吉岡 育哲

Methanol reportedly stimulates citric acid (CA) production by Aspergillus niger and A. tubingensis; however, the underlying mechanisms remain unclear. Here, we elucidated the molecular functions of the citrate exporter gene cexA in relation to CA production by A. tubingensis WU-2223L. Methanol addition to the medium containing glucose as a carbon source markedly increased CA production by strain WU-2223L by 3.38-fold, resulting in a maximum yield of 65.5 g/L, with enhanced cexA expression. Conversely, the cexA-complementing strain with the constitutive expression promoter Ptef1 (strain LhC-1) produced 68.3 or 66.7 g/L of CA when cultivated without or with methanol, respectively. Additionally, strain LhC-2 harboring two copies of the cexA expression cassette produced 80.7 g/L of CA without methanol addition. Overall, we showed that cexA is a target gene for methanol in CA hyperproduction by A. tubingensis WU-2223L. Based on these findings, methanol-independent CA-hyperproducing strains, LhC-1 and LhC-2, were successfully generated.

BACKGROUND: Filamentous fungi of the genus Madurella are the primary causative agents of mycetoma, a disease observed in tropical and subtropical regions. Since early diagnostics based on a morphological approach are difficult and have many shortcomings, a molecular diagnostic method suitable for rural settings is required. In this study, we developed the loop-mediated isothermal amplification (LAMP) method to present a foundational technique of the diagnosis of Madurella spp. (M. mycetomatis, M. pseudomycetomatis, M. tropicana, and M. fahalii), the common causative organisms of eumycetoma. PRINCIPAL FINDINGS: We successfully designed a primer pair targeting the rDNAs of three Madurella spp. excluding M. fahalii, and detected up to 100 fg of genomic DNA extracted from isolates of M. mycetomatis and 1 pg of M. pseudomycetomatis and M. tropicana, within one hour. Second, a primer pair specific to M. mycetomatis, the most common causative species, or M. fahalii, a drug-resistant species, was constructed, and the detection limit of both primer pairs was 1 pg. The designed primers accurately distinguished 16 strains of the genus Madurella from various fungal species known to cause mycetomas. CONCLUSION: In summary, we established the first model of a LAMP detection method that rapidly and sensitively detects and identifies Madurella isolates for clinical diagnostics. Moreover, the combined designed primer sets could identify mycetoma-causing strains simultaneously.

Aspergillus lacticoffeatus WU-2020 is a citric acid hyperproducer that is suitable for solid culture. Here, we present a high-quality draft of its genome sequence (35.9 Mb), which consists of 11 scaffolds and contains 11,490 genes. We also present the mitochondrial genome, which is 31.3 kb in length.

Ethyl α-D-glucopyranoside (α-EG) is detected in sake (Japanese rice wine), that has moisturizing and skin conditioning effects. The production of α-EG by fermentation or enzymatic synthesis to date generates unwanted by-products such as maltooligosaccharides and/or organic acids. In this study, we employed a reaction involving selective α-glucosylation of ethanol by the α-glucosyl transfer enzyme (XgtA) of Xanthomonas campestris WU-9701. Under standard conditions, when 0.80 M ethanol and 1.2 M maltose were used as substrates with XgtA (2.5 units) and incubated in 30 mM HEPES–NaOH buffer (pH 8.0) at 45°C, only one form of ethyl glucopyranoside was selectively obtained as a product. The isolated product was identified as ethyl α-D-glucopyranoside by 1H NMR, 1H–1H COSY, and NOESY analyses. In the reaction mixture, other glucosylated products such as maltotriose and ethylmaltoside were not detected. Under optimum conditions, 180 mM (37.5 g/L) α-EG was produced in one batch production for 80 h. Further, the reaction rate of α-EG production decreased with an increase in glucose, especially more than 500 mM. In contrast, the addition of glucose isomerase decreased the concentration of glucose and was useful for maintaining a glucose concentration of less than 500 mM in the reaction mixture. Thus, owing to the enzymatic reaction with XgtA and glucose isomerase, as much as 260 mM (54.1 g/L) α-EG was produced in one batch production for 100 h. Altogether, this study reports the highest concentration of α-EG produced by enzymatic reaction.

<jats:title>ABSTRACT</jats:title> <jats:p><jats:named-content content-type="genus-species">Aspergillus tubingensis</jats:named-content> WU-2223L, belonging to the section <jats:italic>Nigri</jats:italic>, is a hyperproducer of citric acid. Here, we present the high-quality draft (35 Mb) and mitochondrial (32.4 kb) genome sequences of this strain, which consisted of 16 scaffolds in total. The draft and mitochondrial genome sequences comprised 11,493 and 15 genes, respectively.</jats:p>