were among the predominant phytochemicals found in M. benthamianum. The crude extracts presented significantly higher antioxidant activity than the ascorbic acid standard. At 0.39 mg/mL, acetone extract inhibited the growth of Candida albicans. At lower concentrations (200-50 µg/mL), it significantly inhibited the adherence ability (up to 51%), formation of hyphae (up to 65%), and the production of phospholipase. In conclusion, at high concentrations, M. benthamianum kills C. albicans, and at lower concentrations, it can inhibit the virulence properties of this pathogen. This study on crude extract validates the traditional use of this plant. However, further research is required to establish the anti-virulence activity of the two compounds and their therapeutic potential.

Background and Aim: Candida albicans is a dimorphic fungus that has both yeast and filamentous forms. It is part of the normal flora in the oral and genital areas of mammals. One factor for the pathogenicity of C. albicans is its ability to switch from yeast to hyphae. The hyphal form adheres and penetrates tissues more readily than the yeast form and produces biofilms that are associated with chronic infection. Biofilms are protective niches that enable microorganisms to be more resistant to antibiotic treatment, thus allowing for persistent infection. The first stage in the transition from yeast to hyphae involves the formation of a germ tube, and this transition is triggered by interactions with host cells. Germ tube formation is dependent on serum, pH, temperature, and quorum-sensing molecules (QSMs). Farnesol, which is a QSM in C. albicans, can prevent yeast to hyphae conversion and inhibits the growth of fungal biofilm. Lyticase is a synergistic enzyme complex that catalyzes yeast cell lysis by β-1,3-glucanase and is a highly specific alkaline protease that produces protoplasts or spheroplasts. This study investigated the effect of farnesol and lyticase on the formation of C. albicans biofilms. Materials and Methods: C. albicans ATCC 2091 was cultivated on liquid and solid Sabouraud media. The presence of C. albicans was confirmed using HiCrome Candida Agar chromogenic medium. Enzyme activities were assayed using a HiCandida Identification Kit. The morphology and densitometry parameters of C. albicans biofilms were considered in the presence of farnesol (Sigma-Aldrich, Germany), lyticase (from Arthrobacter luteus; Sigma-Aldrich, Germany), and farnesol–lyticase. Results: This study shows that both farnesol and lyticase possess antifungal activity against C. albicans biofilms. A significant difference among treatment groups (p

Context: Candida is a yeast-like fungus, and it causes candidiasis. Since it is commonly encountered in many cases, the need of the hour is for rapid and reliable method for identification of these fungi in tissue sections. Aim: The aim of this study was to evaluate and compare the staining efficacy of calcofluor white (CFW) and acridine orange (AO) for the detection of Candida species in formalin-fixed paraffin-embedded tissue samples of oral squamous cell carcinoma (OSCC) using fluorescence microscopy. Settings and Design: Sample size comprised forty cases of OSCC. Materials and Methods: Before tissue sampling, a swab of the area was taken, it was immediately inoculated on Sabouraud's dextrose agar media and germ tube test was performed for positive cultures for species identification. Tissue sections were obtained from cases of OSCC from the formalin-fixed paraffin-embedded tissue blocks of the same cases in which microbiological assessment was done at the time of tissue sampling, were stained with CFW and AO stain, respectively, and were examined using a fluorescent microscope. Statistical Analysis Used: Descriptive statistics were expressed in numbers and percentage. Independent t-test (unpaired t-test) and Chi-square test were used. P ≤0.05 was taken to be statistically significant. Results: The mean number of microorganisms per high-power field stained by CFW and AO was 6.35 and 2.57, respectively, and a statistically significant difference (P ≤ 0.001) was observed. CFW compared to swab culture gave P = 0.018, which showed a statistically significant association. Conclusions: CFW is a better fluorescent stain when compared to AO to detect Candida species in tissue sections of OSCC cases.

Bu çalışmada, borik asitin Penicillium expansum’a karşı etkinliği hem in vitro hem de in vivo koşullarda değerlendirilmiştir. In vitro’da, borik asitin %0.125 konsantrasyonu P. expansum’un misel gelişmesini, spor çimlenmesini ve çim tüpü uzunluğunu sırasıyla %68.88, %74.00 ve %85.50 oranında azaltmış, halbuki daha yüksek konsantrasyonlar (%0.25, %0.50, %1.00 ve %2.00)’ı ise tamamen engellemiştir. Ayrıca borik asitin yukarıda belirtilen ilk konsantrasyonu ve sonraki konsantrasyonlarının etkileri arasındaki fark istatistiksel olarak önemli bulunmuştur (P%2 olarak belirlenmiştir. In vivo denemelerde, borik asitin %0.25 konsantrasyonu hariç, diğer tüm konsantrasyonlar (%0.50, %1.00 ve %2.00)’ı hem koruyucu hem de tedavi edici etki göstermişler ve bu konsantrasyonlar mavi küfün hastalık şiddetini patojen ile inokule edilmiş kontrole kıyasla önemli ölçüde azaltmıştır (P

Summary The cell wall of filamentous fungi, comprised of chitin, polysaccharide and glycoproteins, maintains the integrity of hyphae and protect them from defence responses by potential host plants. Here, we report that one polysaccharide deacetylase of Puccinia striiformis f. sp. tritici (Pst), Pst_13661, suppresses Bax‐induced cell death in plants and Pst_13661 is highly induced during early stages of the interaction between wheat and Pst. Importantly, the transgenic wheat expressing the RNA interference (RNAi) construct of Pst_13661 exhibits high resistance to major Pst epidemic races CYR31, CYR32 and CYR33 by inhibiting growth and development of Pst, indicating that Pst_13661 is an available pathogenicity factor and is a potential target for generating broad‐spectrum resistance breeding material of wheat. It forms a homo‐polymer and has high affinity for chitin and germ tubes of Pst compared with the control. Besides, Pst_13661 suppresses chitin‐induced plant defence in plants. Hence, we infer that Pst_13661 may modify the fungal cell wall to prevent recognition by apoplastic surveillance systems in plants. This study opens new approaches for developing durable disease‐resistant germplasm by disturbing the growth and development of fungi and develops novel strategies to control crop diseases.

Filamentous fungi are invasive and multidrug resistant pathogens that commonly contaminate biomedical devices and implants. Once spherical fungal spores attach to a surface, they exhibit germ tube development, hyphal growth, and robust biofilm formation. Nanotopography found on plants, reptiles, and insect wings possess bactericidal properties during prokaryotic cell adhesion. Here, we demonstrate the application of biomimetic nanopillars that inhibit eukaryotic filamentous fungal growth and possess fungicidal properties. Furthermore, many spores on the nanopillars appeared deflated, while those on the flat surfaces remained spherical and intact. These antifungal phenomena provide promising applications in antifouling biointerfaces for biomedical devices and implants.

Penicillium roqueforti is used for the production of blue-veined cheeses but is a spoilage fungus as well. It reproduces asexually by forming conidia. Germination of these spores can start the spoilage process of food. Germination is typically characterized by the processes of activation, swelling and germ tube formation. Here, we studied nutrient requirements for germination of P. roqueforti conidia. To this end, > 300 conidia per condition were monitored in time using an oCelloScope imager and an asymmetric model was used to describe the germination process. Spores were incubated for 72 h in NaNO3, Na2HPO4/NaH2PO4, MgSO4 and KCl with 10 mM glucose or 10 mM of 1 out of the 20 proteogenic amino acids. In the case of glucose, the maximum number of spores (Pmax) that had formed germ tubes was 12.7%, while time needed to reach 0.5 Pmax (τ) was about 14 h. Arginine and alanine were the most inducing amino acids with a Pmax of germ tube formation of 21% and 13%, respectively, and a τ of up to 33.5 h. Contrary to the typical stages of germination of fungal conidia, data show that P. roqueforti conidia can start forming germ tubes without a detectable swelling stage.

Candida spp. is the most common microbial pathogen in fungal infections. There has been a tremendous increase in cases of candidiasis, especially among critically ill non-neutropenic patients. Candida albicans’ isolates were procured from the Prince Sultan Military Hospital, Riyadh, KSA. The isolates were characterized for their identification using CHROMagar, carbohydrate metabolism, germ tube formation, and RAPD-PCR techniques. The essential oil of Thymus vulgaris was obtained by hydro-distillation and characterized to decipher the major bioactive phytoconstituents. The antifungal activity of the thyme essential oil (TEO) was evaluated against fluconazole-resistant C. albicans isolates. The major phytocomponents identified by GC/MS were thymol (68.1%) followed by γ-terpinene (8.9%), cymol (7.7%), caryophyllene (1.1%), linalool (1.4%). The TEO successfully reduced the growth of C. albicans isolates. At very low doses, the TEO proved to be fungi static and fungicidal. TEO also effectively inhibited the germ tube formation and budging of fungal pathogens. The time kill assays have shown that TEO was more effective against drug resistant clinical isolates than fluconazole. This study provides an array of experimental evidence regarding the therapeutic efficacy of TEO against the drug-resistant clinical isolates of C. albicans. The findings may be used in the development of a new antifungal agent accordingly.

Due to their high content of biomolecules, combined with silver’s well known antimicrobial potential, silver nanoparticles biosynthesized using spruce bark (AgNP SBEs) demonstrate antibacterial and antioxidant activity, making them a versatile option for developing new antimicrobial agents that might be used for medical treatment or as adjuvants for the classical agents. This study aims to analyze if silver nanoparticles (AgNPs) mediated by spruce bark extract (SBE) and silver salts (AgNP SBE Acetate, AgNP SBE Nitrate) presents antifungal activity against five different Candida spp., synergistic activity with fluconazole, and if they interact with some virulence factors of C. albicans. AgNP SBEs presented MICs (minimum inhibitory concentrations) for all the five tested Candida spp. AgNP SBEs inhibited the growth of C. parapsilosis, C. krusei, and C. guilliermondii, exerted synergistic activity with fluconazole for C. parapsilosis and C. guilliermondii, and inhibited biofilm production for C. albicans, C. auris, and C. guilliermondii. MICs of AgNP SBE Acetate significantly inhibited the production of germ tubes of C. albicans. The expression of C. albicans&nbsp, SAP2 gene was down-regulated by the short-time treatment with MICs of AgNP SBE Acetate, while ALS3 and HSP70 genes were up-regulated by the AgNPs MICs. These results emphasize the potential of using the AgNP SBEs as treatments/adjuvants options, not only against the redundant C. albicans but also for the non-albicans Candida species (which are not as frequently involved in human pathologies, but, sometimes, can be more aggressive).

Biosynthesis is a green method for the synthesis of silver nanoparticles (AgNPs). This study aimed to assess the antifungal activity of two silver nanoparticle solutions, synthesized using beech bark extract (BBE) and acetate and nitrate silver salts (AgNP Acetate BBE and AgNP Nitrate BBE), their influence on biofilm production, their potential synergistic effects with fluconazole, on different Candida spp., and their influence on virulence factors of C. albicans (germ tube production, gene expression for ALS3, SAP2, HSP70). Both the AgNP BBEs presented different minimum inhibitory concentrations for all the studied Candida spp., but biofilm production was inhibited only for C. albicans and C. guilliermondii. The growth rates of all the studied Candida spp. were inhibited in the presence of both AgNP BBEs, except for C. auris. Synergistic activity was observed for C. parapsilosis and C. guilliermondii, for different combinations of fluconazole with both the AgNP BBEs. The germ tube production of C. albicans was slightly inhibited by the AgNP BBEs. Only AgNP Acetate BBE was able to down-regulate the expression of SAP2. Overall, we can conclude that, even if more studies are necessary, AgNPs synthesized with beech bark extract might be an interesting alternative to classic antifungal treatments.

JRK’s DOCD is a proprietary siddha medicine, indicated for prevention of co-morbidities of diabetes. Earlier studies on the product has proven its effect in boosting the phagocyte mediated immunity. In the current study the ability of the product in preventing pathogenicity of fungi is evaluated. Germ tube formation is proven to increase the pathogenicity of the fungi than the spores alone. For this different species of fungi i.e., Candida albicans, 2 isolates of Candida dubelensis and three isolates of Mucor and 1 isolate of Rhizopus are taken. These fungal cultures when treated with the JRK’s DCOD, there is an inhibition of germ tube formation compared to control. Complete study details are presented in the paper.

A key question that has remained unanswered is how pathogenic fungi switch from vegetative growth to infection-related morphogenesis during a disease cycle. Here, we identify a fungal oxylipin analogous to the phytohormone jasmonic acid (JA), as the principal regulator of such a developmental switch to isotropic growth and pathogenicity in the rice-blast fungus Magnaporthe oryzae. Using specific inhibitors and mutant analyses, we determined the molecular function of intrinsic jasmonates during M. oryzae pathogenesis. Loss of 12-Oxo-phytodienoic Acid (OPDA) Reductase and/or consequent reduction of jasmonate biosynthesis, prolonged germ tube growth and caused delayed initiation and improper development of infection structures in M. oryzae, reminiscent of phenotypic defects upon impaired cyclic AMP (cAMP) signaling. Chemical- or genetic-complementation completely restored proper vegetative growth and appressoria in opr1Δ. Mass spectrometry-based quantification revealed increased OPDA accumulation and significantly decreased jasmonate levels in opr1Δ. Most interestingly, exogenous jasmonate restored proper appressorium formation in pth11Δ that lacks G protein/cAMP signaling, but failed to do so in the Mitogen-activated protein (MAP) kinase mutants. Epistasis analysis placed jasmonate upstream of the cAMP pathway in rice blast. Mechanistically, intrinsic jasmonate orchestrates timely cessation of the vegetative phase and induces pathogenic development via a complex regulatory interaction with the cAMP-PKA cascade and redox signaling in rice blast.

Orobanche cumana WALLR. is a host-specific root parasite of cultivated sunflowers with increasing economic importance in Europe, North Africa, and parts of Asia. While sesquiterpene lactones (STLs) released from sunflower roots were identified as natural germination stimulants of O. cumana seeds in the soil, the chemical nature of the signals guiding the emerging germ tube toward the host root has remained unknown hitherto. Thus, we designed a bioassay that allowed the observation of broomrape germination and subsequent germ tube development in the presence of substances with putative chemotropic activity. Root exudates and sunflower oil extracts, both containing STLs in micromolar concentrations, caused the positive chemotropic orientation of germ tubes. A similar positive chemotropic effect was achieved with costunolide, one of the four STLs of sunflower present in the exudate and oil extracts. In contrast, GR24, a synthetic strigolactone (SL) with germination-inducing activity on O. cumana seeds, showed no effect on the germ tube orientation. The effect of costunolide was concentration-dependent and within the range of its natural micromolar occurrence in roots. We assume that an STL gradient is responsible for the stronger inhibition of elongation growth on the host-facing flank of the germ tube compared with the far side flank. This would confer a double role of STLs from sunflower root exudates in the sunflower–broomrape interaction, namely, as germination stimulants and as chemotropic signals.

Novel agroformulations for simultaneous delivery of chemical and biologically active agents to the plants were prepared by encapsulation of Trichoderma viride spores in calcium alginate microspheres. The impact of calcium ions concentration on the viability and sporulation of T. viride spores as well as on the microsphere important physicochemical properties were investigated. Intermolecular interactions in microspheres are complex including mainly hydrogen bonds and electrostatic interactions. T. viride germination inside matrix and germ tubes penetration out of microspheres revealed calcium alginate microspheres provide a supportive environment for T. viride growth. Differences in physicochemical properties and bioactive agents release behaviour from microspheres were ascribed to the changes in microsphere structure. Fitting to Korsmeyer-Peppas empirical model revealed the underlying T. viride release mechanism as anomalous transport kinetics (a combination of two diffusion mechanisms and the Type II transport (polymer swelling and relaxation of the polymeric matrix)). The increasing amount of T. viride spores in the surrounding medium is closely related to the release from microspheres and germination. The rate controlling mechanism of calcium release is Fickian diffusion. A decrease in the release rate with increasing calcium ion concentrations is in accordance with the calcium ions effect on the strength of the alginate network structure. T. viride germination inside microsphere diminished the amount of released calcium ions and slowed release kinetics in comparison with microspheres prepared without T. viride. The results indicated investigated agroformulations have a great potential to be used for plant protection and nutrition.

Pyropia is a genus of algae used in sushi. Nitrate (NO3−) is essential for its growth. Excess NO3− is flushed into seawater, as a result of anthropogenic activities, increasing the potential for higher than normal concentrations which could be harmful to both algae and humans. This study aims to evaluate the effect of NO3− on the morphology, ultrastructure, spore viability, autofluorescence of chloroplasts, and formation of cell wall, during germination of the red seaweed Pyropia acanthophora var. brasiliensis. To accomplish this, carpospores were cultivated at 24°C, 40 μmol photons m−2 s−1 with photoperiod of 12 h and exposed to 0, 25, 50, and 100 mM of NO3− for 2 days. Samples were cultured for another 5 days to measure the length of the germ tube. Concentrations of NO3− did not significantly influence the development, morphology, viability, or ultrastructure of the spores in the first 2 days. The absence of this nutrient did delay the ultrastructural development of the carpospore, decreasing autofluorescence of the chloroplast and delaying the formation of the cell wall and pyrenoid. Over the seven-day experimental period, higher concentrations of NO3− increased the size of the germ tube, indicating that this extra concentration boosted the development of P. acanthophora.

Mavi küf hastalığına neden olan Penicillium expansum yumuşak çekirdekli meyvelerde hasat sonrasıgörülen önemli patojenlerden biri olup, meyve enfeksiyonu için yaraya ihtiyaç duyan nekrotrofik birfungustur. Bu çalışmada, etidot-67 ve boraks dekahidratın P. expansum’a karşı etkinliği hem in vitrohem de in vivo denemelerle değerlendirilmiştir. In vitro denemelerde, hem etidot-67 hem de boraksdekahidrat P. expansum’un misel gelişmesini, spor çimlenmesini ve çim tüpü uzunluğunu güçlü birşekilde engellemiştir. Her iki tuzun engelleyici etkileri onların artan konsantrasyonları ile yakındanilişkili bulunmuştur. Bor tuzları P. expansum'un misel gelişmesini %0.25 konsantrasyonda tamamenengellerken, bu tuzlar fungusun spor çimlenmesini ve çim tüp uzamasını %0.125 konsantrasyonundatamamen engellemiştir. Ayrıca, test edilen tuzların toksisitelerinin birbirine yakın olduğu, yani etidot-67ve boraks dekahidratın EC50 değerlerinin sırasıyla 0.067 ve 0.071 olduğu belirlenmiştir. Etidot-67 veboraks dekahidratın minimum engelleyici konsantrasyon (MIC) değerleri %0.25 iken, aynı tuzlarınminimum fungisidal konsantrasyon (MFC) değerleri %1'den büyük bulunmuştur. In vivo denemelerde,fungus inokülasyonundan sonra (tedavi edici aktivite), etidot-67 ve boraks dekahidratın %3.0konsantrasyonu uygulanmış elma meyvelerinde mavi küf gelişimi, kontrol uygulaması ilekıyaslandığında, sırasıyla %92.8 ve %78.9’a kadar önemli derecede azalmıştır (P0.05). Bununlabirlikte, aynı konsantrasyonda, fungus (1x104konidi mL-1) ile inokülasyondan önce (koruyucu aktivite)bor tuzları ile muamele edilen elma meyvelerindeki lezyon alanı kontrol uygulaması ile kıyaslandığındasırasıyla %94.3 ve %98.3 azalmıştır. Bu sonuçlar, bor tuzlarının P. expansum'un neden olduğu elmameyvesinin hasat sonrası hastalığının kontrolü için sentetik fungisitlere potansiyel bir alternatif olarakkullanılabileceğini göstermektedir. Penicillium expansum causing blue mold disease is one of the most important pathogens of pome fruitand is a necrotrophic fungus that requires wounds to infect the fruit. In the present study, the efficacy ofetidot-67 and borax decahydrate against P. expansum were evaluated in both in vitro and in in vivo. Inin vitro experiments, both etidot-67 and borax decahydrate strongly inhibited mycelial growth, sporegermination and germ tube elongation of P. expansum. The inhibitory effects of both salts were closelycorrelated with their increasing concentrations. While boron salts completely inhibited the mycelialgrowth of P. expansum at %0.25 concentration, these salts completely inhibited spore germination andgerm tube elongation of the fungus at %0.125 concentration. Additionally, it was determined that thetoxicity of the salts tested were close to each other, namely EC50 values of Etidot-67 and boraxdecahydrate were 0.067 and 0.071, respectively. While the minimum inhibition concentration (MIC)values of the etidot-67 and borax decahydrate were %0.25, the minimum fungicidal concentration(MFC) values of the same salts were found to be greater than 1%. In in vivo experiments, blue molddevelopment in apple fruits treated with 3.0% concentration of etidot-67 and borax decahydrate afterfungal inoculation (curative activity) was significantly reduced by %92.8 and %78.9, respectively,compared with the control treatment (P0.05). However, at same concentration, lesion area on applefruits treated with the salts before inoculation with the fungus (1x104conidia mL-1) (preventive activity)reduced by %94.3 and %98.3, respectively. These results show that both boron salts can be used as apotential alternative to synthetic fungicides for the control of the postharvest disease of apple fruitcaused by P. expansum.

To develop an antimicrobial agent for preventing the devasting damage caused by rice blast, a novel peptide aptamer was identified to interact with calmodulin (CaM) for the inhibition of the spore development in the pathogen Magnaporthe oryzae. A peptide aptamer designated as SNP-D4, consisted of the scaffold protein Staphylococcus aureus nuclease (SN) and an exposed surface loop of 16 random amino acids, was screened from the constructed peptide aptamer libraries by bacterial two-hybrid system using CaM of M. oryzae as the bait. The preliminary inhibition in the sporulation development was observed after treating with the crude extracts expressing SNP-D4. The inhibition efficacies of the purified SNP-D4 were quantified at the stages of conidial germination, germ tube elongation, and appressorium formation in M. oryzae. The binding affinity analysis revealed that SNP-D4 interacted with CaM at a dissociation constant (Kd) of about 20 μM. Moreover, the N-terminus of CaM was identified as the key binding region.

Purpose: The pathogenicity in Candida spp was attributed by several virulence factors such as production of tissue damaging extracellular enzymes, germ tube formation, hyphal morphogenesis and establishment of drug resistant biofilm. The objective of present study was to investigate the effects of silver nanoparticles (AgNPs) on growth, cell morphology and key virulence attributes of Candida species. Methods: AgNPs were synthesized by the using seed extract of Syzygium cumini (Sc), and were characterized by UV-Vis spectrophotometer, Fourier-transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), energy-dispersive X-ray (EDX), and transmission electron microscopy (TEM). ScAgNPs were used to evaluate their antifungal and antibacterial activity as well as their potent inhibitory effects on germ tube and biofilm formation and extracellular enzymes viz. phospholipases, proteinases, lipases and hemolysin secreted by Candida spp. Results: The MICs values of ScAgNPs were ranged from 0.125-0.250 mg/ml, whereas the MBCs and MFCs were 0.250 and 0.500 mg/ml, respectively. ScAgNPs significantly inhibit the production of phospholipases by 82.2, 75.7, 78.7, 62.5, and 65.8%; proteinases by 82.0, 72.0, 77.5, 67.0, and 83.7%; lipase by 69.4, 58.8, 60.0, 42.9, and 65.0%; and hemolysin by 62.8, 69.7, 67.2, 73.1, and 70.2% in C. albicans, C. tropicalis, C. dubliniensis, C. parapsilosis and C. krusei, respectively, at 500 μg/ml. ScAgNPs inhibit germ tube formation in C. albicans up to 97.1% at 0.25 mg/ml. LIVE/DEAD staining results showed that ScAgNPs almost completely inhibit biofilm formation in C. albicans. TEM analysis shows that ScAgNPs not only anchored onto the cell surface but also penetrated and accumulated in the cytoplasm that causes severe damage to the cell wall and cytoplasmic membrane. Conclusion: To summarize, the biosynthesized ScAgNPs strongly suppressed the multiplication, germ tube and biofilm formation and most importantly secretion of hydrolytic enzymes (viz. phospholipases, proteinases, lipases and hemolysin) by Candia spp. The present research work open several avenues of further study, such as to explore the molecular mechanism of inhibition of germ tubes and biofilm formation and suppression of production of various hydrolytic enzymes by Candida spp.

Curcumin is a promising antifungal agent because of its wide range of antifungal activity. In the present study, the effect of curcumin on gray mold (Botrytis cinerea) in kiwifruit was evaluated. Results showed that spore germination, germ tube elongation, and mycelial growth of B. cinerea were significantly suppressed by curcumin in a dose dependent manner. Mycelium treated with curcumin exhibited a higher sensitivity to osmotic stress, and the ability of hyphae to penetrate plant cell walls was also inhibited. Three mitogen-activated protein kinase (MAPK) genes, Bmp1, Bmp3, and Sak1, involved in vegetative growth, osmotic stress response, and pathogenicity, were down-regulated by curcumin. The application of curcumin up to 400 mg L−1 significantly delayed disease development in kiwifruit. These results indicate that curcumin can effectively inhibit gray mold in kiwifruit; and this effect involves alterations in the sensitivity of B. cinerea to pathogenicity-associated MAPK cascades.

This study investigated the effect of surface wettability, hardness and surface contact on the germination and subsequent development of Botryosphaeriaceae species conidia. Mean percent germination for isolates Neofusicoccum luteum MM558, N. luteum CC445, N. parvum G652 and Botryosphaeria dothidea 007 was highest on cellulose which was the most hydrophilic surface (72.2%, 76.9%, 52.4% and 74.9%, respectively), but did not differ between the other surfaces tested (glass, polystyrene and Parafilm M™). Germ tube growth for all isolates was least on cellulose and similar on the other surfaces. Cellulose agar assays indicated that mycelia could be utilising the cellulose as a food source. Germination assays on surfaces of different hardness showed no discernible patterns of germination or growth related to hardness. Further, there was no effect on germination of surface contact versus continuous shaking of conidial suspensions for isolate N. luteum MM558. These assays demonstrated that conidia of the Botryosphaeriaceae were able to germinate on a variety of surfaces with this flexibility indicative of their reported pathogenicity of different host tissues.

As a powerful sanitizer, chlorine dioxide (ClO2) has mainly focused on control of bacteria, but little is known about its antifungal effects, especially on postharvest fungi found in vegetable and fruit. This study investigated the effect of ClO2 on Botrytis cinerea growth in vitro and on its disease severity in green pepper and winter jujube. Result showed that ClO2 fumigation reduced disease development of grey mold caused by B. cinerea on green pepper and winter jujube, and the efficiency was positively correlated with ClO2 concentrations. ClO2 treatment directly inhibited mycelial growth, spore germination and germ tube elongation of B. cinerea. To further survey the mechanisms by which ClO2 treatment inhibited hypha growth of B. cinerea, we observed the mycelial morphology and the integrity of the plasma membrane; the results showed that ClO2 treatment resulting in B. cinerea mycelial morphology was strongly affected, and leading to the loss of plasma membrane integrity, causing the cellular leakage. These data showed that ClO2 was effective for controlling grey mold caused by B. cinerea on green pepper and winter jujube, and the antifungal activity of ClO2 could be attributed to the disruption of cell membrane of the fungal pathogen.

Epsilon-polylysine (e-PL) is a natural antimicrobial and has been widely used as a food preservative. Previous studies showed e-PL being effective for control of foodborne bacteria; however, little is known about its control effects on postharvest pathogenic fungi and the possible modes of action. Here, we found that e-PL could significantly inhibit mycelial growth, spore germination and germ tube elongation of Botrytis cinerea in vitro, as well as effectively control gray mold in harvested jujube fruit. Further investigation indicates that the modes of action of e-PL against B. cinerea include (i) stimulating the accumulation of intracellular reactive oxygen species (ROS) and reducing the expression of pathogenesis-related genes of the pathogen, (ii) resulting in the leakage of soluble carbohydrates and nucleic acids, as well as the damage of plasma membrane integrity of the fungal cells, (iii) inducing the expression of respiratory burst oxidase homolog (RBOH) genes in e-PL-treated jujube fruit.

This study examines the ability of the quorum-sensing molecules (QSMs) farnesol and tryptophol to induce programmed cell death of the pathogenic fungus Candida albicans, to alter the expression of apoptosis-related genes, and to reduce the pathogenicity and virulence of C. albicans in Galleria mellonella. Our results showed that both farnesol and tryptophol inhibited C. albicans germ tube formation. In the QSM-treated group, the expression levels of the apoptosis genes increased, whereas the expression level of the anti-apoptosis gene decreased. Further, pretreatment of C. albicans with tryptophol or farnesol prior to G. mellonella larval infection significantly enhanced host survival compared with larvae infected with untreated C. albicans. Thus, farnesol and tryptophol may trigger apoptosis of C. albicans in vitro and reduce the virulence of C. albicans in vivo. Although further study is needed to identify the precise mechanisms underlying the antifungal properties of farnesol and tryptophol, these results suggest that QSMs may be effective agents for controlling fungal infection.

Satureja khuzistanica Jamzad is known as antiseptic and analgesic agent in folk medicine. The aim of this investigation was to evaluate the anti-candidal activity of S. khuzistanica aerial parts essential oil against clinical isolates of Candida albicans, which were isolated from women with chronic recurrent candidiasis. For this purpose, the chemical composition of hydro-distilled essential oil was determined by GC and GC-MS analysis. Then, the anti-candidal activity of essential oil and its main component (carvacrol) were determined. Carvacrol (94.1%) was the main component of essential oil, followed by β-bisabolene, p-cymene and γ-terpinene. S. khuzistanica essential oil had strong anti-candidal activity against clinical isolates of C. albicans via inhibition of germ tube formation and induction the huge punctures in the cytoplasmic structures. The cell membranes were intact in presence of essential oil or carvacrol. S. khuzistanica essential oil as the main source of carvacrol can be used for treatment of C. albicans related infections. Resumen Satureja khuzistanica Jamzad es conocido como analgésico y antiséptico en la medicina tradicional. El objetivo de esta investigación fue evaluar el efecto anti- Candida de los aceites esenciales obtenidos de las partes aéreas de S. khuzistanica sobre aislados clínicos de Candida albicans, obtenidos de mujeres con candidadiasis crónica recurrente. Para este propósito la composición química de aceites esenciales hidrodestilados fueron determinados por análisis GC y GC-MS. Luego la actividad anti-candidasica de los aceites esenciales y de su componente principal (carvacrol) fue determiando. Carvacrol (94.1%) fue el principal compuesto del aceite esencial seguido por β-bisaboleno, p-cimeno and γ-terpineno. El aceite esencial de S. khuzistanica tuvo fuerte actividad anti-candida contra aislados clínicos de C. albicans via la inhibicion de tubo germinal y la inducción de estructuras puntiformes en la membrana citoplásmatica. Las membranas celulares quedaron intactas en presencia del aceite esencial o del carvacrol. El aceite esencial de S. khuzistanica como fuente principal de carvacrol podría ser usado como tratamiento de infecciones relacionadas con Candida albicans.

BACKGROUND Sanguinarine (SAN) is a benzophenanthridine alkaloid that broadly targets a range of pathways in mammalian and fungal cells. In this study we set out to explore the molecular mechanism of sanguinarine inhibition of the fungal development and pathogenicity of Magnaporthe oryzae with the hope that sanguinarine will bolster the development of antiblast agents. RESULTS We found that the fungus exhibited a significant reduction in vegetative growth and hyphal melanization while the spores produced long germ tubes on the artificial hydrophobic surface characteristic of a defect in thigmotropic sensing when exposed to 4, 8 and 0.5 μm sanguinarine, respectively. Consistent with these findings, we observed that the genes involved in melanin biosynthesis and the fungal hydrophobin MoMPG1 were remarkably suppressed in mycelia treated with 8 μm sanguinarine. Additionally, sanguinarine inhibited appressorium formation at a dose of 1.0 μm and this defect was restored by supplementing 5 mM of exogenous cAMP. By qRT-PCR assay we found cAMP pathway signalling genes such as MoCAP1 and MoCpkA were significantly repressed whereas MoCDTF1 and MoSOM1 were upregulated in sanguinarine-treated strains. Furthermore, we showed that sanguinarine does not selectively inhibit vegetative growth and appressorium formation of Guy11 but also other strains of M. oryzae. Finally, treatment of sanguinarine impaired the appressorium-mediated penetration and pathogenicity of M. oryzae in a dose-dependent manner. CONCLUSION Based on our results we concluded that sanguinarine is an attractive antimicrobial candidate for fungicide development in the control of rice blast disease. © 2021 Society of Chemical Industry.

Chlorogenic acid is a plant polyphenol with antioxidant and antimicrobial activities. Fusarium fujikuroi is a fungal pathogen that causes many vegetables and fruits, including tomato, to rot. The effects of chlorogenic acid on the development of Fusarium rot of cherry tomato fruit were examined in the present study. Results showed that conidial germination, germ tube elongation, cell viability, and mycelial growth of F. fujikuroi were all significantly inhibited by chlorogenic acid. Chlorogenic acid stimulated the accumulation of reactive oxygen species (ROS), leading to cell apoptosis in F. fujikuroi. The addition of N-acetylcysteine partially recovered the mycelial growth, implying the antifungal activity of chlorogenic acid is related to a ROS burst. The application of chlorogenic acid decreased disease incidence and severity in cherry tomato fruit in a concentration-dependent manner. Taken together, these results suggest that chlorogenic acid inhibits the postharvest rot of cherry tomato fruit caused by F. fujikuroi by inducing cellular oxidative stress in the pathogen.

Candida albicans infection mainly occurs in patients with suppressed immune function, and it is also the main pathogen of hospital infection. The new strategies are needed to treat the existing resistance of antifungal drugs. The use of natural products aimed at controlling fungal diseases is considered an interesting alternative to synthetic fungicides due to their lower adverse reactions, lower cost to plant preparations compared to modern conventional pharmaceuticals. Natural borneol has a long history of treating ulcers and local infections. In this study, the minimum inhibitory concentration of natural borneol on ATCC10231 and 10 clinically isolated Candida albicans was determined by vapor phase method and dilution method, and the influence of sub-minimum inhibitory concentration on the formation of Candida albicans hyphae was observed. We found that the minimum inhibitory concentration of ATCC10231 and 10 clinically the isolates in the vapor phase were both 0.4 mg/cm3, agar and broth dilution methods were 2 mg/mL. The vapor phase of natural borneol has a better inhibitory effect on Candida albicans, Sub-mic concentration of borneol (0.125-1mg/ml) in the liquid phase inhibits the 60%-99% formation of Candida albicans germ tube. Natural borneol is a potential natural medicine for the treatment and prevention of Candida albicans infection. It brings new insights into the development of novel effective antifungal drugs.

La eficacia antifúngica de los extractos acuosos de mesocarpio de coco (Cocos nucifera L.) (EAC) y el quitosano comercial (QC) aplicados de manera individual y en combinación se evaluó contra Rhizopus stolonifer en términos de crecimiento micelial, esporulación y germinación de las esporas, así como la obtención de los modelos de crecimiento primario en dos temperaturas de almacenamiento (15-25 ºC). La aplicación de los EAC al 10% redujo significativamente el crecimiento micelial de R. stolonifer (58.81 ± 6.48%); por otro lado, el QC (1.5%) mostró un efecto de control de hasta un 87%, sin embargo, la combinación de los EAC (10%) con el QC (1.5%) fue más efectiva al reducir el crecimiento micelial (> 93 %). Todos los tratamientos fueron efectivos para disminuir la producción de esporas (> 94 %) en comparación con el control (agar). El QC al combinarse con los EAC fue más efectivo inhibiendola elongación del tubo germinal (> 98 %) comparado con los tratamientos individuales (< 48 %). El modelo modificado de Gompertz mostró un ajuste adecuado para ambos rangos de temperatura (> 98-99 %) observándose diferencias significativas (p ≤ 0.05) entre las variables de velocidad máxima (Vmáx) y periodo de latencia (ƛ), no obstante, todos los tratamientos mostraron un efecto fungistático sobre el desarrollo micelial de R. stolonifer. La combinación de los EAC y el QC puede ser una alternativa eco-amigable contra la pudrición suave de los frutos de guanábana.

The exopolysaccharide galactosaminogalactan (GAG) has been well characterized in Aspergilli, especially the human pathogen Aspergillus fumigatus. It has been found that a five-gene cluster is responsible for GAG biosynthesis in Aspergilli to mediate fungal adherence, biofilm formation, immunosuppression or induction of host immune defences. Herein, we report the presence of the conserved GAG biosynthetic gene cluster in the insect pathogenic fungus Metarhizium robertsii to mediate either similar or unique biological functions. Deletion of the gene cluster disabled fungal ability to produce GAG on germ tubes, mycelia and appressoria. Relative to the wild type strain, null mutant was impaired in topical infection but not injection of insect hosts. We found that GAG production by Metarhizium is partially acetylated and could mediate fungal adherence to hydrophobic insect cuticles, biofilm formation, and penetration of insect cuticles. In particular, it was first confirmed that this exopolymer is responsible for the formation of appressorium mucilage, the essential extracellular matrix formed along with the infection structure differentiation to mediate cell attachment and expression of cuticle degrading enzymes. In contrast to its production during A. fumigatus invasive growth, GAG is not produced on the Metarhizium cells harvested from insect hemocoels; however, the polymer can glue germ tubes into aggregates to form mycelium pellets in liquid culture. The results of this study unravel the biosynthesis and unique function of GAG in a fungal system apart from the aspergilli species., Author summary Fungal cell wall associated exopolymer GAG was first identified from Aspergillus niger decades ago and has been well characterized in terms of its biosynthetic mechanism, pathological and biological functions in Aspergillus fungi, especially the human pathogen A. fumigatus. With the accomplishment of diverse fungal genomes, the conserved GAG biosynthetic gene cluster is found to patchily distribute in fungal species beyond Aspergilli. However, until this study, the contribution of the clustered genes to GAG production has not been investigated in any non-Aspergillus fungi. We performed gene deletions and functional characterizations of GAG in an insect pathogenic fungus Metarhizium robertsii, and found that the partially acetylated GAG can be produced by the fungus to contribute to fungal virulence during topical infection of insect hosts. This exopolymer can be produced on germ tubes, mycelia and appressoria to mediate cell adherence, cuticle penetration, biofilm formation, mycelium aggregation in liquid culture and in particular, appressorium mucilaginous matrix formation. In addition, it was found that loss of GAG production could alter appressorium protein expressions including the enzymes involved in degrading insect cuticles. Our data advance the understanding of GAG biosynthesis and functions in an animal pathogenic fungus that is highly divergent from Aspergilli.

Silver nanoparticles (AgNPs) are metallic zero-dimensional nanomaterials that have attracted a lot of attention due to their conductivity, chemical resistance, antibacterial, antiviral, antifungal, antiangiogenic, and antiinflammatory properties. Applying Ag as a microbiocidal agent involves oxidation Ag0 to the Ag+. AgNPs are well-known for their excellent antimicrobial effects against numerous organisms such as bacteria, fungi, and viruses. Direct contact of AgNPs with large surfaces on the cell wall of bacteria can result in damage to the membrane, causing leakage of cell contents and death of cells. When pathogenic cells are exposed to the AgNPs, a significantly large number of ROS was generated. AgNPs can inhibit the production of different kinds of extracellular hydrolytic enzymes as well as germ tube formation of fungi. Also, AgNPs influence the biosynthesis of organic acid in some fungi and alter the profile of extracellular enzyme, although the total enzymatic activity is increased. Furthermore, AgNPs can disturb the integrity of cell wall and cell regulatory mechanisms, leading to pore formation and cell collapse. The antiviral properties of AgNPs have been reported recently. AgNPs are recorded to associate with glycoproteins on the viral surface and have links to the host cells and then introduce their virus behavior through contact with the viral genome.

Infections caused by Scedosporium species present a wide range of clinical manifestations, from superficial to disseminated, especially in immunocompromised patients. Glucosylceramides (GlcCer) are glycosphingolipids found on the fungal cell surface and play an important role in growth and pathogenicity processes in different fungi. The present study aimed to evaluate the structure of GlcCer and its role during growth in two S. aurantiacum isolates. Purified GlcCer from both isolates were obtained and its chemical structure identified by mass spectrometry. Using ELISA and immunofluorescence techniques it was observed that germination and NaOH-treatment of conidia favor GlcCer exposure. Monoclonal anti-GlcCer antibody reduced germination when cultivated with the inhibitor of melanin synthesis tricyclazole and also reduced germ tube length of conidia, both cultivated or not with tricyclazole. It was also demonstrated that anti-GlcCer altered lipid rafts organization, as shown by using the fluorescent stain filipin, but did not affect the susceptibility of the cell surface to damaging agents. Anti-GlcCer reduced total biomass and viability in biofilms formed on polystyrene plates. In the presence of anti-GlcCer, germinated S. aurantiacum conidia and biofilms could not adhere to polystyrene with the same efficacy as control cells. These results highlight the relevance of GlcCer in growth processes of S. aurantiacum.

Thymus albicans is an endemic species of the Iberian Peninsula with a vulnerable conservation status. In an attempt to contribute to the valorization of this species, the present study brings new insights on the antifungal and anti-inflammatory mechanism of action of T. albicans essential oil. The antifungal activity of the oil and its major compounds was assessed for the first time against standard and clinically isolated strains of yeasts and filamentous fungi. The effect on the two major virulence factors of Candida albicans (germ tube formation and biofilm disruption) was considered in more detail. At 0.08 μL/mL, the oil inhibited C. albicans germ tube formation by more than 40% and decreased biofilm biomass at MIC values, thus pointing out its antivirulent potential. The anti-inflammatory activity of the essential oil was investigated on LPS-stimulated mouse macrophages (RAW 264.7) by evaluating the levels of several pro-inflammatory mediators, namely nitric oxide (NO), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). T. albicans oil reduced the production of nitrites, a NO derived sub-product, at non-cytotoxic concentrations of 0.32 and 0.64 μL/mL, by 27 and 41%, respectively. In addition, the iNOS protein levels of essential oil pre-treated cells were reduced by 14%. Overall, the high essential oil yield of T. albicans as well as its bioactive effects at concentrations without cytotoxicity, encourage further studies on the potential pharmacological applications of this species. Furthermore, these results raise awareness for the need to preserve endangered species that may hold relevant medicinal value.

Aspergillosis cases by Aspergillus fumigatus have increased, along with fungal resistance to antifungals, urging the development of new therapies. Passive immunization targeting common fungal antigens, such as chitin and &beta, glucans, are promising and would eliminate the need of species-level diagnosis, thereby expediting the therapeutic intervention. However, these polysaccharides are poorly immunogenic. To overcome this drawback, we developed the lectin-Fc(IgG) fusion proteins, Dectin1-Fc(IgG2a), Dectin1-Fc(IgG2b) and wheat germ agglutinin (WGA)-Fc(IgG2a), based on their affinity to &beta, 1,3-glucan and chitooligomers, respectively. The WGA-Fc(IgG2a) previously demonstrated antifungal activity against Histoplasma capsulatum, Cryptococcus neoformans and Candida albicans. In the present work, we evaluated the antifungal properties of these lectin-Fc(s) against A. fumigatus. Lectin-Fc(IgG)(s) bound in a dose-dependent manner to germinating conidia and this binding increased upon conidia germination. Both lectin-Fc(IgG)(s) displayed in vitro antifungal effects, such as inhibition of conidia germination, a reduced length of germ tubes and a diminished biofilm formation. Lectin-Fc(IgG)(s) also enhanced complement deposition on conidia and macrophage effector functions, such as increased phagocytosis and killing of fungi. Finally, administration of the Dectin-1-Fc(IgG2b) and WGA-Fc(IgG2a) protected mice infected with A. fumigatus, with a 20% survival and a doubled life-span of the infected mice, which was correlated to a fungal burden reduction in lungs and brains of treated animals. These results confirm the potential of lectin-Fc(IgGs)(s) as a broad-spectrum antifungal therapeutic.

Background: Catheter associated urinary tract infection (CAUTI) is a crucial attribute to virulence of organisms. Emergency of non albicans Candida (NAC) species become common due to several virulence factors, which play an important role in pathogenicity and their resistance to treatment. The aim of work: To investigate different virulence factors of Candida spp. isolated from CAUTI patients and evaluate their antifungal susceptibility. Patients and methods: Urine samples collected from 219 indwelling catheters of ICU patients. Candida isolates identified using Germ tube test, Chrom ID agar, Corn meal tween agar, VITEK 2 system and Integral System Yeast Plus (ISYP) used to detect antifungal susceptibility. Enzymatic, haemolytic activity and biofilm formation assessed by standard methods. Results: The incidence of candiduria was 50/219 (22.8%). Candida tropicalis was predominant (54.0%) followed by C. albicans (26.0%) and C. glabrata (20.0%). Females gender (64.0%), Diabetes (50.0%) associated with positive candiduria, while antibiotic therapy was predominant risk factor (78.0%). C. albicans have highest enzymatic activity for phospholipase (84.6 %), proteinase (84.6%) and haemolytic activity (100%), while C. tropicalis have highest biofilm formation (100%). NAC spp. showed higher rates of resistance to azole drugs (Clotrimazole 77.6%, Miconazole 70.3%, Econazole 63.2%, Nystatine 44.3%), while C. albicans didn’t show any resistance. Conclusion: Candiduria is prevalent among catheterized patients with shift toward NAC spp. causing nosocomial UTI. A strong relationship between host risk factors (old age, antibiotic use, catheterization, female sex, ICU stay, diabetes) and expression of various virulence factors of Candida spp. causing candiduria and resistance to antifungal drugs.

BACKGROUND Essential oils (EO) extracted from Cinnamomum verum has been used as an antimicrobial agents for centuries. The effects of C. verum leaf oil against virulence of microorganisms is not well studied yet. OBJECTIVES This study evaluates the effect of C. verum leaf oil against three virulence factors of Candida albicans, C. tropicalis and C. dubliniensis and its in-vivo toxicity. METHODS Chemical composition of EO was determined using gas chromatography-mass spectrometry (GC-MS). Minimum inhibitory concentration (MIC) was determined using clinical and laboratory standards institute (CLSI) M27-A3 broth microdilution. Effect of EO on initial adhesion was quantified using XTT assay after allowing Candida cells to adhere to the polystyrene surface for 2 h. Biofilm formation of Candida in the presence of EO was quantified using XTT viability assay. Efficacy on reduction of germ tube formation was evaluated using standard protocol. Visualisation of biofilm formation and progression under the EO treatment were done using scanning electron microscope (SEM) and Time lapses microscope respectively. In-vivo toxicity of EO was determined using Galleria mellonella larvae. Chlorhexidine digluconate: positive control. RESULTS Eugenol was the main compound of EO. MIC was 1.0 mg/mL. 50% reduction in initial adhesion was achieved by C. albicans, C. tropicalis and C. dubliniensis with 1.0, > 2.0 and 0.34 mg/mL respectively. 0.5 and 1.0 mg/mL significantly inhibit the germ tube formation. MBIC50 for forming biofilms were ≤ 0.35 mg/mL. 1.0 mg/mL prevent biofilm progression of Candida. SEM images exhibited cell wall damages, cellular shrinkages and decreased hyphal formation. No lethal effect was noted with in-vivo experiment model at any concentration tested. CONCLUSION C. verum leaf oil acts against virulence factors of Candida and does not show any toxicity.

The prevalence and fatality rates with fungal biofilm-associated infections urgently need to develop targeted therapeutic approaches to augment the action of antifungal drugs. This study developed amphotericin B-loaded PLGA-PEG nanoparticles (AmB-NPs) with AD1 aptamer conjugation on its surface via an EDC/NHS technique. Their high nuclease resistance of the conjugation was confirmed by PAGE gel electrophoresis. The targeting and toxicity of AD1-AmB-NPs in the subcutaneous C. albicans infection model were evaluated. AD1-AmB-NPs can bind to different morphological forms(including yeast cells, germ tubes, hyphae) of C. albicans biofilms and extracellular matrix material. Low-frequency and low-intensity ultrasound (LFU, with a fixed frequency of 42 kHz, at the intensity of 0.30 W/cm2 for 15 min) significantly promoted permeability of the biofilm and allowed AD1-AmB-NPs into the deepest layers of the biofilm. After 7 days of treatment, the combination treatment of AD1-AmB-NPs and LFU, kills at least 99% of the biofilm fungal population in vivo comparison with ultrasound alone or AD1-AmB-NPs alone, and returned to normal subcutaneously. Our data suggest that the combined strategy of AD1-AmB-NPs and ultrasound treatment selective delivered of therapeutic drugs to the infection site and exhibited significant synergistic antifungal effects.

Candida albicans is a commensal yeast that may become pathogenic and even lethal to the host. Over the last few decades, antifungal resistance has increased, promoting screening of the antifungal potential of old and new substances. This study investigates the antifungal potential of isothiocyanates (ITCs) against C. albicans oral isolates. A preliminary susceptibility disk diffusion test (DD) was performed using allyl isothiocyanate (AITC), benzyl isothiocynanate (BITC) and phenyl ethyl isothiocyanate (PEITC) at a fixed concentration range (0.001–0.1 M). Because C. albicans isolates were more susceptible to BITC and PEITC, their effect on cell size and on germ tube formation (GTF) were tested. The most promising molecule, BITC, was further tested for effects on cell viability, oxidative stress and for ultrastructure. ITCs, especially the aromatic ones, had a significant type-, dose- and isolate-dependent anti-Candida activity. Although BITC and PEITC had similar activity against the yeast cells, BITC had a more pronounced effect on cell size and GTF. Furthermore, BITC appears to induce oxidative stress and promote changes in the cell ultrastructure, interfering with cell wall structure. Our work showed that aromatic ITCs have the potential to effect C. albicans cells in multiple ways, including size, shape and GTF (BITC and PEITC), oxidative stress, and ultrastructure (BITC). Overall, our results suggest that BITC may be effectively used against C. albicans to modulate its growth, and control or suppress its invasive potential.

This work was carried out to investigate the incidence of the invasiveCandida albicansfungus in oral swabs collected from a private primary school and to confirm its invasiveness by RT-PCR. Accordingly, the total of (368) students in the age range of (7-12) years from both sexes was orally examined for the incidence ofCandida albicansfrom October 2019 to March 2020. Wet oral swabs were directly cultured on Sabouraud agar medium, and positive cultures were examined microbiologically. Results of Gram's staining revealed the isolation of the invasiveCandida albicansconfirmed by the germ tube test. This fungus was significantly (P