Paraben hydrolase and tannase catalyze the hydrolysis of parabens (4-hydroxybenzoic acid esters) and gallic acid (3,4,5-trihydroxybenzoic acid) esters, respectively. Paraben hydrolase (AoPrbA) and tannase (AoTanB) from Aspergillus oryzae belong to the tannase family in the ESTHER database. However, the substrate specificities of AoPrbA and AoTanB are narrow. Based on structural information of Aspergillus niger tannase (PDB code 7k4o), we constructed five single variants of AoPrbA (Thr200Glu, Phe231Gln, Leu232Gln, Ile361Tyr, and Leu428Ser) and four of AoTanB (Glu203Asp, Glu203Thr, His237Ala, and Ser440Leu) to investigate substrate discrimination between AoPrbA and AoTanB. Each variant was expressed in Pichia pastoris and were purified from the culture supernatant. Five purified variants of AoPrbA and four variants of AoTanB showed reduced paraben hydrolase and tannase activities compared with AoPrbA and AoTanB wild types, respectively. Interestingly, the AoPrbA wild type did not hydrolyze gallic acid methyl ester, whereas the Thr200Glu, Leu232Gln, and Leu428Ser variants did, indicating that these three variants acquired tannase activity. In particular, the Leu428Ser variant exhibited considerably greater hydrolysis of gallic acid and protocatechuic acid methyl esters. Meanwhile, the AoTanB wild type, and Glu203Asp, His237Ala and Ser440Leu variants hydrolyzed the protocatechuate methyl and 4-hydroxybenzoate ethyl esters; however, the Glu203Thr variant did not hydrolyze above-mentioned substrates. Additionally, the ratio of paraben hydrolase activity to tannase activity in Ser440Leu was markedly elevated.

Asperalins represent a novel class of viridicatin natural products with potent inhibitory activities against fish pathogens. In this study, we elucidated the biosynthesis of asperalins in the Aspergillus oryzae NSAR1 heterologous host and identified the FAD-dependent monooxygenase AplB stereoselectively hydroxylates viridicatin to yield a unique 3R,4S configuration. The monomodular NRPS AplJ catalyzes a rare intramolecular ester bond formation reaction using dihydroquinoline as a nucleophile. Subsequent modifications by cytochrome P450 AplF, chlorinase AplN, and prenyltransferase AplE tailor the anthranilic acid portion, leading to the formation of asperalins. Additionally, we explored the potential of AplB for the hydroxylation of viridicatin analogs, demonstrating its relaxed substrate specificity. This finding suggests that AplB could be developed as a biocatalyst for the synthesis of viridicatin derivatives.

Aspergillus flavus produces aflatoxin, a carcinogenic fungal toxin that poses a threat to the agricultural and food industries. There is a concern that the distribution of aflatoxin-producing A. flavus is expanding in Japan due to climate change, and it is necessary to understand what types of strains inhabit. In this study, we sequenced the genomes of four Aspergillus strains isolated from agricultural fields in the Ibaraki prefecture of Japan and identified their genetic variants. Phylogenetic analysis based on single-nucleotide variants revealed that the two aflatoxin-producing strains were closely related to A. flavus NRRL3357, whereas the two non-producing strains were closely related to the RIB40 strain of Aspergillus oryzae, a fungus widely used in the Japanese fermentation industry. A detailed analysis of the variants in the aflatoxin biosynthetic gene cluster showed that the two aflatoxin-producing strains belonged to different morphotype lineages. RT-qPCR results indicated that the expression of aflatoxin biosynthetic genes was consistent with aflatoxin production in the two aflatoxin-producing strains, whereas the two non-producing strains expressed most of the aflatoxin biosynthetic genes, unlike common knowledge in A. oryzae, suggesting that the lack of aflatoxin production was attributed to genes outside of the aflatoxin biosynthetic gene cluster in these strains.

DNA-binding transcription factors (TFs) play a central role in transcriptional regulation mechanisms, mainly through their specific binding to target sites on the genome and regulation of the expression of downstream genes. Therefore, a comprehensive analysis of the function of these TFs will lead to the understanding of various biological mechanisms. However, the functions of TFs in vivo are diverse and complicated, and the identified binding sites on the genome are not necessarily involved in the regulation of downstream gene expression. In this study, we investigated whether DNA structural information around the binding site of TFs can be used to predict the involvement of the binding site in the regulation of the expression of genes located downstream of the binding site. Specifically, we calculated the structural parameters based on the DNA shape around the DNA binding motif located upstream of the gene whose expression is directly regulated by one TF AoXlnR from Aspergillus oryzae, and showed that the presence or absence of expression regulation can be predicted from the sequence information with high accuracy ([Formula: see text]-1.0) by machine learning incorporating these parameters.

Aspergillus species are common hyphal fungi. In addition to allergies and mycotoxicosis, Aspergillus species can cause various infections known as aspergillosis. Aspergillosis of the respiratory tract, central nervous system, skin and soft tissues is well described. However, musculoskeletal infections due to invasive aspergillosis are not well described. Fungal joint infection due to invasive aspergillosis is a rare form of septic arthritis. In this case report, a patient who admitted to our hospital for liver transplantation and developed knee joint arthritis caused by Aspergillus flavus/Aspergillus oryzae during this process was presented. A 28-year-old male patient with autoimmune hepatitis was admitted to hospital with decompensated liver cirrhosis and encephalopathy. The patient, who was awaiting an emergency liver transplant, developed pain, swelling and limitation of movement in his right knee and appropriate consultations and tests were requested. Three joint fluid cultures taken one day apart and nine days later were positive for fungal growth. Macroscopic examination of the mould growth and microscopic examination with lactophenol cotton blue suggested a species belonging to the A.flavus complex and the isolate was identified as A.flavus/A.oryzae by matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS) (EXS 2600, Zybio, China). As a result of ITS gene sequencing, the species was determined to be A.oryzae. As cases have been reported where A.flavus and A.oryzae species could not be distinguished by ITS gene sequencing, the pathogen was defined as A.flavus/oryzae. The patient died of liver disease during treatment with amphotericin B. There are few cases of arthritis caused by Aspergillus species in the literature. Aspergillus species found in joint infections are, Aspergillus fumigatus, A.flavus, Aspergillus niger and Aspergillus terreus species complexes, in order of frequency. A.flavus and A.oryzae are closely related. They are difficult to distinguish by conventional methods, MALDI-TOF MS or ITS region sequencing, which is commonly used for genus/species identification in fungi. The number of Aspergillus arthritis cases is low and the identification methods applied to the species reported as causative agents in most studies can identify at the species complex level. In addition, it can be assumed that species not previously reported as causative agents may be encountered as a result of developments in identification methods. In the few publications in the literature where A.flavus complex was reported as the causative agent of joint infections, it seems possible that some of the agents may be A.flavus and some may be A.oryzae, since the agents were identified at the complex level. There are a limited number of cases in the literature where A.oryzae is the causative agent, particularly in the respiratory tract. A PubMed search using the keywords \"A.oryzae infections, arthritis, osteomyelitis\" did not reveal any literature on joint infections caused by A.oryzae.

This study addresses the challenge of enhancing gamma-aminobutyric acid (GABA) content in soy sauce through optimized fermentation condition. Using a multiple starter culture, consisting of Aspergillus oryzae strain NSK, Bacillus cereus strain KBC and Tetragenococcus halophilus strain KBC, the incubation conditions including the percentage of bacterial inoculum (10, 15 and 20 %), pH (3, 5 and 7) and agitation speed (100, 150 and 200 rpm) were optimized through Response Surface Methodology (RSM). Under the optimal conditions (20 % inoculum, pH 7 and stirring at 100 rpm), the multiple starter culture generated 128.69 mg/L of GABA after 7 days and produced 239.08 mg/L of GABA after 4 weeks of fermentation, which is 36 % higher than under non-optimized conditions (153.48 mg/L). Furthermore, sensory analysis revealed high consumer acceptance of the fermented soy sauce than the control (soy sauce without any treatment and additional bacteria) and commercial soy sauce. Consumers indicated that the starter culture offered an improved umami taste and reduced bitter, sour and salty flavours compared to the commercial product. Under optimal fermentation conditions determined by RSM statistical analysis, the multiple starter culture is able to produce high levels of GABA and is more likely to be accepted by consumers. The findings of this research have the potential to impact the food sector by offering a functional soy sauce with added health benefits and also being well-received by consumers.

Solid-state fermentation (SSF) and extrusion are effective methods to improve the nutritional and sensory quality of rice bran. The effect of the processing sequence of SSF and extrusion and microbial strains on the quality of rice bran was studied. The results showed that the first SSF followed by extrusion increased the contents of phenolic, flavonoid and γ-oryzanol, but the color changed to brown. The first extrusion followed by SSF caused damage to bioactive components and antioxidant activity, but significantly increased the content of arabinoxylans. The difference between the two processing sequences may be related to the process time and the effect of substrate on microbial induction. Aspergillus oryzae and Neurospora sitophila were suitable for increasing the bioactive components of rice bran, while Lactiplantibacillus plantarum was suitable for increasing water-extractable arabinoxylan content. Different processing sequences and microbial strains have their advantages, and these results can provide reference for rice bran processing.

Fungi produce various bioactive secondary metabolites (SMs) as protective and weaponized tools to enhance survival in shared ecological niches. By mimicking a competitive ecosystem, cocultivation has been proven to be particularly successful in stimulating SM discovery. Here, we reported the identification of four novel metabolites, epiclactones A and B, epioxochromane and aoergostane, from the coculture of two biotechnologically important strains, Aspergillus oryzae and Epicoccum dendrobii. Transcriptome and metabolome analyses revealed widespread silent gene activation during fungal-fungal interaction. The majority of differentially expressed gene clusters were summarized for both strains. Based on these highly activated biosynthetic pathways, we suggested that a bidirectional chemical defense occurred under cocultivation. E. dendrobii enhanced the production of the spore inhibitor, fumigermin. Moreover, A. oryzae highly accumulated the antifungal agent kojic acid with a yield of up to 1.10 g/L. This study provides an excellent example for the discovery of hidden natural products by cocultivation.

Aflatoxins (AFs) are carcinogenic fungal toxins contaminating up to 25% of the global food supply. Over half of the world\'s population is exposed to unmonitored levels of AFs, mostly aflatoxin B1 (AFB1). Despite numerous efforts over the past 60 years, there are no solutions to remove AFs safely from food. Here, we present a safe and effective AF-degrading product called \"D-Tox\", a filtered culture broth of Aspergillus oryzae grown in a food-grade liquid medium. When 5 ppm of AFB1 is added to D-Tox, ∼90% is degraded at 48 and 24 hr at room temperature and 50°C, respectively. Moreover, when varying amounts (0.1 ppm ∼ 100 ppm) of AFB1 are added to D-Tox at 100°C, over 95% of AFB1 is degraded in 1 hr, suggesting a nonenzymatic process. Examining degradation of 100 ppm AFB1 reveals that aflatoxin D1 (AFD1) is the major transient degradant of AFB1, indicating that degradation occurs irreversibly by lactone ring hydrolysis followed by decarboxylation. D-Tox further degrades AFD1 to unknown fragmented products. Importantly, the practical application of D-Tox is also demonstrated, as more than 70% of AFB1 is degraded when wheat, corn, and peanuts naturally contaminated with high levels of AFB1 (0.3 ∼ 4.5 ppm) are boiled in D-Tox for 1 hr. Additionally, D-Tox can degrade other lactone-ring containing mycotoxins, including patulin and ochratoxin. D-Tox exhibits no cytotoxicity under the conditions tested in MCF-7 breast cancer cell lines. In summary, D-Tox is a safe and effective AF-detoxifying product that can enhance global food safety.

Amazake is a traditional, sweet, non-alcoholic Japanese beverage typically produced through koji fermentation by the fungus Aspergillus oryzae. However, alternative microorganisms such as Bacillus amyloliquefaciens offer potential advantages and novel possibilities for producing similar fermented beverages. This study aimed to replicate the ancestral beverage of amazake by replacing A. oryzae (W-20) with B. amyloliquefaciens (NCIMB 12077) and comparing their fermentation processes and resulting products. Our results show that the production of amazake with B. amyloliquefaciens (ABA) is not only possible but also results in a beverage that is otherwise distinct from traditional amazake (AAO). Saccharification was achievable in ABA at higher temperatures than in AAO, albeit with lower reducing sugar and enzymatic activity values. Amino acids and organic acids were more abundant in AAO, with cysteine being uniquely present in AAO and shikimic acid only being present in ABA. The volatile aroma compound profiles differed between the two beverages, with AAO exhibiting a greater abundance of aldehydes, and ABA a greater abundance of ketones and alcohols. Interestingly, despite these compositional differences, the two beverages showed similar consumer panel acceptance rates. An analysis of their microbial communities revealed pronounced differences between the amazakes, as well as temporal changes in ABA but not in AAO. This study provides promising insights into harnessing the potential of B. amyloliquefaciens as the primary microorganism in the fermentation process of amazake-like beverages, marking an important advancement in the field of fermented low-alcohol beverage production, with possible applications in other fermented foods.