Document Type

Thesis - Open Access

Award Date

2020

Degree Name

Master of Science (MS)

Department / School

Biology and Microbiology

First Advisor

Bishnu Karki

Keywords

A. sojae, glyceollin, natural antimicrobial, pullulans, soybeans, T. reesei

Abstract

Glyceollin is a secondary metabolite produced under stress conditions by soybean to act as a plant defender to pathogen attack. The antimicrobial nature of glyceollin makes it a promising natural alternative to antibiotics if incorporated into livestock feed. However, production of glyceollin is highly variable, dependent on soybean variety, and fungal elicitor used as inoculum. Our study compares two fungal spore elicitors, Trichoderma reesei and Aspergillus sojae on soybeans of diverse maturity level, and fungal susceptibility to determine highest glyceollin titer. Spore inoculation with T. reesei elicited highest glyceollin titers of 2.42±0.20 mg/g at 120 h, while A. sojae elicited 1.45±0.49 mg/g at 96 h. Additionally, our results demonstrated decreasing trends in glucoside isoflavones, with increase in aglycone precursor daidzein for all soybean varieties. These results indicate the production of high level of glyceollin by T. reesei and A. sojae, which can be incorporated into livestock feed as a natural antimicrobial. To further enhance glyceollin production, a co-treatment of UV light and inoculation with A. sojae fungal spores was applied to soybeans. UV light treatment alone showed significantly lower total glyceollin as compared to fungal spore inoculum. When UV light treatment was applied for 5, 10, or 5 minutes along with A. sojae fungal spores higher average glyceollin titers were apparent compared to the fungal only treatment. Additionally, the UV light and A. spore fungal treatment did not show a significant effect to total isoflavones when compared to fungal only treatment. UV light and fungal spore elicitor treatment showed higher glyceollin titer compared to conventional fungal elicitor treatment. This method has potential to be utilized at industrial scale to drive research into the antimicrobial and human health benefits of glyceollin. Aureobasidium pullulans is a morphologically diverse fungus with industrial significance including enzymes, and pullulan production. Sucrose is known to support A. pullulans growth and pullulan synthesis. Our goal was to determine if growth rate of A. pullulans can be enhanced by varying the level of initial sucrose concentration. Three batch mode of fermentations (0.25, 0.5, and 5 L) were conducted. The preliminary trials conducted using sucrose concentrations of 5, 10, 15, 20, 25, and 50 g/L did not show any significant difference in growth kinetics of A. pullulans for 24 h fermentation. After 24 h, cell counts, pH, and dry cell mass showed significant difference in growth rate of A. pullulans for 5 and 10 g/L compared to the 50 g/L sucrose. Scaled-up batch fermentation (5 L Bioflo) showed similar results, and significantly increased pullulan production at 72 h for 50 g/L sucrose. These results indicated that initial sucrose concentration may not influence the growth rate in the first 24 h of incubation; however, it can significantly impact the cell viability and pullulan production capacity of A. pullulans during long term fermentation. Hence, high sucrose level could potentially increase the overall fermentation efficiency of A. pullulans for pullulan production.

Library of Congress Subject Headings

Soybean -- Disease and pest resistance.
Anti-infective agents.
Sucrose.
Polysaccharides.
Aspergillus.
Trichoderma reesei.

Format

application/pdf

Number of Pages

186

Publisher

South Dakota State University

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Rights Statement

In Copyright