The rapid and sensitive classification of bacteria is the first step of bacterial community research and the treatment of infection. Herein, a fluorescent probe BacGO is presented, which shows the ...best universal selectivity for Gram‐positive bacteria among known probes with a minimum staining procedure for sample detection and enrichment of the live bacteria. BacGO could also be used to assess of the Gram status in the bacterial community from wastewater sludge. Furthermore, BacGO could sensitively and selectively detect a Gram‐positive bacterial infection, not only in vitro but also using an in vivo keratitis mouse model. BacGO provides an unprecedented research tool for the study of dynamic bacterial communities and for clinical application.
BacGO, a novel Gram‐positive bacterial probe, was developed from a library of fluorescent molecules with a boronic‐acid motif that binds to peptidoglycan on the Gram‐positive bacterial cell wall. BacGO can be used to identify Gram‐positive bacteria in diverse, highly complex samples, and is an attractive alternative to Gram staining.
Microorganisms produce diverse polymers for various purposes such as storing genetic information, energy, and reducing power, and serving as structural materials and scaffolds. Among these polymers, ...polyhydroxyalkanoates (PHAs) are microbial polyesters synthesized and accumulated intracellularly as a storage material of carbon, energy, and reducing power under unfavorable growth conditions in the presence of excess carbon source. PHAs have attracted considerable attention for their wide range of applications in industrial and medical fields. Since the first discovery of PHA accumulating bacteria about 100 years ago, remarkable advances have been made in the understanding of PHA biosynthesis and metabolic engineering of microorganisms toward developing efficient PHA producers. Recently, nonnatural polyesters have also been synthesized by metabolically engineered microorganisms, which opened a new avenue toward sustainable production of more diverse plastics. Herein, the current state of PHAs and nonnatural polyesters is reviewed, covering mechanisms of microbial polyester biosynthesis, metabolic pathways, and enzymes involved in biosynthesis of short‐chain‐length PHAs, medium‐chain‐length PHAs, and nonnatural polyesters, especially 2‐hydroxyacid‐containing polyesters, metabolic engineering strategies to produce novel polymers and enhance production capabilities and fermentation, and downstream processing strategies for cost‐effective production of these microbial polyesters. In addition, the applications of PHAs and prospects are discussed.
Polyhydroxyalkanoates (PHAs) are biodegradable and bio‐based polymers that can substitute petroleum‐based plastics currently in use. A comprehensive overview of the mechanisms and metabolism of PHA biosynthesis, and strategies for strain development, fermentation, and downstream processing toward the cost‐effective production of natural and nonnatural polyesters having diverse material properties is provided. Additionally, applications of PHAs and future prospects are discussed.
Plastics, including poly(ethylene terephthalate) (PET), possess many desirable characteristics and thus are widely used in daily life. However, non-biodegradability, once thought to be an advantage ...offered by plastics, is causing major environmental problem. Recently, a PET-degrading bacterium, Ideonella sakaiensis, was identified and suggested for possible use in degradation and/or recycling of PET. However, the molecular mechanism of PET degradation is not known. Here we report the crystal structure of I. sakaiensis PETase (IsPETase) at 1.5 Å resolution. IsPETase has a Ser-His-Asp catalytic triad at its active site and contains an optimal substrate binding site to accommodate four monohydroxyethyl terephthalate (MHET) moieties of PET. Based on structural and site-directed mutagenesis experiments, the detailed process of PET degradation into MHET, terephthalic acid, and ethylene glycol is suggested. Moreover, other PETase candidates potentially having high PET-degrading activities are suggested based on phylogenetic tree analysis of 69 PETase-like proteins.
Abstract
Background
Detection of active pulmonary tuberculosis on chest radiographs (CRs) is critical for the diagnosis and screening of tuberculosis. An automated system may help streamline the ...tuberculosis screening process and improve diagnostic performance.
Methods
We developed a deep learning–based automatic detection (DLAD) algorithm using 54c221 normal CRs and 6768 CRs with active pulmonary tuberculosis that were labeled and annotated by 13 board-certified radiologists. The performance of DLAD was validated using 6 external multicenter, multinational datasets. To compare the performances of DLAD with physicians, an observer performance test was conducted by 15 physicians including nonradiology physicians, board-certified radiologists, and thoracic radiologists. Image-wise classification and lesion-wise localization performances were measured using area under the receiver operating characteristic (ROC) curves and area under the alternative free-response ROC curves, respectively. Sensitivities and specificities of DLAD were calculated using 2 cutoffs (high sensitivity 98% and high specificity 98%) obtained through in-house validation.
Results
DLAD demonstrated classification performance of 0.977–1.000 and localization performance of 0.973–1.000. Sensitivities and specificities for classification were 94.3%–100% and 91.1%–100% using the high-sensitivity cutoff and 84.1%–99.0% and 99.1%–100% using the high-specificity cutoff. DLAD showed significantly higher performance in both classification (0.993 vs 0.746–0.971) and localization (0.993 vs 0.664–0.925) compared to all groups of physicians.
Conclusions
Our DLAD demonstrated excellent and consistent performance in the detection of active pulmonary tuberculosis on CR, outperforming physicians, including thoracic radiologists.
A deep learning–based algorithm outperformed radiologists in detecting active pulmonary tuberculosis on chest radiographs and thus may play an important role in diagnosis and screening of tuberculosis in select situations, contributing to the reduction of the high burden of tuberculosis worldwide.
Widespread utilization of polyethylene terephthalate (PET) has caused a variety of environmental and health problems; thus, the enzymatic degradation of PET can be a promising solution. Although ...PETase from Ideonalla sakaiensis (IsPETase) has been reported to have the highest PET degradation activity under mild conditions of all PET-degrading enzymes reported to date, its low thermal stability limits its ability for efficient and practical enzymatic degradation of PET. Using the structural information on IsPETase, we developed a rational protein engineering strategy using several IsPETase variants that were screened for high thermal stability to improve PET degradation activity. In particular, the IsPETaseS121E/D186H/R280A variant, which was designed to have a stabilized β6-β7 connecting loop and extended subsite IIc, had a T m value that was increased by 8.81 °C and PET degradation activity was enhanced by 14-fold at 40 °C in comparison with IsPETaseWT. The designed structural modifications were further verified through structure determination of the variants, and high thermal stability was further confirmed by a heat-inactivation experiment. The proposed strategy and developed variants represent an important advancement for achieving the complete biodegradation of PET under mild conditions.
Herein, we investigate the effects of catholyte feed method and anode binder content on the characteristics of anion exchange membrane water electrolysis (AEMWE) to construct a high-performance ...electrolyzer, revealing that the initial AEMWE performance is significantly improved by pre-feeding 0.5 M aqueous KOH to the cathode. The highest long-term activity during repeated voltage cycling is observed for AEMWE operation in the dry cathode mode, for which the best long-term performance among membrane electrode assemblies (MEAs) featuring polytetrafluoroethylene (PTFE) binder–impregnated (5–20 wt%) anodes is detected for a PTFE content of 20 wt%. MEAs with low PTFE content (5 and 9 wt%) demonstrate high initial performance, rapid performance decay, and significant catalyst loss from the electrode during long-term operation, whereas the MEA with 20 wt% PTFE allows stable water electrolysis for over 1600 voltage cycles. Optimization of cell operating conditions (i.e., operation in dry cathode mode at an optimum anode binder content following an initial solution feed) achieves an enhanced water splitting current density (1.07 A cm−2 at 1.8 V) and stable long-term AEMWE performance (0.01% current density reduction per voltage cycle).
•Catholyte feed method and anode binder content in an MEA for AEMWE were examined.•Dry cathode operation was beneficial for long-term performance.•Optimal PTFE binder content in the anode was 20 wt% for dry cathode operation.
Houttuynia cordata (HC) is a traditional oriental herbal medicinal plant widely used as a component of complex prescriptions in Asia for alopecia treatment. The effect of HC on hair growth and its ...underlying mechanism, however, have not been demonstrated or clarified. In this study, we investigated the hair growth promoting effect of HC in cultured human dermal papilla cells (hDPCs). HC extract was found to stimulate the proliferation of hDPCs and this stimulation might be in part a consequence of activated cellular energy metabolism, because treatment of HC extract increased the generation of nicotinamide adenine dinucleotide (NADH) and ATP through increasing the mitochondrial membrane potential (ΔΨ). In the context of cell cycle, HC extract increased the expression of CDK4 and decreased the expression of CCNA2 and CCNB1, implying that HC extract might induce G1 phase progression of DPCs which resulted in enhanced proliferation. HC extract increased the expression of Bcl2 essential for maintaining hair follicle anagen stage and cell survival. On the contrary, the expression of p16 and p21 was down-regulated by HC extract. In addition, HC extract enhanced the secretion of platelet-derived growth factor (PDGF)-aa and vascular endothelial growth factor (VEGF) and induced phosphorylation of extracellular signal-regulated kinase (ERK) and AKT. Furthermore, HC extract prolonged anagen stage in organ cultured human hair follicles. Our data strongly suggest that HC extract could support hair growth by stimulating proliferation of DPCs and elongating anagen stage, resulted from enhanced cellular energy metabolism and modulation of gene expression related to cell cycle, apoptosis, and growth factors.
Interpretation of chest radiographs is a challenging task prone to errors, requiring expert readers. An automated system that can accurately classify chest radiographs may help streamline the ...clinical workflow.
To develop a deep learning-based algorithm that can classify normal and abnormal results from chest radiographs with major thoracic diseases including pulmonary malignant neoplasm, active tuberculosis, pneumonia, and pneumothorax and to validate the algorithm's performance using independent data sets.
This diagnostic study developed a deep learning-based algorithm using single-center data collected between November 1, 2016, and January 31, 2017. The algorithm was externally validated with multicenter data collected between May 1 and July 31, 2018. A total of 54 221 chest radiographs with normal findings from 47 917 individuals (21 556 men and 26 361 women; mean SD age, 51 16 years) and 35 613 chest radiographs with abnormal findings from 14 102 individuals (8373 men and 5729 women; mean SD age, 62 15 years) were used to develop the algorithm. A total of 486 chest radiographs with normal results and 529 with abnormal results (1 from each participant; 628 men and 387 women; mean SD age, 53 18 years) from 5 institutions were used for external validation. Fifteen physicians, including nonradiology physicians, board-certified radiologists, and thoracic radiologists, participated in observer performance testing. Data were analyzed in August 2018.
Deep learning-based algorithm.
Image-wise classification performances measured by area under the receiver operating characteristic curve; lesion-wise localization performances measured by area under the alternative free-response receiver operating characteristic curve.
The algorithm demonstrated a median (range) area under the curve of 0.979 (0.973-1.000) for image-wise classification and 0.972 (0.923-0.985) for lesion-wise localization; the algorithm demonstrated significantly higher performance than all 3 physician groups in both image-wise classification (0.983 vs 0.814-0.932; all P < .005) and lesion-wise localization (0.985 vs 0.781-0.907; all P < .001). Significant improvements in both image-wise classification (0.814-0.932 to 0.904-0.958; all P < .005) and lesion-wise localization (0.781-0.907 to 0.873-0.938; all P < .001) were observed in all 3 physician groups with assistance of the algorithm.
The algorithm consistently outperformed physicians, including thoracic radiologists, in the discrimination of chest radiographs with major thoracic diseases, demonstrating its potential to improve the quality and efficiency of clinical practice.
We examined the effects of empagliflozin, a selective inhibitor of Na
-glucose cotransporter 2, on mitochondrial quality control and autophagy in renal tubular cells in a diabetic environment in vivo ...and in vitro. Human renal proximal tubular cells (hRPTCs) were incubated under high-glucose conditions. Diabetes was induced with streptozotocin in male C57BL/6J mice. Improvements in mitochondrial biogenesis and balanced fusion-fission protein expression were noted in hRPTCs after treatment with empagliflozin in high-glucose media. Empagliflozin also increased autophagic activities in renal tubular cells in the high-glucose environment, which was accompanied with mammalian target of rapamycin inhibition. Moreover, reduced mitochondrial reactive oxygen species production and decreased apoptotic and fibrotic protein expression were observed in hRPTCs after treatment with empagliflozin, even in the hyperglycemic circumstance. Importantly, empagliflozin restored AMP-activated protein kinase-α phosphorylation and normalized levels of AMP-to-ATP ratios in hRPTCs subjected to a high-glucose environment, which suggests the way that empagliflozin is involved in mitochondrial quality control. Empagliflozin effectively suppressed Na
-glucose cotransporter 2 expression and ameliorated renal morphological changes in the kidneys of streptozotocin-induced diabetic mice. Electron microscopy analysis showed that mitochondrial fragmentation was decreased and 8-hydroxy-2'-deoxyguanosine content was low in renal tubular cells of empagliflozin treatment groups compared with those of the diabetic control group. We suggest one mechanism related to the renoprotective actions of empagliflozin, which reverse mitochondrial dynamics and autophagy.
The experimental animal model is still essential in the development of new anticancer drugs. We characterized mouse tumors derived from two-dimensional (2D) monolayer cells or three-dimensional (3D) ...spheroids to establish an in vivo model with highly standardized conditions. Primary cancer-associated fibroblasts (CAFs) were cultured from head and neck squamous cell carcinoma (HNSCC) tumor tissues and co-injected with monolayer cancer cells or spheroids into the oral mucosa of mice. Mice tumor blood vessels were stained, followed by tissue clearing and 3D Lightsheet fluorescent imaging. We compared the effect of exosomes secreted from 2D or 3D culture conditions on the angiogenesis-related genes in HNSCC cells. Our results showed that both the cells and spheroids co-injected with primary CAFs formed tumors. Interestingly, vasculature was abundantly distributed inside the spheroid-derived but not the monolayer-derived mice tumors. In addition, cisplatin injection more significantly decreased spheroid-derived but not monolayer-derived tumor size in mice. Additionally, exosomes isolated from co-culture media of FaDu spheroid and CAF upregulated angiogenesis-related genes in HNSCC cells as compared to exosomes from FaDu cell and CAF co-culture media under in vitro conditions. The mouse tumor xenograft model derived from 3D spheroids of HNSCC cells with primary CAFs is expected to produce reliable chemotherapy drug screening results given the robust angiogenesis and lack of necrosis inside tumor tissues.
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