Mutation breeding of high-stress resistant strains for succinic acid production from corn straw.

The production of succinic acid from corn stover is a promising and sustainable route; however, during the pretreatment stage, byproducts such as organic acids, furan-based compounds, and phenolic compounds generated from corn stover inhibit the microbial fermentation process. Selecting strains that are resistant to stress and utilizing nondetoxified corn stover hydrolysate as a feedstock for succinic acid production could be effective. In this study, A. succinogenes CICC11014 was selected as the original strain, and the stress-resistant strain A. succinogenes M4 was obtained by atmospheric and room temperature plasma (ARTP) mutagenesis and further screening. Compared to the original strain, A. succinogenes M4 exhibited a twofold increase in stress resistance and a 113% increase in succinic acid production when hydrolysate was used as the substrate. By conducting whole-genome resequencing of A. succinogenes M4 and comparing it with the original strain, four nonsynonymous gene mutations and two upstream regions with base losses were identified. KEY POINTS: • A high-stress-resistant strain A. succinogenes M4 was obtained by ARTP mutation •  The production of succinic acid increased by 113% • The mutated genes of A. succinogenes M4 were detected and analyzed.

Enhancing high-efficiency breeding and microbial microdroplet cultivation techniques for Ganoderma lucidum.

Ganoderma lucidum is known for its bioactive compounds, such as polysaccharides and triterpenoids, which are crucial in food and medicine. However, liquid fermentation encounters challenges in terms of strain differentiation and stability. In this research, we employed atmospheric room temperature plasma mutation and a microbial microdroplet culture system to identify strains with enhanced biomass and triterpenoid production. The three mutant strains, YB05, YB09, and YB18, exhibited accelerated growth rates and antagonized the initial strain G0023 more effectively than the controls. Notably, YB18 displayed the fastest growth, with a 17.25% increase in colony radius. Shake flask cultivation demonstrated that, compared with the initial strain, YB05 and YB18 had 26.33% and 17.85% greater biomass, respectively. Moreover, the triterpenoid production of YB05 and YB18 surpassed that of the control by 32.10% and 15.72%, respectively, as confirmed by colorimetric detection. Importantly, these mutant strains remained stable for five generations. This study revealed a comprehensive screening system utilizing atmospheric pressure, room temperature plasma mutation technology and microbial droplet cultivation. This innovative approach offers a promising pathway for obtaining advantageous Ganoderma strains for liquid fermentation. The methodology of atmospheric room temperature plasma mutation and microbial microdroplet culture systems is detailed for better comprehension.

Enhancing (2S)-naringenin production in Saccharomyces cerevisiae by high-throughput screening method based on ARTP mutagenesis.

(2S)-Naringenin, a dihydro-flavonoid, serves as a crucial precursor for flavonoid synthesis due to its extensive medicinal values and physiological functions. A pathway for the synthesis of (2S)-naringenin from glucose has previously been constructed in Saccharomyces cerevisiae through metabolic engineering. However, this synthetic pathway of (2S)-naringenin is lengthy, and the genes involved in the competitive pathway remain unknown, posing challenges in significantly enhancing (2S)-naringenin production through metabolic modification. To address this issue, a novel high-throughput screening (HTS) method based on color reaction combined with a random mutagenesis method called atmospheric room temperature plasma (ARTP), was established in this study. Through this approach, a mutant (B7-D9) with a higher titer of (2S)-naringenin was obtained from 9600 mutants. Notably, the titer was enhanced by 52.3% and 19.8% in shake flask and 5 L bioreactor respectively. This study demonstrates the successful establishment of an efficient HTS method that can be applied to screen for high-titer producers of (2S)-naringenin, thereby greatly improving screening efficiency and providing new insights and solutions for similar product screenings.

Mutation breeding of Aspergillus niger by atmospheric room temperature plasma to enhance phosphorus solubilization ability.

Background: Phosphorus (P) is abundant in soils, including organic and inorganic forms. Nevertheless, most of P compounds cannot be absorbed and used by plants. Aspergillus niger v. Tiegh is a strain that can efficiently degrade P compounds in soils. Methods: In this study, A. niger xj strain was mutated using Atmospheric Room Temperature Plasma (ARTP) technology and the strains were screened by Mo-Sb Colorimetry with strong P-solubilizing abilities. Results: Compared with the A. niger xj strain, setting the treatment time of mutagenesis to 120 s, four positive mutant strains marked as xj 90-32, xj120-12, xj120-31, and xj180-22 had higher P-solubilizing rates by 50.3%, 57.5%, 55.9%, and 61.4%, respectively. Among them, the xj120-12 is a highly efficient P solubilizing and growth-promoting strain with good application prospects. The growth characteristics such as plant height, root length, and dry and fresh biomass of peanut (Arachis hypogaea L.) increased by 33.5%, 43.8%, 43.4%, and 33.6%, respectively. Besides available P, the chlorophyll and soluble protein contents also vary degrees of increase in the P-solubilizing mutant strains. Conclusions: The results showed that the ARTP mutagenesis technology can improve the P solubilization abilities of the A. niger mutant strains and make the biomass of peanut plants was enhanced of mutant strains.

Rapid improvement in the macrolactins production of Bacillus sp. combining atmospheric room temperature plasma with the specific growth rate index.

Macrolactins (MLNs) have attracted considerable attention due to their antibacterial and antiviral properties. Here, the MLN production of Bacillus sp. strain IMDGX0108 was improved using a breeding strategy of atmospheric room temperature plasma (ARTP) technique. Combining with a selection procedure based on the colony morphology and specific growth rate index (SGRI), two genetically stable mutants A29 and A72 were identified. The MLN production of A29 and A72 was 35.2% and 52.8% greater than that of the parent strain, respectively. The best-performing mutant A72 was subjected to RNA-sequence analysis. Five pathways were significantly enriched, and fatty acid bioprocesses might play an important role in improving the production of MLNs. The combined strategy developed herein (i.e., ARTP mutation plus an efficient screening procedure) might be an appropriate method by which to obtain strains overproducing MLNs.