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Molecules of natural origin often possess useful biological activities. For instance, the natural peptide Tilapia Piscidin 4 (TP4) exhibits potent antimicrobial activity against a broad spectrum of pathogens. In this study, we explored the potential application of TP4 as a food preservative, asking whether it can prevent spoilage due to microbial contamination. A preliminary in silico analysis indicated that TP4 should interact strongly with fungal cell membrane components. Hence, we tested the activity of TP4 toward Candida albicans within fruit juice and found that the addition of TP4 could abolish fungal growth. We further determined that the peptide acts via a membranolytic mechanism and displays concentration-dependent killing efficiency. In addition, we showed that TP4 inhibited growth of Rhizopus oryzae in whole fruit (tomato) samples. Based on these findings, we conclude that TP4 should be further evaluated as a potentially safe and green solution to prevent food spoilage., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Jyh-Yih Chen reports financial support was provided by Higher Education Sprout Project from the Ministry of Education (MOE-110-S-0023-A) Taiwan. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

Background: Plukenetia volubilis Linneo is an oleaginous plant belonging to the family Euphorbiaceae. Due to its seeds containing a high content of edible oil and rich in vitamins, P. volubilis is cultivated as an economical plant worldwide. However, the cultivation and growth of P. volubilis is challenged by phytopathogen invasion leading to production loss., Methods: In the current study, we tested the pathogenicity of fungal pathogens isolated from root rot infected P. volubilis plant tissues by inoculating them into healthy P. volubilis seedlings. Metagenomic sequencing was used to assess the shift in the fungal community of P. volubilis rhizosphere soil after root rot infection., Results: Four Fusarium isolates and two Rhizopus isolates were found to be root rot causative agents of P. volubilis as they induced typical root rot symptoms in healthy seedlings. The metagenomic sequencing data showed that root rot infection altered the rhizosphere fungal community. In root rot infected soil, the richness and diversity indices increased or decreased depending on pathogens. The four most abundant phyla across all samples were Ascomycota, Glomeromycota, Basidiomycota, and Mortierellomycota. In infected soil, the relative abundance of each phylum increased or decreased depending on the pathogen and functional taxonomic classification., Conclusions: Based on our results, we concluded that Fusarium and Rhizopus species cause root rot infection of P. volubilis. In root rot infected P. volubilis, the shift in the rhizosphere fungal community was pathogen-dependent. These findings may serve as a key point for a future study on the biocontrol of root rot of P. volubilis., (© 2023. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

The soft rot disease caused by Rhizopus stolonifer is an important disease in cherry tomato fruit. In this study, the effect of iturin A on soft rot of cherry tomato and its influence on the storage quality of cherry tomato fruit were investigated. The results showed that 512 μg/mL of iturin A could effectively inhibit the incidence of soft rot of cherry tomato fruit. It was found that iturin A could induce the activity of resistance-related enzymes including phenylalanine ammonia lyase (PAL), polyphenol oxidase (PPO), peroxidase (POD), glucanase (GLU), and chitinase (CHI), and active oxygen-related enzymes including ascorbate peroxidases (APX), superoxide dismutases (SOD), catalases (CAT), and glutathione reductase (GR) of cherry tomato fruit. In addition, iturin A treatment could slow down the weight loss of cherry tomato and soften the fruit. These results indicated that iturin A could retard the decay and improve the quality of cherry tomato fruit by both the inhibition growth of R. stolonifera and the inducing the resistance.

Aims: With a goal to identify specific essential oils that can control postharvest Rhizopus rot on strawberry and peach fruits, we performed screening for 26 essential oils based on their antifungal activity against Rhizopus stolonifer in vitro and investigated the underlying mechanism., Methods and Results: Mentha spicata (Ms), Mentha piperita (Mp), Thymus vulgaris CT carvacrol (Tc) and Thymus vulgaris CT thymol (Tt) exhibited strong inhibition on R. stolonifer growth in the screening. These essential oils increased plasma membrane permeability of R. stolonifer and resulted in the outflow of intercellular electrolyte, nucleic acid, protein and soluble sugar. Morphology of R. stolonifer mycelia was greatly altered by these essential oils. Hyphae treated by these essential oils exhibited high accumulation of superoxide anion and malonaldehyde. Combination of these essential oils in commercial package reduced Rhizopus rot on strawberry and peach fruits, with Mp showing the strongest efficiency., Conclusion: Ms, Mp, Tc and Tt essential oils inhibited R. stolonifer growth by targeting its plasma membrane and reduced Rhizopus rot on strawberry and peach fruits., Significance and Impact of the Study: This study provides potential applications of natural plant extract, as alternatives to tradition fungicides, to control postharvest decay on fruits and vegetables., (© 2020 The Society for Applied Microbiology.)

Background: Synthetic fungicides are most commonly used for controlling postharvest disease of fruit, although they can cause the emergence of drug-resistant strains, environmental pollution and fruit safety issues. Bacillomycin D (BD), a novel antifungal lipopeptide, and chitosan (CTS) are applied for the preservation of cherry tomato., Results: The combination of BD and CTS showed an additive inhibition on the growth of Rhizopus stolonifer and Botrytis cinerea compared to that of its individual compound. In addition, BD + CTS reduced the incidence of soft rot and gray mold in cherry tomato caused by R. stolonifer and B. cinerea, respectively. Tomato treated with BD + CTS exhibited a lower weight loss and higher firmness and higher contents of total soluble solids, titratable acidity and ascorbic acid compared to those treated with sterile water (control). The kinetics models demonstrated that the shelf life of cherry tomato treated with BD + CTS could be extended by approximately 15 days longer than the control., Conclusion: The utilization of BD + CTS provided a novel strategy for reducing postharvest fungal rot and maintaining the storage quality of cherry tomato. © 2020 Society of Chemical Industry., (© 2020 Society of Chemical Industry.)

The order Mucorales is an ancient group of fungi classified in the subphylum Mucoromycotina. Mucorales are mainly fast-growing saprotrophs that belong to the first colonizers of diverse organic materials and represent a permanent part of the human environment. Several species are able to cause human infections (mucormycoses) predominantly in patients with impaired immune system, diabetes, or deep trauma. In this review, we compiled 32 reports on community- and hospital-acquired outbreaks caused by Mucorales. The most common source of mucoralean outbreaks was contaminated medical devices that are responsible for 40.7% of the outbreaks followed by contaminated air (31.3%), traumatic inoculation of soil or foreign bodies (9.4%), and the contact (6.2%) or the ingestion (6.2%) of contaminated plant material. The most prevalent species were Rhizopus arrhizus and R. microsporus causing 57% of the outbreaks. The genus Rhizomucor was dominating in outbreaks related to contaminated air while outbreaks of Lichtheimia species and Mucor circinelloides were transmitted by direct contact. Outbreaks with the involvement of several species are reported. Subtyping of strains revealed clonality in two outbreaks and no close relation in two other outbreaks. Based on the existing data, outbreaks of Mucorales can be caused by heterogeneous sources consisting of different strains or different species. Person-to-person transmission cannot be excluded because Mucorales can sporulate on wounds. For a better understanding and prevention of outbreaks, we need to increase our knowledge on the physiology, ecology, and population structure of outbreak causing species and more subtyping data.

This study attempted to characterize the involvement of a change in the redox status and subcellular localization in the BABA-induced priming resistance of peach fruit against Rhizopus rot. Specifically, 50 mM BABA primed the peaches for the enhanced disease resistance against R. stolonifer, as demonstrated by suppression of the disease development upon pathogen challenge accompanied by the clearly elevated level of TGA transcription factor (PpTGA1) and NPR1 gene (PpNPR1). In addition, the BABA elicitation enhanced the activities of a series of critical enzymes in the PPP and AsA-GSH cycle, and eventually promoted the NADPH and GSH pools, which altered the intracellular redox state towards a highly reductive condition. Additionally, PpTGA1-GFP was localized in the cytoplasm in the absence of BABA treatment or R. stolonifer inoculation, while BABA elicitation plus R. stolonifer inoculation caused PpTGA1-GFP to specifically translocate to the nucleus, where it interacted with PpNPR1 and regulated the positive expression of PR genes. Therefore, the observations implied that BABA could promote the reduction of the redox state, resulting in the translocation of PpTGA1 to the nucleus, which was a prerequisite for the induction of a priming defence against Rhizopus rot in peach.

Antibacterial activities against Staphylococcus aureus and Bacillus subtilis were found in an ethanol fraction of tempe, an Indonesian fermented soybean produced using Rhizopus oligosporus . The ethanol fraction contained free fatty acids, monoglycerides, and fatty acid ethyl esters. Among these substances, linoleic acid and α-linolenic acid exhibited antibacterial activities against S. aureus and B. subtilis , whereas 1-monolinolenin and 2-monolinolenin exhibited antibacterial activity against B. subtilis . The other free fatty acids, 1-monoolein, monolinoleins, ethyl linoleate, and ethyl linolenate did not exhibit bactericidal activities. These results revealed that R. oligosporus produced the long-chain polyunsaturated fatty acids and monolinolenins as antibacterial substances against the Gram-positive bacteria during the fungal growth and fermentation of heat-processed soybean.

The microbial interaction between Salmonella enterica and the main postharvest fungal pathogens of strawberries was evaluated. Inoculation of fungal suspension was done 2 (D2) and 1 (D1) day(s) before and at the same time (D0) as S. enterica. Fruits were stored at 20 °C and 4 °C. At both temperatures, Botrytis cinerea and Rhizopus stolonifer caused a decrease in S. enterica population. Treatments where the mould was inoculated (D2, D1 and D0) achieved a significant logarithmic reduction (P < 0.05) of S. enterica populations after 48 h (20 °C) and 14 days (4 °C) compared to fungal-uninoculated fruits (CK). Regarding temperature, average reductions were significantly higher at 4 °C (3.38 log 10  CFU/wound) than at 20 °C (1.16 log 10  CFU/wound) (P < 0.05). Average reductions comprising all treatments were 1.91 and 0.41 log 10  CFU/wound for B. cinerea and R. stolonifer at 20 °C, and 3.39 and 3.37 log 10  CFU/wound for B. cinerea and R. stolonifer at 4 °C. A linear log 10 model was fitted in order to predict the inactivation rate (k max , log 10  CFU/h) of S. enterica. Inactivation rates were higher at 20 °C for D2 treatments than at 4 °C throughout the running time. The main inactivation rate was obtained for B. cinerea at 20 °C (0.160 ± 0.027/h), which was found to have stronger inhibitory activity against S. enterica than R. stolonifer. Univariate analysis ANOVA was carried out to evaluate the effect of different external variables on the inhibition of S. enterica. Results found that single effects were significant (P < 0.05) except for the pH. The inhibitory effect caused by the action of moulds in conjunction with some environmental factors could indicate the potential interactions between strawberry fungal pathogens and S. enterica., Competing Interests: Conflict of interest The authors declare no conflict of interest., (Copyright © 2019. Published by Elsevier B.V.)

This paper addresses the issue of combining the usage of waste frying oil (WFO), as a feedstock, and a lipase produced in solid-state fermentation (SSF), as a biocatalyst, for semi-pilot scale production of biodiesel as fatty acid methyl esters (FAME). Two fungal mutants namely; Rhizopus stolonifer 1aNRC11 mutant F (1F) and Aspergillus tamarii NDA03a mutant G (3G) were used as a cocatalyst. The two mutants were cultivated separately by SSF in a tray bioreactor. The dried fermented solid of 1F and 3G mutants were used in a ratio of 3:1, respectively, for WFO transesterification. Optimization of several semi-pilot process stages including SSF and WFO transesterification reaction conditions resulted in 92.3% conversion of WFO to FAME. This FAME yield was obtained after 48 h using 10% cocatalyst (w/w of WFO), 10% water (w/w of WFO) and 3:1 methanol/ WFO molar ratio at 30 °C and 250 rpm. A preliminary economic evaluation of produced biodiesel price (190 $/Ton) is less than half the price of petroleum diesel in Egypt (401$/Ton) and is about 40.3% the price of biodiesel produced using a pure enzyme, which is a promising result. This strategy makes the biodiesel synthesis process greener, economical and sustainable.

Rhizopus arrhizus ( R. arrhizus ) is a common causative agent of mucormycosis that usually enters the human body through the respiratory tract and skin. Both these sites harbor staphylococci as a part of the normal microflora, indicating the possibility of interspecies interactions. We aimed to elucidate this interaction and identify the molecular mechanisms involved. Both Staphylococcus aureus ( S. aureus ) and Staphylococcus epidermidis ( S. epidermidis ) substantially hindered R. arrhizus radial growth, spore germination, and liquid culture biomass. Secreted components in the stationary-phase supernatant were responsible for this activity. The active components, based on molecular weight-based fractionation, mass spectrometry, and ion exclusion chromatography, were identified as a truncated version of phenol soluble modulin α2 (Δ1Δ2PSMα2) and PSMα3 in S. aureus , PSMδ in S. epidermidis , and organic acids in both the species. Exposure to the phenol soluble modulins (PSMs) extensively damaged the fungal spores and pre-existing hyphae, leading to bleb formation, shriveling, hyphal shrinkage, and cell distortion.

The aim of this work was to investigate the early detection of anthracnose and soft rot diseases in cold stored strawberry fruit by evaluating the CO 2 and volatile organic compounds (VOCs) released by the fungi Colletotrichum fragariae and Rhizopus stolonifer. Strawberries were stored at 5, 10 and 21 °C (control group) and the VOCs and CO 2 production of inoculated and non-inoculated strawberries were followed by gas chromatography. To evaluate and estimate the growth of both fungi, the CO 2 data were fitted to the Gompertz model. Data of the VOCs released at the end of the fungal growth were analyzed using principal components analysis (PCA) to discriminate between infected and non-infected strawberries. The results showed that fungal growth was affected by temperature and C. fragariae had a maximum growth after 14.6 h at 5 °C and R. stolonifer at 21 °C after 45.2 h. On the other hand, through VOCs released by C. fragariae and R. stolonifer and PCA, four groups were obtained: a) strawberry infected with C. fragariae, stored at 10 °C, b) strawberry infected with R. stolonifer, stored at 21 °C, c) control group kept at 10 °C and, d) strawberry infected with C. fragariae and control group (5 and 21 °C), and strawberry infected with R. stolonifer at 5 and 10 °C. In conclusion, CO 2 and VOCs released by C. fragariae and R. stolonifer on strawberries could infer the presence of anthracnose and soft rot during storage of the fruit at low temperature., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

DDT (dichlorodiphenyltrichloroethane) was used worldwide as an organochlorine insecticide to control agricultural pests and vectors of several insect-borne human diseases. It was banned in most industrialized countries; however, due to its persistence in the environment, DDT residues remain in environmental compartments, becoming long-term sources of exposure. To identify and select fungal species suitable for bioremediation of DDT-contaminated sites, soil samples were collected from DDT-contaminated agricultural soils in Poland, and 38 fungal taxa among 18 genera were isolated. Two of them, Trichoderma hamatum FBL 587 and Rhizopus arrhizus FBL 578, were tested for tolerance in the presence of 1-mg liter -1 DDT concentration by using two indices based on fungal growth rate and biomass production (the tolerance indices Rt:Rc and TI), showing a clear tolerance to DDT. The two selected strains were studied to evaluate catabolic versatility on 95 carbon sources with or without DDT by using the Phenotype MicroArray system and to investigate the induced oxidative stress responses. The two strains were able to use most of the substrates provided, resulting in both high metabolic versatility and ecological functionality in the use of carbon sources, despite the presence of DDT. The activation of specific metabolic responses with species-dependent antioxidant enzymes to cope with the induced chemical stress has been hypothesized, since the presence of DDT promoted a higher formation of reactive oxygen species in fungal cells than the controls. The tested fungi represent attractive potential candidates for bioremediation of DDT-contaminated soil and are worthy of further investigations. IMPORTANCE The spread and environmental accumulation of DDT over the years represent not only a threat to human health and ecological security but also a major challenge because of the complex chemical processes and technologies required for remediation. Saprotrophic fungi, isolated from contaminated sites, hold promise for their bioremediation potential toward toxic organic compounds, since they might provide an environment-friendly solution to contamination. Once we verified the high tolerance of autochthonous fungal strains to high concentrations of DDT, we showed how fungi from different phyla demonstrate a high metabolic versatility in the presence of DDT. The isolates showed the singular ability to keep their functionality, despite the DDT-induced production of reactive oxygen species., (Copyright © 2019 American Society for Microbiology.)

Background: Mucormycosis is a rare, worldwide fungal infection with high mortality, which mostly affects immunocompromised patients. Compared to large parts of Asia, Europe, and the USA, information on clinical expression and fungal species distribution in mucormycosis in Turkey is limited. Objectives and methods: The main aim of this study was to evaluate the demographic features of mucormycosis cases, identify the isolates at the species level by using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF), compare culture results with histopathological examination and determine the antifungal susceptibility patterns of the pathogens. Results: Between 2016 and 2018, 10 mucormycosis cases (six female, four male; age range: 35-74 years) were evaluated retrospectively. The predominance of the cases were in late autumn and winter. Diabetes mellitus was the most common underlying condition. Seven patients had rhinoorbitocerebral, two had pulmonary and one had cutaneous mucormycosis. By mycological culture and direct microscopic examination nine strains were identified as Rhizopus spp. and one as Mucor spp. Seven of these strains were identified at the species level by MALDI-TOF. Histopathological examination of eight tissues were reported as compatible with mucormycosis. All isolates were resistant to azoles and echinocandins. Two isolates were resistant to Amphotericin B and one was resistant to posaconazole. Surgical debridement combined with antifungal therapy was the main treatment option. The mortality rate was 40% ( n  = 4) and the mean number of days between the onset of complaints and the initiation of treatment was 9.25. Conclusions: Early, rapid and accurate diagnosis of mucormycosis is of critical importance in the treatment of immunosuppressed patients.

The inhibitory effect of Bacillomycin D, a cyclic lipopeptide, on Rhizopus stolonifer colonization of cherry tomato was studied, and its possible mechanism of action was explored. Bacillomycin D showed a direct inhibitory effect on R. stolonifer spore germination and mycelial growth in vitro. It conferred both a direct inhibitory effect on R. stolonifer growth in cherry tomato in vivo and induced host resistance in cherry tomato. Moreover, Bacillomycin D treatment significantly increased the activities of plant defense-related enzymes, including chitinase (CHI), β-1,3-glucanase (GLU), phenylalanine ammonia-lyase (PAL), and peroxidase (POD). Real-time PCR (RT-PCR) showed that defense-related genes involved in the salicylic acid defense signaling pathway and genes encoding pathogenesis-related proteins were up-regulated in Bacillomycin D treatment. Furthermore, Bacillomycin D-C16 resulted in direct inhibition and a remarkable induced resistance to R. stolonifer which was higher than as induced by Bacillomycin D-C14. Together, the data indicated that Bacillomycin D can control the growth of R. stolonifer through both the direct inhibition of the fungus and the activation of defense-related genes and enzymes in cherry tomato.

Background: One of the most important species used to ferment Chinese turbid rice wine (CTRW) at an industrial-scale level is Rhizopus oryzae, although the flavor of CTRW fermented by pure R. oryzae is inferior to that of traditional CTRW., Results: Mucor indicus was used as a cooperative species to improve the flavor of CTRW presented by R. oryzae. The flavor compounds in different fermentation stages were determined by headspace solid-phase microextraction-gas chromatography-mass spectrometry and high-performance liquid chromatography. It was noted that the M. indicus and R. oryzae co-culture changed the profiles of flavor compounds in CTRW, including esters, higher alcohols, amino acids and organic acids, and also significantly enhanced the concentration of sweet amino acids, fruity and floral esters, and higher alcohols. Sensory evaluation demonstrated that the CTRW fermented by M. indicus and R. oryzae had a more intense aroma, harmonious taste, continuation and full body mouth-feel because of more abundant flavor compounds., Conclusion: Mucor indicus is a promising species for co-culture with R. oryzae to improve the flavor of CTRW. © 2019 Society of Chemical Industry., (© 2019 Society of Chemical Industry.)

Submerged fermentation (SmF) is an attractive biotechnological option for waste treatment, generating fungal bioprotein from food industry by-products. Using different Rhizopus sp. strains as fermentation agents, this paper describes a global strategy to identify interactions between cultivation parameters (pH 4.75-7.75, 7.5-82.5 g/l glucose, 0.75-3.75 g/l nitrogen, incubation time up to 5 days) for valorization of food industry by-products. Selected parameters and/or their interactions are critical for most of the proposed resulting values, giving the opportunity to optimize the process depending on the objective and making an "in silico" pre-evaluation of the process conditions. SmF of orange molasses leads to a high biomass and protein yield (11.9 g biomass/l and 4.0 g protein/l), with 43.1 ± 0.1% of essential amino acids and chemical oxygen demand (COD) reduction of almost 50%. Experiments with cheese whey result in 76.3 ± 1.1% of glucose consumption and a biomass production of 5.6 ± 2.2 g/l with 48.9 ± 5.1% of essential amino acids. SmF of orange molasses and cheese whey could contribute to promoting a sustainable feed industry while reducing by-product treatment.

Evolution of Rhizopus oryzae and Trichoderma reesei biomass in rice bran, their enzyme activity, and the profile of phenolic compounds released from the lignocellulosic matrices were determined and correlated by principal component analysis (PCA). PCA analysis confirms that cultivation of rice bran affected the release of methanol-soluble phenolic compounds (MSPC), ethanol-soluble phenolic compounds (ESPC), and bound phenolic compounds (BPC) positively, due to their enzymatic activity. The release of MSPC was influenced by the activity of cellulase and endoglucanase, which increased 110.6% and 136.3%, respectively, for Rhizopus oryzae and Trichoderma reesei. Gallic acid was the main component in the MSPC and ESPC compound fractions. Ferulic and syringic acids were found in its bound (BPC) form in the biomass. This study showed that bioactive compounds be released from lignocellulosic materials by fungus action and this process can be conducted to obtain specific phenolic compounds. PRACTICAL APPLICATION: Due the demand by natural compounds with biological activity, such as phenolic compounds, it is interesting to purpose alternatives to enhance their yield, like for instance, by fungal fermentation of lignocellulosic material. Therefore, understanding the relations among different phenolic compounds released and the production of fungal hydrolases during growth of Rhizopus oryzae and Trichoderma reesei in solid state cultivation using rice bran as a substrate is fundamental to control the process. This knowledge gets viable scale up to apply the phenolic compounds as preservative in food chain, because this becomes possible directing the process to obtain specific bioactive compounds in less time of cultivation and with low cost., (© 2019 Institute of Food Technologists®.)

Rhizopus spp are the most common etiological agents of mucormycosis, causing over 90% mortality in disseminated infection. Key to pathogenesis is the ability of fungal spores to swell, germinate, and penetrate surrounding tissues. Antibiotic treatment in at-risk patients increases the probability of the patient developing mucormycosis, suggesting that bacteria have the potential to control the growth of the fungus. However, research into polymicrobial relationships involving Rhizopus spp has not been extensively explored. Here we show that co-culturing Rhizopus microsporus and Pseudomonas aeruginosa results in the inhibition of spore germination. This inhibition was mediated via the secretion of bacterial siderophores, which induced iron stress on the fungus. Addition of P. aeruginosa siderophores to R. microsporus spores in the zebrafish larval model of infection resulted in inhibition of fungal germination and reduced host mortality. Therefore, during infection antibacterial treatment may relieve bacterial imposed nutrient restriction resulting in secondary fungal infections.

Mucormycosis is an aggressive and high-mortality opportunistic fungal infection, especially in immunocompromised patients. Conventional antifungals or surgery showed a limited effect on this disease. The antimicrobial photodynamic therapy (aPDT) has been proven to be a promising therapeutic choice against multiple pathogenic fungi. We evaluated the effect of aPDT by using methylene blue (MB) combined with a light emitting diode (LED) on the viability of Rhizopus oryzae, as well as the antifungal susceptibility after aPDT treatment in vitro. A total of six strains were included in this study; MB (8, 16, and 32 μg/ml) was chosen for the photosensitizer, and a light source of LED (635 ± 10 nm, 12 J/cm 2 ) device was used to active it. aPDT with MB (32 μg/ml) and LED was highly effective in cell growth inhibition and exhibited colony-forming unit reductions of up to 4.3log 10 . The minimal inhibitory concentration ranges of itraconazole, posaconazole, and amphotericin B decreased from > 32 μg/ml to 4-8 μg/ml, 8-16 μg/ml to 0.5-2 μg/ml, and 2-4 μg/ml to 0.25-0.5 μg/ml, respectively, after pre-treatment with MB (8 μg/ml) and LED. In conclusion, aPDT with MB and LED was a promising therapeutic option against R. oryzae infections alone or combined with antifungal agents. However, further investigation is needed to determine the potential for clinic therapy and to elucidate the underlying mechanism.

Rhizopus microsporus var. oligosporus is a fungus that belongs to the Mucoraceae family that is used for the preparation of some soy-fermented foods. Microbial biotransformation of progesterone by R. microsporus var. oligosporus afforded some monohydroxylated and dihydroxylated metabolites. The main product was purified using chromatographic methods and identified as 11α-hydroxyprogesterone on the basis of its spectroscopic features. Time course studies by high-performance thin-layer chromatography demonstrated that this fungi efficiently hydroxylated progesterone at the 11α-position for 3 days with a yield of 76.48%, but beyond this time, the microorganism transformed 11α-hydroxyprogesterone into dihydroxylated metabolites. 11α-Hydroxyprogesterone is widely used as a precursor in the synthesis of hydrocortisone and other steroidal anti-inflammatory agents.

The effect of temperature on the mycelium growth kinetics of four postharvest fungal isolates (i.e., Penicillium expansum, Alternaria alternata, Botrytis cinerea and Rhizopus stolonifer) was assessed. A cardinal model with inflection (CMI) was used to describe the effect of the temperature on the growth rate (μ) and the lag time (λ) of each isolate. Cardinal temperature values such as T min , T max and T opt were estimated and isolates were sorted according to their growth rate and lag time duration. Additionally, model validation was performed on a medium prepared from mashed pear pulp and on artificially wound-inoculated pear fruits. P. expansum was shown to be the most psychotrophic fungus with the lowest estimated T min  = -8.78. Model validation on pear pulp agar showed growth rate over-prediction in the case of R. stolonifer and B. cinerea but a good correlation in the case of P. expansum and A. alternata. In vivo experiments on pear fruits showed discrepancies from the synthetic and the simulated counterparts for all the fungi with the only exception of P. expansum., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Mold specific T-cells have been described as a supportive biomarker to monitor invasive mycoses and mold exposure. This study comparatively evaluated frequencies and cytokine profiles of Aspergillus fumigatus and Mucorales reactive T-cells depending on environmental mold exposure. Peripheral blood mononuclear cells (PBMCs) obtained from 35 healthy donors were stimulated with mycelial lysates of A. fumigatus and three human pathogenic Mucorales species. CD154 + specific T-cells were quantified by flow cytometry. In a second cohort of 20 additional donors, flow cytometry was complemented by 13-plex cytokine assays. Mold exposure of the subjects was determined using a previously established questionnaire. Highly exposed subjects exhibited significantly greater CD154 + A. fumigatus and Mucorales specific naïve and memory T-helper cell frequencies. Significant correlation (r = 0.48 - 0.79) was found between A. fumigatus and Mucorales specific T-cell numbers. Logistic regression analyses revealed that combined analysis of mold specific T-cell frequencies and selected cytokine markers (A. fumigatus: IL-5 and TNF-α, R. arrhizus: IL-17A and IL-13) significantly improves classification performance, resulting in 75-90 % predictive power using 10-fold cross-validation. In conclusion, mold specific T-cell frequencies and their cytokine signatures offer promising potential in the assessment of environmental mold exposure. The cytokines identified in this pilot study should be validated in the clinical setting, e. g. in patients with hypersensitivity pneumonitis., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

Enhanced application of solid-state fermentation (SSF) in industrial production and the influence of SSF of Rhizopus K1 on glucoamylase productivity were analyzed using the flat band method. A growth model was implemented through SSF of Rhizopus K1 in this experiment, and spectrophotometric method was used to determine glucoamylase activity. Results showed that in bran and potato culture medium with 70% moisture in a loose state, μ of mycelium reached to 0.15 h -1 after 45 h of culture in a thermostatic water bath incubator at 30°C. Under a low-magnification microscope, mycelial cells appeared uniform, bulky with numerous branches, and were not easily ruptured. The generated glucoamylase activity reached to 55 U/g (dry basis). This study has good utilization value for glucoamylase production by Rhizopus in SSF.

A non-destructive method for detection of fungal contamination in peaches using an electronic nose (E-nose) is presented. Peaches were inoculated with three common spoilage fungi, Botrytis cinerea, Monilinia fructicola and Rhizopus stolonifer and then stored for various periods. E-nose was then used to analyze volatile compounds generated in the fungi-inoculated peaches, which was then compared with the growth data (colony counts) of the fungi. The results showed that changes in volatile compounds in fungi-inoculated peaches were correlated with total amounts and species of fungi. Terpenes and aromatic compounds were the main contributors to E-nose responses. While principle component analysis (PC1) scores were highly correlated with fungal colony counts, Partial Least Squares Regression (PLSR) could effectively be used to predict fungal colony counts in peach samples. The results also showed that the E-nose had high discrimination accuracy, demonstrating the potential use of E-nose to discriminate among fungal contamination in peaches., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Rhizopus delemar is an invasive fungal pathogen responsible for the frequently fatal disease mucormycosis. Germination, a crucial mechanism by which infectious spores of Rhizopus delemar cause disease, is a key developmental process that transforms the dormant spore state into a vegetative one. The molecular mechanisms that underpin this transformation may be key to controlling mucormycosis; however, the regulation of germination remains poorly understood. This study describes the phenotypic and transcriptional changes that take place over the course of germination. This process is characterized by four distinct stages: dormancy, isotropic swelling, germ tube emergence, and hyphal growth. Dormant spores are shown to be transcriptionally unique, expressing a subset of transcripts absent in later developmental stages. A large shift in the expression profile is prompted by the initiation of germination, with genes involved in respiration, chitin, cytoskeleton, and actin regulation appearing to be important for this transition. A period of transcriptional consistency can be seen throughout isotropic swelling, before the transcriptional landscape shifts again at the onset of hyphal growth. This study provides a greater understanding of the regulation of germination and highlights processes involved in transforming Rhizopus delemar from a single-cellular to multicellular organism. IMPORTANCE Germination is key to the growth of many organisms, including fungal spores. Mucormycete spores exist abundantly within the environment and germinate to form hyphae. These spores are capable of infecting immunocompromised individuals, causing the disease mucormycosis. Germination from spore to hyphae within patients leads to angioinvasion, tissue necrosis, and often fatal infections. This study advances our understanding of how spore germination occurs in the mucormycetes, identifying processes we may be able to inhibit to help prevent or treat mucormycosis., (Copyright © 2018 Sephton-Clark et al.)

The Fenton reaction as an oxidative degradation process was used for industrial chemical wastewater (ICW) pretreatment. The biodegradation of pretreated ICW was performed, in aqueous environment under aerobic condition, by a defined fungal consortium. The central composite design (CCD) was used to study the effect of nitrogen and phosphorus addition and the concentration of the pollution on the removal of polyvinyl alcohol (PVA) and organic compounds. The interaction between parameters was modeled using the response surface methodology (RSM). Results of optimization showed COD, PVA and color removal yields of 97.8%, 98.5% and 99.75%, respectively with a supplementof 1.4 gL -1 of (NH 4 ) 2 SO 4 , 1.2 gL -1 of KH 2 PO 4 and 75% of concentrated ICW. Enzymatic analysis proved that laccase and lignin peroxidase were involved in the biodegradation with 45 UIL -1 and 450 UIL -1 , respectively. Furthermore, the analysis of metabolic products using Fourier transforms infrared spectroscopy (FTIR) and nuclear magnetic resonance ( 1 HNMR) showed clearly the mineralization of organic compounds and the formation of formic acid and ethanol. Therefore, the effective treatment of ICW was achieved by developing an integrated chemical and biological process which met the requirement for a safety effluent respectful for environment without risks for public health., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Mucormycosis is a life-threatening respiratory fungal infection predominantly caused by Rhizopus species. Mucormycosis has incompletely understood pathogenesis, particularly how abnormalities in iron metabolism compromise immune responses. Here we show how, as opposed to other filamentous fungi, Rhizopus spp. establish intracellular persistence inside alveolar macrophages (AMs). Mechanistically, lack of intracellular swelling of Rhizopus conidia results in surface retention of melanin, which induces phagosome maturation arrest through inhibition of LC3-associated phagocytosis. Intracellular inhibition of Rhizopus is an important effector mechanism, as infection of immunocompetent mice with swollen conidia, which evade phagocytosis, results in acute lethality. Concordantly, AM depletion markedly increases susceptibility to mucormycosis. Host and pathogen transcriptomics, iron supplementation studies, and genetic manipulation of iron assimilation of fungal pathways demonstrate that iron restriction inside macrophages regulates immunity against Rhizopus. Our findings shed light on the pathogenetic mechanisms of mucormycosis and reveal the role of macrophage-mediated nutritional immunity against filamentous fungi.

DKxanthenes are a class of yellow secondary metabolites produced by myxobacterial genera Myxococcus and Stigmatella . We identified a putative 49.5 kb DKxanthene biosynthetic gene cluster from Myxococcus stipitatus DSM 14675 by genomic sequence and mutational analysis. The cluster was comprisedof 15 genes (MYSTI&#95;06004-MYSTI&#95;06018) encoding polyketide synthases, non-ribosomal peptide synthases, and proteins with unknown functions. Disruption of the genes by plasmid insertion resulted in defects in the production of yellow pigments. High-performance liquid chromatography and liquid chromatography-tandem mass spectrometry analysis indicated that the yellow pigments produced by M. stipitatus DSM 14675 might be noble DKxanthene derivatives. M. stipitatus did not require DKxanthenes for the formation of heat-resistant viable spores, unlike Myxococcus xanthus . Furthermore, DKxanthenes showed growth inhibitory activity against the fungi Aspergillus niger , Candida albicans , and Rhizopus stolonifer .

Direct measurement of cell biomass is difficult in a solid-state fermentation (SSF) process involving filamentous fungi since the mycelium and the solid substrate are often inseparable. However, respiratory data are rich in information for real-time monitoring of microbial biomass production. In this regard, a correlation was obtained between oxygen uptake rate (OUR) and biomass concentration (X) of Rhizopus oryzae MTCC 1987, during phytase production, in an intermittently mixed novel SSF bioreactor. To obtain the correlation, various models describing sigmoidal growth were tested, namely the logistic, Gompertz, Stannard, and Schnute models. Regression analysis of experimental results, at different operating conditions of inlet air flow rate and relative humidity suggested that OUR and X were correlated well by the logistic model (R 2  > 0.90). To corroborate the use of respiratory data for on-line measurement of metabolic activity, OUR was related to metabolic heat generation rate (R q ), and the logistic model was found to satisfactorily correlate R q and X as well. The model parameter, Y Q/X , when substituted into a heat transfer design equation, along with the values of other parameters and operating variables, gave reliable estimates of bed temperature. The correlations developed in the present study, between respiratory activity and biomass concentration may be extended on to other SSF processes for further validation and real-time monitoring of cell biomass and bed temperature.

Rapid assessment of fungal growth and screening antifungal compounds, such as nanoparticles (NPs), for effectiveness is a challenging procedure because no primary standards exist as they do for yeasts and bacteria. Because fungi do not grow as single cells, but as hyphal filaments, they cannot be quantified by the usual enumeration techniques used in bacteriology. The growth of three postharvest fungal isolates ( Alternaria alternata, Rhizopus stolonifer, and Botrytis cinerea) was investigated at different inoculum concentrations and in three nutrient media (Sabouraud dextrose agar, potato dextrose agar, and yeast extract dextrose agar [YED]) with a turbidimetric assay. Sequential measurements were performed to generate optical density versus time plots, whereas the growth responses were expressed quantitatively as the generated trapezoidal area. YED medium showed the lowest variation among replicated experiments; potato dextrose agar showed the next lowest, but there was no significant difference. The inoculum size had a minimal effect on the variation of the fungal dynamics. Microscopic assessment of the fungal growth confirmed that YED medium allowed the most homogeneous development of the studied fungi. Therefore, we developed a rapid and reliable technique to evaluate the efficacy of novel antifungal compounds such as zinc oxide NPs. Turbidimetric assessment showed that these NPs were able to inhibit the growth of all three isolates. A. alternata and B. cinerea did not show a significant difference in the level of inhibition at 15 mM, whereas R. stolonifer showed the highest inhibition at the same concentration.

In previous work, we proposed a novel modified one-step fermentation fed-batch strategy to efficiently generate L-lactic acid (L-LA) using Rhizopus oryzae. In this study, to further enhance efficiency of L-LA production through one-step fermentation in fed-batch cultures, we systematically investigated the initial peptone- and glucose-feeding approaches, including different initial peptone and glucose concentrations and maintained residual glucose levels. Based on the results of this study, culturing R. oryzae with initial peptone and glucose concentrations of 3.0 and 50.0 g/l, respectively, using a fed-batch strategy is an effective approach of producing L-LA through one-step fermentation. Changing the residual glucose had no obvious effect on the generation of L-LA. We determined the maximum LA production and productivity to be 162 g/l and 6.23 g/(l·h), respectively, during the acid production stage. Compared to our previous work, there was almost no change in L-LA production or yield; however, the productivity of L-LA increased by 14.3%.

A fermented solid containing lipases was produced by solid-state fermentation of Rhizopus microsporus on sugarcane bagasse enriched with urea, soybean oil, and a mineral solution. The dry fermented solid produced using R. microsporus (RMFS) was used to catalyze the synthesis of alkyl-esters by esterification in a solvent-free system containing ethanol and oleic acid (as a model system) or a mixture of fatty acids obtained from the physical hydrolysis of soybean soapstock acid oil (FA-SSAO) in subcritical water. The conversions were 93.5 and 84.1%, for oleic acid and FA-SSAO, respectively, at 48 h and 40 °C, at a molar ratio (MR) of ethanol to fatty acid of 5:1. A further increase in the MR to 10:1 improved the production of ethylic-esters, giving conversions at 48 h of 98 and 86% for oleic acid and FA-SSAO, respectively. The results obtained in this work foster further studies on scaling-up of an environmentally friendly process to produce biofuels.

Consolidated bioprocessing (CBP) is a potential breakthrough technology for reducing costs of biochemical production from lignocellulosic biomass. Production of cellulase enzymes, saccharification of lignocellulose, and conversion of the resulting sugars into a chemical of interest occur simultaneously within a single bioreactor. In this study, synthetic fungal consortia composed of the cellulolytic fungus Trichoderma reesei and the production specialist Rhizopus delemar demonstrated conversion of microcrystalline cellulose (MCC) and alkaline pre-treated corn stover (CS) to fumaric acid in a fully consolidated manner without addition of cellulase enzymes or expensive supplements such as yeast extract. A titer of 6.87 g/L of fumaric acid, representing 0.17 w/w yield, were produced from 40 g/L MCC with a productivity of 31.8 mg/L/hr. In addition, lactic acid was produced from MCC using a fungal consortium with Rhizopus oryzae as the production specialist. These results are proof-of-concept demonstration of engineering synthetic microbial consortia for CBP production of naturally occurring biomolecules., (© 2017 Wiley Periodicals, Inc.)

Forty Bacillus isolates obtained from maari (used as condiment in Burkina Faso) including 17 B. subtilis, 4 B. circulans, 7 B. pumilus and 6 B. licheniformis were investigated for use as starter cultures in maari production. The isolates were screened by PCR for the sfp gene responsible for the production of the lipopeptide biosurfactant, surfactin. The sfp gene was detected in all of the seventeen B. subtilis isolates, in 2 out of 7 B. pumilus, in 4 out of 6 B. licheniformis whereas no B. circulans was positive for the sfp gene by PCR screening. Furthermore, all the 40 Bacillus spp. were screened for biosurfactant production and inhibitory activity against Aspergillus flavus, A. niger, A. versicolor and Rhizopus oryzae. Results demonstrated a relationship between the presence of the sfp gene and the antifungal activity and biosurfactant production of Bacillus isolates. In addition, molecular typing of the 17 B. subtilis isolates by Multilocus Sequence Typing (MLST) resulted in 15 Sequence Types, one of them included three strains. Randomly Amplified Polymorphic DNA-PCR (RAPD-PCR), used for B. licheniformis, B. megaterium, B. circulans and B. pumilus revealed that the inhibitory activity and biosurfactant production were strain-dependent. Finally, the detection of chitinase (chi) and β-glucanase (glu) biosynthesis genes was found to be associated with the antifungal activity for 16 B. subtilis isolates. The present work provides a greater understanding of the antifungal activity and biosurfactant production ability within the Bacillus spp. isolated from maari and contributes to the selection of Bacillus isolates to be used as starter cultures for controlled production of maari., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

The use of microorganism fermentation for production of fumaric acid (FA), which is widely used in food, medicine, and other fields, can provide technical support for the FA industry. In this study, we aimed to increase the titer of FA production by using an improved Rhizopus oryzae WHT5, which was domesticated to obtain a furfural-resistant strain in corncob hydrolytes. The metabolic pathways and metabolic network of this strain were investigated, and the related enzymes and metabolic flux were analyzed. Metabolic pathway analysis showed that the R. oryzae WHT5 strain produced FA mainly through two pathways. One occurred in the cytoplasm and the other was a mitochondrial pathway. The key parameters of the fermentation process were analyzed. The FA titer was 49.05 g/L from corncob hydrolytes using R. oryzae WHT5 in a 7-L bioreactor. The use of a furfural-resistant strain developed through domestication effectively increased the titer of FA. This capacity of the microorganisms to produce high amounts of FA by bioconverting corncob hydrolyte can be further applied for industrial production of FA.

Aims: This study aimed to investigate the antifungal mechanism of carvacrol and eugenol to inhibit Rhizopus stolonifer and the control of postharvest soft rot decay in peaches., Methods and Results: To investigate the antifungal mechanism, the effects of carvacrol and eugenol on the mycelium growth, leakages of cytoplasmic contents, mycelium morphology, cell membrane and membrane composition of R. stolonifer were studied. Carvacrol and eugenol both exhibited dose-dependent antifungal activity against R. stolonifer, carvacrol at a concentration of 2 μl per plant and eugenol at a concentration of 4 μl per plant inhibited fungal growth completely. The two essential oils (EOs) increased cell membrane penetrability and caused the leakage of cytoplasm, nucleic acid and protein content. The observation using scanning electron microscopy and fluorescent microscopy showed modification of the hyphal morphology and breakage of the cell plasma membrane. Decreased ergosterol contents confirmed that the two EOs could destroy the membrane of R. stolonifer. For the in vivo test, the inhibition of soft rot disease and the induction of defence-related enzymes were investigated. Carvacrol and eugenol significantly reduced the incidence and severity of soft rot decay in inoculated peaches. The best treatments for controlling soft rot decay were obtained at 0·5 μl l -1 for carvacrol and 1 μl l -1 for eugenol. The activities of defence-related enzymes in peaches were also enhanced by fumigation with two EOs., Conclusion: This study showed that carvacrol and eugenol could effectively inhibit the growth of R. stolonifer in vitro and successfully control the incidence of soft rot decay in honey peaches., Significance and Impact of the Study: The above findings may be the main antifungal mechanism of carvacrol and eugenol on R. stolonifer. Furthermore, carvacrol and eugenol are helpful for their commercial application on the preservation of fresh fruit., (© 2017 The Society for Applied Microbiology.)

The aim of this study was to evaluate the antifungal activity (in vitro) of thymol and carvacrol alone or in mixtures against Fusarium verticillioides and Rhizopus stolonifer, and to obtain primary growth models. Minimal inhibitory concentration (MIC) was evaluated with fungal radial growth with thymol or carvacrol concentrations (0-1600mg/l). Mixtures were evaluated using concentrations below MIC values. Radial growth curves were described by the modified Gompertz equation. MIC values of carvacrol were 200mg/l for both fungi. Meanwhile, MIC values of thymol were between 500 and 400mg/l for F. verticillioides and R. stolonifer, respectively. A synergistic effect below MIC concentrations for carvacrol (100mg/l) and thymol (100-375mg/l) was observed. Significant differences (p<0.05) between the Gompertz parameters for the antimicrobial concentrations and their tested mixtures established an inverse relationship between antimicrobial concentration and mycelial development of both fungi. Modified Gompertz parameters can be useful to determine fungistatic concentrations., (Copyright © 2017 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Soil salinization, as the combination of primary and secondary events, can adversely affect organisms inhabiting this compartment. In the present study, the effects of increased salinity were assessed in four species of terrestrial fungi: Lentinus sajor caju, Phanerochaete chrysosporium, Rhizopus oryzae and Trametes versicolor. The mycelial growth and biochemical composition of the four fungi were determined under three exposure scenarios: 1) exposure to serial dilutions of natural seawater (SW), 2) exposure to serial concentrations of NaCl (potential surrogate of SW); and 3) exposure to serial concentrations of NaCl after a period of pre-exposure to low levels of NaCl. The toxicity of NaCl was slightly higher than that of SW, for all fungi species: the conductivities causing 50% of growth inhibition (EC 50 ) were within 14.9 and 22.0 mScm -1 for NaCl and within 20.2 and 34.1 mScm -1 for SW. Phanerochaete chrysosporium showed to be the less sensitive species, both for NaCl and SW. Exposure to NaCl caused changes in the biochemical composition of fungi, mainly increasing the production of polysaccharides. When fungi were exposed to SW this pattern of biochemical response was not observed. Fungi pre-exposed to low levels of salinity presented higher EC 50 than fungi non-pre-exposed, though 95% confidence limits overlapped, with the exception of P. chrysosporium. Pre-exposure to low levels of NaCl also induced changes in the biochemical composition of the mycelia of L. sajor caju and R. oryzae, relatively to the respective control. These results suggest that some terrestrial fungi may acquire an increased tolerance to NaCl after being pre-exposed to low levels of this salt, thus, suggesting their capacity to persist in environments that will undergo salinization. Furthermore, NaCl could be used as a protective surrogate of SW to derive safe salinity levels for soils, since it induced toxicity similar or higher than that of SW., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Metabolic control improves outcomes associated with mucormycosis. The aim of this study was to compare the in vitro proliferation of Rhizopus oryzae in blood of individuals with and without diabetes at different glycaemic levels. Ninety-five individuals were included. Blood samples from each participant were incubated with sporangiospores of R. oryzae. The germination, filamentation and growth of R. oryzae were compared at different time points. Four groups were defined, one without (group A, n = 30) and three with diabetes: group B (HbA1c ≤7%, N = 24), group C (HbA1c 7.1-9%, N = 20) and group D (HbA1c > 9%, N = 21). The percentage of germinated sporangiospores was higher in the group A after 6 h (group A 56% ± 3, group B 35% ± 4, group C 48% ± 4, group D 46% ± 1.4, p = 0.01), 12 h (group A 54% ± 1.4, group B 19% ± 4, group C 16% ± 1, group D 9.5% ± 5, p < 0.001) and 24 h (group A 29% ± 1, group B 12% ± 4, group C 13.5% ± 3.5, group D 12% ± 1, p < 0.01). The filamentation was higher in groups with diabetes. Group B showed higher filamentation grade than group A at 6 h (0.4 ± 0.04 vs 1 ± 0.09, p < 0.001) and 24 h (1.6 ± 0.05 vs 2.1 ± 0.1, p = 0.05). In conclusion, R. oryzae proliferation was higher among diabetic individuals, including good glycaemic control, than among non-diabetic individuals.

Increasing concerns continue to be expressed about health hazards and environmental pollution resulting from the use of conventional fungicides for postharvest disease control. Nanoparticles represent an alternative solution for postharvest disease management. The objective of this work was to assess the physiological effects and the antifungal efficiency of ZnO nanoparticles (ZnO NPs) against a number of fungal contaminants. The efficacy of ZnO NPs was qualitatively and quantitatively assessed against: Penicillium expansum, Alternaria alternata, Botrytis cinerea and Rhizopus stolonifer. Mycelium growth diameters were measured onto Potato Dextrose Agar (PDA) plates loaded with different ZnO NPs concentrations (from 0mM to 15mM). Hereafter, the rate of the fungal diameter increase was quantified by linear regression modelling. Microscopic analysis was performed by scanning electron microscopy (SEM) images of agar plugs excised from plates with 0mM and 12mM ZnO. All the fungi were inhibited by ZnO NPs at concentrations higher than 6mM. SEM images showed clear morphological aberrations in the fungal structures of all the isolates grown in presence of ZnO. Additionally, knowing that the chelating agent EDTA sequesters metal ions, it was added to fungal inoculated PDA plates with ZnO to study the NPs' mode of action. Cultures where ZnO was mixed with EDTA showed a decrease in the antifungal effect of the nanoparticles. In conclusion, ZnO NPs are therefore a good candidate as an effective postharvest disease control antifungal agent., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Cellulolytic fungi have evolved a sophisticated genetic regulatory network of cellulase synthesis to adapt to the natural environment. Even in the absence of lignocellulose, it still secretes low levels of "constitutive" cellulase for standby application. However, the mechanisms of this constitutive expression remain incompletely understood. Here we identified a cellobiose synthetase (CBS) from Rhizopus stolonifer, which has the capacity to catalyse the synthesis of cellobiose from uridine diphosphate glucose (UDPG). Through the construction of R. stolonifer Δcbs strain, we found that CBS plays a key role in the synthesis of cellulase. Further analysis of cellulase synthesis under glucose culture reveals that the cellobiose-responsive regulator CLR1 was activated by CBS-synthesized cellobiose, thereby promoting the expression of CLR2 and finally opening the transcription of cellulase genes. Our results suggest that R. stolonifer can be induced by self-synthesized cellobiose to produce cellulase, which can be used to reconstruct the expression regulation network to achieve rapid production of cellulase using simple carbon source. Based on our data, the "constitutive expression" of cellulase actually derives from the induction of cellobiose that synthesized by CBS from carbohydrate metabolites, which updates our knowledge of cellulase, and provides a novel insight into the regulation of cellulase synthesis.

Water extracts from solid-state fermentation (SSF) on rapeseed meal using filamentous fungi exhibit interesting immunomodulatory activities in vitro. Immunomodulation was determined by the capacity of the compounds to activate blood neutrophils and to influence cytokine production in human peripheral blood mononuclear cells (PBMC) and mouse bone marrow-derived macrophages (BMDM). Among the strains tested, Aspergillus sojae mycelium and SSF extracts were the most promising in terms of enhancing the immune response. The filamentous fungus was also successfully cultivated in a pre-pilot bioreactor with forced aeration. The results indicated that the extracts not only activated blood neutrophils but also significantly modulated IL-1β cytokine levels with lipopolysaccharide (LPS)-stimulated PBMC and BMDM without any cytotoxicity in immune cells. IL-1β was down-regulated in a dose-dependent manner in the presence of A. sojae crude mycelium and SSF extract with PBMC, which indicated that there was an anti-inflammatory activity, whereas IL-1β secretion was up-regulated in the presence of stimulated BMDM with the highest concentration that was tested (100 μg/mL). The non-fermented rapeseed had no effect at the same concentration. SSF culture, as a natural product, may be a good source for the development of functional feed with an immunostimulating effect or could potentially be used in medicinal applications.

Evolutions of phenolic contents and antioxidant activities during solid-state fermentation (SSF) of plum pomaces (from the juice industry) and brandy distillery wastes with Aspergillus niger and Rhizopus oligosporus were investigated. The effect of fermentation time on the oil content and major lipid classes in the plum kernels was also studied. Results showed that total phenolic (TP) amounts increased by over 30% for SSF with Rhizopus oligosporus and by >21% for SSF with A. niger. The total flavonoid contents presented similar tendencies to those of the TPs. The free radical scavenging activities of methanolic extracts were also significantly enhanced. The HPLC-MS analysis showed that quercetin-3-glucoside was the major phenolic compound in both fermented plum by-products. The results also demonstrated that SSF not only helped to achieve higher lipid recovery from plum kernels, but also resulted in oils with better quality attributes (high sterol ester and n-3 PUFA-rich polar lipid contents)., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Background: Oregano essential oil (EO) was incorporated into film-forming dispersions (FFDs) based on biopolymers (chitosan and/or methylcellulose) at two different concentrations. The effect of the application of the FFDs was evaluated on tomato plants (cultivar Micro-Tom) at three different stages of development, and on pre-harvest and postharvest applications on tomato fruit., Results: The application of the FFDs at '3 Leaves' stage caused phytotoxic problems, which were lethal when the EO was applied without biopolymers. Even though plant growth and development were delayed, the total biomass and the crop yield were not affected by biopolymer-EO treatments. When the FFDs were applied in the 'Fruit' stage the pre-harvest application of FFDs had no negative effects. All FFDs containing EO significantly reduced the respiration rate of tomato fruit and diminished weight loss during storage. Moreover, biopolymer-EO FFDs led to a decrease in the fungal decay of tomato fruit inoculated with Rhizopus stolonifer spores, as compared with non-treated tomato fruit and those coated with FFDs without EO., Conclusion: The application of biopolymer-oregano essential oil coatings has been proven to be an effective treatment to control R. stolonifer in tomato fruit. © 2016 Society of Chemical Industry., (© 2016 Society of Chemical Industry.)

Mucorales are saprobes, ubiquitously distributed and able to infect a heterogeneous population of human hosts. The fungi require robust stress responses to survive in human host. We tested the growth of Mucorales in the presence of different abiotic stress. Eight pathogenic species of Mucorales, including Rhizopus arrhizus, Rhizopus microsporus, Rhizomucor pusillus, Apophysomyces elegans, Licthemia corymbifera, Cunninghamella bertholletiae, Syncephalastrum racemosum and Mucor racemosus, were exposed to different stress inducers: osmotic (sodium chloride and d-sorbitol), oxidative (hydrogen peroxide and menadione), pH, cell wall and metal ions (Cu, Zn, Fe and Mg). Wide variation in stress responses was noted: R. arrhizus showed maximum resistance to both osmotic and oxidative stresses, whereas R. pusillus and M. indicus were relatively sensitive. Rhizopus arrhizus and R. microsporus showed maximum resistance to alkaline pH, whereas C. bertholletiae, L. corymbifera, M. racemosus and A. elegans were resistant to acidic pH. Maximum tolerance was noted in R. microsporus to Cu, R. microsporus and R. arrhizus to Fe and C. bertholletiae to Zn. In contrast, L. corymbifera, A. elegans and M. indicus were sensitive to Cu, Zn and Fe respectively. In conclusion, R. arrhizus showed high stress tolerance in comparison to other species of Mucorales, and this could be the possible reason for high pathogenic potential of this fungi., (© 2016 Blackwell Verlag GmbH.)

A new chitosan producing fungus was locally isolated from soil samples collected around Erzurum, Turkey and identified as Rhizopus oryzae PAS 17 (GenBank accession number KU318422.1). Cultivation in low cost non-sterile conditions was achieved by exploiting its ability to grow at low temperature and pH, thus, undesired microbial contamination could be eliminated when appropriate culture conditions (incubation temperature as 15°C and initial pH of the medium as 4.5) were selected. Medium composition and culture conditions were optimized using Taguchi orthogonal array (OA) design of experiment (DOE). An OA layout of L16 (4(5)) was constructed with five most influensive factors at four levels on chitosan production like, carbon source (molasses), metal ion (Mg(2+)), inoculum amount, agitation speed and incubation time. The optimal combinations of factors (molasses, 70ml/l; MgSO4·7H2O, 0.5g/l; inoculum, 6.7×10(6) spores/disc; agitation speed, 150rpm and incubation time, 8days) obtained from the proposed DOE methodology was further validated by analysis of variance (ANOVA) test and the results revealed the increment of chitosan and biomass yields of 14.45 and 8.58 folds from its unoptimized condition, respectively., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The potential of two zygomycetes fungi, Mucor indicus and Rhizopus oryzae, in assimilating citrus waste free sugars (CWFS) and producing fungal chitosan, oil, and protein as well as ethanol was investigated. Extraction of free sugars from citrus waste can reduce its environmental impact by decreasing the possibility of wild microorganisms growth and formation of bad odors, a typical problem facing the citrus industries. A total sugar concentration of 25.1 g/L was obtained by water extraction of citrus waste at room temperature, used for fungal cultivation in shake flasks and airlift bioreactor with no additional nutrients. In shake flasks cultivations, the fungi were only able to assimilate glucose, while fructose remained almost intact. In contrast, the cultivation of M. indicus and R. oryzae in the four-liter airlift bioreactor resulted in the consumption of almost all sugars and production of 250 and 280 g fungal biomass per kg of consumed sugar, respectively. These biomasses correspondingly contained 40% and 51% protein and 9.8% and 4.4% oil. Furthermore, the fungal cell walls, obtained after removing the alkali soluble fraction of the fungi, contained 0.61 and 0.69 g chitin and chitosan per g of cell wall for M. indicus and R. oryzae, respectively. Moreover, the maximum ethanol yield of 36% and 18% was obtained from M. indicus and R. oryzae, respectively. Furthermore, that M. indicus grew as clump mycelia in the airlift bioreactor, while R. oryzae formed spherical suspended pellets, is a promising feature towards industrialization of the process.

Lactic acid has a wide industrial application area and can be produced by fungal strains. However, excessive bulk growth form of fungi during the fermentations is a major problem, which limits the fermentation performance. Microparticles are excellent tools to prevent bulk fungal growth and provide homogenized fermentation broth to increase uniformity and the prediction performance of the models. Therefore, in this study, addition of aluminum oxide and talcum microparticles into fermentations was evaluated to enhance the production of lactic acid by Rhizopus oryzae. The results showed that the bulk fungal growth was prevented and the lactic acid concentration increased from 6.02 to 13.88 and 24.01 g/L, when 15 g/L of aluminum oxide or 10 g/L of talcum was used, respectively, in the shake-flask fermentations. Additionally, substrate concentration, pH, and agitation were optimized in the bioreactors using response surface methodology, and optimum values were determined as 126 g/L of glucose, 6.22 pH, and 387 rpm, respectively. Under these conditions, lactic acid production further increased to 75.1 ± 1.5 g/L with 10 g/L of talcum addition. Also, lactic acid production and glucose consumption in the batch fermentation were successfully modeled with modified Gompertz model and modified logistic model. RMSE and MAE values for lactic acid production were calculated as 2.279 and 1.498 for the modified Gompertz model; 3.6 and 4.056 for the modified logistic model. Additionally, modified logistic model predicted glucose consumption with -2.088 MAE and 2.868 RMSE, whereas these values were calculated as 2.035 and 3.946 for the modified Gompertz model.

The effects of germination and elicitation on (iso)flavonoid composition of extracts from three edible lupine species (Lupinus luteus, Lupinus albus, Lupinus angustifolius) were determined by RP-UHPLC-MS(n). The total (iso)flavonoid content of lupine increased over 10-fold upon germination, with the total content and composition of isoflavonoids more affected than those of flavonoids. Glycosylated isoflavones were the most predominant compounds found in lupine seedlings. Lesser amounts of isoflavone aglycones, including prenylated ones, were also accumulated. Elicitation with Rhizopus oryzae, in addition to germination, raised the content of isoflavonoids further: the total content of 2'-hydroxygenistein derivatives was increased considerably, without increasing that of genistein derivatives. Elicitation by fungus triggered prenylation of isoflavonoids, especially of the 2'-hydroxygenistein derivatives. The preferred positions of prenylation differed among the three lupine species. The change in isoflavone composition increased the agonistic activity of the extracts towards the human estrogen receptors, whereas no antagonistic activity was observed., (Copyright © 2015 Elsevier Ltd. All rights reserved.)