Adaptive laboratory evolution of Bacillus subtilis to overcome toxicity of lignocellulosic hydrolysate derived from Distiller's dried grains with solubles (DDGS)

Driessen, Jasper L.S.P.; Johnsen, Josefin; Pogrebnyakov, Ivan; Mohamed, Elsayed Tharwat Tolba; Mussatto, Solange I.; Feist, Adam M.; Jensen, Sheila Ingemann; Nielsen, Alex Toftgaard

doi:10.1016/j.mec.2023.e00223

Cited by 5 publications

(8 citation statements)

References 40 publications

(52 reference statements)

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“…In our pursuit to harness this adaptability for biotechnological applications, we employed ALE to improve the galactose utilization capability of B. subtilis. There were several trials to engineer B. subtilis with ALE (Driessen et al 2023;Li et al 2023;Zeigler and Nicholson 2017). Bo Zhang et al have reported that B. subtilis was adapted under xylose as a selection pressure and validated several key mutations (Zhang et al 2015).…”

Section: Discussionmentioning

confidence: 99%

“…Uncovering significant mutations that enhance cell growth under selective pressure is a pivotal aspect of ALE (Driessen et al 2023;Kang et al 2019;Kim et al 2022). The technique of reverse engineering is employed to discern causative mutations from incidental ones.…”

Section: Genomic Resequencing Of Strains Bsga14mentioning

confidence: 99%

“…Notable studies have explored growth under glucose as the primary carbon source (Liu et al 2020;Yuan et al 2019), while others have delved into growth on xylose as the sole carbon substrate (Zhang et al 2015). Additionally, investigations have been carried out with NH4Cl serving as the exclusive nitrogen source (Li et al 2023), and within the milieu of lignocellulosic hydrolysate (Driessen et al 2023). Through these varied explorations, ALE has emerged as a powerful tool in fine-tuning the metabolic and growth attributes of B. subtilis to adapt to distinct nutrient and environmental regimes.…”

Section: Introductionmentioning

confidence: 99%

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Engineering Bacillus subtilis J46 for efficient utilization of galactose through adaptive laboratory evolution

Choi,

Song,

Hong

et al. 2023

Preprint

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Efficient utilization of galactose by microorganisms can lead to the production of valuable bio-products and improved metabolic processes. While Bacillus subtilis has inherent pathways for galactose metabolism, there is potential for enhancement via evolutionary strategies. This study aimed to boost galactose utilization in B. subtilis using adaptive laboratory evolution (ALE) and to elucidate the genetic and metabolic changes underlying the observed enhancements. B. subtilis strains were subjected to successive rounds (~ 5000 generations) of ALE under conditions promoting galactose utilization with a higher specific growth rate of 0.319 h-1 compared to wild type (0.03 h-1). Resultant evolved strains, BSGA14 was analyzed for their galactose conversion rates. After genome resequencing, 63 SNPs were identified in BSGA14 strain. Two of them, located in the coding sequences of the genes araR and glcR, were found to be the advantageous mutations after reverse engineering. The strain with these two accumulated mutations, BSGALE4, exhibited similar specific growth rate on galactose to the evolved strain BSGA14 (0.296 h-1). Furthermore, evolved strain showed higher productivity of protease and β-galactosidase in mock soybean biomass medium. ALE proved to be a potent tool for enhancing galactose metabolism in B. subtilis. The findings offer valuable insights into the potential of evolutionary strategies in microbial engineering and pave the way for industrial applications harnessing enhanced galactose conversion.

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Section: Discussionmentioning

confidence: 99%

Section: Genomic Resequencing Of Strains Bsga14mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Engineering Bacillus subtilis J46 for efficient utilization of galactose through adaptive laboratory evolution

Choi,

Song,

Hong

et al. 2023

Preprint

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“…This versatile approach finds wide applicability across various microorganisms that can be cultured in a laboratory setting. Notably, ALE has been effective for optimizing Bacillus species specifically in the past for enhancing tolerance towards inhibitors in hydrolyzed biomass 33 , as well as stress tolerance to low pressures 34 , 35 , and establishing a non-native pathway 36 . Thus, ALE is a well-suited strategy to augment the heat tolerance of Bacillus spores 37 , 38 , thereby enabling their use as a functional living additive material.…”

Section: Introductionmentioning

confidence: 99%

Biocomposite thermoplastic polyurethanes containing evolved bacterial spores as living fillers to facilitate polymer disintegration

Kim,

Noh,

White

et al. 2024

Nat Commun

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The field of hybrid engineered living materials seeks to pair living organisms with synthetic materials to generate biocomposite materials with augmented function since living systems can provide highly-programmable and complex behavior. Engineered living materials have typically been fabricated using techniques in benign aqueous environments, limiting their application. In this work, biocomposite fabrication is demonstrated in which spores from polymer-degrading bacteria are incorporated into a thermoplastic polyurethane using high-temperature melt extrusion. Bacteria are engineered using adaptive laboratory evolution to improve their heat tolerance to ensure nearly complete cell survivability during manufacturing at 135 °C. Furthermore, the overall tensile properties of spore-filled thermoplastic polyurethanes are substantially improved, resulting in a significant improvement in toughness. The biocomposites facilitate disintegration in compost in the absence of a microbe-rich environment. Finally, embedded spores demonstrate a rationally programmed function, expressing green fluorescent protein. This research provides a scalable method to fabricate advanced biocomposite materials in industrially-compatible processes.

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“…Notable studies have explored growth under glucose as the primary carbon source (Liu et al 2020 ; Yuan et al 2019 ), while others have delved into growth on xylose as the sole carbon substrate (Averesch and Rothschild 2019 ; Zhang et al 2015 ). Additionally, investigations have been carried out with NH 4 Cl serving as the exclusive nitrogen source (Li et al 2023 ), and within the milieu of lignocellulosic hydrolysate (Driessen et al 2023 ). Through these varied explorations, ALE has emerged as a powerful tool in fine-tuning the metabolic and growth attributes of B. subtilis to adapt to distinct nutrient and environmental regimes.…”

Section: Introductionmentioning

confidence: 99%

Engineering Bacillus subtilis J46 for efficient utilization of galactose through adaptive laboratory evolution

Choi,

Song,

Hong

et al. 2024

AMB Expr

View full text Add to dashboard Cite

Efficient utilization of galactose by microorganisms can lead to the production of valuable bio-products and improved metabolic processes. While Bacillus subtilis has inherent pathways for galactose metabolism, there is potential for enhancement via evolutionary strategies. This study aimed to boost galactose utilization in B. subtilis using adaptive laboratory evolution (ALE) and to elucidate the genetic and metabolic changes underlying the observed enhancements. The strains of B. subtilis underwent multiple rounds of adaptive laboratory evolution (approximately 5000 generations) in an environment that favored the use of galactose. This process resulted in an enhanced specific growth rate of 0.319 ± 0.005 h−1, a significant increase from the 0.03 ± 0.008 h−1 observed in the wild-type strains. Upon selecting the evolved strain BSGA14, a comprehensive whole-genome sequencing revealed the presence of 63 single nucleotide polymorphisms (SNPs). Two of them, located in the coding sequences of the genes araR and glcR, were found to be the advantageous mutations after reverse engineering. The strain with these two accumulated mutations, BSGALE4, exhibited similar specific growth rate on galactose to the evolved strain BSGA14 (0.296 ± 0.01 h−1). Furthermore, evolved strain showed higher productivity of protease and β-galactosidase in mock soybean biomass medium. ALE proved to be a potent tool for enhancing galactose metabolism in B. subtilis. The findings offer valuable insights into the potential of evolutionary strategies in microbial engineering and pave the way for industrial applications harnessing enhanced galactose conversion.

show abstract

Adaptive laboratory evolution of Bacillus subtilis to overcome toxicity of lignocellulosic hydrolysate derived from Distiller's dried grains with solubles (DDGS)

Cited by 5 publications

References 40 publications

Engineering Bacillus subtilis J46 for efficient utilization of galactose through adaptive laboratory evolution

Engineering Bacillus subtilis J46 for efficient utilization of galactose through adaptive laboratory evolution

Biocomposite thermoplastic polyurethanes containing evolved bacterial spores as living fillers to facilitate polymer disintegration

Engineering Bacillus subtilis J46 for efficient utilization of galactose through adaptive laboratory evolution

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