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Aspergillus oryzae is a filamentous fungus widely used in food industry and as a microbial cell factory for recombinant protein production. Due to the inherent resistance of A. oryzae to common antifungal compounds, genetic transformation of this mold usually requires auxotrophic mutants. In this study, we show that Agrobacterium tumefaciens-mediated transformation (ATMT) method is very efficient for deletion of the pyrG gene in different Aspergillus oryzae wild-type strains to generate uridine/uracil auxotrophic mutants. Our data indicated that all the obtained uridine/uracil auxotrophic transformants, which are 5- fluoroorotic acid (5-FOA) resistant, exist as the pyrG deletion mutants. Using these auxotrophic mutants and the pyrG selectable marker for genetic transformation via A. tumefaciens, we could get about 1060 transformants per 10 fungal spores. In addition, these A. oryzae mutants were also used successfully for expression of the DsRed fluorescent reporter gene under control of the A. oryzae amyB promoter by the ATMT method, which resulted in obvious red transformants on agar plates. Our work provides a new and effective approach for constructing the uridine/uracil auxotrophic mutants in the importantly industrial fungus A. oryzae. This strategy appears to be applicable to other filamentous fungi to develop similar genetic transformation systems based on auxotrophic/nutritional markers for food-grade recombinant applications.

Medicinal plants play important roles in traditional medicine, and numerous compounds among them have been recognized for their antimicrobial activity. However, little is known about the potential of Vietnamese medicinal plants for antifungal activity. In this study, we examined the antagonistic activity of twelve medicinal plant species collected in Northern Vietnam against Penicillium digitatum, Aspergillus flavus, Aspergillus fumigatus, and Candida albicans. The results showed that the antifungal activities of the crude extracts from Mahonia bealei, Ficus semicordata, and Gnetum montanum were clearly detected with the citrus postharvest pathogen P. digitatum. These extracts could fully inhibit the growth of P. digitatum on the agar medium, and on the infected citrus fruits at concentrations of 300–1000 µg/mL. Meanwhile, the other tested fungi were less sensitive to the antagonistic activity of the plant extracts. In particular, we found that the ethanolic extract of M. bealei displayed a broad-spectrum antifungal activity against all four pathogenic fungi. Analysis of this crude extract by enrichment coupled with high-performance liquid chromatography revealed that berberine and palmatine are major metabolites. Additional inspections indicated berberine as the key compound responsible for the antifungal activity of the M. bealei ethanolic extract. Our study provides a better understanding of the potential of Vietnamese medicinal plant resources for combating fungal pathogens. This work also highlights that the citrus pathogen P. digitatum can be employed as a model fungus for screening the antifungal activity of botanicals.

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Porcine epidemic diarrhea virus (PEDV) causes diarrhea in pigs leading to severe illnesses and high mortality rates. The development of medicinal agents to treat PEDV infection is therefore crucial. In this study, antiviral activities against PEDV of ethanol and aqueous extracts of 17 Vietnamese traditional medicinal plants were evaluated using the cytopathic effect-based assay. The results showed that 14 out of 17 medicinal plants could inhibit the cytopathic effect of PEDV. The ethanol extract of Stixis scandens was identified as the most active extract with its MIC (minimum inhibitory concentration) being 0.15 lg/ mL. Other plant extracts also displayed strong antiviral activity against PEDV, including Anisomeles indica, Pericampylus glaucus and Croton kongensis. The results demonstrate that certain medicinal plants have a high antiviral potential and may serve as a lead to develop novel pharmaceutical agents to cure PED as well as the diseases caused by other coronaviruses.