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Pseudomonas putida KT2440 is a saprophytic, generally recognized as safe microorganism that plays important roles in the biodegradation and production of value-added chemicals. Chromosomal gene deletion of P. putida KT2440 usually involves time-consuming gene coning, conjugal transfer and counterselection. Recently, we developed a P. putida KT2440 markerless gene deletion method based on recombineering and Cre/loxP site-specific recombination. PCR-based λ Red recombineering circumvents the tedious cloning steps and is more amenable to high-throughput manipulation. Herein we report an improved scarless gene deletion strategy based on recombineering and I-SceI-mediated double-strand break repair. Sixteen drug exporter gene(s) were deleted and the minimal inhibition concentrations of the mutants to a variety of antibiotics were determined. The robustness of the procedure was also demonstrated by sequential deletion of five large genomic regions. Up to 59% recombination efficiency was achieved for 54.8 kb deletion, and the efficiency of RecA mediated double-strand break repair, which was boosted by λ Red recombinase, was nearly 100%. The strain with a 3.76% genome reduction showed an improved growth rate and transformation efficiency. The straightforward, time-saving and highly efficient scarless deletion approach has the potential to facilitate the genetic study and biotechnological and environmental applications of P. putida KT2440.

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Zircon has been widely applied in various traditional and emerging fields due to its distinct physiochemical advantages while monodisperse spherical zircon (MSZ) powders hitherto have not been explored for the tremendous challenges in the control of zircon crystallization and growth. Herein, systematic investigations are presented to reveal the paramount role of halide mineralizers upon the crystallization and growth of MSZ powders synthesized under highly acidic hydrothermal environments (pH <0). A formation mechanism is revealed revolving around the complexation of central Zr4+ with the Lewis basic ligand, fluorine, which forms the F‐containing species of [(OH)1–y·Ry] (R = F, Cl) to replace Zr and generates Zr‐deficient zircons as (ZrO4)1‐xSi[(OH)1‐y·Ry]4x. This finding differs much from conventional views on common fluoro‐hydroxylated hydrothermal zircons, which were generally regarded as Si‐deficient silicates from the substitution of fluoro‐hydroxyls for the [SiO4] tetrahedra. A parameter, Kr=4xy, is first proposed as an index to evaluate the stability of hydrothermal zircons. The MSZ powders demonstrate superb thermal stability at high temperature, superior photo‐reflectance, and thermal insulation over normal zircons, promising attractive prospects to extend zircon applications.

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Background and Purpose: The purpose of the study was to evaluate the usefulness of thromboelastography with platelet mapping (TEG-PM) for predicting hematoma expansion (HE) and poor functional outcome in patients with intracerebral hemorrhage (ICH).Methods: Patients with primary ICH who underwent baseline computed tomography (CT) and TEG-PM within 6 h after symptom onset were enrolled in the observational cohort study. We performed univariate and multivariate logistic regression models to assess the association of admission platelet function with HE and functional outcome. In addition, a receiver operating characteristic (ROC) curve analysis investigated the accuracy of platelet function in predicting HE. A mediation analysis was undertaken to determine causal associations among platelet function, HE, and outcome.Results: Of 142 patients, 37 (26.1%) suffered HE. Multivariate logistic regression identified arachidonic acid (AA) and adenosine diphosphate (ADP) inhibition as significant independent predictors of HE. The area under the ROC curves was 0.727 for AA inhibition and 0.721 for ADP inhibition. Optimal threshold for AA inhibition was 41.75% (75.7% sensitivity; 67.6% specificity) and ADP inhibition was 65.8% (73.0% sensitivity; 66.7% specificity). AA and ADP inhibition were also associated with worse 3-month outcomes after adjusting for age, admission Glasgow Coma Scale score, intraventricular hemorrhage, baseline hematoma volume, and hemoglobin. The mediation analysis showed that the effect of higher platelet inhibition with poor outcomes was mediated through HE.Conclusions: These findings suggest that the reduced platelet response to ADP and AA independently predict HE and poor outcome in patients with ICH. Platelet function may represent a modifiable target of ICH treatment.

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