Designing glucose utilization "highway" for recombinant biosynthesis.

cAMP receptor protein (CRP) is known as a global regulatory factor mainly mediating carbon source catabolism. Herein, we successfully engineered CRP to develop microbial chassis cells with improved recombinant biosynthetic capability in minimal medium with glucose as single carbon source. The obtained best-performing cAMP-independent CRPmu9 mutant conferred both faster cell growth and a 133-fold improvement in expression level of lac promoter in presence of 2% glucose, compared with strain under regulation of CRPwild-type. Promoters free from "glucose repression" are advantageous for recombinant expression, as glucose is a frequently used inexpensive carbon source in high-cell-density fermentations. Transcriptome analysis demonstrated that the CRP mutant globally rewired cell metabolism, displaying elevated tricarboxylic acid cycle activity; reduced acetate formation; increased nucleotide biosynthesis; and improved ATP synthesis, tolerance, and stress-resistance activity. Metabolites analysis confirmed the enhancement of glucose utilization with the upregulation of glycolysis and glyoxylate-tricarboxylic acid cycle. As expected, an elevated biosynthetic capability was demonstrated with vanillin, naringenin and caffeic acid biosynthesis in strains regulated by CRPmu9. This study has expanded the significance of CRP optimization into glucose utilization and recombinant biosynthesis, beyond the conventionally designated carbon source utilization other than glucose. The Escherichiacoli cell regulated by CRPmu9 can be potentially used as a beneficial chassis for recombinant biosynthesis.

Robust Carbon Nitride Homojunction Photoelectrode for Solar-Driven Water Splitting.

Achieving intimate particle-to-particle and particle-to-substrate contacts is the first priority for fabricating high-quality photoelectrodes to ensure sufficient visible light absorption and efficient charge separation/transport. To achieve this goal, a seeding strategy is designed to construct a robust carbon nitride (CN) homojunction photoelectrode, in which vaporized precursors are condensed into a compact seeding layer at low temperatures, inducing the further deposition of the top layer. This optimized photoelectrode displays an excellent photocurrent density of 320 µA cm-2 in 0.1 M NaOH electrolyte at 1.23 VRHE (V vs reversible hydrogen electrode) under AM 1.5G illumination, with H2 and O2 evolution rates of 2.98 and 1.47 µmol h-1 cm-2, respectively. Characterizations show that both the robust contact and the homojunction of the double-layered CN film contribute to enhanced photoelectrochemical performance. This work may provide a new strategy for the design of high-performing CN photoelectrodes.

Engineering an SspB-mediated degron for novel controllable protein degradation.

Elegant controllable protein degradation tools have great applications in metabolic engineering and synthetic biology designs. SspB-mediated ClpXP proteolysis system is well characterized, and SspB acts as an adaptor tethering ssrA-tagged substrates to the ClpXP protease. This degron was applied in metabolism optimization, but the efficiency was barely satisfactory. Limited high-quality tools are available for controllable protein degradation. By coupling structure-guided modeling and directed evolution, we establish state-of-the-art high-throughput screening strategies for engineering both degradation efficiency and SspB-ssrA binding specificity of this degron. The reliability of our approach is confirmed by functional validation of both SspB and ssrA mutants using fluorescence assays and metabolic engineering of itaconic acid or ferulic acid biosynthesis. Isothermal titration calorimetry analysis and molecular modeling revealed that an appropriate instead of excessively strong interaction between SspB and ssrA benefited degradation efficiency. Mutated SspB-ssrA pairs with 7-22-fold higher binding KD than the wild-type pair led to higher degradation efficiency, revealing the advantage of directed evolution over rational design in degradation efficiency optimization. Furthermore, an artificial SspB-ssrA pair exhibiting low crosstalk of interactions with the wild-type SspB-ssrA pair was also developed. Efforts in this study have demonstrated the plasticity of SspB-ssrA binding pocket for designing high-quality controllable protein degradation tools. The obtained mutated degrons enriched the tool box of metabolic engineering designs.

Levosimendan versus dobutamine for sepsis-induced cardiac dysfunction: a systematic review and meta-analysis.

Levosimendan and dobutamine are extensively used to treat sepsis-associated cardiovascular failure in ICU. Nevertheless, the role and mechanism of levosimendan in patients with sepsis-induced cardiomyopathy remains unclear. Moreover, previous studies on whether levosimendan is superior to dobutamine are still controversial. More importantly, these studies did not take changes (before-after comparison to the baseline) in quantitative parameters such as ejection fraction into account with the baseline level. Here, we aimed to determine the pros and cons of the two medicines by assessing the changes in cardiac function and blood lactate, mortality, with the standardized mean difference used as a summary statistic. Relevant studies were obtained by a thorough and disciplined literature search in several notable academic databases, including Google Scholar, PubMed, Cochrane Library and Embase until November 2020. Outcomes included changes in cardiac function, lactic acid, mortality and length of hospital stay. A total of 6 randomized controlled trials were included in this study, including 192 patients. Compared with dobutamine, patients treated with levosimendan had a greater improvement of cardiac index (ΔCI) (random effects, SMD = 0.90 [0.20,1.60]; I2 = 76%, P < 0.01) and left ventricular stroke work index (ΔLVSWI) (random effects, SMD = 1.56 [0.90,2.21]; I2 = 65%, P = 0.04), a significant decrease of blood lactate (Δblood lactate) (random effects, MD = - 0.79 [- 1.33, - 0.25]; I2 = 68%, P < 0.01) at 24-h after drug intervention, respectively. There was no significant difference between levosimendan and dobutamine on all-cause mortality in ICU (fixed effect, OR = 0.72 [0.39,1.33]; I2 = 0%, P = 0.99). We combine effect sizes related to different measurement parameters to evaluate cardiac function, which implied that septic patients with myocardial dysfunction might have a better improvement of cardiac function by levosimendan than dobutamine (random effects, SMD = 1.05 [0.69,1.41]; I2 = 67%, P < 0.01). This study suggested a significant improvement of CI, LVSWI, and decrease of blood lactate in septic patients with myocardial dysfunction in ICU after 24-h administration of levosimendan than dobutamine. However, the administration of levosimendan has neither an impact on mortality nor LVEF. Septic patients with myocardial dysfunction may partly benefit from levosimendan than dobutamine, mainly embodied in cardiac function improvement.

Evaluation on the effect of acupuncture on patients with sepsis-induced myopathy (ACU-SIM pilot study): A single center, propensity-score stratified, assessor-blinded, prospective pragmatic controlled trial.

BACKGROUND: Sepsis-induced myopathy (SIM) is a disease that causes motor dysfunction in patients with sepsis. There is currently no targeted treatment for this disease. Acupuncture has shown considerable efficacy in the treatment of sepsis and muscle weakness. Therefore, our research aims to explore the effects of acupuncture on the improvement of muscle structure and function in SIM patients and on activities of daily living. METHODS: The ACU-SIM pilot study is a single-center, propensity-score stratified, assessor-blinded, prospective pragmatic controlled trial (pCT) with a 1-year follow-up period. This study will be deployed in a multi-professional critical care department at a tertiary teaching hospital in Guangzhou, China. Ninety-eight intensive care unit subjects will be recruited and assigned to either the control group or the acupuncture group. Both groups will receive basic treatment for sepsis, and the acupuncture group will additionally receive acupuncture treatment. The primary outcomes will be the rectus femoris cross-sectional area, the Medical Research Council sum-score and time-to-event (defined as all-cause mortality or unplanned readmission to the intensive care unit due to invasive ventilation). The activities of daily living will be accessed by the motor item of the Functional Independence Measure. Recruitment will last for 2 years, and each patient will have a 1-year follow-up after the intervention. DISCUSSION: There is currently no research on the therapeutic effects of acupuncture on SIM. The results of this study may contribute to new knowledge regarding early muscle atrophy and the treatment effect of acupuncture in SIM patients, and the results may also direct new approaches and interventions in these patients. This trial will serve as a pilot study for an upcoming multicenter real-world study. TRIAL REGISTRATION: Chinese Clinical Trials Registry: ChiCTR-1900026308, registered on September 29th, 2019.

Dynamic Effects of Initial pH of Substrate on Biological Growth and Metamorphosis of Black Soldier Fly (Diptera: Stratiomyidae).

Edible insects have become a recognized alternative and sustainable source of high-quality proteins and fats for livestock or human consumption. In the production process of black soldier fly (BSF), (Hermetia illucens L. [Diptera: Stratiomyidae]), initial substrate pH is a critical parameter to ensure the best value of insect biomass, life history traits, and quality bio-fertilizer. This study examined the impact of initial pH values on BSF larvae production, development time, and adult longevity. The BSF were reared on artificial diet with initial pH of 2.0, 4.0, 6.0, 8.0, and 10.0; the control was set at 7.0. Final BSF larval weight was significantly greater in substrates having initial pH 6.0 (0.21 g), control 7.0 (0.20 g), and 10.0 (0.20 g) with no significant difference among them, whereas larval weight reared with initial pH 2.0 and 4.0 were lowest at 0.16 g (-23%). Prepupal weight was greatest when larvae were reared on substrates with initial pH 6.0 (0.18 g), control 7.0 (0.19 g), 8.0 (0.18 g), and 10.0 (0.18 g). In contrast, the prepupal weight of larvae reared on diets with initial pH 2.0 was lowest at 0.15 g (-22%). Larval development time was 21.19 d at pH 8.0, about 3 d (12.5%) shorter than that of those reared on diets with initial pH 6.0, 7.0 control, and 10.0. In all treatments, pH shifted to 5.7 after 3-4 d and 8.5 after 16-17 d except for two groups (2.0 and 4.0) where the pH remained slightly acidic 5.0 and 6.5, respectively.

Tuning to optimize SVM approach for assisting ovarian cancer diagnosis with photoacoustic imaging.

Support vector machine (SVM) is one of the most effective classification methods for cancer detection. The efficiency and quality of a SVM classifier depends strongly on several important features and a set of proper parameters. Here, a series of classification analyses, with one set of photoacoustic data from ovarian tissues ex vivo and a widely used breast cancer dataset- the Wisconsin Diagnostic Breast Cancer (WDBC), revealed the different accuracy of a SVM classification in terms of the number of features used and the parameters selected. A pattern recognition system is proposed by means of SVM-Recursive Feature Elimination (RFE) with the Radial Basis Function (RBF) kernel. To improve the effectiveness and robustness of the system, an optimized tuning ensemble algorithm called as SVM-RFE(C) with correlation filter was implemented to quantify feature and parameter information based on cross validation. The proposed algorithm is first demonstrated outperforming SVM-RFE on WDBC. Then the best accuracy of 94.643% and sensitivity of 94.595% were achieved when using SVM-RFE(C) to test 57 new PAT data from 19 patients. The experiment results show that the classifier constructed with SVM-RFE(C) algorithm is able to learn additional information from new data and has significant potential in ovarian cancer diagnosis.