Fungi of the order Mucorales express a "sealing-only" tRNA ligase.

Some eukaryotic pre-tRNAs contain an intron that is removed by a dedicated set of enzymes. Intron-containing pre-tRNAs are cleaved by tRNA splicing endonuclease, followed by ligation of the two exons and release of the intron. Fungi use a "heal and seal" pathway that requires three distinct catalytic domains of the tRNA ligase enzyme, Trl1. In contrast, humans use a "direct ligation" pathway carried out by RTCB, an enzyme completely unrelated to Trl1. Because of these mechanistic differences, Trl1 has been proposed as a promising drug target for fungal infections. To validate Trl1 as a broad-spectrum drug target, we show that fungi from three different phyla contain Trl1 orthologs with all three domains. This includes the major invasive human fungal pathogens, and these proteins can each functionally replace yeast Trl1. In contrast, species from the order Mucorales, including the pathogens Rhizopus arrhizus and Mucor circinelloides, have an atypical Trl1 that contains the sealing domain but lacks both healing domains. Although these species contain fewer tRNA introns than other pathogenic fungi, they still require splicing to decode three of the 61 sense codons. These sealing-only Trl1 orthologs can functionally complement defects in the corresponding domain of yeast Trl1 and use a conserved catalytic lysine residue. We conclude that Mucorales use a sealing-only enzyme together with unidentified nonorthologous healing enzymes for their heal and seal pathway. This implies that drugs that target the sealing activity are more likely to be broader-spectrum antifungals than drugs that target the healing domains.

Characterization of tRNA splicing enzymes RNA ligase and tRNA 2'-phosphotransferase from the pathogenic fungi Mucorales.

Fungal Trl1 is an essential trifunctional tRNA splicing enzyme that heals and seals tRNA exons with 2',3'-cyclic-PO4 and 5'-OH ends. Trl1 is composed of C-terminal cyclic phosphodiesterase and central polynucleotide kinase end-healing domains that generate the 3'-OH,2'-PO4 and 5'-PO4 termini required for sealing by an N-terminal ATP-dependent ligase domain. Trl1 enzymes are present in many human fungal pathogens and are promising targets for antifungal drug discovery because their domain structures and biochemical mechanisms are unique compared to the mammalian RtcB-type tRNA splicing enzyme. Here we report that Mucorales species (deemed high-priority human pathogens by WHO) elaborate a noncanonical tRNA splicing apparatus in which a monofunctional RNA ligase enzyme is encoded separately from any end-healing enzymes. We show that Mucor circinelloides RNA ligase (MciRNL) is active in tRNA splicing in vivo in budding yeast in lieu of the Trl1 ligase domain. Biochemical and kinetic characterization of recombinant MciRNL underscores its requirement for a 2'-PO4 terminus in the end-joining reaction, whereby the 2'-PO4 enhances the rates of RNA 5'-adenylylation (step 2) and phosphodiester synthesis (step 3) by ∼125-fold and ∼6200-fold, respectively. In the canonical fungal tRNA splicing pathway, the splice junction 2'-PO4 installed by RNA ligase is removed by a dedicated NAD+-dependent RNA 2'-phosphotransferase Tpt1. Here we identify and affirm by genetic complementation in yeast the biological activity of Tpt1 orthologs from three Mucorales species. Recombinant M. circinelloides Tpt1 has vigorous NAD+-dependent RNA 2'-phosphotransferase activity in vitro.

Light irradiation changes the regulation pattern of BtCrgA on carotenogenesis in Blakeslea trispora.

CrgA has been shown to be a negative regulator of carotenogenesis in some filamentous fungi, while light irradiation is an inducible environmental factor for carotenoid biosynthesis. To clarify the relationship between CrgA and light-inducible carotenogenesis in Blakeslea trispora, the cis-acting elements of the btcrgA promoter region were investigated, followed by the analyses of correlation between the expression of btcrgA and carotenoid structural genes under different irradiation conditions. A variety of cis-acting elements associated with light response was observed in the promoter region of btcrgA, and transcription of btcrgA and carotenoid structural genes under different irradiation conditions was induced by white light with a clear correlation. Then, RNA interference and overexpression of btcrgA were performed to investigate their effects on carotenogenesis at different levels under irradiation and darkness. The analyses of transcription and enzyme activities of carotenoid structural gene, and accumulation of carotenoids among btcrgA-interfered, btcrgA-overexpressed, and wild-type strains under irradiation and darkness indicate that btcrgA negatively regulates the synthesis of carotenoid in darkness, while promotes the carotenogenesis under irradiation regardless of reduced or overexpression of btcrgA .

Combating Black Fungus: Using Allicin as a Potent Antifungal Agent against Mucorales.

Invasive fungal (IF) diseases are a leading global cause of mortality, particularly among immunocompromised individuals. The SARS-CoV-2 pandemic further exacerbated this scenario, intensifying comorbid IF infections such as mucormycoses of the nasopharynx. In the work reported here, it is shown that zygomycetes, significant contributors to mycoses, are sensitive to the natural product allicin. Inhibition of Mucorales fungi by allicin in solution and by allicin vapor was demonstrated. Mathematical modeling showed that the efficacy of allicin vapor is comparable to direct contact with the commercially available antifungal agent amphotericin B (ampB). Furthermore, the study revealed a synergistic interaction between allicin and the non-volatile ampB. The toxicity of allicin solution to human cell lines was evaluated and it was found that the half maximal effective concentration (EC50) of allicin was 25-72 times higher in the cell lines as compared to the fungal spores. Fungal allicin sensitivity depends on the spore concentration, as demonstrated in a drop test. This study shows the potential of allicin, a sulfur-containing defense compound from garlic, to combat zygomycete fungi. The findings underscore allicin's promise for applications in infections of the nasopharynx via inhalation, suggesting a novel therapeutic avenue against challenging fungal infections.

Heterochromatin and RNAi act independently to ensure genome stability in Mucorales human fungal pathogens.

Chromatin modifications play a fundamental role in controlling transcription and genome stability and yet despite their importance, are poorly understood in early-diverging fungi. We present a comprehensive study of histone lysine and DNA methyltransferases across the Mucoromycota, emphasizing heterochromatin formation pathways that rely on the Clr4 complex involved in H3K9-methylation, the Polycomb-repressive complex 2 driving H3K27-methylation, or DNMT1-like methyltransferases that catalyze 5mC DNA methylation. Our analysis uncovered H3K9-methylated heterochromatin as the major chromatin modification repressing transcription in these fungi, which lack both Polycomb silencing and cytosine methylation. Although small RNAs generated by RNA interference (RNAi) pathways facilitate the formation of heterochromatin in many eukaryotic organisms, we show that RNAi is not required to maintain either genomic or centromeric heterochromatin in Mucor. H3K9-methylation and RNAi act independently to control centromeric regions, suggesting a functional subspecialization. Whereas the H3K9 methyltransferase Clr4 and heterochromatin formation are essential for cell viability, RNAi is dispensable for viability yet acts as the main epigenetic, regulatory force repressing transposition of centromeric GremLINE1 elements. Mutations inactivating canonical RNAi lead to rampant transposition and insertional inactivation of targets resulting in antimicrobial drug resistance. This fine-tuned, Rdrp2-dependent RNAi activity is critical for genome stability, restricting GremLINE1 retroelements to the centromeres where they occupy long heterochromatic islands. Taken together, our results suggest that RNAi and heterochromatin formation are independent genome defense and regulatory mechanisms in the Mucorales, contributing to a paradigm shift from the cotranscriptional gene silencing observed in fission yeasts to models in which heterochromatin and RNAi operate independently in early-diverging fungi.

SR5AL serves as a key regulatory gene in lycopene biosynthesis by Blakeslea trispora.

BACKGROUND: Trisporic acids are considered to be key regulators of carotenoid biosynthesis and sexual reproduction in zygomycetes, but the mechanisms underlying this regulation have not been fully elucidated. RESULTS: In this study, the relationships between trisporic acids and lycopene synthesis were investigated in Blakeslea trispora. The lycopene concentration in single fermentation by the (-) strain with the addition of 24 µg/L trisporic acids was slightly higher than that observed in mated fermentation. After transcriptomic analysis, a steroid 5α-reductase-like gene, known as SR5AL in B. trispora, was first reported. 5α-Reductase inhibitors reduced lycopene biosynthesis and downregulated the expression of sex determination and carotenoid biosynthesis genes. Overexpression of the SR5AL gene upregulated these genes, regardless of whether trisporic acids were added. CONCLUSION: These findings indicated that the SR5AL gene is a key gene associated with the response to trisporic acids.

Gene manipulation in the Mucorales fungus Rhizopus oryzae using TALENs with exonuclease overexpression.

The Mucorales fungal genus Rhizopus is used for the industrial production of organic acids, enzymes and fermented foods. The metabolic engineering efficiency of Rhizopus could be improved using gene manipulation; however, exogenous DNA rarely integrates into the host genome. Consequently, a genetic tool for Mucorales fungi needs to be developed. Recently, programmable nucleases that generate DNA double-strand breaks (DSBs) at specific genomic loci have been used for genome editing in various organisms. In this study, we examined gene disruption in Rhizopus oryzae using transcription activator-like effector nucleases (TALENs), with and without exonuclease overexpression. TALENs with an overexpressing exonuclease induced DSBs, followed by target site deletions. Although DSBs are repaired mainly by nonhomologous end joining in most organisms, our results suggested that in R. oryzae microhomology-mediated end joining was the major DSB repair system. Our gene manipulation method using TALENs coupled with exonuclease overexpression contributes to basic scientific knowledge and the metabolic engineering of Rhizopus.

Biotransformation of ursolic acid by Circinella muscae and their anti-neuroinflammatory activities of metabolites.

In this study, the biotransformation of ursolic acid by Circinella muscae CGMCC 3.2695 was investigated. Scaled-up biotransformation reactions yielded ten metabolites. Their structures were established based on extensive NMR and HR-ESI-MS data analyses, and four of them are new compounds. C. muscae could selectively catalyze hydroxylation, lactonisation, carbonylation and carboxyl reduction reactions. Furthermore, all the identified metabolites were evaluated for their anti-neuroinflammatory activities in LPS-induced BV-2 cells. Most metabolites displayed pronounced inhibitory effect on nitric oxide (NO) production. The results suggested that biotransformed derivatives of ursolic acid might be served as potential neuroinflammatory inhibitors.

Synthesis, Biological Evaluation, and Structure-Activity Relationships of 4-Aminopiperidines as Novel Antifungal Agents Targeting Ergosterol Biosynthesis.

The aliphatic heterocycles piperidine and morpholine are core structures of well-known antifungals such as fenpropidin and fenpropimorph, commonly used as agrofungicides, and the related morpholine amorolfine is approved for the treatment of dermal mycoses in humans. Inspired by these lead structures, we describe here the synthesis and biological evaluation of 4-aminopiperidines as a novel chemotype of antifungals with remarkable antifungal activity. A library of more than 30 4-aminopiperidines was synthesized, starting from N-substituted 4-piperidone derivatives by reductive amination with appropriate amines using sodium triacetoxyborohydride. Antifungal activity was determined on the model strain Yarrowia lipolytica, and some compounds showed interesting growth-inhibiting activity. These compounds were tested on 20 clinically relevant fungal isolates (Aspergillus spp., Candida spp., Mucormycetes) by standardized microbroth dilution assays. Two of the six compounds, 1-benzyl-N-dodecylpiperidin-4-amine and N-dodecyl-1-phenethylpiperidin-4-amine, were identified as promising candidates for further development based on their in vitro antifungal activity against Candida spp. and Aspergillus spp. Antifungal activity was determined for 18 Aspergillus spp. and 19 Candida spp., and their impact on ergosterol and cholesterol biosynthesis was determined. Toxicity was determined on HL-60, HUVEC, and MCF10A cells, and in the alternative in vivo model Galleria mellonella. Analysis of sterol patterns after incubation gave valuable insights into the putative molecular mechanism of action, indicating inhibition of the enzymes sterol C14-reductase and sterol C8-isomerase in fungal ergosterol biosynthesis.

Lactoferrin, a potential iron-chelator as an adjunct treatment for mucormycosis - A comprehensive review.

Mucormycosis is a deadly infection which is caused by fungi of the order Mucorales including species belonging to the genus Rhizopus, Mucor, Mycocladus, Rhizomucor, Cunninghamella, and Apophysomyces. Despite antifungal therapy and surgical procedures, the mortality rate of this disease is about 90-100% which is exceptionally high. The hypersensitivity of patients with raised available serum iron indicates that the Mucorales are able to use host iron as a critical factor of virulence. This is because iron happens to be a crucial element playing its role in the growth of cells and development. In this review, we have described Lactoferrin (Lf) as a potential iron-chelator. Lf is a naturally occurring glycoprotein which is expressed in most of the biological fluids. Moreover, Lf possesses exclusive anti-inflammatory effects along with several anti-fungal effects that could prove to be helpful to the pathological physiology of inexorable mucormycosis cases. This literature summarises the biological insights into the Lf being considered as a potential fungistatic agent and an immune regulator. The review also proposes that unique potential of Lf as an iron-chelator can be exploited as the adjunct treatment for mucormycosis infection.

Neuroprotective Potency of Tofu Bio-Processed Using Actinomucor elegans against Hypoxic Injury Induced by Cobalt Chloride in PC12 Cells.

Fermented soybean products have attracted great attention due to their health benefits. In the present study, the hypoxia-injured PC12 cells induced by cobalt chloride (CoCl2) were used to evaluate the neuroprotective potency of tofu fermented by Actinomucor elegans (FT). Results indicated that FT exhibited higher phenolic content and antioxidant activity than tofu. Moreover, most soybean isoflavone glycosides were hydrolyzed into their corresponding aglycones during fermentation. FT demonstrated a significant protective effect on PC12 cells against hypoxic injury by maintaining cell viability, reducing lactic dehydrogenase leakage, and inhibiting oxidative stress. The cell apoptosis was significantly attenuated by the FT through down-regulation of caspase-3, caspases-8, caspase-9, and Bax, and up-regulation of Bcl-2 and Bcl-xL. S-phase cell arrest was significantly inhibited by the FT through increasing cyclin A and decreasing the p21 protein level. Furthermore, treatment with the FT activated autophagy, indicating that autophagy possibly acted as a survival mechanism against CoCl2-induced injury. Overall, FT offered a potential protective effect on nerve cells in vitro against hypoxic damage.

Protoplast fusion between Blakeslea trispora 14,271 (+) and 14,272 (-) enhanced the yield of lycopene and ß-carotene.

Blakeslea trispora, a heterothallic Zygomycota with two mating types (termed "plus" and "minus"), is an ideal source of lycopene and ß-carotene. The lycopene and ß-carotene yields when the two type strains are used for fermentation separately are lower than those when they are joint together. To enhance the yield of lycopene and ß-carotene in B. trispora, protoplast fusion technology was carried out between ATCC 14,271 (+) and ATCC 14,272 (-). After protoplast preparation, protoplast fusion, fusion sorting, fusion regeneration, and high-throughput screening, two fusions (Fu-1and Fu-2) with high lycopene and ß-carotene yields were obtained. The lycopene yields of Fu-1 and Fu-2 were increased to 0.60 mg/gDW and 0.90 mg/gDW, which were respectively 3.62- and 5.44-fold those of 14,271 and 1.76- and 2.64-fold those of 14,272. The ß-carotene yields of Fu-1 and Fu-2 were increased to 22.07 mg/gDW and 36.93 mg/gDW, which were respectively 1.72- and 2.89-fold those of 14,271 and 1.23- and 2.06-fold those of 14,272. In this study, the protoplast fusion technique was successfully used in Blakeslea trispora, providing new ideas for improving lycopene and ß-carotene production.

Iron Assimilation during Emerging Infections Caused by Opportunistic Fungi with emphasis on Mucorales and the Development of Antifungal Resistance.

Iron is a key transition metal required by most microorganisms and is prominently utilised in the transfer of electrons during metabolic reactions. The acquisition of iron is essential and becomes a crucial pathogenic event for opportunistic fungi. Iron is not readily available in the natural environment as it exists in its insoluble ferric form, i.e., in oxides and hydroxides. During infection, the host iron is bound to proteins such as transferrin, ferritin, and haemoglobin. As such, access to iron is one of the major hurdles that fungal pathogens must overcome in an immunocompromised host. Thus, these opportunistic fungi utilise three major iron acquisition systems to overcome this limiting factor for growth and proliferation. To date, numerous iron acquisition pathways have been fully characterised, with key components of these systems having major roles in virulence. Most recently, proteins involved in these pathways have been linked to the development of antifungal resistance. Here, we provide a detailed review of our current knowledge of iron acquisition in opportunistic fungi, and the role iron may have on the development of resistance to antifungals with emphasis on species of the fungal basal lineage order Mucorales, the causative agents of mucormycosis.

Novel Monomeric Fungal Subtilisin Inhibitor from a Plant-Pathogenic Fungus, Choanephora cucurbitarum: Isolation and Molecular Characterization.

The bacterial protease inhibitor domains known as Streptomyces subtilisin inhibitors (SSI) are rarely found in fungi. Genome analysis of a fungal pathogen, Choanephora cucurbitarum KUS-F28377, revealed 11 SSI-like domains that are horizontally transferred and sequentially diverged during evolution. We investigated the molecular function of fungal SSI-like domains of C. cucurbitarum, designated "choanepins." Among the proteins tested, only choanepin9 showed inhibitory activity against subtilisin as the target protease, accounting for 47% of the inhibitory activity of bacterial SSI. However, the binding affinity (expressed as the dissociation constant [Kd ]) of choanepin9 measured via microscale thermophoresis was 21 nM, whereas that for bacterial SSI is 34 nM. The trend of binding and inhibitory activity suggests that the two inhibitors exhibit different inhibitory mechanisms for subtilisin protease. Interestingly, choanepin9 was identified as a monomer in studies in vitro, whereas bacterial SSI is a homodimer. Based on these observations, we constructed a monomeric bacterial SSI protein with decreased binding affinity to abrogate its inhibitory activity. By altering the reactive sites of choanepin9 deduced from the P1 and P4 sites of bacterial SSI, we reestablished that these residues in choanepins are also crucial for modulating inhibitory activity. These findings suggest that the fungal SSI evolved to target specific cognate proteases by altering the residues involved in inhibitory reactivity (reactive sites) and binding affinity (structural integrity). The function of fungal SSI proteins identified in this study provides not only a clue to fungal pathogenesis via protease inhibition but also a template for the design of novel serine protease inhibitors.IMPORTANCE Until recently, Streptomyces subtilisin inhibitors (SSI) were reported and characterized only in bacteria. We found SSI-like domains in a plant-pathogenic fungus, Choanephora cucurbitarum KUS-F28377, which contains 11 sequentially diverged SSI-like domains. None of these fungal SSI-like domains were functionally characterized before. The active form of fungal SSI-like protein is a monomer, in contrast to the homodimeric bacterial SSI. We constructed a synthetic monomer of bacterial SSI to demonstrate the modulation of its activity based on structural integrity and not reactive sites. Our results suggest the duplication and divergence of SSI-like domains of C. cucurbitarum within the genome to inhibit various cognate proteases during evolution by modulating both binding and reactivity. The molecular functional characterization of fungal SSI-like domains will be useful in understanding their biological role and future biotechnological applications.

NADP-dependent glutamate dehydrogenases in a dimorphic zygomycete Benjaminiella poitrasii: Purification, characterization and their evaluation as an antifungal drug target.

BACKGROUND: It has been reported that the genes coding for NADP-dependent glutamate dehydrogenases (NADP-GDHs) showed a cause-effect relationship with Yeast-Hypha (YH) reversible transition in a zygomycete Benjaminiella poitrasii. As YH transition is significant in human pathogenic fungi for their survival and proliferation in the host, the NADP-GDHs can be explored as antifungal drug targets. METHODS: The yeast-form specific BpNADPGDH I and hyphal-form specific BpNADPGDH II of B. poitrasii were purified by heterologous expression in E. coli BL-21 cells and characterized. The structural analogs of L-glutamate, dimethyl esters of isophthalic acid (DMIP) and its derivatives were designed, synthesized and screened for inhibition of NADP-GDH activity as well as YH transition in B. poitrasii, and also in human pathogenic Candida albicans strains. RESULTS: The BpNADPGDH I and BpNADPGDH II were found to be homo-hexameric proteins with native molecular mass of 282 kDa and 298 kDa, respectively and subunit molecular weights of 47 kDa and 49 kDa, respectively. Besides the distinct kinetic properties, BpNADPGDH I and BpNADPGDH II were found to be regulated by cAMP-dependent- and Calmodulin (CaM) dependent- protein kinases, respectively. The DMIP compounds showed a more pronounced effect on H-form specific BpNADPGDH II and inhibited YH transition as well as growth in B. poitrasii and C. albicans strains. CONCLUSION: The present study will be useful to design and develop antifungal drugs against dimorphic human pathogens using glutamate dehydrogenase as a target. SIGNIFICANCE: Glutamate dehydrogenases can be explored as a target against human pathogenic fungi.

New anti-neuroinflammatory steroids against LPS induced NO production in BV2 microglia cells by microbial transformation of isorhodeasapogenin.

Microbial transformation of isorhodeasapogenin (1), the major steroidal sapogenin of Tupistra chinensis, was performed with the fungus Syncephalastrum racemosum (AS 3.264). As a result, nine new biotransformation metabolites (2-10) were isolated and their structures were elucidated by spectroscopic analysis. Hydroxylation, oxidation and glycosylation reactions were observed on the B, C, D and F rings of steroidal skeleton. Substrate (1) and its biotransformed metabolites 2-6, 8-10 were evaluated for their anti-neuroinflammatory effect on the NO accumulation induced by LPS in BV-2 cells. All the tested metabolites were found to have more potential anti-neuroinflammatory activity than the substrate. Especially, metabolites 2, 5 and 6 exhibited significant inhibition on NO production after hydroxylation at C-12 or C-15. Moreover, metabolite 2 dose-dependently reduced the LPS-induced protein expression of iNOS and COX-2.

Functional surface proteomic profiling reveals the host heat-shock protein A8 as a mediator of Lichtheimia corymbifera recognition by murine alveolar macrophages.

Mucormycosis is an emergent, fatal fungal infection of humans and warm-blooded animals caused by species of the order Mucorales. Immune cells of the innate immune system serve as the first line of defence against inhaled spores. Alveolar macrophages were challenged with the mucoralean fungus Lichtheimia corymbifera and subjected to biotinylation and streptavidin enrichment procedures followed by LC-MS/MS analyses. A total of 28 host proteins enriched for binding to macrophage-L. corymbifera interaction. Among those, the HSP70-family protein Hspa8 was found to be predominantly responsive to living and heat-killed spores of a virulent and an attenuated strain of L. corymbifera. Confocal scanning laser microscopy of infected macrophages revealed colocalization of Hspa8 with phagocytosed spores of L. corymbifera. The amount of detectable Hspa8 was dependent on the multiplicity of infection. Incubation of alveolar macrophages with an anti-Hspa8 antibody prior to infection reduced their capability to phagocytose spores of L. corymbifera. In contrast, anti-Hspa8 antibodies did not abrogate the phagocytosis of Aspergillus fumigatus conidia by macrophages. These results suggest an important contribution of the heat-shock family protein Hspa8 in the recognition of spores of the mucoralean fungus L. corymbifera by host alveolar macrophages and define a potential immunomodulatory therapeutic target.

Biotechnological Strategies for Chitosan Production by Mucoralean Strains and Dimorphism Using Renewable Substrates.

We investigated the influence of corn steep liquor (CSL) and cassava waste water (CWW) as carbon and nitrogen sources on the morphology and production of biomass and chitosan by Mucor subtilissimus UCP 1262 and Lichtheimia hyalospora UCP 1266. The highest biomass yields of 4.832 g/L (M. subtilissimus UCP 1262) and 6.345 g/L (L. hyalospora UCP 1266) were produced in assay 2 (6% CSL and 4% CWW), factorial design 22, and also favored higher chitosan production (32.471 mg/g) for M. subtilissimus. The highest chitosan production (44.91 mg/g) by L. hyalospora (UCP 1266) was obtained at the central point (4% of CWW and 6% of CSL). The statistical analysis, the higher concentration of CSL, and lower concentration of CWW significantly contributed to the growth of the strains. The FTIR bands confirmed the deacetylation degree of 80.29% and 83.61% of the chitosan produced by M. subtilissimus (UCP 1262) and L. hyalospora (UCP 1266), respectively. M. subtilissimus (UCP 1262) showed dimorphism in assay 4-6% CSL and 8% CWW and central point. L. hyalospora (UCP 1266) was optimized using a central composite rotational design, and the highest yield of chitosan (63.18 mg/g) was obtained in medium containing 8.82% CSL and 7% CWW. The experimental data suggest that the use of CSL and CWW is a promising association to chitosan production.

Comparative aroma and taste profiles of oil furu (soybean curd) fermented with different mucor strains.

The effects of different mucor strains (Mucor racemosus, Actinomucor, and Mucor wutungkiao) on aroma and taste profiles based on proteolysis, lipolysis, and their catabolism in oil furu were studied. Gas chromatography-mass spectrometry and relative odor activity were used to monitor the changes of key volatile compounds and the differences in the characteristic aroma contents of oil furu. Using principal component analysis, the different fermentation strains had an effect on aroma profiles. The volatile compounds from metabolism of protein and fatty acid contributed to the aroma of oil furu with different contribution from the different strains, presumably due to their different enzymes. The electronic tongue and free amino acid profiles also showed strain differences of taste. Based on these results, optimization of the amount of each of the different mucor strains during cofermentation might achieve better flavor.