Search Results (19)

Poly(ε-caprolactone) (PCL) is a synthetic plastic known for its excellent physicochemical properties and a wide range of applications in packaging, coatings, foaming, and agriculture. In medicine, its versatility allows it to function as a scaffold for drug delivery, sutures, implants, tissue engineering, and 3D printing. In addition to its biocompatibility, PCL’s most notable characteristic is its biodegradability. However, this property is affected by temperature, microbial activity, and environmental conditions, which means PCL can sometimes remain in nature for long periods. This review shows that various types of microorganisms can efficiently degrade PCL, including different strains of Pseudomonas spp., Streptomyces spp., Alcaligenes faecalis, and fungi like Aspergillus oryzae, Fusarium spp., Rhizopus delemar, and Thermomyces lanuginosus. These microorganisms produce enzymes such as lipases, esterases, and cutinases that break down PCL into smaller molecules that act as substrates. The review also examines the phylogenetic diversity of organisms capable of biodegrading PCL, the biochemical pathways involved in this process, and specific aspects of the genetic framework responsible for the expression of the enzymes that facilitate degradation. Targeted research on microbial PCL biodegradation and its practical applications could significantly aid in reducing and managing plastic waste on a global ecological scale. Full article

The growing aquaculture industry has an increasing demand for novel, sustainably produced protein sources for aquafeed. This study aimed to determine the apparent digestibility (AD%), pellet quality, and protein score of four novel fungal proteins in rainbow trout (Oncorhynchus mykiss), namely, PEKILO^® (PEK) derived from Paecilomyces variotii, Aspergillus oryzae (AO), Rhizopus oligosporus (RO), and Rhizopus delemar (RD). All fungi were grown on various side-streams, such as beet vinasse, thin stillage, and whole stillage. The diets were produced by extrusion technology and consisted of control and test diets with a 30:70 test ingredient/control ratio. Feeding lasted for 39 days. Each tank had 20 fish, with three replicates per dietary treatment. One-way ANOVA was performed to compare the means of the groups with each other. The dry matter (DM) digestibility of PEK was significantly higher than that of AO, RD, and RO, all with similar digestibility. The crude protein AD% for PEK was 86.5%, which is significantly higher than that of the other fungal sources. AO, PEK, RD, and RO had similar crude fat AD% compared to each other, at 83.8%, 87.4%, 90.5%, and 88.5%, respectively. The pellet quality was found to deteriorate with addition of fungal proteins. PEK had high AD% for most of the macronutrients tested and better pellet quality. Full article

Certain species of Mucorales have been identified as causative agents of mucormycosis, a rare yet often lethal fungal infection. Notably, these fungi exhibit intrinsic resistance to common azole drugs, which target lipids. Given the pivotal role of lipids in drug resistance and their contribution to innate resistance to azoles, this study provides a comprehensive overview of key lipid classes, including sphingolipids (SLs), glycerophospholipids (GPLs), and sterols, in Rhizopus delemar 99-880, a well-characterized reference strain among Mucorales. Using shotgun lipidomics as well as liquid- and gas-chromatography-based mass spectrometric analyses, we identified the lipid intermediates and elucidated the biosynthetic pathways of SLs, PGLs, and sterols. The acidic SLs were not found, probably because the acidic branch of the SL biosynthesis pathway terminates at α-hydroxy phytoceramides, as evident by their high abundance. Intermediates in the neutral SL pathway incorporated higher levels of 16:0 fatty acid compared to other pathogenic fungi. A strikingly high phosphatidylethanolamine (PE)/phosphatdylcholine (PC) ratio was observed among GPLs. Ergosterol remains the major sterol, similar to other fungi, and our analysis confirms the existence of alternate ergosterol biosynthesis pathways. The total lipidomic profile of R. delemar 99-880 offers insights into its lipid metabolism and potential implications for studying pathogenesis and drug resistance mechanisms. Full article

Rice-flavor baijiu is a distilled Chinese spirit prepared from Xiaoqu culture. However, its dull taste may be a market limitation. In order to enhance the flavor profile of rice-flavor baijiu, two ester-producing yeast strains (Saccharomyces cerevisiae and Cyberlindnera fabianii) were inoculated for fermentation. At the end of the fermentation, the total alcohol and ester contents had also increased by 43.3% and 29.8%, respectively, and the number of ester species had increased by eight. Additionally, eleven flavor substances had significant contributions in the inoculated fermentation process, including several different esters and alcohols. A macrogenomic analysis revealed that the majority of the gene abundances associated with the alcohol, acid, and ester pathways were elevated by the third day of inoculated fermentation, and greater abundances of Saccharomyces cerevisiae, Cyberlindnera fabianii, Lichtheimia ramosa, Rhizopus delemar, and Rhizopus oryzaefive, annotated with these genes, were observed from either the pre-fermentation stage or post-fermentation stage. The results demonstrate that two added strains are associated with an increase in the content of the flavor substances. These findings may prove beneficial in enhancing the quality of rice-flavor baijiu through using inoculated fermentation with ester-producing yeast. Full article

Lipases have valuable potential for industrial use, particularly those mostly active against water-insoluble substrates, such as triglycerides composed of long-carbon chain fatty acids. However, in most cases, engineered variants often need to be constructed to achieve optimal performance for such substrates. Protein engineering techniques have been reported as strategies for improving lipase characteristics by introducing specific mutations in the cap domain of esterases or in the lid domain of lipases or through lid domain swapping. Here, we improved the lipase activity of a lipase (WP_075743487.1, or Lip_MRD) retrieved from the Marine Metagenomics MarRef Database and assigned to the Actinoalloteichus genus. The improvement was achieved through site-directed mutagenesis and by substituting its lid domain (FRGTEITQIKDWLTDA) with that of Rhizopus delemar lipase (previously R. oryzae; UniProt accession number, I1BGQ3) (FRGTNSFRSAITDIVF). The results demonstrated that the redesigned mutants gain activity against bulkier triglycerides, such as glyceryl tridecanoate and tridodecanoate, olive oil, coconut oil, and palm oil. Residue W89 (Lip_MRD numbering) appears to be key to the increase in lipase activity, an increase that was also achieved with lid swapping. This study reinforces the importance of the lid domains and their amino acid compositions in determining the substrate specificity of lipases, but the generalization of the lid domain swapping between lipases or the introduction of specific mutations in the lid domain to improve lipase activity may require further investigation. Full article

The first genome sequenced of a eukaryotic organism was for Saccharomyces cerevisiae, as reported in 1996, but it was more than 10 years before any of the zygomycete fungi, which are the early-diverging terrestrial fungi currently placed in the phyla Mucoromycota and Zoopagomycota, were sequenced. The genome for Rhizopus delemar was completed in 2008; currently, more than 1000 zygomycete genomes have been sequenced. Genomic data from these early-diverging terrestrial fungi revealed deep phylogenetic separation of the two major clades—primarily plant—associated saprotrophic and mycorrhizal Mucoromycota versus the primarily mycoparasitic or animal-associated parasites and commensals in the Zoopagomycota. Genomic studies provide many valuable insights into how these fungi evolved in response to the challenges of living on land, including adaptations to sensing light and gravity, development of hyphal growth, and co-existence with the first terrestrial plants. Genome sequence data have facilitated studies of genome architecture, including a history of genome duplications and horizontal gene transfer events, distribution and organization of mating type loci, rDNA genes and transposable elements, methylation processes, and genes useful for various industrial applications. Pathogenicity genes and specialized secondary metabolites have also been detected in soil saprobes and pathogenic fungi. Novel endosymbiotic bacteria and viruses have been discovered during several zygomycete genome projects. Overall, genomic information has helped to resolve a plethora of research questions, from the placement of zygomycetes on the evolutionary tree of life and in natural ecosystems, to the applied biotechnological and medical questions. Full article

The top-gray chalkiness of hybrid rice (Oryza sativa L.) seeds is a typical phenomenon in hybrid rice seeds. The chalky part of the grain is infected and is the inoculum to infect the normal seeds during storage and soaking. These seed-associated microorganisms were cultivated and sequenced using metagenomics shotgun sequencing to obtain more comprehensive information on the seed-associated microorganisms in this experiment. The results showed that fungi could grow well on the rice flour medium, similar to the ingredients of rice seed endosperms. After the assembly of metagenomic data, a gene catalog was established, comprising 250,918 genes. Function analysis showed that glycoside hydrolases were the dominant enzymes, and the genus Rhizopus accounted for the dominant microorganisms. The fungal species R. microspores, R. delemar, and R. oryzae were likely to be the candidate pathogens in the top-gray chalky grains of hybrid rice seeds. These results will provide a reference for improving hybrid rice processing after harvest. Full article

Carrot pomace (CP) which is generated in a large volume in the juice production process, is rich in cellulose, hemicellulose, sugars, pectin, and minerals. However, in many previous investigations, only cellulose was purified and utilized while other components of CP were discarded as waste. Here, CP was valorized into fungal biomass and cellulose with the aim of utilizing all the CP components. Enzymatic pretreatments were applied to solubilize the digestible fraction of CP including hemicellulose, pectin, sucrose, and other sugars for fungal cultivation, while cellulose remained intact in the solid fraction. The dissolved fraction was utilized as a substrate for the cultivation of an edible fungus (Rhizopus delemar). Fungal cultivation was performed in shake flasks and bench-scale bioreactors. The highest fungal biomass concentration was obtained after pretreatment with invertase (5.01 g/L) after 72 h of cultivation (36 and 42% higher than the concentrations obtained after hemicellulase and pectinase treatments, respectively). Invertase pretreatment resulted in the hydrolysis of sucrose, which could then be taken up by the fungus. Carbohydrate analysis showed 28–33% glucan, 4.1–4.9% other polysaccharides, 0.01% lignin, and 2.7–7% ash in the CP residues after enzymatic pretreatment. Fourier transform infrared spectroscopy and thermogravimetric analysis also confirmed the presence of cellulose in this fraction. The obtained fungal biomass has a high potential for food or feed applications, or as a raw material for the development of biomaterials. Cellulose could be purified from the solid fraction and used for applications such as biobased-textiles or membranes for wastewater treatment, where pure cellulose is needed. Full article

The antifungal effect of metabolites produced by a new strain of Lactiplantibacillus (Lpb.) plantarum LPP703, isolated from naturally fermented yak yogurt, was investigated. The results showed that Lpb. plantarum LPP703 significantly inhibited four fungal species, including Penicillium sp., Rhizopus delemar, Aspergillus flavus, and Aspergillus niger. The metabolites produced after 20 h of Lpb. plantarum LPP703 fermentation showed the highest antifungal activity against Penicillium sp. Compared with the control group, the Lpb. plantarum LPP703 metabolites-treated Penicillium sp. spores were stained red by propidium iodide, indicating that the cell membrane of the fungal spores was damaged. Moreover, the antifungal effect of the Lpb. plantarum LPP703 metabolites on Penicillium sp. was not changed after heating or treatment with various proteases, but showed a sharp decrease when the pH value was regulated to 5.0 or above. The oleamide, trans-cinnamic acid, and citric acid were the three most abundant in the Lpb. plantarum LPP703 metabolites. Molecular docking predicated that the oleamide interacted with the active site of lanosterol 14-alpha-demethylase (CYP51, a crucial enzyme for fungal membrane integrity) through hydrogen bonds and had the lowest docking score, representing the strongest binding affinity to CYP51. Taken together, the metabolites from a new strain of Lpb. plantarum, LPP703, had potent antifungal activity against Penicillium sp., which might be associated with the damage of the active ingredient to fungal membrane integrity. This study indicated that Lpb. plantarum LPP703 and its metabolites might act as biological control agents to prevent fungal growth in the food industry. Full article

Mucormycosis is a highly aggressive angio-invasive disease of humans caused by fungi in the zygomycete order, Mucorales. Though a number of different species can cause mucormycosis, the principal agent of the disease worldwide is Rhizopus arrhizus, which accounts for the majority of rhino-orbital-cerebral, pulmonary, and disseminated infections in immunocompromised individuals. It is also the main cause of life-threatening infections in patients with poorly controlled diabetes mellitus, and in corticosteroid-treated patients with SARS-CoV-2 infection, where it causes the newly described disease, COVID-19-associated mucormycosis (CAM). Diagnosis currently relies on non-specific CT, a lengthy and insensitive culture from invasive biopsy, and a time-consuming histopathology of tissue samples. At present, there are no rapid antigen tests for the disease that detect biomarkers of infection, and which allow point-of-care diagnosis. Here, we report the development of an IgG1 monoclonal antibody (mAb), KC9, which is specific to Rhizopus arrhizus var. arrhizus (syn. Rhizopus oryzae) and Rhizopus arrhizus var. delemar (Rhizopus delemar), and which binds to a 15 kDa extracellular polysaccharide (EPS) antigen secreted during hyphal growth of the pathogen. Using the mAb, we have developed a competitive lateral-flow device (LFD) that allows rapid (30 min) and sensitive (~50 ng/mL running buffer) detection of the EPS biomarker, and which is compatible with human serum (limit of detection of ~500 ng/mL) and bronchoalveolar lavage fluid (limit of detection of ~100 ng/mL). The LFD, therefore, provides a potential novel opportunity for the non-invasive detection of mucormycosis caused by Rhizopus arrhizus. Full article

Mucormycoses are rare but serious opportunistic fungal infections caused by filamentous organisms of the order Mucorales. Here we report the first molecular identification of Rhizopus oryzae (heterotypic synonym Rhizopus arrhizus), R. delemar, and Apophysomyces ossiformis as the etiological agents of three cases of severe mucormycosis in Honduras. Conventional microbiological cultures were carried out, and DNA was extracted from both clinical samples and axenic cultures. The ITS ribosomal region was amplified and sequenced. Molecular tools are suitable strategies for diagnosing and identifying Mucorales in tissues and cultures, especially in middle-income countries lacking routine diagnostic strategies. Full article

Mucormycosis (a.k.a. zygomycosis) is an often-life-threatening disease caused by fungi from the ancient fungal division Mucoromycota. Globally, there are nearly a million people with the disease. Rhizopus spp., and R. delemar (R. oryzae, R. arrhizus) in particular, are responsible for most of the diagnosed cases. Pulmonary, rhino-orbito-cerebral, and invasive mucormycosis are most effectively treated with amphotericin B (AmB) and particularly with liposomal formulations (e.g., AmBisome^®). However, even after antifungal therapy, there is still a 50% mortality rate. Hence, there is a critical need to improve therapeutics for mucormycosis. Targeting AmB-loaded liposomes (AmB-LLs) with the pathogen receptor Dectin-1 (DEC1-AmB-LLs) to the beta-glucans expressed on the surface of Aspergillus fumigatus and Candida albicans lowers the effective dose required to kill cells relative to untargeted AmB-LLs. Because Dectin-1 is an immune receptor for R. delemar infections and may bind it directly, we explored the Dectin-1-mediated delivery of liposomal AmB to R. delemar. DEC1-AmB-LLs bound 100- to 1000-fold more efficiently to the exopolysaccharide matrix of R. delemar germlings and mature hyphae relative to AmB-LLs. DEC1-AmB-LLs delivering sub-micromolar concentrations of AmB were an order of magnitude more efficient at inhibiting and/or killing R. delemar than AmB-LLs. Targeted antifungal drug-loaded liposomes have the potential to improve the treatment of mucormycosis. Full article

Calcium carbonate has been extensively used as a neutralising agent in acid-forming microbial processes. The effect of increasing calcium carbonate concentrations on Rhizopus delemar has not been previously investigated. In this study, an evaluation of fumaric acid (FA) and malic acid (MA) production was conducted at three CaCO₃ concentrations in shake flask cultivations. Increased CaCO₃ concentrations resulted in the co-production of FA and MA in the first 55 h of the fermentation (regime 1), and the subsequent depletion of FA thereafter (regime 2). Three factors were highlighted as likely causes of this response: insoluble solids, metal ion concentrations, and pH. Further shake flask cultivations as well as a continuous fermentation with immobilised R. delemar were used to explore the effect of the three factors on regime 1 and 2. Insoluble solids were found to have no effect on the response in either regime 1 or 2. Increasing the aqueous calcium ion concentrations to 10 g L⁻¹ resulted in a three-fold increase in MA titres (regime 1). Moreover, an increase in pH above 7 was associated with a drop in FA concentrations in regime 2. Further tests established that this was due to the hydration of FA to MA, influenced by high pH conditions (7 or higher), nitrogen starvation, and glucose depletion. Anaerobic conditions were also found to significantly improve the hydration process. This study presents the first investigation in which the production of FA followed by in situ hydration of FA to MA with R. delemar has been achieved. Full article

The in vitro interaction of amphotericin B in combination with colistin was evaluated against a total of 86 strains comprising of 47 Candida species (10 Candida albicans, 15 Candida auris, five Candida glabrata, three Candida kefyr, five Candida krusei, four Candida parapsilosis and five Candida tropicalis), 29 Aspergillus species (five Aspergillus flavus, 10 Aspergillus fumigatus, four Aspergillus nidulans, five Aspergillus niger, and five Aspergillus terreus), and 10 Rhizopus species (seven Rhizopus arrhizus, one Rhizopus delemar and two Rhizopus microsporus) strains. For the determination of the interaction, a microdilution checkerboard technique based on the European Committee on Antimicrobial Susceptibility Testing (EUCAST) reference method for antifungal susceptibility testing was used. Results of the checkerboard technique were evaluated by the fractional inhibitory concentration index (FICI) based on the Loewe additivity model for all isolates. Different inhibition endpoints were used to capture both the interaction at MIC and sub-MIC levels. Additionally, checkerboard technique results for Candida species were evaluated by response surface analysis based on the Bliss independence model. Against common Candida species, the combination was synergistic for 75% of the strains by FICI and for 66% of the strains by response surface analysis. For C. tropicalis, the interaction was antagonistic for three isolates by FICI, but antagonism was not confirmed by response surface analysis. Interestingly, synergistic and antagonistic FICIs were simultaneously present on checkboard microplates of all three strains. Against C. auris the combination was synergistic for 73% of the strains by response surface analysis and for 33% of the strains by FICI. This discrepancy could be related to the insensitivity of the FICI to detect weak interactions. Interaction for all other strains was indifferent. For Aspergillus and Rhizopus species combination exhibited only indifferent interactions against all tested strains. Full article

Fungal fermented foods are nutritious, environmentally friendly and sustainable protein sources. To develop fungal fermented food with acceptable sensory characteristics, it is important to assess factors that can affect the sensory characteristics of the product. The current study aimed to investigate the correlations between the chemical and microbiological characteristics and sensory characteristics of fungal fermented food. Soybeans were fermented using five local Indonesian strains of the genus Rhizopus sp. and one strain of industrial starter to mimic traditional Indonesian tempe. The chemical (amino acid and ammonia content), microbiological (lactic acid bacteria, proteolytic bacteria and yeast) and sensory characteristics of the fermented products were examined. The results showed that there is a correlation between the chemical properties, particularly glutamic acid and aspartic acid, and the overall liking of different types of tempe. In general, Rhizopus oligosporus-fermented products had better sensory characteristics than those fermented with Rhizopus oryzae and Rhizopus delemar. The sensory characteristics of the fermented products in this work made from isolates are comparable to those made with an industrial starter culture. In addition, taste and texture affect the overall liking of the product. The results of this study contribute to the development of acceptable sensory fungal fermented food and, in particular, the screening of potential starters. Full article