Positional cloning identified HvTUBULIN8 as the candidate gene for round lateral spikelet (RLS) in barley (Hordeum vulgare L.).

KEY MESSAGE: Map-based cloning, subcellular localization, virus-induced-gene-silencing and transcriptomic analysis reveal HvTUB8 as a candidate gene with pleiotropic effects on barley spike and leaf development via ethylene and chlorophyll metabolism. Barley lateral spikelet morphology and grain shape play key roles in grain physical quality and yield. Several genes and QTLs for these traits have been cloned or fine mapped previously. Here, we report the phenotypic and genotypic analysis of a barley mutant with round lateral spikelet (rls) from cv. Edamai 934. rls had round lateral spikelet, short but round grain, shortened awn, thick glume and dark green leaves. Histocytologic and ultrastructural analysis revealed that the difference of grain shape of rls was caused by change of cell arrangement in glume, and the dark leaf color resulted from enlarged chloroplast. HvTUBULIN8 (HvTUB8) was identified as the candidate gene for rls by combination of RNA-Seq, map-based-cloning, virus-induced-gene-silencing (VIGS) and protein subcellular location. A single G-A substitution at the third exon of HvTUB8 resulted in change of Cysteine 354 to tyrosine. Furthermore, the mutant isoform Hvtub8 could be detected in both nucleus and cytoplasm, whereas the wild-type protein was only in cytoplasm and granular organelles of wheat protoplasts. Being consistent with the rare phenotype, the "A" allele of HvTUB8 was only detected in rls, but not in a worldwide barley germplasm panel with 400 accessions. VIGS confirmed that HvTUB8 was essential to maintain spike integrity. RNA-Seq results suggested that HvTUB8 may control spike morphogenesis via ethylene homeostasis and signaling, and control leaf color through chlorophyll metabolism. Collectively, our results support HvTUB8 as a candidate gene for barley spike and leaf morphology and provide insight of a novel mechanism of it in barley development.

Discovery of a novel analogue of FR901533 and the corresponding biosynthetic gene cluster from Streptosporangium roseum No. 79089.

FR901533 (1, also known as WS79089B), WS79089A (2), and WS79089C (3) are polycyclic aromatic natural products with promising inhibitory activity to endothelin-converting enzymes. In this work, we isolated five tridecaketide products from Streptosporangium roseum No. 79089, including 1-3, benaphthamycin (4) and a novel FR901533 analogue (5). The structure of 5 was characterized based on spectroscopic data. Compared with the major product 2, the new compound 5 has an additional hydroxyl group at C-12 and an extra methyl group at the 13-OH. The configuration of C-19 of these compounds was determined to be R using Mosher's method. A putative biosynthetic gene cluster for compounds 1-5 was discovered by analyzing the genome of S. roseum No. 79089. This 38.6-kb gene cluster contains 38 open reading frames, including a minimal polyketide synthase (wsaA-C), an aromatase (wsaD), three cyclases (wsaE, F, and W), and a series of tailoring enzymes such as monooxygenases (wsaO1-O7) and methyltransferases (wsaM1 and M2). Disruption of the ketosynthase gene (wsaA) in this gene cluster abolished the production of 1-5, confirming that this gene cluster is indeed responsible for the biosynthesis of 1-5. A type II polyketide biosynthetic pathway was proposed for this group of natural endothelin-converting enzyme inhibitors. KEY POINTS: • Five aromatic tridecaketides were isolated from Streptosporangium roseum No. 79089. • A novel FR901533 analogue, 12-hydroxy-13-O-methyl-WS79089A, was characterized. • The absolute configuration of C-19 of FR901533 and analogues was determined. • The biosynthetic gene cluster of FR901533 and analogues was discovered.

Simultaneous determination of 14 urinary biomarkers of exposure to organophosphate flame retardants and plasticizers by LC-MS/MS.

Organophosphate flame retardants and plasticizers (PFRs) are a group of chemicals widely added to consumer products. PFRs are quickly metabolized in the human body into two types of metabolites, (1) dialkyl and diaryl phosphate esters (DAPs), such as diphenyl phosphate (DPHP) and bis(1,3-dichloro-2-propyl) phosphate (BDCIPP); and (2) hydroxylated PFRs (HO-PFRs), such as 1-hydroxy-2-propyl bis(1-chloro-2-propyl) phosphate (BCIPHIPP) and 2-hydroxyethyl bis(2-butoxyethyl) phosphate (BBOEHEP). Existing analytical methods usually focus on DAPs; therefore, human biomonitoring data on HO-PFRs remain scarce. In this study, an analytical procedure was developed for the simultaneous quantification of multiple PFR metabolites in human urine, covering eight DAPs and six HO-PFRs. Sample preparation was optimized to include all target compounds using Bond-Elut C18 solid-phase extraction cartridges, followed by instrumental analysis based on liquid-chromatography coupled to tandem mass spectrometry (LC-MS/MS). Method performance was validated according to established guidelines and satisfactory results were obtained for all metabolites in terms of recovery, linearity, limits of quantification, precision, and accuracy. Recoveries ranged from 87 to 112%. Method detection limits from 0.002 ng/mL for 2-ethyl-5-hydroxyhexyl diphenyl phosphate (5-HO-EHDPHP) to 0.66 ng/mL for 4-hydroxyphenyl phenyl phosphate (4-HO-DPHP). Seven PFR metabolites were frequently detected in a small biomonitoring study (n = 14), among them bis(1,3-dichloro-2-propyl) phosphate (BDCIPP), di-n-butyl phosphate (DNBP), 5-HO-EHDPHP, and BBOEHEP. Highest mean concentrations were found for DPHP, 2-ethylhexyl phenyl phosphate (EHPHP), and BCIPHIPP, while 4-HO-DPHP, 5-HO-EHDPHP, and EHPHP were detected in urine for the first time. Overall, the obtained results demonstrate that the developed method can be used for the simultaneous determination of 14 urinary biomarkers of exposure to PFRs. Graphical abstract ᅟ.

New insights into pradimicin biosynthesis revealed by two O-methyltransferases.

Pradimicins are a group of antiviral and antifungal natural products from Actinomadura hibisca. Two putative O-methyltransferase genes, pdmF and pdmT, are present in the pradimicin biosynthetic gene cluster. However, there is only one methoxy group (11-OCH3) in pradimicins. Through heterologous expression and in vitro reactions with various substrates, PdmF was characterized as the C-11 O-methyltransferase with a relatively broad substrate specificity. To probe the role of PdmT in pradimicin biosynthesis, the corresponding gene was disrupted through homologous recombination, leading to the production of pradimicinone II. This enzyme was then expressed in Escherichia coli with an N-terminal His6 tag and purified by Ni-NTA chromatography. Reaction of pradimicinone II with PdmT generated 7-O-methylpradimicinone II, confirming that this enzyme is a C-7 O-methyltransferase. Characterization of PdmT suggests a novel pathway that leads to the "flip" of 7-OH to C-14 in pradimicin biosynthesis.

Identification of Cyclic Depsipeptides and Their Dedicated Synthetase from Hapsidospora irregularis.

Seven cyclic depsipeptides were isolated from Hapsidospora irregularis and structurally characterized as the calcium channel blocker leualacin and six new analogues based on the NMR and HRESIMS data. These new compounds were named leualacins B-G. The absolute configurations of the amino acids and 2-hydroxyisocaproic acids were determined by recording the optical rotation values. Biological studies showed that calcium influx elicited by leualacin F in primary human lobar bronchial epithelial cells involves the TRPA1 channel. Through genome sequencing and targeted gene disruption, a noniterative nonribosomal peptide synthetase was found to be involved in the biosynthesis of leualacin. A comparison of the structures of leualacin and its analogues indicated that the A2 and A4 domains of the leualacin synthetase are substrate specific, while A1, A3, and A5 can accept alternative precursors to yield new molecules.

Comprehensive Study of Human External Exposure to Organophosphate Flame Retardants via Air, Dust, and Hand Wipes: The Importance of Sampling and Assessment Strategy.

We compared the human exposure to organophosphate flame retardants (PFRs) via inhalation, dust ingestion, and dermal absorption using different sampling and assessment strategies. Air (indoor stationary air and personal ambient air), dust (floor dust and surface dust), and hand wipes were sampled from 61 participants and their houses. We found that stationary air contains higher levels of ΣPFRs (median = 163 ng/m(3), IQR = 161 ng/m(3)) than personal air (median = 44 ng/m(3), IQR = 55 ng/m(3)), suggesting that the stationary air sample could generate a larger bias for inhalation exposure assessment. Tris(chloropropyl) phosphate isomers (ΣTCPP) accounted for over 80% of ΣPFRs in both stationary and personal air. PFRs were frequently detected in both surface dust (ΣPFRs median = 33 100 ng/g, IQR = 62 300 ng/g) and floor dust (ΣPFRs median = 20 500 ng/g, IQR = 30 300 ng/g). Tris(2-butoxylethyl) phosphate (TBOEP) accounted for 40% and 60% of ΣPFRs in surface and floor dust, respectively, followed by ΣTCPP (30% and 20%, respectively). TBOEP (median = 46 ng, IQR = 69 ng) and ΣTCPP (median = 37 ng, IQR = 49 ng) were also frequently detected in hand wipe samples. For the first time, a comprehensive assessment of human exposure to PFRs via inhalation, dust ingestion, and dermal absorption was conducted with individual personal data rather than reference factors of the general population. Inhalation seems to be the major exposure pathway for ΣTCPP and tris(2-chloroethyl) phosphate (TCEP), while participants had higher exposure to TBOEP and triphenyl phosphate (TPHP) via dust ingestion. Estimated exposure to ΣPFRs was the highest with stationary air inhalation (median =34 ng·kg bw(-1)·day(-1), IQR = 38 ng·kg bw(-1)·day(-1)), followed by surface dust ingestion (median = 13 ng·kg bw(-1)·day(-1), IQR = 28 ng·kg bw(-1)·day(-1)), floor dust ingestion and personal air inhalation. The median dermal exposure on hand wipes was 0.32 ng·kg bw(-1)·day(-1) (IQR = 0.58 ng·kg bw(-1)·day(-1)) for ΣTCPP. The selection of sampling and assessment strategies could significantly affect the results of exposure assessment.

Characterization and fine mapping of a novel barley Stage Green-Revertible Albino Gene (HvSGRA) by Bulked Segregant Analysis based on SSR assay and Specific Length Amplified Fragment Sequencing.

BACKGROUND: Leaf color variations are common in plants. Herein we describe a natural mutant of barley cultivar Edamai No.6, whs18, whose leaf color showed stable and inheritable stage-green-revertible-albino under field condition. METHODS: Bulked Segregant Analysis (BSA) based on SSR assay and Specific Length Amplified Fragment Sequencing (SLAF-seq) was used to map the candidate gene for this trait. RESULTS: We found that leaf color of whs18 was green at seedling stage, while the seventh or eighth leaf began to show etiolation, and albino leaves emerged after a short period. The newly emerged leaves began to show stripe white before jointing stage, and normal green leaves emerged gradually. The duration of whs18 with abnormal leaf color lasted for about 3 months, which had some negative impacts on yield-related-traits. Further investigations showed that the variation was associated with changes in chlorophyII content and chloroplast development. Genetic analysis revealed that the trait was controlled by a single recessive nuclear gene, and was designed as HvSGRA in this study. Based on the F2 population derived from Edamai No.9706 and whs18, we initially mapped the HvSGRA gene on the short arm of chromosome 2H using SSR and BSA. GBMS247 on 2HS showed co-segregation with HvSGRA. The genetic distance between the other marker GBM1187 and HvSGRA was 1.2 cM. Further analysis using BSA with SLAF-seq also identified this region as candidate region. Finally, HvSGRA interval was narrowed to 0.4 cM between morex_contig_160447 and morex_contig_92239, which were anchored to two adjacent FP contigs, contig_34437 and contig_46434, respectively. Furthermore, six putative genes with high-confidence in this interval were identified by POPSEQ. Further analysis showed that the substitution from C to A in the third exon of fructokinase-1-like gene generated a premature stop codon in whs18, which may lead to loss function of this gene. CONCLUSIONS: Using SSR and SLAF-seq in conjunction with BSA, we mapped HvSGRA within two adjacent FP contigs of barley. The mutation of fructokinase-1-like gene in whs18 may cause the stage green-revertible albino of barley. The current study lays foundation for hierarchical map-based cloning of HvSGRA and utilizing the gene/trait as a visualized maker in molecular breeding in future.

Seasonal and Particle Size-Dependent Variations of Hexabromocyclododecanes in Settled Dust: Implications for Sampling.

Particle size is a significant parameter which determines the environmental fate and the behavior of dust particles and, implicitly, the exposure risk of humans to particle-bound contaminants. Currently, the influence of dust particle size on the occurrence and seasonal variation of hexabromocyclododecanes (HBCDs) remains unclear. While HBCDs are now restricted by the Stockholm Convention, information regarding HBCD contamination in indoor dust in China is still limited. We analyzed composite dust samples from offices (n = 22), hotels (n = 3), kindergartens (n = 2), dormitories (n = 40), and main roads (n = 10). Each composite dust sample (one per type of microenvironment) was fractionated into 9 fractions (F1-F9: 2000-900, 900-500, 500-400, 400-300, 300-200, 200-100, 100-74, 74-50, and <50 µm). Total HBCD concentrations ranged from 5.3 (road dust, F4) to 2580 ng g(-1) (dormitory dust, F4) in the 45 size-segregated samples. The seasonality of HBCDs in indoor dust was investigated in 40 samples from two offices. A consistent seasonal trend of HBCD levels was evident with dust collected in the winter being more contaminated with HBCDs than dust from the summer. Particle size-selection strategy for dust analysis has been found to be influential on the HBCD concentrations, while overestimation or underestimation would occur with improper strategies.

Efficient production of indigoidine in Escherichia coli.

Indigoidine is a bacterial natural product with antioxidant and antimicrobial activities. Its bright blue color resembles the industrial dye indigo, thus representing a new natural blue dye that may find uses in industry. In our previous study, an indigoidine synthetase Sc-IndC and an associated helper protein Sc-IndB were identified from Streptomyces chromofuscus ATCC 49982 and successfully expressed in Escherichia coli BAP1 to produce the blue pigment at 3.93 g/l. To further improve the production of indigoidine, in this work, the direct biosynthetic precursor L-glutamine was fed into the fermentation broth of the engineered E. coli strain harboring Sc-IndC and Sc-IndB. The highest titer of indigoidine reached 8.81 ± 0.21 g/l at 1.46 g/l L-glutamine. Given the relatively high price of L-glutamine, a metabolic engineering technique was used to directly enhance the in situ supply of this precursor. A glutamine synthetase gene (glnA) was amplified from E. coli and co-expressed with Sc-indC and Sc-indB in E. coli BAP1, leading to the production of indigoidine at 5.75 ± 0.09 g/l. Because a nitrogen source is required for amino acid biosynthesis, we then tested the effect of different nitrogen-containing salts on the supply of L-glutamine and subsequent indigoidine production. Among the four tested salts including (NH4)2SO4, NH4Cl, (NH4)2HPO4 and KNO3, (NH4)2HPO4 showed the best effect on improving the titer of indigoidine. Different concentrations of (NH4)2HPO4 were added to the fermentation broths of E. coli BAP1/Sc-IndC+Sc-IndB+GlnA, and the titer reached the highest (7.08 ± 0.11 g/l) at 2.5 mM (NH4)2HPO4. This work provides two efficient methods for the production of this promising blue pigment in E. coli.

A specific cytochrome P450 hydroxylase in herboxidiene biosynthesis.

The anti-cholesterol natural product herboxidiene is synthesized by a noniterative modular polyketide synthase (HerB, HerC and HerD) and three tailoring enzymes (HerE, HerF and HerG) in Streptomyces chromofuscus A7847. In this work, the putative monooxygenase HerG was expressed in Escherichia coli and the purified enzyme was subjected to biochemical studies. It was identified as a cytochrome P450 enzyme responsible for the stereospecific hydroxylation at C-18. This enzyme is highly substrate-specific as it efficiently hydroxylates 18-deoxy-25-demethyl-herboxidiene, but showed no activity towards 18-deoxy-herboxidiene. The kcat/Km value for the HerG-catalyzed hydroxylation of 18-deoxy-25-demethyl-herboxidiene was determined to be 1669.70±47.36 M(-1) s(-1). In vitro co-reaction of HerG with the methyltransferase HerF and analysis of the product formation in S. chromofuscus A7847 revealed that the biosynthetic intermediate 18-deoxy-25-demethyl-herboxidiene is successively hydroxylated at C-18 by HerG and methylated at 17-OH to yield the final product herboxidiene. The minor metabolite 18-deoxy-hereboxidiene is a byproduct of the biosynthetic pathway.

Distribution patterns of brominated, chlorinated, and phosphorus flame retardants with particle size in indoor and outdoor dust and implications for human exposure.

Dust samples were collected in Beijing, China, from four different indoor microenvironments (office, hotel, kindergarten, and student dormitory) and one outdoor (road dust) microenvironment. These five composite samples were fractionated into 13 sequential size fractions and an individual fraction of <50 µm for further analysis. In the fractions of <50 µm, nine phosphorus flame retardants (∑PFRs), four novel brominated flame retardants (∑NBFRs), and two Dechlorane Plus isomers (DPs) showed the highest concentrations in hotel dust (124,000 ng g(-1)), dormitory dust (14,200 ng g(-1)), and kindergarten dust (231 ng g(-1)), respectively. Nevertheless, nine polybrominated diphenyl ethers (∑PBDEs) were the dominant flame retardants (FRs) (96% of total FRs) in road dust, with the maximum concentration of 23,700 ng g(-1), higher than in any indoor dust. The FR contamination varied strongly among different types of microenvironments, leading to high human exposure to various FRs. Concentrations of FRs did not increase constantly with a particle size decrease. Fractions with a particle size around 900, 100, and 10 µm could represent peak values, while valley values were commonly detected around fractions with a particle size around 40 µm. Large differences were found between indoor dust and road dust. In road dust, FRs were mainly enriched in fractions of <50 µm. The organic content of dust, FR application, and consequent abrasion processes of FR-containing materials might be the determinants of the FR concentrations. Volatilization and abrasion were considered to be important migration pathways for FRs. DPs and BDE-209 were sought to be mainly applied in abrasion-proof materials, while most phosphorus flame retardants (PFRs) were probably added in a large proportion in materials easy to wear.

Human biomonitoring of emerging pollutants through non-invasive matrices: state of the art and future potential.

Human biomonitoring (HBM) is a scientific technique that allows us to assess whether and to what extent environmental pollutants enter humans. We review here the current HBM efforts for organophosphate esters, emerging flame retardants, perfluoroalkyl substances, and phthalate esters. Use of some of these chemicals has already been banned or restricted; they are regularly detected in the environment, wildlife, and human matrices. Traditionally, blood and urine collection have been widely used as sampling methods. New non-invasive approaches (e.g., saliva, hair, nails) are emerging as valid alternatives since they offer advantages with respect to sampling, handling, and ethical aspects, while ensuring similar reliability and sensitivity. Nevertheless, the identification of biomarkers of exposure is often difficult because chemicals may be metabolized in the human body. For many of the above-mentioned compounds, the mechanisms of the favorable metabolization pathways have not been unraveled, but research on important metabolites that could be used as biomarkers of exposure is growing. This review summarizes the state of the art regarding human exposure to, (non-invasive) HBM of, and metabolism of major organophosphate esters, emerging flame retardants, perfluoroalkyl substances, and phthalate esters currently detected in the environment.

Efficacy and Safety of Ketamine Compared with Placebo and Other Medications for Preventing Propofol Injection Pain in Adults: A Systematic Review and Meta-Analysis.

Purpose: To systematically evaluate the effectiveness and safety of ketamine in preventing propofol injection pain (PIP). Patients and Methods: The electronic databases including PubMed, Embase, Web of Science, and Cochrane Library were searched from their inception until 2 August 2023. Randomized controlled trials (RCT) comparing ketamine with placebo or other interventions to alleviate PIP in adults were included. Fixed-effects or random-effects models were used to calculate pooled risk ratios (RR) and corresponding 95% confidence intervals (CI) based on the heterogeneity of the studies included. Results: Thirteen RCTs involving 2105 patients were included. In terms of reducing the incidence of PIP, ketamine is more effective than placebo (RR = 0.43, 95% CI = [0.34, 0.55], P < 0.00001), lidocaine (RR = 0.70, 95% CI = [0.55, 0.90], P = 0.005), dexmedetomidine (RR = 0.52, 95% CI = [0.40, 0.66], P < 0.00001), and thiopental (RR = 0.25, 95% CI = [0.08, 0.83], P = 0.02). In reducing the incidence of severe PIP, ketamine is superior to placebo (RR = 0.12, 95% CI = [0.08, 0.19], P < 0.00001), and lidocaine (RR = 0.34, 95% CI = [0.21, 0.56], P < 0.0001), except dexmedetomidine (RR = 0.20, 95% CI = [0.04, 1.13], P = 0.07), and thiopental (RR = 0.33, 95% CI = [0.04, 3.10], P = 0.33). Compared with mixed injection, separate injection of ketamine and propofol showed no significant difference in the incidence of PIP (RR = 0.96, 95% CI = [0.31, 3.00], P = 0.95) and severe PIP (RR = 1.19, 95% CI = [0.07, 21.29], P = 0.90). Based solely on the reports from the studies included, subanesthetic doses of ketamine are generally safe in preventing PIP. Conclusion: A subanesthetic dose of ketamine can effectively and safely reduce the incidence of PIP and severe PIP in adults, and is more effective than lidocaine, dexmedetomidine, and thiopental. Registration: PROSPERO CRD42023455093.

Early versus delayed enteral nutrition in ICU patients with sepsis: a propensity score-matched analysis based on the MIMIC-IV database.

Background: Early enteral nutrition (EN) is recommended for sepsis management, but its optimal timing and clinical benefits remain uncertain. This study evaluates whether early EN improves outcomes compared to delayed EN in patients with sepsis. Methods: We analyzed data of septic patients from the MIMIC-IV 2.2 database, focusing on those in the Medical Intensive Care Unit (MICU) and Surgical Intensive Care Unit (SICU). Patients who initiated EN within 3 days were classified into the early EN group, while those who started EN between 3 and 7 days were classified into the delayed EN group. Propensity score matching was used to compare outcomes between the groups. Results: Among 1,111 patients, 786 (70.7%) were in the early EN group and 325 (29.3%) were in the delayed EN group. Before propensity score matching, the early EN group demonstrated lower mortality (crude OR = 0.694; 95% CI: 0.514-0.936; p = 0.018) and shorter ICU stays (8.3 [5.2, 12.3] vs. 10.0 [7.5, 14.2] days; p < 0.001). After matching, no significant difference in mortality was observed. However, the early EN group had shorter ICU stays (8.3 [5.2, 12.4] vs. 10.1 [7.5, 14.2] days; p < 0.001) and a lower incidence of AKI stage 3 (49.3% vs. 55.5%; p = 0.030). Subgroup analysis revealed that early EN significantly reduced the 28-day mortality rate in sepsis patients with lactate levels ≤4 mmol/L, with an adjusted odds ratio (aOR) of 0.579 (95% CI: 0.361, 0.930; p = 0.024). Conclusion: Early enteral nutrition may not significantly reduce overall mortality in sepsis patients but may shorten ICU stays and decrease the incidence of AKI stage 3. Further research is needed to identify specific patient characteristics that benefit most from early EN.

Engineered production of fungal anticancer cyclooligomer depsipeptides in Saccharomyces cerevisiae.

Two fungal cyclooligomer depsipeptide synthetases(CODSs), BbBEAS (352 kDa) and BbBSLS (348 kDa) from Beauveria bassiana ATCC7159, were reconstituted in Saccharomyces cerevisiae BJ5464-NpgA, leading to the production of the corresponding anticancer natural products, beauvericins and bassianolide, respectively. The titers of beauvericins (33.8 ± 1.4 mg/l) and bassianolide (21.7± 0.1 mg/l) in the engineered S. cerevisiae BJ5464-NpgA strains were comparable to those in the native producer B. bassiana. Feeding D-hydroxyisovaleric acid (D-Hiv) and the corresponding L-amino acid precursors improved the production of beauvericins and bassianolide. However, the high price of D-Hiv limits its application in large-scale production of these cyclooligomer depsipeptides. Alternatively, we engineered another enzyme, ketoisovalerate reductase (KIVR) from B. bassiana, into S. cerevisiae BJ5464-NpgA for enhanced in situ synthesis of this expensive substrate. Co-expression of BbBEAS and KIVR in the yeast led to significant improvement of the production of beauvericins.The total titer of beauvericin and its congeners (beauvericins A-C) was increased to 61.7 ± 3.0 mg/l and reached 2.6-fold of that in the native producer B. bassiana ATCC7159. Supplement of L-Val at 10 mM improved the supply of ketoisovalerate, the substrate of KIVR, which consequently further increased the total titer of beauvericins to 105.8 ± 2.1 mg/l. Using this yeast system,we functionally characterized an unknown CODS from Fusarium venenatum NRRL 26139 as a beauvericin synthetase, which was named as FvBEAS. Our work thus provides a useful approach for functional reconstitution and engineering of fungal CODSs for efficient production of this family of anticancer molecules.

Characterization of a methyltransferase involved in herboxidiene biosynthesis.

The herboxidiene biosynthetic gene cluster contains a regulatory gene and six biosynthetic genes that encode three polyketide synthases (HerB, HerC and HerD) and three tailoring enzymes (HerE, HerF and HerG). Through single crossover recombination, an integrative plasmid was inserted into the genome of Streptomyces chromofuscus ATCC 49982 between herE and herF, resulting in low-level expression of herF and the downstream herG. The mutant strain produced two new compounds, 18-deoxy-25-demethyl-herboxidiene and 25-demethyl-herboxidiene. HerF was expressed in Escherichia coli and biochemically characterized as the dedicated methyltransferase in herboxidiene biosynthesis. It prefers 25-demethyl-herboxidiene to 18-deoxy-25-demethyl-herboxidiene, suggesting that C-25 methylation is the last tailoring step.

An indigoidine biosynthetic gene cluster from Streptomyces chromofuscus ATCC 49982 contains an unusual IndB homologue.

A putative indigoidine biosynthetic gene cluster was located in the genome of Streptomyces chromofuscus ATCC 49982. The silent 9.4-kb gene cluster consists of five open reading frames, named orf1, Sc-indC, Sc-indA, Sc-indB, and orf2, respectively. Sc-IndC was functionally characterized as an indigoidine synthase through heterologous expression of the enzyme in both Streptomyces coelicolor CH999 and Escherichia coli BAP1. The yield of indigoidine in E. coli BAP1 reached 2.78 g/l under the optimized conditions. The predicted protein product of Sc-indB is unusual and much larger than any other reported IndB-like protein. The N-terminal portion of this enzyme resembles IdgB and the C-terminal portion is a hypothetical protein. Sc-IndA and/or Sc-IndB were co-expressed with Sc-IndC in E. coli BAP1, which demonstrated the involvement of Sc-IndB, but not Sc-IndA, in the biosynthetic pathway of indigoidine. The yield of indigoidine was dramatically increased by 41.4 % (3.93 g/l) when Sc-IndB was co-expressed with Sc-IndC in E. coli BAP1. Indigoidine is more stable at low temperatures.

Comparison of Remimazolam-Flumazenil versus Propofol for Recovery from General Anesthesia: A Systematic Review and Meta-Analysis.

(1) Purpose: to systematically evaluate the recovery following sedation and anesthesia with remimazolam combined with flumazenil in comparison to propofol. (2) Methods: Electronic databases, including PubMed, Embase, Web of Science, and the Cochrane Library, were systematically searched from their inception up to 22 October 2023. Included in this analysis were randomized controlled trials (RCT) that compared remimazolam-flumazenil with propofol for the recovery from sedation and anesthesia in adults. The risk of bias was assessed using the Cochrane risk of bias tool. Pooled risk ratios (RR) or mean differences (MD) along with their corresponding 95% confidence intervals (CI) were calculated using either fixed-effects or random-effects models, and the results were visualized in forest plots. (3) Results: Nine RCTs involving 745 patients who underwent general anesthesia in three different countries were included. Compared to propofol, the remimazolam-flumazenil combination shortened the emergence time (MD = -4.34 min, 95% CI = [-6.88, -1.81], p = 0.0008, low certainty), extubation time (MD = -4.26 min, 95% CI = [-6.81, -1.7], p = 0.0011, low certainty), and the post-anesthesia care unit (PACU) stay (MD = -4.42 min, 95% CI = [-7.45, -1.38], p = 0.0044, low certainty), while reducing the incidence of respiratory depression (RR = 0.2, 95% CI = [0.04, 0.89], p = 0.03, high certainty) after general anesthesia. However, this combination was associated with a higher incidence of re-sedation (RR = 4.15, 95% CI = [1.31, 13.13], p = 0.01, moderate certainty). (4) Conclusions: Based on the existing evidence, the combination of remimazolam and flumazenil accelerates recovery from general anesthesia and lowers the risk of respiratory depression compared to propofol. However, it is important to consider the higher risk of re-sedation when using this combination in clinical practice. Due to limitations in the quality of the evidence, it is advisable to interpret the results of meta-analyses with caution.

Prognostic value of plasma 7-ketocholesterol in sepsis.

BACKGROUND: Early evaluation of the severity of sepsis and estimation of its prognosis remains one of the main challenges in current therapeutic strategies. This study aimed to evaluate the prognostic value of plasma 7-ketocholesterol (7-KC) in sepsis. METHODS: We retrospectively measured by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) the plasma 7-KC concentration in 176 patients with sepsis and 90 healthy volunteers. A multivariate Cox proportional hazard model was introduced to identify independent factors, including plasma 7-KC and clinical features, for the 28-day mortality of sepsis, and a nomogram for predicting the 28-day mortality of sepsis was established. Decision curve analysis (DCA) was performed to assess the prediction model of death risk of sepsis. RESULTS: The area under the curve (AUC) of plasma 7-KC in diagnosing sepsis was 0.899 (95% CI = 0.862-0.935, P < 0.001), while it was 0.830 (95% CI = 0.764-0.894, P < 0.001) in diagnosing septic shock. The AUCs of plasma 7-KC in predicting the survival of sepsis patients in the training cohort and the test cohort were 0.770 (95% CI = 0.692-0.848, P < 0.05) and 0.869 (95% CI = 0.763-0.974, P < 0.05), respectively. In addition, high plasma 7-KC expression predicts poor prognosis in sepsis. Then, 7-KC and platelet count were identified as the two factors with significant differences by a multivariate Cox proportional hazard model, and the 28-day mortality probability ranged from 0.002 to 0.985 and was assessed using a nomogram. DCA results revealed that the combination of plasma 7-KC and platelet count showed the best prognostic efficiency of the risk threshold compared to a single factor in both the training cohort and test cohort. CONCLUSIONS: Collectively, the elevated plasma 7-KC level is an indicator of sepsis and was identified as a prognostic indicator for sepsis patients, providing a landscape for predicting survival in early sepsis with potential clinical utility.

Characterization of a barley (Hordeum vulgare L.) mutant with multiple stem nodes and spikes and dwarf (msnsd) and fine-mapping of its causal gene.

Introduction: Multiple nodes and dwarf mutants in barley are a valuable resource for identifying genes that control shoot branching, vegetative growth and development. Methods: In this study, physiological, microscopic and genetic analysis were conducted to characterize and fine-map the underling gene of a barley mutant with Multiple Stem Nodes and Spikes and Dwarf (msnsd), which was selected from EMS- and 60Co-treated barley cv. Edamai 934. Results and discussion: The msnsd mutant had more stem nodes, lower plant height and a shorter plastochron than Edamai 934. Moreover, the mutant had two or more spikes on each tiller. Microscopic analysis showed that the dwarf phenotype of msnsd resulted from reduced cell lengths and cell numbers in the stem. Further physiological analysis showed that msnsd was GA3-deficient, with its plant height increasing after external GA3 application. Genetic analysis revealed that a single recessive nuclear gene, namely, HvMSNSD, controlled the msnsd phenotype. Using a segregating population derived from Harrington and the msnsd mutant, HvMSNSD was fine-mapped on chromosome 5H in a 200 kb interval using bulked segregant analysis (BSA) coupled with RNA-sequencing (BSR-seq), with a C-T substitution in the exon of HvTCP25 co-segregating with the msnsd phenotype. RNA-seq analysis showed that a gene encoding gibberellin 2-oxidase 8, a negative regulator of GA biosynthesis, was upregulated in the msnsd mutant. Several known genes related to inflorescence development that were also upregulated and enriched in the msnsd mutant. Collectively, we propose that HvMSNSD regulates the plastochron and morphology of reproductive organs, likely by coordinating GA homeostasis and changed expression of floral development related genes in barley. This study offers valuable insights into the molecular regulation of barley plant architecture and inflorescence development.