Enhancement of Bacillus subtilis Growth and Sporulation by Two-Stage Solid-State Fermentation Strategy

Zhao Zhao,Ma Ma,Xu Xu,Xi Xi

doi:10.3390/pr7100644

Abstract

Two-stage solid-state fermentation strategy was exploited and systematically optimized to enhance Bacillus subtilis growth and sporulation for increasing effective cell number in B. subtilis microbial ecological agents. The first stage focused on improving cell growth followed by the second stage aiming to enhance both cell growth and sporulation. The optimal fermentation condition was that temperature changed from 37 °C to 47 °C at a fermentation time of 48 h and Mn2+ content in medium was 4.9 mg MnSO4/g dry medium. Solid medium properties were improved by the optimal two-stage fermentation. HPLC results demonstrated that glucose utilization was facilitated and low-field nuclear magnetic resonance (LF-NMR) results showed that more active sites in medium for microbial cells were generated during the optimal two-stage fermentation. Moreover, microbial growth and sporulation were enhanced simultaneously during the second stage of fermentation through delaying microbial decline phase and increasing sporulation rate. As a result, effective cell number of B. subtilis reached 1.79 × 1010/g dry medium after fermentation for 72 h, which was 29.7% and 8.48% higher than that of conventional fermentation for 72 h and 48 h, respectively. Therefore, the optimal two-stage fermentation could increase the effective cell number of B. subtilis microbial ecological agents efficiently.

Highlights

Microbial ecological agents are environmental friendly alternatives to antibiotics, which can be utilized as feed additives for preventing disease and promoting growth of animals [1]
Because the optimal conditions are different between microbial growth and sporulation, a two-stage fermentation has been considered as an effective strategy to promote both cell growth and spore formation
Combining with the previous research, this study presented a promising way to increase the effective cell number of microbial ecological agents, which should facilitate the cleaner production of alternatives to antibiotics utilizing agro-industrial residues by solid-state fermentation (SSF)

Summary

Introduction

Microbial ecological agents are environmental friendly alternatives to antibiotics, which can be utilized as feed additives for preventing disease and promoting growth of animals [1]. Bacillus subtilis can generate spores, which have strong resistance to environmental stress including heat in drying process, mechanical force in granulation process, and acid in digestive tract. Effective cell number involving spore number and viable cell number is the key measurement to evaluate the quality of B. subtilis microbial ecological agents [2,3]. Because the optimal conditions are different between microbial growth and sporulation, a two-stage fermentation has been considered as an effective strategy to promote both cell growth and spore formation. The first stage aimed at improving the cell growth followed by the second stage which was under the optimal conditions for sporulation. Sen and Babu employed the response surface methodology to optimize the operation parameters including pH, Processes 2019, 7, 644; doi:10.3390/pr7100644 www.mdpi.com/journal/processes

Methods

Results

Conclusion