Inhibition of Aspergillus oryzae Mycelium Growth and Conidium Production by Irradiation with Light at Different Wavelengths and Intensities.

Aspergillus oryzae is a safe filamentous fungus widely used in the food, medicine, and feed industries, but there is currently not enough research on the light response of A. oryzae. In this study, 12 different light conditions were set and A. oryzae GDMCC 3.31 was continuously irradiated for 72 h to investigate the effect of light on mycelial growth and conidium production. Specifically, each light condition was the combination of one light wavelength (475, 520, or 630 nm) and one light intensity (20, 40, 60, or 80 µmol photon m-2 s-1). The results show that mycelium growth was inhibited significantly by green light (wavelength of 520 nm and intensities of 20 and 60 µmol photon m-2 s-1) and blue light (wavelength of 475 nm and intensity of 80 µmol photon m-2 s-1). The production of conidia was suppressed only by blue light (wavelength of 475 nm and intensities of 40, 60, and 80 µmol photon m-2 s-1), and those levels of inhibition increased when the intensity of blue light increased. When the strain was irradiated by blue light (80 µmol photon m-2 s-1), the number of conidia was 57.4% less than that of the darkness group. However, within our set range of light intensities, A. oryzae GDMCC 3.31 was insensitive to red light (wavelength of 630 nm) in terms of mycelium growth and conidium production. Moreover, interaction effects between light wavelength and intensity were found to exist in terms of colony diameter and the number of conidia. This research investigated the light response of A. oryzae, which may provide a new method to regulate mixed strains in fermented foods by light. IMPORTANCE Studies on the monochromatic light response of Aspergillus nidulans and Neurospora crassa have gone deep into the molecular mechanism. However, research methods for the light response of A. oryzae remain in the use of white light sources. In this study, we first demonstrated that A. oryzae GDMCC 3.31 was sensitive to light wavelength and intensity. We have observed that blue light inhibited its growth and sporulation and the inhibitory effect increased with intensity. This research not only adds new content to the study of the photoreaction of Aspergillus but also brings new possibilities for the use of light to regulate mixed strains and ultimately improve the flavor quality of fermented foods.

Optimization of liquid-state fermentation conditions for the glyphosate degradation enzyme production of strain Aspergillus oryzae by ultraviolet mutagenesis.

This study aimed to obtain strains with high glyphosate-degrading ability and improve the ability of glyphosate degradation enzyme by the optimization of fermentation conditions. Spore from Aspergillus oryzae A-F02 was subjected to ultraviolet mutagenesis. Single-factor experiment and response surface methodology were used to optimize glyphosate degradation enzyme production from mutant strain by liquid-state fermentation. Four mutant strains were obtained and named as FUJX 001, FUJX 002, FUJX 003, and FUJX 004, in which FUJX 001 gave the highest total enzyme activity. Starch concentration at 0.56%, GP concentration at 1,370 mg/l, initial pH at 6.8, and temperature at 30°C were the optimum conditions for the improved glyphosate degradation endoenzyme production of A. oryzae FUJX 001. Under these conditions, the experimental endoenzyme activity was 784.15 U/100 ml fermentation liquor. The result (784.15 U/100 ml fermentation liquor) was approximately 14-fold higher than that of the original strain. The result highlights the potential of glyphosate degradation enzyme to degrade glyphosate.

Enhanced ß-galactosidase production of Aspergillus oryzae mutated by UV and LiCl.

In order to breed a high-yield ß-galactosidase-producing strain, Aspergillus oryzae was used as the parent strain and mutagenized with ultraviolet (UV) and UV plus lithium chloride (LiCl), respectively. After being mutagenized by UV, the ß-galactosidase activity of mutant UV-15-20 reached 114.08 U/mL, which revealed a 49.22% increase compared with the original strain. A mutant UV-LiCl-38 with high ß-galactosidase activity (121.42 U/mL) was obtained after compound mutagenesis of UV and LiCl; the ß-galactosidase activity of this mutant was 58.82% higher than that of the parent strain. Subculture testing indicated that UV-15-20 and UV-LiCl-38 had good hereditary stability and may be ideal strains for the production of ß-galactosidase. Additionally, it was demonstrated that compound mutagenesis with UV and LiCl is an effective mutation method for breeding industrially interesting strains.

Lethal and mutagenic effects of ion beams and γ-rays in Aspergillus oryzae.

Aspergillus oryzae is a fungus that is used widely in traditional Japanese fermentation industries. In this study, the lethal and mutagenic effects of different linear energy transfer (LET) radiation in freeze-dried conidia of A. oryzae were investigated. The lethal effect, which was evaluated by a 90% lethal dose, was dependent on the LET value of the ionizing radiation. The most lethal ionizing radiation among that tested was (12)C(5+) ion beams with an LET of 121keV/µm. The (12)C(5+) ion beams had a 3.6-times higher lethal effect than low-LET (0.2keV/µm) γ-rays. The mutagenic effect was evaluated by the frequency of selenate resistant mutants. (12)C(6+) ion beams with an LET of 86keV/µm were the most effective in inducing selenate resistance. The mutant frequency following exposure to (12)C(6+) ion beams increased with an increase in dose and reached 3.47×10(-3) at 700Gy. In the dose range from 0 to 700Gy, (12)C(5+) ion beams were the second most effective in inducing selenate resistance, the mutant frequency of which reached a maximum peak (1.67×10(-3)) at 400Gy. To elucidate the characteristics of mutation induced by ionizing radiation, mutations in the sulphate permease gene (sB) and ATP sulfurylase gene (sC) loci, the loss of function of which results in a selenate resistant phenotype, were compared between (12)C(5+) ion beams and γ-rays. We detected all types of transversions and transitions. For frameshifts, the frequency of a +1 frameshift was the highest in all cases. Although the incidence of deletions >2bp was generally low, deletions >20bp were characteristic for (12)C(5+) ion beams. γ-rays had a tendency to generate mutants carrying a multitude of mutations in the same locus. Both forms of radiation also induced genome-wide large-scale mutations including chromosome rearrangements and large deletions. These results provide new basic insights into the mutation breeding of A. oryzae using ionizing radiation.

Responses of Aspergillus oryzae to stimuli from near-UV light irradiation in the presence or absence of titanium dioxide.

The responses of Aspergillus oryzae to the stimuli from near-UV irradiation were investigated by using a black light fluorescent lamp in the presence or absence of TiO2 particles. Light irradiation at an intensity of 6 W/m2 strongly inhibited the growth of germinated pellets of A. oryzae. This growth inhibition was weakened by TiO2 particles (0.05 g/l), especially in the initial growth phase, in which the expression level of catalase gene (catB) was approximately three times higher than that in the absence of TiO2 particles. However, the initial induction of catB expression by H2O2 pretreatment did not restore the growth under the black light irradiation. The weakening of growth inhibition is thought to result from alternative physiological responses of A. oryzae against stimulus by photo-excited TiO2.

Radiobiological investigations of soft X-rays near carbon, nitrogen, oxygen K-shell edges on Aspergillus oryzae spores.

Soft X-rays at carbon, nitrogen, oxygen K-shell edges have special radiobiological effects. Using Aspergillus oryzae spores as sample, the radiation effects of soft X-rays near the K-shell edges of C, N and O elements from synchrotron radiation were investigated. Also the dose depositions of different X-ray energies in spore were discussed. At the same time, the spores were irradiated by gamma rays from 60Co and relative biological effects were compared with those produced by soft X-rays. The results showed that soft X-rays near K-shell edges of O element had higher ability of radiation damage than that of X-rays near K-shell edges of C and N elements as compared with one another. But they all had higher killing abilities per unit dose than that of gamma rays from 60Co. The relative biological effects (RBEs), the comparison of dose to gamma rays at 10% survival level, of the three soft X-rays were 1.65, 1.73 and 1.91, respectively.

Light represses conidiation in koji mold Aspergillus oryzae.

In the filamentous fungus Aspergillus oryzae, there has been no report on photoreaction. Here we investigated the effect of light in A. oryzae and found that conidiation was repressed by white light. This reaction is contrary to that of other Aspergilli, which show abundant conidiation under light. Moreover, red light also caused reduced conidiation. Genome sequencing of A. oryzae indicated the existence of homologs of some light-related genes in other filamentous fungi. To approach the molecular mechanism of this photoresponse, the effect of red light on the expression level of several genes putatively responsible for conidiation or photoperception, i.e., brlA, a gene known to be required for conidiation, AofphA, the putative homolog of the A. nidulans phytochrome gene fphA, and AoveA, the putative homolog of the negative regulator gene in conidiation in A. nidulans, was examined. These three genes showed no significant response to red light at the transcriptional level. The results indicate that A. oryzae perceives and responds to red light in a manner independent of the transcriptional regulation of these genes.

Visualizing nuclear migration during conidiophore development in Aspergillus nidulans and Aspergillus oryzae: multinucleation of conidia occurs through direct migration of plural nuclei from phialides and confers greater viability and early germination in Aspergillus oryzae.

Nuclear migration is indispensable for normal growth, differentiation, and development, and has been studied in several fungi including Aspergillus nidulans and Neurospora crassa. To better characterize nuclear movement and its consequences during conidiophore development, conidiation, and conidial germination, we performed confocal microscopy and time-lapse imaging on A. nidulans and Aspergillus oryzae strains expressing the histone H2B-EGFP fusion protein. Active trafficking of nuclei from a vesicle to a phialide and subsequently into a conidium provided the mechanistic basis for the formation of multinucleate conidia in A. oryzae. In particular, the first direct visual evidence on multinucleate conidium formation by the migration of nuclei from a phialide into the conidium, rather than by mitotic division in a newly formed conidium, was obtained. Interestingly, a statistical analysis on conidial germination revealed that conidia with more nuclei germinated earlier than those with fewer nuclei. Moreover, multinucleation of conidia conferred greater viability and resistance to UV-irradiation and freeze-thaw treatment.