Evaluation of an oxygen‐dependent self‐inducible surfactin synthesis in B. subtilis by substitution of native promoter PsrfA by anaerobically active PnarG and PnasD

A novel approach targeting self-inducible surfactin synthesis under oxygen-limited conditions is presented. Because both the nitrate (NarGHI) and nitrite (NasDE) reductase are highly expressed during anaerobic growth of B. subtilis, the native promoter PsrfA of the surfactin operon in strain B. subtilis JABs24 was replaced by promoters PnarG and PnasD to induce surfactin synthesis anaerobically. Shake flask cultivations with varying oxygen availabilities indicated no significant differences in native PsrfA expression. As hypothesized, activity of PnarG and PnasD increased with lower oxygen levels and surfactin was not produced by PsrfA::PnarG as well as PsrfA::PnasD mutant strains under conditions with highest oxygen availability. PnarG showed expressions similar to PsrfA at lowest oxygen availability, while maximum value of PnasD was more than 5.5-fold higher. Although the promoter exchange PsrfA::PnarG resulted in a decreased surfactin titer at lowest oxygen availability, the strain carrying PsrfA::PnasD reached a 1.4-fold increased surfactin concentration with 696 mg/L and revealed an exceptional high overall YP/X of 1.007 g/g. This value also surpassed the YP/X of the reference strain JABs24 at highest and moderate oxygen availability. Bioreactor cultivations illustrated that significant cell lysis occurred when the process of “anaerobization” was performed too fast. However, processes with a constantly low agitation and aeration rate showed promising potential for process improvement, especially by employing the strain carrying PsrfA::PnasD promoter exchange. Additionally, replacement of other native promoters by nitrite reductase promoter PnasD represents a promising tool for anaerobic-inducible bioprocesses in Bacillus.

Influence of carbon source, agitation and aeration rates for production yeast biomass which potential of use for biological control

The fermentation processes are used to promote better results when it is desired to increase the biomass of organisms that can be used in biological control. Thus, the aim of this work was to expand the scale of production of biomass of yeasts that have potential for biological control. A total of five strains of yeasts isolated from natural microflora of tropical fruits, a strain was selected and evaluated for the influence of the carbon source, cultivation orbital shaker and bioreactor. The tests performed demonstrate that the use of invert sugar as carbon source favored production of biomass material when compared to that obtained with glucose. The bioreactor cultivation of the yeast strain has enhanced performance, since it favors an increase of 24.12% (6.47 to 8.03 g.L-1) in biomass production when compared to the fermentation orbital shaker and that may be associated with increased agitation and aeration rate.

Development of a Cost Effective Medium for Enhanced Production of Bacillus thuringiensis ?-endotoxin

The present study was carried out to develop a sustainable production medium using locally available cheap raw materials for biopesticide production by Bacillus thuringiensis subsp. kurstaki (Btk) HD-73. In submerged fermentation (SmF) condition, the conventional Luria-Bertani (LB) medium which was enriched with nitrogen source (10% defatted soybean meal) supported 28.57% sporulation and 125% endotoxin increase over LB (alone). The effect of cystine on sporulation and endotoxin synthesis was highly pronounced in LB-soybean medium (LBS) with a range of 19.54% and 131.35% higher endotoxin yield respectively in SmF condition. While basal salts supplemented in soybean-cystine (SMc) medium, it resulted in 7.65% endotoxin yield compared to LB-soybean-cystine (LBSc) medium. Addition of molasses balanced the C: N ratio in the SMc medium thus helping 84.85% higher endotoxin synthesis after 24 hours fermentation. Substitution of basal salts with cost effective sea water yielded about 19.27% less endotoxin. The optimum medium thus obtained consisting of soybean extract-molasses-cystine with sea water was used in 3.0 L bioreactor cultivation for endotoxin synthesis by Btk HD-73 under 30% saturation of dO2 through cascade of agitation and aeration. The production rate obtained was 1.67 fold higher in bioreactor than in shake flask culture.Bangladesh J Microbiol, Volume 32, Number 1-2,June-Dec 2015, pp 1-6