A cell-free transcription-translation pipeline for recreating methylation patterns boosts DNA transformation in bacteria.

The bacterial world offers diverse strains for understanding medical and environmental processes and for engineering synthetic biological chassis. However, genetically manipulating these strains has faced a long-standing bottleneck: how to efficiently transform DNA. Here, we report imitating methylation patterns rapidly in TXTL (IMPRINT), a generalized, rapid, and scalable approach based on cell-free transcription-translation (TXTL) to overcome DNA restriction, a prominent barrier to transformation. IMPRINT utilizes TXTL to express DNA methyltransferases from a bacterium's restriction-modification systems. The expressed methyltransferases then methylate DNA in vitro to match the bacterium's DNA methylation pattern, circumventing restriction and enhancing transformation. With IMPRINT, we efficiently multiplex methylation by diverse DNA methyltransferases and enhance plasmid transformation in gram-negative and gram-positive bacteria. We also develop a high-throughput pipeline that identifies the most consequential methyltransferases, and we apply IMPRINT to screen a ribosome-binding site library in a hard-to-transform Bifidobacterium. Overall, IMPRINT can enhance DNA transformation, enabling the use of sophisticated genetic manipulation tools across the bacterial world.

[The expression and secretion of human lactoferrin in Bacillus subtilis].

Human lactoferrin (HLF), an essential nutrient found in breast milk, possesses antibacterial, anti-inflammatory, and immune-enhancing properties. In this study, the effects of three constitutive promoters (P21, P43, and Pveg) and three inducible promoters (Pgrac100, PxylA, and Ptet*) on the expression of HLF were compared using Bacillus subtilis G601 as the host strain. The results showed that the highest expression of HLF, reaching 651.57 µg/L, was achieved when regulated by the Ptet* promoter. Furthermore, the combinational optimization of ribosome binding site (RBS) and signal peptides was investigated, and the optimal combination of RBS6 and SPyycP resulted in increased HLF expression to 1 099.87 µg/L, with 498.68 µg/L being secreted extracellularly. To further enhance HLF secretion, the metal cations-related gene dltD was knocked out, leading to an extracellular HLF level of 637.28 µg/L. This study successfully demonstrated the secretory expression of HLF in B. subtilis through the selection and optimization of expression elements, laying the foundation for the development of efficient B. subtilis cell factories for lactoprotein synthesis.

Association of Glycated Hemoglobin With Acute Ischemic Stroke in a Tertiary Care Center in a Tribal Region of Jharkhand.

BACKGROUND: Increased glycated hemoglobin (HbA1c) levels have shown an association with an increased risk of stroke in patients admitted to a tertiary care center in Jharkhand. OBJECTIVES: To find out and estimate the risk of acute ischemic stroke in patients with increased HbA1c levels compared with controls. METHODS: This observational case-control study was conducted on patients admitted to the department of general medicine at a tertiary care center in Ranchi from June 2021 to November 2022. The patients included in this study were those aged 18 years or older and who were clinically and radiologically diagnosed with acute ischemic stroke. Only patients with a first episode of stroke were included, and patients with hemorrhagic stroke or transient ischemic attack were excluded from this study. An equal number of control participants were also included. Ion exchange high-performance liquid chromatography was used to perform the HbA1c tests. The same method was used to measure HbA1c levels in the controls. All findings were recorded in a Microsoft Excel sheet (Microsoft Corporation, Redmond, WA), and the data were analyzed using SPSS version 22.0 software (IBM Corp., Armonk, NY). After performing a descriptive statistical analysis, the findings were classified over a range of values and described accordingly. For each variable, an independent t-test was performed to compare the cases with the controls. A multivariable logistic regression analysis was used to choose the appropriate potential factors to determine the association in the multivariable analysis. RESULTS: A total of 185 cases and 185 controls were included. The mean age of the cases with ischemic stroke was 63.77 ± 10.312, and that of the controls was 53.18 ± 11.35 (p < 0.01). The mean HbA1c level in the patients of acute ischemic stroke was 6.97 ± 1.84, and that of the controls was 5.99 ± 1.69 (p < 0.01). The mean random blood sugar (RBS) value in the ischemic stroke cases was 170.21 ± 84.16, and that of the controls was 150.03 ± 82.25 (p = 0.02). To compare the factors that were determined to be statistically significant between ischemic stroke cases and controls, a multivariable logistic regression analysis was performed. The HbA1c p-value was 0.01, the odds ratio (OR) was 1.280, and the 95% CI was 1.11-1.48. The other variables apart from HbA1c that were statistically significant between the ischemic stroke cases and the controls were age (p < 0.01, OR: 1.280, 95% CI: 1.06-1.11), hypertension (p = 0.618, OR: 1.130, 95% CI: 0.70-1.83), and high-density lipoprotein (p = 0.055, OR: 0.975, 95% CI: 0.95-1.00). When other cofounders were considered, it was concluded that with a 1% increase in HbA1c, the risk of stroke increases by 28% (OR: 1.28, 95% CI: 1.11-1.48). To compare the variables that were determined to be statistically significant between the control and ischemic stroke case groups, a multivariable logistic regression was used. The area under the receiver operating characteristic curve for HbA1c was 0.773 and RBS was 0.600. CONCLUSION: This study shows that higher HbA1c levels in patients increase the risk of ischemic stroke. This study brings to light the need to screen the population periodically for diabetes by routinely testing for Hba1c in those who are at high risk of diabetes. Stroke risk can be reduced with early management and intervention. This study also concludes that HbA1c is a better predictor for assessing the risk of ischemic stroke than RBS levels.

Cross validation of the response bias scale and the response bias scale-19 in active-duty personnel: use on the MMPI-2-RF and MMPI-3.

The Response Bias Scale (RBS) is the central measure of cognitive over-reporting in the MMPI-family of instruments. Relative to other clinical populations, the research evaluating the detection of over-reporting is more limited in Veteran and Active-Duty personnel, which has produced some psychometric variability across studies. Some have suggested that the original scale construction methods resulted in items which negatively impact classification accuracy and in response crafted an abbreviated version of the RBS (RBS-19; Ratcliffe et al., 2022; Spencer et al., 2022). In addition, the most recent edition of the MMPI is based on new normative data, which impacts the ability to use existing literature to determine effective cut-scores for the RBS (despite all items having been retained across MMPI versions). To date, no published research exists for the MMPI-3 RBS. The current study examined the utility of the RBS and the RBS-19 in a sample of Active-Duty personnel (n = 186) referred for neuropsychological evaluation. Using performance validity tests as the study criterion, we found that the RBS-19 was generally equitably to RBS in classification. Correlations with other MMPI-2-RF over- and under-reporting symptom validity tests were slightly stronger for RBS-19. Implications and directions for research and practice with RBS/RBS-19 are discussed, along with implications for neuropsychological assessment and response validity theory.

The Impact of Clinical Factors and SARS-CoV-2 Variants on Antibody Production in Vaccinated German Healthcare Professionals Infected Either with the Delta or the Omicron Variant.

BACKGROUND: The aim of the rapid introduction of vaccines during the COVID-19 pandemic was a reduction in SARS-CoV-2 transmission and a less frequent occurrence of severe COVID-19 courses. Thus, we evaluated COVID-19 severity in vaccinated individuals to examine variant-specific symptom characteristics and their clinical impact on the serological immune response. METHODS: A total of 185 individuals previously vaccinated against and infected with the SARS-CoV-2 Delta (B.1.617.2) or Omicron (BA.4 and BA.5) variant, were enrolled for anti-SARS-CoV-2 anti-N- and anti-RBD/S1-Ig level detection. A structured survey regarding medical history was conducted. RESULTS: In 99.5 percent of cases, outpatient treatment was satisfactory. Specific symptoms associated with variants included ageusia and anosmia in patients with Delta infections and throat pain in Omicron infections. Among Delta-infected individuals with specific symptoms, significantly higher levels of anti-N antibodies were observed. CONCLUSION: Our study identified variant-specific differences in the amount of SARS-CoV-2 antibody production and COVID-19 symptoms. Despite this, vaccinated individuals with Omicron or Delta infections generally experienced mild disease courses. Additionally, asymptomatic individuals exhibit lower anti-SARS-CoV-2 antibody levels, indicating a clinical correlation between disease-specific antibodies and distinct symptoms, particularly in the case of the Delta variant. In follow-up studies, exploring post-COVID syndrome and focusing on cognitive symptoms in the acute phase of Omicron infections is crucial as it has the potential to longitudinally impact the lives of those affected.

Study of the effect of implantation temperature on the migration behaviour of Xe implanted into glassy carbon.

The effect of implantation temperature on the migration behaviour of xenon (Xe) implanted into glassy carbon and the effect of annealing on radiation damage retained by ion implantation were investigated. Glassy carbon substrates were implanted with 320 keV Xe+ to a fluence of 2 × 1016 cm-2. The implantation process was performed at room temperature (RT) and 100 °C Some of the as-implanted samples were isochronally annealed in vacuum at temperatures ranging from 300 °C to 700 °C in steps of 100 °C for 10 h. The as-implanted and annealed samples were characterized using Rutherford backscattering spectrometry (RBS) and Raman spectroscopy. The RT implanted depth profiles indicated that the migration of Xe towards the surface of glassy carbon was accompanied by a loss of Xe ions. The samples implanted at 100 °C indicated no diffusion or loss of Xe after annealing at 300 °C. However, annealing at temperatures ranging from 400 °C to 700 °C resulted in a slight shift in the Xe profile tail-end towards the bulk of glassy carbon. The diffusion coefficients (D) in the temperature range of 300 °C-700 °C for the RT and 100 °C implanted samples, activation energies (Ea), and pre-exponential factors (Do), were extracted. The values of D ranged from (9.72 ± 0.48) × 10-21 to (1.87 ± 0.09) × 10-20 m2/s with an activation energy of (6.25 ± 0.31) × 10-5 eV for RT implanted samples, and the samples implanted at 100 °C, D ranged from (3.85 ± 0.19) × 10-21 to (6.96 ± 0.34) × 10-20 m2/s with activation energy of (4.10 ± 0.02) × 10-5 eV. The Raman analysis revealed that implantation at the RT amorphised the glassy carbon structure while the samples implanted at 100 °C showed mild damage compared to RT implantation. Annealing of the RT-implanted sample resulted in some recovery of the damaged region as a function of increasing annealing temperature.

A smart RBS library and its prediction model for robust and accurate fine-tuning of gene expression in Bacillus species.

Bacillus species, such as Bacillus subtilis and Bacillus licheniformis, are important industrial bacteria. However, there is a lack of standardized and predictable genetic tools for convenient and reproducible assembly of genetic modules in Bacillus species to realize their full potential. In this study, we constructed a Ribosome Binding Site (RBS) library in B. licheniformis, which provides incremental regulation of expression levels over a 104-fold range. Additionally, we developed a model to quantify the resulting translation rates. We successfully demonstrated the robust expression of various target genes using the RBS library and showed that the model accurately predicts the translation rates of arbitrary coding genes. Importantly, we also extended the use of the RBS library and prediction model to B. subtilis, B. thuringiensis, and B. amyloliquefacie. The versatility of the RBS library and its prediction model enables quantification of biological behavior, facilitating reliable forward engineering of gene expression.

Glucose indices as inflammatory markers in children with acute surgical abdomen: a cross-sectional study.

BACKGROUND: Glycaemic dysregulation potentiates the pro-inflammatory response and increases oxidative injury; therefore, preoperative hyperglycaemia is linked to increased mortalities. In addition, inflammation is accompanied by higher glycated haemoglobin (HbA1c) levels, and the relationship between this and random blood sugar (RBS) could be non-linear. METHODS: This is a cross-sectional study. Non-diabetic paediatric patients with acute surgical abdomen, presenting to the emergency surgical services were enrolled, over a period of 6 months. They were all screened for their random blood sugar and HbA1c levels. RESULTS: Fifty-three cases were studied. The prevalence of glycaemic dysregulation in the enrolled children was high. Abnormal HbA1c was observed in 66% of the study group. Stress hyperglycaemia was observed in 60% of the enrolled children. There was a significant correlation (r = 0.770, p-value: < 0.001) between RBS and the total leucocytic count (TLC). The TLC cutoff value for predicting stress hyperglycaemia was 13,595 cells/mm3. The cutoff value of RBS for predicting leukocytosis was 111.5 mg/dl. Median RBS level was significantly higher in complicated appendicitis (169.5 mg/dl), compared to uncomplicated appendicitis (118.0 mg/dl). CONCLUSION: HbA1c and RBS could be used as inflammatory markers for surgical acute abdomen and its degree of severity, respectively. HbA1c rises in a considerable number of cases with surgical acute abdomen, irrespective of the disease stage. However, as the disease progresses, the random blood sugar rises due to stress hyperglycaemia, thus becoming a surrogate inflammatory marker.

What is already known on this topic? Preoperative stress hyperglycaemia is common in children, and it is linked to adverse postoperative outcomes. HbA1c could be a marker for inflammation and oxidative stress.What does this study add? HbA1c could be an inflammatory marker for surgical acute abdomen, irrespective of the disease stage, as it had a high prevalence in the enrolled children with an acute surgical abdomen. However, as the disease progresses, the random blood sugar rises due to stress hyperglycaemia, thus becoming a surrogate inflammatory marker, as there is a significant correlation between it and the TLC.How might this study affect research, practice, or policy? The novelty in this study centers around the role of glucose metabolism, as evaluated by random blood sugar and HBA1c, in the diagnostic evaluation and prognostication of inflammation, represented by the surgical acute abdomen. This may invite further research into understanding the underlining mechanisms. The outcome of the clinical management of conditions involving inflammation can be improved by using the proposed biomarkers, as peri/preoperative hyperglycaemia could lead to morbidity and mortality, consequently, as proven, the reliability of those biomarkers facilitates risk assessment and stratification. As both tests are cost-effective and universally available, they can be readily implemented in practice guidelines and departmental policies.

[Rational metabolic engineering of Corynebacterium glutamicum for efficient synthesis of L-glutamate].

L-glutamic acid is the world's largest bulk amino acid product that is widely used in the food, pharmaceutical and chemical industries. Using Corynebacterium glutamicum G01 as the starting strain, the fermentation by-product alanine content was firstly reduced by knocking out the gene encoding alanine aminotransferase (alaT), a major by-product related to alanine synthesis. Secondly, since the α-ketoglutarate node carbon flow plays an important role in glutamate synthesis, the ribosome-binding site (RBS) sequence optimization was used to reduce the activity of α-ketoglutarate dehydrogenase and enhance the glutamate anabolic flow. The endogenous conversion of α-ketoglutarate to glutamate was also enhanced by screening different glutamate dehydrogenase. Subsequently, the glutamate transporter was rationally desgined to improve the glutamate efflux capacity. Finally, the fermentation conditions of the strain constructed using the above strategy were optimized in 5 L fermenters by a gradient temperature increase combined with a batch replenishment strategy. The glutamic acid production reached (135.33±4.68) g/L, which was 41.2% higher than that of the original strain (96.53±2.32) g/L. The yield was 55.8%, which was 11.6% higher than that of the original strain (44.2%). The combined strategy improved the titer and the yield of glutamic acid, which provides a reference for the metabolic modification of glutamic acid producing strains.

Two-Step Electrochemical Au Nanoparticle Formation in Polyaniline.

In this work, we use a two-step cyclic electrochemical process to insert Au into polyaniline (PANI). It was suggested previously that this method would lead to the formation of atomic Au clusters with controlleds number of Au atoms without providing morphological proof. In each cycle, tetrachloroaurate anions (AuCl4-) are attached on the protonated imine sites of PANI, followed by a controlled reduction using cyclic voltammetry (CV). In contrast to previous work, we demonstrate that the reduction leads to the nucleation and growth of an Au nanoparticle (NP) whose density and size dispersion depend on the Au loading in PANI. Adding more deposition cycles increases the Au NP density and size. Transmission electron microscopy (TEM) and corresponding energy dispersive X-ray spectroscopy (EDS) indicate a homogeneous distribution of Au elements in the PANI matrix before CV reduction, while Au elements are aggregated and clearly localized in the NPs positions after CV reduction. We further use Rutherford backscattering spectrometry (RBS) to quantify the Au uptake in PANI. The Au distribution is verified to be initially homogeneous across the PANI layer whereas the increasing number of deposition cycles leads to a surface segregation of Au. We propose a two-step growth model based on our experimental results. Finally, we discuss the results with respect to the formation of atomic Au clusters reported previously using the same deposition method.

Enhanced extracellular production of raw starch-degrading α-amylase in Bacillus subtilis through expression regulatory element modification and fermentation optimization.

BACKGROUND: Raw starch-degrading α-amylase (RSDA) can hydrolyze raw starch at moderate temperatures, thus contributing to savings in starch processing costs. However, the low production level of RSDA limits its industrial application. Therefore, improving the extracellular expression of RSDA in Bacillus subtilis, a commonly used industrial expression host, has great value. RESULTS: In this study, the extracellular production level of Pontibacillus sp. ZY raw starch-degrading α-amylase (AmyZ1) in B. subtilis was enhanced by expression regulatory element modification and fermentation optimization. As an important regulatory element of gene expression, the promoter, signal peptide, and ribosome binding site (RBS) sequences upstream of the amyZ1 gene were sequentially optimized. Initially, based on five single promoters, the dual-promoter Pveg-PylB was constructed by tandem promoter engineering. Afterward, the optimal signal peptide SPNucB was obtained by screening 173 B. subtilis signal peptides. Then, the RBS sequence was optimized using the RBS Calculator to obtain the optimal RBS1. The resulting recombinant strain WBZ-VY-B-R1 showed an extracellular AmyZ1 activity of 4824.2 and 41251.3 U/mL during shake-flask cultivation and 3-L fermenter fermentation, which were 2.6- and 2.5-fold greater than those of the original strain WBZ-Y, respectively. Finally, the extracellular AmyZ1 activity of WBZ-VY-B-R1 was increased to 5733.5 U/mL in shake flask by optimizing the type and concentration of carbon source, nitrogen source, and metal ions in the fermentation medium. On this basis, its extracellular AmyZ1 activity was increased to 49082.1 U/mL in 3-L fermenter by optimizing the basic medium components as well as the ratio of carbon and nitrogen sources in the feed solution. This is the highest production level reported to date for recombinant RSDA production. CONCLUSIONS: This study represents a report on the extracellular production of AmyZ1 using B. subtilis as a host strain, and achieved the current highest expression level. The results of this study will lay a foundation for the industrial application of RSDA. In addition, the strategies employed here also provide a promising way for improving other protein production in B. subtilis.

Addressing forensic science challenges with nuclear analytical techniques - A review.

We review the application of Nuclear Analytical Techniques (NATs) to forensic problems for the first time. NATs include neutron activation analysis (NAA), carried out in nuclear reactors for elemental analysis; accelerator-based techniques, mainly Ion Beam Analysis (IBA) for elemental and molecular analysis; and Accelerator Mass Spectrometry (AMS) for dating of traces of forensic interest by "radiocarbon dating" and other related methods. Applications include analysis of drugs of abuse, food fraud, counterfeit medicine, gunshot residue, glass fragments, forgery of art objects and documents, and human material. In some applications only the NATs are able to provide relevant information for forensic purposes. This review not only includes a wide collection of forensic applications, but also illustrates the wide availability worldwide of NATs, opening up opportunities for an increased use of NATs in routine forensic casework.

The Impact of Rayleigh Scattering in UWFBG Array-Based Φ-OTDR and Its Suppression Method.

Ultra-weak fiber Bragg grating (UWFBG) array-based phase-sensitive optical time-domain reflectometry (Φ-OTDR) utilizes the interference interaction between the reference light and the reflected light from the broadband gratings for sensing. It significantly improves the performance of the distributed acoustic sensing system (DAS) because the intensity of the reflected signal is much higher than that of the Rayleigh backscattering. This paper shows that Rayleigh backscattering (RBS) has become one of the primary noise sources in the UWFBG array-based Φ-OTDR system. We reveal the impact of the Rayleigh backscattering signal on the intensity of the reflective signal and the precision of the demodulated signal, and we suggest reducing the pulse duration to improve the demodulation accuracy. Experimental results demonstrate that using light with a 100 ns pulse duration can improve the measurement precision by three times compared with the use of a 300 ns pulse duration.

Modulating Sensitivity of an Erythromycin Biosensor for Precise High-Throughput Screening of Strains with Different Characteristics.

Genetically encoded biosensors are powerful tools for product-driven high-throughput screening in synthetic biology and metabolic engineering. However, most biosensors can only properly function in a limited concentration cutoff, and the incompatible performance characteristics of biosensors will lead to false positives or failure in screening. The transcription factor (TF)-based biosensors are usually organized in modular architecture and function in a regulator-depended manner, whose performance properties can be fine-tuned by modifying the expression level of the TF. In this study, we modulated the performance characteristics, including sensitivity and operating range, of an MphR-based erythromycin biosensor by fine-adjusting regulator expression levels via ribosome-binding site (RBS) engineering and obtained a panel of biosensors with varied sensitivities by iterative fluorescence-assisted cell sorting (FACS) in Escherichia coli to accommodate different screening purposes. To exemplify their application potential, two engineered biosensors with 10-fold different sensitivities were employed in the precise high-throughput screening by microfluidic-based fluorescence-activated droplet sorting (FADS) of Saccharopolyspora erythraea mutant libraries with different starting erythromycin productions, and mutants representing as high as 6.8 folds and over 100% of production improvements were obtained starting from the wild-type strain and the high-producing industrial strain, respectively. This work demonstrated a simple strategy to engineer biosensor performance properties, which was significant to stepwise strain engineering and production improvement.

Efficient Production of 3-Amino-2-Hydroxy Acetophenone by Multi-Enzyme Biosynthesis.

We developed a synthetic route for producing 3-amino-2-hydroxy acetophenone (3AHAP) from m-nitroacetophenone (3NAP) using an in vitro approach. Various reaction systems were evaluated, and a direct reaction method with crude enzyme and supersaturated substrates for optimal catalytic efficiency was chosen. The reaction system included three enzymes and was enhanced by adjusting enzyme molar ratios and optimizing ribosomal binding sites. We performed substrate docking and alanine scanning to identify key sites in the enzymes nitrobenzene nitroreductase (nbzA) and hydroxylaminobenzene mutase (habA). The optimal mutant was obtained through site-directed mutagenesis, and incorporated into the reaction system, resulting in increased product yield. After optimization, the yield of 3AHAP increased from 75 mg/L to 580 mg/L within 5 hours, the highest reported yield using biosynthesis. This work provides a promising strategy for the efficient and sustainable production of 3AHAP, which has critical applications in the chemical and pharmaceutical industries.

Direct Imaging of Band Structure for Powdered Rhombohedral Boron Monosulfide by Microfocused ARPES.

Boron-based two-dimensional (2D) materials are an excellent platform for nanoelectronics applications. Rhombohedral boron monosulfide (r-BS) is attracting particular attention because of its unique layered crystal structure suitable for exploring various functional properties originating in the 2D nature. However, studies to elucidate its fundamental electronic states have been largely limited because only tiny powdered crystals were available, hindering a precise investigation by spectroscopy such as angle-resolved photoemission spectroscopy (ARPES). Here we report the direct mapping of the band structure with a tiny (∼20 × 20 µm2) r-BS powder crystal by utilizing microfocused ARPES. We found that r-BS is a p-type semiconductor with a band gap of >0.5 eV characterized by the anisotropic in-plane effective mass. The present results demonstrate the high applicability of micro-ARPES to tiny powder crystals and widen an opportunity to access the yet-unexplored electronic states of various novel materials.

Effect of precursor concentration on stoichiometry and optical properties of spray pyrolyzed nanostructured NiO thin films.

In this study, spray pyrolysis was used to produce nanostructured NiO thin films from high purity nickel acetate (Ni(CH3COO)2.3H2O) precursors on pre-heated ultrasonically cleaned soda-lime glass substrates. The metallic constituent concentrations in the films were varied, and the precursors were produced in distilled water at various molarities ranging from 0.1 to 0.4 M. In the study, the field-emission scanning electron microscope (FESEM) results strongly confirmed adherence of the films to the glass substrate at 350 °C. The presence of Ni and O in the samples was confirmed using Rutherford backscattering spectroscopy (RBS), X-ray diffractometry (XRD) and energy dispersive X-ray spectroscopy (EDX). For the 0.1 M NiO thin films, the thickness was approximately 43 nm, and for the 0.2 M, 0.3 M, and 0.4 M films, the thickness was 46 nm, 47 nm, and 49 nm, respectively. The XRD findings were supported by the increased Raman intensity peaks with increased precursor concentration, which confirmed the films' improved crystallinity. For the same number of passes of films deposition, as the molar concentration increases, the films thickness increases. The amount of nickel in NiO thin films increases as the molarity increases, but the amount of oxygen in NiO thin films decreases as the molarity increases. It was discovered that as molarity increases, the optical transmittance decreases and the optical band gap narrows. The qualities of NiO discovered in this study suggest the films' potentials for usage as window layer and buffer material in thin film solar cells.

Real-time monitoring of subcellular states with genetically encoded redox biosensor system (RBS) in yeast cell factories.

During industrial fermentation, microbial cell factories are usually confronted with environmental or metabolic stresses, leading to the imbalance of intracellular redox and the reduction of cell metabolic capacity. Here, we constructed the genetically encoded redox biosensor system (RBS) based on redox-sensitive fluorescent proteins to detect redox metabolites, including reactive oxygen species (ROS), oxidized glutathione, NADH, and NADPH in Saccharomyces cerevisiae. The functional biosensors were quantitatively characterized and the orthogonal redox biosensor system (oRBS) was designed for detecting multiple redox metabolites. Furthermore, the compartment targeted redox biosensor system (ctRBS) was constructed to detect ROS and NADPH, revealing the distribution and spatiotemporal dynamics of ROS in yeast under various stress conditions. As a proof-of-concept, RBS was applied to evaluate the redox states of engineered yeast with stress resistance and heterogenous triterpene synthesis in vivo, elucidating the redox balance significantly affecting the growth and production phenotypes. The RBS in this study allowed the exploration of the diversity of compartmental redox state and real-time monitoring of the production process of yeast, providing a reliable and effective approach for accurate and in-depth profiling of bottlenecks of yeast cell factories.

Rational metabolic engineering of Corynebacterium glutamicum for efficient synthesis of L-glutamate / 生物工程学报

L-glutamic acid is the world's largest bulk amino acid product that is widely used in the food, pharmaceutical and chemical industries. Using Corynebacterium glutamicum G01 as the starting strain, the fermentation by-product alanine content was firstly reduced by knocking out the gene encoding alanine aminotransferase (alaT), a major by-product related to alanine synthesis. Secondly, since the α-ketoglutarate node carbon flow plays an important role in glutamate synthesis, the ribosome-binding site (RBS) sequence optimization was used to reduce the activity of α-ketoglutarate dehydrogenase and enhance the glutamate anabolic flow. The endogenous conversion of α-ketoglutarate to glutamate was also enhanced by screening different glutamate dehydrogenase. Subsequently, the glutamate transporter was rationally desgined to improve the glutamate efflux capacity. Finally, the fermentation conditions of the strain constructed using the above strategy were optimized in 5 L fermenters by a gradient temperature increase combined with a batch replenishment strategy. The glutamic acid production reached (135.33±4.68) g/L, which was 41.2% higher than that of the original strain (96.53±2.32) g/L. The yield was 55.8%, which was 11.6% higher than that of the original strain (44.2%). The combined strategy improved the titer and the yield of glutamic acid, which provides a reference for the metabolic modification of glutamic acid producing strains.

Study of HbA1c levels in non-diabetic individuals recovered from Covid-19.

Objectives: To study the glycemic control by estimating HbA1c levels among previously non-diabetic individuals aged above 40 years, who recovered from coronavirus disease 2019 (Covid-19) infection. Materials and Methods: This is an analytical cross-sectional study of people affected with Covid-19 infection, who were home-isolated, hospitalized, and discharged. The simple random sampling technique was used for the study group. Tirupati City, Andhra Pradesh State, India. We assessed socio-demographic profiles, hba1c levels, RBS, and severity of infection. The obtained laboratory reports were analyzed by using means and proportions after entering data in MS Excel and converted to SPSS software new version. Results: The study participants comprised mostly middle age group males and females (mostly 40-60 years, with 60% of them being females). Most of the study subjects were healthy, and 58% of them have normal hba1c levels. 32% of them are in border line diabetic. The prevalence of diabetes post Covid-19 infection is 10%. The majority of participants have normal random blood sugar levels (around 92%), and very less individuals have abnormal random blood sugar levels (2% of the subjects). There is no significant association between hypertension and raised blood sugar levels post Covid-19 infection. Conclusions: Based on the study, Covid-19 infection has an impact on blood sugar levels and altered glycemic control, switching a few individuals into diabetic and a significant number of individuals into the pre-diabetic state, who are non-diabetic individuals before Covid-19 infection.