Synergistic application of atmospheric and room temperature plasma mutagenesis and adaptive laboratory evolution improves the tolerance of Escherichia coli to L-cysteine.

L-Cysteine production through fermentation stands as a promising technology. However, excessive accumulation of L-cysteine poses a challenge due to the potential to inflict damage on cellular DNA. In this study, we employed a synergistic approach encompassing atmospheric and room temperature plasma mutagenesis (ARTP) and adaptive laboratory evolution (ALE) to improve L-cysteine tolerance in Escherichia coli. ARTP-treated populations obtained substantial enhancement in L-cysteine tolerance by ALE. Whole-genome sequencing, transcription analysis, and reverse engineering, revealed the pivotal role of an effective export mechanism mediated by gene eamB in augmenting L-cysteine resistance. The isolated tolerant strain, 60AP03/pTrc-cysEf , achieved a 2.2-fold increase in L-cysteine titer by overexpressing the critical gene cysEf during batch fermentation, underscoring its enormous potential for L-cysteine production. The production evaluations, supplemented with L-serine, further demonstrated the stability and superiority of tolerant strains in L-cysteine production. Overall, our work highlighted the substantial impact of the combined ARTP and ALE strategy in increasing the tolerance of E. coli to L-cysteine, providing valuable insights into improving L-cysteine overproduction, and further emphasized the potential of biotechnology in industrial production.

Sperm retrieval rate and patient factors in azoospermia factor c microdeletion azoospermia: a systematic review.

OBJECTIVE: To reveal the overall sperm retrieval rate (SRR) and range in patients with azoospermia factor c (AZFc) microdeletion azoospermia by microdissection testicular sperm extraction (mTESE) and discuss the differences of preoperative patient factors among studies with various SRRs. PATIENTS AND METHODS: We searched PubMed, Web of Science and Embase until February 2023. All studies reporting SRRs by mTESE and required parameters of patients with AZFc microdeletions were included. The primary outcome was the SRR and, if available, the pregnancy rate (PR) and live-birth rate (LBR) after intracytoplasmic sperm injection were also investigated as secondary outcomes. RESULTS: Eventually 11 cohort studies were included in this review. A total number of 441 patients underwent mTESE and in 275 of them sperm was obtained, reaching an overall SRR of 62.4%. The SRRs among studies had a wide range from 25.0% to 85.7%. The studies reporting higher SRRs generally had older mean ages, and higher follicle-stimulating hormone and testosterone levels. Only four studies provided practical data on pregnancies and live-born children of patients with AZFc microdeletions, so the overall PR and LBR were unavailable. CONCLUSIONS: The overall SRR of patients with AZFc microdeletion azoospermia was 62.4%. The effect of patient factors in SR needs further evidence in future work.

Combinatorial Metabolic Engineering of Escherichia coli for Enhanced L-Cysteine Production: Insights into Crucial Regulatory Modes and Optimization of Carbon-Sulfur Metabolism and Cofactor Availability.

Microbial production of valuable compounds can be enhanced by various metabolic strategies. This study proposed combinatorial metabolic engineering to develop an effective Escherichia coli cell factory dedicated to L-cysteine production. First, the crucial regulatory modes that control L-cysteine levels were investigated to guide metabolic modifications. A two-stage fermentation was achieved by employing multi-copy gene expression, improving the balance between production and growth. Subsequently, carbon flux distribution was further optimized by modifying the C1 unit metabolism and the glycolytic pathway. The modifications of sulfur assimilation demonstrated superior performance of thiosulfate utilization pathways in enhancing L-cysteine titer. Furthermore, the studies focusing on cofactor availability and preference emphasized the vital role of synergistic enhancement of sulfur-carbon metabolism in L-cysteine overproduction. In a 5 L bioreactor, the strain BW15-3/pED accumulated 12.6 g/L of L-cysteine. This work presented an effective metabolic engineering strategy for the development of L-cysteine-producing strains.

Plant growth-promoting rhizobacteria are important contributors to rice yield in karst soils.

The difficulty of releasing nutrients from soils in karst areas limits the yield of local crops and leads to poverty. In this study, two strains of plant growth-promoting rhizobacteria (PGPR) were isolated from the rhizosphere soil of typical plants in karst areas, which were both identified as Bacillus sp. and named GS1 and N1. And two isolates were used to construct a composite PGPR named MC1. These three strains of PGPR were used for soil inoculation in the pot experiment and field trial and their capacity to promote rice development was assessed. The results showed that MC1 inoculation exhibited notable rice growth-promoting ability in pot experiments, and, respectively, had an increment of 16.96, 18.74, and 11.50% in shoot biomass, total biomass, and rice height compared with control. This is largely attributed to PGPR's capacity to secrete phytohormones and soil enzymes, particularly urease (UE) in GS1, whose secreted UE content was significantly higher by 12.18% compared to the control. When applied to the field, MC1 inoculation not only increased rice yield by 8.52% and the available nutrient content in rice rhizosphere soil, such as available phosphorus (AP) and exchangeable magnesium (EMg); but also improved the abundance of beneficial rhizobacteria and the diversity of microbial communities in rice rhizosphere soil. Results in this study revealed that inoculated PGPR played a major role in promoting rice growth and development, and a new strategy for facilitating the growth of rice crops in agriculture was elucidated. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03593-0.

Spatiotemporal Gene Expression by a Genetic Circuit for Chemical Production in Escherichia coli.

Gene expression in spatiotemporal distribution improves the ability of cells to respond to changing environments. For microbial cell factories in artificial environments, reconstruction of the target compound's biosynthetic pathway in a new spatiotemporal dimension/scale promotes the production of chemicals. Here, a genetic circuit based on the Esa quorum sensing and lac operon was designed to achieve the dynamic temporal gene expression. Meanwhile, the pathway was regulated by an l-cysteine-specific sensor and relocalized to the plasma membrane for further flux enhancement to l-cysteine and toxicity reduction on a spatial scale. Finally, the integrated spatiotemporal regulation circuit for l-cysteine biosynthesis enabled a 14.16 g/L l-cysteine yield in Escherichia coli. Furthermore, this spatiotemporal regulation circuit was also applied in our previously constructed engineered strain for pantothenic acid, methionine, homoserine, and 2-aminobutyric acid production, and the titer increased by 29, 33, 28, and 41%, respectively. These results highlighted the applicability of our spatiotemporal regulation circuit to enhance the performance of microbial cell factories.

Clostridium weizhouense sp. nov., an anaerobic bacterium isolated from activated sludge of petroleum wastewater.

A Gram-stain-positive, anaerobic, spore-forming, rod-shaped (0.4-0.6 µm×2.5-3.2 µm), flagellated bacterium, designated strain YB-6T, was isolated from activated sludge of an anaerobic tank at Weizhou marine oil mining wastewater treatment plant in Beihai, Guangxi, PR China. The culture conditions were 25-45 °C (optimum, 37 °C), pH 4-12 (pH 7.0) and NaCl concentration of 0-7 % w/v (0%). Strain YB-6T grew slowly in petroleum wastewater and removed 68.2 % of the total organic carbon in petroleum wastewater within 10 days. Concentrations of naphthalene, anthracene and phenanthrene at an initial concentration of 50 mg l-1 were reduced by 32.8, 40.4 and 14.6 %, respectively, after 7 days. Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain YB-6T belongs to Clostridium cluster I and is most closely related to Clostridium uliginosum CK55T (98.5 % similarity). The genome size of strain YB-6T was 3.96 Mb, and the G+C content was 26.5 mol%. The average nucleotide identity value between strain YB-6T and C. uliginosum CK55T was 81.9 %. The major fatty acids in strain YB-6T were C14 : 0 FAME, C16 : 0 FAME and summed feature 4 (unknown 14.762 and/or C15 : 2 FAME). The main polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, five unidentified aminophospholipids, one unidentified glycolipid and one unidentified aminolipid. Diaminopimelic acid was not detected in the strain YB-6T cell walls. Whole-cell sugars mainly consisted of ribose and galactose. Based on the results of phenotypic and genotypic analyses, strain YB-6T represents a novel species of the genus Clostridium, for which the name Clostridium weizhouense sp. nov. is proposed. The type strain is YB-6T (=GDMCC 1.2529T=JCM 34754T).