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Previous studies have demonstrated that the levels of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide (NO) synthesis, are strongly associated with hypertension, diabetes, and cardiovascular diseases. Profilin‐1, an actin‐binding protein, has been documented to be involved in endothelial injury and in the proliferation of vascular smooth muscle cells resulting from hypertension. However, the role of profilin‐1 in ADMA‐induced vascular injury in hypertension remains largely unknown. Forty healthy subjects and forty‐two matched patients with essential hypertension were enrolled, and the related indexes of vascular injury in plasma were detected. Rat aortic smooth muscle cells (RASMCs) were treated with different concentrations of ADMA for different periods of time and transfected with profilin‐1 small hairpin RNA to interrupt the expression of profilin‐1. To determine the role of the Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) pathway, RASMCs were pretreated with AG490 or rapamycin. The expression of profilin‐1 was tested using real‐time polymerase chain reaction (PCR) and western blot analysis. Cell proliferation was measured by flow cytometry and 3‐(4,5‐dimethylthiazol‐2yl)‐2,5‐diphenyltetrazoliumbromide assays. Compared with healthy subjects, the levels of ADMA and profilin‐1 were markedly elevated in hypertensive individuals, while the levels of NO were significantly decreased (p

The steviol glycosides (SGs) in stevia (Stevia rebaudiana Bertoni) leaves are becoming increasingly valuable due to its high sweetness but low calorific value, which is driving the development of stevia commercial cultivation. Optimizing fertilization management can effectively increase SGs productivity, but knowledge on the relationship between potassium (K) fertilization and SGs production is still lacking. In this study, pot experiments were conducted in order to investigate the effect of K deficiency on SGs synthesis in stevia leaves, as well as the underlying mechanisms. Our results showed that when compared with standard K fertilization, K deficiency treatment has no significant effect on the biomass of stevia plant grown in a given soil with high K contents. However, K deficiency critically decreased leaf SGs contents as well as the expression of SGs synthesis-related genes. The contents of different sugar components decreased and the activities of sugar metabolism-related enzymes were inhibited under the K deficiency condition. Moreover, spraying sucrose on the leaves of stevia seedlings diminished the inhibitory effect caused by K deficiency. Our results also revealed the significant positive correlations between sucrose, glucose and SGs contents. Overall, our results suggest that K deficiency would suppress the synthesis of SGs in stevia leaves, and this effect may be mediated by the leaf sugar metabolism. Our findings provide new insights into the improvement of SGs production potential.

Background Nova Circular RNA (circRNA) of non-coding RNA has gradually become an important regulatory factor, and it has made people attach great concern over the occurrence and development of many diseases, particularly carcinomas. circ_RPPH1 is a newly discovered circRNA. Gene Expression Omnibus (GEO) analysis showed that there are high contents of circ_RPPH1 in breast cancer (BC), but the mechanism of circRNA in BC remains unclear. Methods Real-time quantitative PCR (qRT-PCR) was applied to test the role of circ_RPPH1 in BC patients, and functional experiments were applied to test the role of circ_RPPH1 on BC tumor. Fluorescence in situ hybridization, double luciferase reporter gene analysis, RNA pull-down and RNA immunoprecipitation experiments were performed to explore the correlation of circ_RPPH1 with miR-146b-3p/E2F2 in BC. Results circ_RPPH1 was evidently enhanced in BC, and its content was related to the clinical stage and pathological grade. circ_RPPH1 can accelerate the proliferation, migration and invasion, and promote tumorigenesis and metastasis. Mechanism exploration indicated that circ_RPPH1 acted as ceRNA (competing endogenous RNA) of miR-146b-3p, so as to reduce the inhibitory role of miR-146b-3p on its target E2F2. Conclusion Circ_RPPH1/miR-146b-3p/E2F2 axis can promote the progression of BC, and it might be a latent therapeutic target for clinical BC.

Species interactions and mechanisms affect plant coexistence and community assembly. Despite increasing knowledge of kin recognition and allelopathy in regulating interspecific and intraspecific interactions among plants, little is known about whether kin recognition mediates allelopathic interference. We used allelopathic rice cultivars with the ability for kin recognition grown in kin vs. non-kin mixtures to determine their impacts on paddy weeds in field trials and a series of controlled experiments. We experimentally tested potential mechanisms of the interaction via altered root behavior, allelochemical production, and soil microbial community composition, as well as carbon and nitrogen partitioning in the weeds. We consistently found that the establishment and growth of paddy weeds were more inhibited by kin mixtures compared to non-kin mixtures. The effect was driven by kin recognition that induced altered root placement, established similar soil microbial communities, and altered weed carbon and nitrogen partitioning. Importantly, genetic relatedness enhanced the production of intrusive roots towards weeds and reduced the production of rice allelochemicals. These findings suggest that relatedness allows allelopathic plants to discriminate their neighboring collaborators (kin) or competitors and then adjust their growth, competitiveness and chemical defense accordingly.

To screen for specific transcription factors (TFs) that induce expression of the HMGB1 promoter in response to stimulation by Ang-II. A HMGB1 overexpressing vector and small interfering (si)RNA were constructed and used to transfect the three HCC cell lines used in scratched monolayer wound healing and Transwell assays. Chromatin immunoprecipitation (ChIP) assays were used to confirm the relationship between a specific TF and the HMGB1 promoter. Invasion and migration by HMGB1 overexpressing HCC cells after treatment with Ang-II were significantly increased compared to negative controls (NC); E-cadherin was down-regulated while vimentin was up-regulated. However, compared with NC, invasion and migration by HMGB1 siRNA HCC cells stimulated by Ang-II were not altered; the expression of E-cadherin and vimentin was also unaltered. Nineteen TFs were predicted by Promoter 2.0 Prediction Server and TFsitescan. Real-time qPCR was used to evaluate TF expression levels. E4F1 was the only TF abnormally elevated in all three HCC cell lines when stimulated by Ang-II. WB and ChIP assays revealed high expression of E4F1 compared to other TFs in cells stimulated by Ang-II. E4F1 is activated by Ang-II and binds to the HMGB1 promoter region to promote HMGB1 expression; it then enhances Ang-II to induce HCC cell invasion and migration, and EMT.

It has been hypothesized that the human brain has less redundancy than animals, but the structural evidence has not been identified to confirm this claim. Here, we report three redundancy circuits of the commissural pathways in primate brains, namely the orbitofrontal, temporal, and occipital redundancy circuits of the anterior commissure and corpus callosum. Each redundancy circuit has two distinctly separated routes connecting a common pair of cortical regions. We mapped their trajectories in human and rhesus macaque brains using individual and population‐averaged tractography. The dissection results confirmed the existence of these redundancy circuits connecting the orbitofrontal lobe, amygdala, and visual cortex. The volume analysis showed a significant reduction in the orbitofrontal and occipital redundancy circuits of the human brain, whereas the temporal redundancy circuit had a substantial organizational difference between the human and rhesus macaque. Our results support the hypothesis that the human brain has less redundancy in the commissural pathways than that of the rhesus macaque brain. Further studies are needed to explore its neuropathological implications., It has been hypothesized that the human brain has less redundancy than animals, but the structural evidence has not been identified to confirm this claim. Here, we report three redundancy circuits of the commissural pathways in primate brains, namely the orbitofrontal, temporal, and occipital redundancy circuits of the anterior commissure and corpus callosum. Each redundancy circuit has two distinctly separated routes connecting a common pair of cortical regions. Our results support the hypothesis that the human brain has less redundancy in the commissural pathways than that of the rhesus macaque brain.

Flowering time, a key transition point from vegetative to reproductive growth, is regulated by an intrinsic complex of endogenous and exogenous signals including nutrient status. For hundreds of years, nitrogen has been well known to modulate flowering time, but the molecular genetic basis on how plants adapt to ever-changing nitrogen availability remains not fully explored. Here we explore how Arabidopsis natural variation in flowering time responds to nitrate fluctuation. Upon nitrate availability change, we detect accession- and photoperiod-specific flowering responses, which also feature a accession-specific dependency on growth traits. The flowering time variation correlates well with the expression of floral integrators, SOC1 and FT, in an accession-specific manner. We find that gene expression variation of key hub genes in the photoperiod-circadian-clock (GI), aging (SPLs) and autonomous (FLC) pathways associates with the expression change of these integrators, hence flowering time variation. Our results thus shed light on the molecular genetic mechanisms on regulation of accession- and photoperiod-specific flowering time variation in response to nitrate availability.

Major depressive disorder (MDD) is associated with reductions in white matter microstructural integrity as measured by fractional anisotropy (FA), an index derived from diffusion tensor imaging (DTI). The neurotropic herpesvirus, human cytomegalovirus (HCMV), is a major cause of white matter pathology in immunosuppressed populations but its relationship with FA has never been tested in MDD despite the presence of inflammation and weakened antiviral immunity in a subset of depressed patients. We tested the relationship between FA and HCMV infection in two independent samples consisting of 176 individuals with MDD and 44 healthy controls (HC) (Discovery sample) and 88 participants with MDD and 48 HCs (Replication sample). Equal numbers of HCMV positive (HCMV+) and HCMV negative (HCMV−) groups within each sample were balanced on ten different clinical/demographic variables using propensity score matching. Anti-HCMV IgG antibodies were measured using a solid-phase ELISA. In the Discovery sample, significantly lower FA was observed in the right inferior fronto-occipital fasciculus (IFOF) in HCMV+ participants with MDD compared to HCMV− participants with MDD (cluster size 1316 mm3; pFWE d = −0.58). This association was confirmed in the replication sample by extracting the mean FA from this exact cluster and applying the identical statistical model (p d = −0.45). There was no significant effect of diagnosis or interaction between diagnosis and HCMV in either sample. The effect of chronic HCMV infection on white matter integrity may—in at-risk individuals—contribute to the psychopathology of depression. These findings may provide a novel target of intervention for a subgroup of patients with MDD.

For active targeting nanodrug delivery systems conjugated with antibodies, both lack of control of the antibody at the molecular level and protein corona formation remarkably decreases targeting efficacy. Herein, we designed a series of silica nanoparticles toward HER2-positive breast cancer cells, with an anti-HER2 Fab-6His density ranging from 50 to 180 molecules per nanoparticle. Through the site-directed immobilization method we developed, the antigen-binding domain of anti-HER2 Fab was mostly accessible to the HER2 receptor. Both polyethylene glycol (PEG) chains and a high density of Fab were shown to suppress protein corona formation and macrophage uptake. The dependency of targeting efficacy and cytotoxicity on Fab density was shown using a series of breast cancer cell lines with different levels of the HER2 expression. The high density of Fab stimulates quick responses from HER2-positive cells. Combined with PEG chains, conjugated antibodies with a well-controlled orientation and density significantly improves delivery performance and sheds light on the design and preparation of an improved active targeting nanodrug delivery system.

Organ transplantation has been employed upon serious injuries, but a T-cell-mediated potent inflammatory immune response often leads to graft rejection. Immunosuppressive drugs such as rapamycin (RAPA) have to be taken after organ transplantation, but long-term use of these drugs causes severe adverse effects. Immune checkpoint pathways such as the programmed death-receptor 1/programmed death-ligand 1 (PD-1/PD-L1) provides an immunosuppressive environment, preventing excessive tissue destruction due to inflammatory immune responses. In this study, we bioengineered cell membrane-derived PD-L1 nanovesicles (PD-L1 NVs) to carry low doses of RAPA. These NVs inhibited T-cell activation and proliferation in vitro, by enhancing the PD-1/PD-L1 immune co-inhibitory signaling axis and inhibiting the mTOR pathway. Importantly, PD-L1 NVs encapsulated with rapamycin exerted stronger effects on inhibiting T-cell proliferation than PD-L1 NVs or rapamycin alone. This can be recapitulated in a mouse skin transplantation model, leading to the weakened alloimmune response and allograft tolerance. We also found that PD-L1/rapamycin vesicles have additional function to induce regulatory T cells in the recipient spleens. Our study highlighted the power of combining low-dose rapamycin and PD-L1 in the nanovesicles as immunosuppressants to promote allograft acceptance.

Adipose stem cell-derived exosomes have great potential in accelerating cutaneous wound healing by optimizing fibroblast activities. Recent studies have demonstrated that exosomes play an active role in the transport of functional cytoskeletal proteins such as vimentin. Previously we showed that vimentin serves as a coordinator of the healing process. Therefore, we hypothesized that vimentin incorporated into the exosomes may contribute to mediate fibroblast activities in wound healing. Our results revealed that exosomal vimentin from adipocyte progenitor cells acts as a promoter of fibroblast proliferation, migration, and ECM secretion. Furthermore, our in vitro and in vivo experiments provide evidence that exosomal vimentin shortens the healing time and reduces scar formation. These findings suggest the reciprocal roles of exosomes and vimentin in accelerating wound healing. Exosomes can serve as an efficient transportation system to deliver and internalize vimentin into target cells, while vimentin could have an impact on exosome transportation, internalization, and cell communication. This article is protected by copyright. All rights reserved.

Continuous cropping obstacles seriously affect the sustainable production of tomatoes (Solanum lycopersicum L.). Researchers have found that intercropping with garlic (Allium sativum L.) could alleviate tomato continuous cropping obstacles. Diallyl disulfide (DADS) is the main allelochemical in garlic. However, the mechanism of DADS in alleviating tomato continuous cropping obstacles is still unknown. In this research, aqueous extracts of tomato continuous cropping soil were used to simulate the continuous cropping condition of tomato. Our results showed that DADS increased root activity and chlorophyll content and improved the activity of antioxidant enzymes (superoxide dismutase (SOD), peroxidase (POD), and phenylalanine ammonia-lyase (PAL)) and the metabolism of nonenzymatic antioxidants (glutathione (GSH) and oxidized glutathione (GSSG)) in tomato plants. DADS treatment reduced the content of fatty acid esters in tomato root exudates (e.g., palmitate methyl ester, palmitoleic acid methyl ester, oleic acid methyl ester) and increased the level of substances such as dibutyl phthalate and 2,2'-methylenebis(6-tert-butyl-4-methylphenol). The higher concentrations of palmitate methyl ester inhibited tomato hypocotyl growth, while oleic acid methyl ester inhibited tomato root growth. Moreover, the application of DADS significantly inhibited the secretion of these esters in the root exudates. Therefore, it suggests that DADS may increase tomato resistance and promote tomato plant growth by increasing root activity and photosynthetic capacity and development to reduce autotoxicity of tomato.

Studies have confirmed that Angelica sinensis, which is a famous medicinal food in China, can effectively alleviate the symptoms of ulcerative colitis (UC) in rats. However, as the major water-soluble ingredient, the specific effects of A. sinensis polysaccharide (ASP) on UC and potential mechanisms were uncertain. In this study, we aimed to elucidate the protective effects of ASP on dextran sulfate sodium (DSS)-induced UC and to further explore the mechanisms. ASP could significantly ameliorate the symptoms of weight loss, disease activity index score, and colon shortening caused by DSS. ASP treatment also significantly suppressed the myeloperoxidase activity in colon tissues. Furthermore, after ASP administration, the expression of the proinflammatory cytokines (interleukin [IL]-6, IL-1β, and tumor necrosis factor alpha) induced by DSS was remarkably suppressed, and there was a definite improvement in the expressions of tight junction proteins, such as zona occludens 1, occludin, and claudin-1. In addition, the results of apoptosis experiments showed that the apoptotic events were noticeably reduced after ASP treatment. Taken together, these results suggested that ASP may be a potential natural agent against UC.

Switchgrass (Panicum virgatum), a C4 perennial grass native to North America and developed as a sustainable biofuel feedstock, occurs in two ecotypes, lowland and upland, which vary in their architecture as well as their range of adaptation. In this study, we assessed single nucleotide polymorphism (SNP) variation in 372 switchgrass genotypes for nine genes involved in lignin and pectin biosynthesis. STRUCTURE results at K = 3 differentiated the genotypes into three genetic subpopulations that corresponded largely to the previously characterized upland C1 and lowland C2 and C3 subpopulations. Out of the 146 SNPs identified, 19 SNPs were non-synonymous, including two non-conservative and common SNPs in cinnamyl alcohol dehydrogenase (CAD, Chr01N) and p-coumarate3-hydroxylase (C3H, Chr09N), two genes in the lignin biosynthesis pathway. Allele status at seven of the 19 non-synonymous SNPs, including the non-conservative SNP in C3H, was significantly associated with four dry matter traits within subpopulations. Dry matter traits appeared to be mostly dominant and three of them (acid detergent fiber, non-fiber carbohydrate, water-soluble carbohydrates) were the most frequently differentiated traits. In addition, a heterosis effect was detected at the non-conservative SNP in phenylalanine ammonia-lyase (PAL) for neutral detergent fiber. Association analysis revealed the CAD gene on Chr01N, its homoeolog on Chr01K and cinnamate 4-hydroxylase (C4H, Chr03K) as potential candidates associated with dry matter traits. Further analyses are needed to determine whether these candidate genes play a role in switchgrass lignin content and could be exploited to reduce recalcitrance in this bioenergy crop.

Fatty alcohols (FOHs) are important feedstocks in the chemical industry to produce detergents, cosmetics, and lubricants. Microbial production of FOHs has become an attractive alternative to production in plants and animals due to growing energy demands and environmental concerns. However, inhibition of cell growth caused by intracellular FOH accumulation is one major issue that limits FOH titers in microbial hosts. In addition, identification of FOH‐specific exporters remains a challenge and previous studies towards this end are limited. To alleviate the toxicity issue, we exploited nonionic surfactants to promote the export of FOHs in Rhodosporidium toruloides, an oleaginous yeast that is considered an attractive next‐generation host for the production of fatty acid‐derived chemicals. Our results showed FOH export efficiency was dramatically improved and the growth inhibition was alleviated in the presence of small amounts of tergitol and other surfactants. As a result, FOH titers increase by 4.3‐fold at bench scale to 352.6 mg/L. With further process optimization in a 2‐L bioreactor, the titer was further increased to 1.6 g/L. The method we show here can potentially be applied to other microbial hosts and may facilitate the commercialization of microbial FOH production., Microbial production of fatty alcohols has gained broad interests, however one major challenge for commercialization of this technology is inhibition of cell growth caused by intracellular fatty alcohol accumulation. This study develops a novel strategy that exploits nonionic surfactants to promote the export of fatty alcohols from R. toruloides, an oleaginous yeast that is considered an attractive host for fatty acid‐derived chemicals. The method significantly alleviated growth inhibition and enhanced fatty alcohol titers and yields.

Astragalus membranaceus is the most popular traditional Chinese medicine for managing vital energy deficiency. Its injectable polysaccharide PG2 has been used for relieving cancer-related fatigue, and PG2 has immune-modulatory and anti-inflammatory effects. In this study, we explored the effects of PG2 in lung adenocarcinoma A549 and CL1-2 cells and investigated its anticancer activity, and the results were validated in severe combined immunodeficiency (SCID) mice. Although PG2 did not inhibit the growth of these cells, it dose-dependently suppressed their migration and invasion, accompanied by reduced vimentin and AXL and induced epithelial cadherin (E-cadherin) expression. Regarding the underlying molecular mechanism, PG2 treatment reduced the macrophage migration inhibitory factor (MIF), an inflammatory cytokine that promotes the epithelial–mesenchymal transition and aggressiveness of cancer cells. Consistent with the previous finding that MIF regulates matrix metalloproteinase-13 (MMP-13) and AMP-activated protein kinase (AMPK), treatment with PG2 reduced MMP-13 and activated AMPK in A549 and CL1-2 cells in this study. In SCID mice injected with A549 cells through the tail vein, intraperitoneal injection with PG2 reduced lung and abdominal metastases in parallel with decreased immunohistochemical staining of AXL, vimentin, MMP-13, and MIF in the tumor. Collectively, data revealed a potential application of PG2 in integrative cancer treatment through the suppression of MIF in cancer cells and their aggressiveness.

The development of a universal coating strategy for the construction of functional surfaces and modulation of surface properties is of great research interest. Tannic acid (TA) could serve as a sole precursor for the deposition of colorless coatings on substrate surfaces. However, the deposition of TA requires a high salt concentration (0.6 M), which may limit its practical application. Herein, primary amine moieties were introduced on the gallic acid groups in TA. The resultant amine-containing TA derivative (TAA) can self-polymerize under mild conditions (10 mM, Tris buffer), and form uniform and colorless coatings in a material-independent manner. In comparison with the TA coating under the same preparation conditions, the TAA coating exhibits an increased thickness as measured by ellipsometry. The TAA coating is adapted for secondary surface functionalization. The hydrophilic mPEG brushes can be grafted on the TAA coating to inhibit non-specific protein adsorption. A biotin probe can be immobilized on the TAA coating to promote specific binding with avidin. In addition, the TAA coating can be utilized for in situ reduction of silver ions to AgNPs. The resulting AgNP-loaded TAA coating can inhibit bacterial adhesion and prevent biofilm formation.

Cellular membranes contain many lipids, some of which, such as sphingolipids, have important structural and signaling functions. The common sphingolipid glucosylceramide (GlcCer) is present in plants, fungi, and animals. As a major plant sphingolipid, GlcCer is involved in the formation of lipid microdomains, and the regulation of GlcCer is key for acclimation to stress. Although the GlcCer biosynthetic pathway has been elucidated, little is known about GlcCer catabolism, and a plant GlcCer-degrading enzyme (glucosylceramidase (GCD)) has yet to be identified. Here, we identified AtGCD3, one of four Arabidopsis thaliana homologs of human nonlysosomal glucosylceramidase, as a plant GCD. We found that recombinant AtGCD3 has a low Km for the fluorescent lipid C6-NBD GlcCer and preferentially hydrolyzes long acyl-chain GlcCer purified from Arabidopsis leaves. Testing of inhibitors of mammalian glucosylceramidases revealed that a specific inhibitor of human β-glucosidase 2, N-butyldeoxynojirimycin, inhibits AtGCD3 more effectively than does a specific inhibitor of human β-glucosidase 1, conduritol β-epoxide. We also found that Glu-499 and Asp-647 in AtGCD3 are vital for GCD activity. GFP-AtGCD3 fusion proteins mainly localized to the plasma membrane or the endoplasmic reticulum membrane. No obvious growth defects or changes in sphingolipid contents were observed in gcd3 mutants. Our results indicate that AtGCD3 is a plant glucosylceramidase that participates in GlcCer catabolism by preferentially hydrolyzing long-acyl-chain GlcCers.

BackgroundVarious bacterial putative virulence factors are involved in the pathogenesis of bacterial infection. However, the effect of comorbidities or infection syndrome in the association of virulence factors and mortality remains inconclusive.MethodThis study addressed whether specific sequence type (ST) and virulence factors of extended-spectrum beta-lactamase–producingEscherichia coli(ESBL-EC) are associated with different outcomes in patients with bloodstream infection.121 adults from southern Taiwan with ESBL-producingE. colibloodstream infections were enrolled during a 6-year period. Demographic data, including infection syndromes, underlying disease and outcomes, were collected. The virulence factors in isolates were analyzed by PCR and multilocus sequence typing.ResultPositivity for the virulence genes iha,hlyD,sat,iut,fyu,malX,ompT,uspandtraTwas associated with ST131 positivity (PiroNandisswas significantly associated with increased 30-day mortality (death within 30 days) on univariate analysis (PiroNpositivity.ConclusionIn bacteremic patients with UTI, and the ST131 clone was borderline associated with mortality. Positivity for the virulence geneiroNmay be linked to mortality in bacteremic patients without UTI.

Porcine circovirus type 3 (PCV3), a virus belonging to the Circoviridae family, is considered to be associated with respiratory and neurological signs, cardiac and multisystemic inflammation, reproductive failure, and porcine dermatitis and nephropathy syndrome-like disease in pigs (Sus scrofa). In this study, epidemiological and serological investigations of PCV3 in clinically healthy pigs from different regions of China were performed. Overall, 42.87% (1,101/2,568) of pigs were positive for PCV3 Cap antibody via indirect enzyme-linked immunosorbent assay, with a higher prevalence of PCV3 in multiparous sows (62.22%, 881/1,416) and fattening pigs (28.96%, 159/549) than in suckling piglets (8.96%, 32/357) and nursery pigs (11.79%, 29/246). Of the 2,568 samples, 255 were further tested for PCV3 DNA using real-time polymerase chain reaction, and 63.14% of these were positive, with nearly half having

Colistin is the last resort antimicrobial for treating multidrug-resistant gram-negative bacterial infections. The plasmid-mediated colistin resistance gene, mcr-1, crucially influences colistin’s resistance transmission. Human fecal carriages of mcr-1-positive Escherichia coli (E. coli) were detected in many regions worldwide; however, only a few studies have focused on children. Therefore, we identified the prevalence and risk factors of mcr-1-positive E. coli in fecal carriages among community children in Southern Taiwan. In this study, 510 stool samples were collected from April 2016 to August 2019 from the pediatric department at a medical center in Southern Taiwan. These samples were collected within 3 days after admission and were all screened for the presence of the mcr-1 gene. Diet habits, travel history, pet contact, and medical history were also obtained from participants to analyze the risk factors of their fecal carriages to mcr-1-positive E. coli. Antimicrobial susceptibility testing was determined using the VITEK 2 system and the broth microdilution test. Twelve mcr-1-positive E. coli. were isolated from 2.4% of the fecal samples. Through multivariate analysis, frequent chicken consumption (at least 3 times per week) had a significantly positive association with the presence of mcr-1-positive E. coli in fecal carriages (adjust odds ratio 6.60, 95% confidence interval1.58– 27.62, p = 0.033). Additionally, multidrug resistance was more common in mcr-1-positive E. coli. (75.0% vs. 39.5%, p = 0.031) than in non-mcr-1-positive Escherichia coli. Furthermore, the percentage of extraintestinal pathogenic E. coli in mcr-1-positive isolates was 83.3%. Some multi-locus sequence types in our mcr-1-positive E. coli were also similar to those isolated from food animals in the literature. The prevalence of fecal carriages of mcr-1-positive E. coli was low among community children in Southern Taiwan. Our data shows that chicken consumption with a higher frequency increases the risk of mcr-1-positive E. coli. in fecal carriages.

Tigecycline serves as one of the last-resort antibiotics to treat severe infections caused by carbapenem-resistant Enterobacterales. Recently, a novel plasmid-mediated resistance-nodulation-division (RND)-type efflux pump gene cluster, TmexCD1-ToprJ1, and its variants, TmexCD2-ToprJ2 and TmexCD3-ToprJ3, encoding tetracyclines and tigecycline resistance, were revealed. In this study, we reported three TmexCD2-ToprJ2-harboring Klebsiella species strains, collected from two teaching tertiary hospitals in China, including one K. quasipneumoniae, one K. variicola, and one K. michiganensis. The three strains were characterized by antimicrobial susceptibility testing (AST), conjugation assay, WGS, and bioinformatics analysis. AST showed that K. variicola and K. quasipneumoniae strains were resistant to tigecycline with MIC values of 4μg/ml, whereas the K. michiganensis was susceptible to tigecycline with an MIC value of 1μg/ml. The TmexCD2-ToprJ2 clusters were located on three similar IncHI1B plasmids, of which two co-harbored the metallo-β-lactamase gene blaNDM-1. Conjugation experiments showed that all three plasmids were capable of self-transfer via conjugation. Our results showed, for the first time, that this novel plasmid-mediated tigecycline resistance mechanism TmexCD2-ToprJ2 has spread into different Klebsiella species, and clinical susceptibility testing may fail to detect. The co-occurrence of blaNDM-1 and TmexCD2-ToprJ2 in the same plasmid is of particular public health concern as the convergence of “mosaic” plasmids can confer both tigecycline and carbapenem resistance. Its further spread into other clinical high-risk Klebsiella clones will likely exacerbate the antimicrobial resistance crisis. A close monitoring of the dissemination of TmexCD-ToprJ encoding resistance should be considered.

Color is an essential agronomic trait and the consumption of high anthocyanin containing vegetables in daily diet does provide benefits to human health, but the mechanisms on anthocyanin accumulation in tender pods of okra (Abelmoschus esculentus L.) were totally unknown. In this study, a wide characterization and quantitation of anthocyanins and flavonols in tender pods of 15 okra varieties were performed by UHPLC-Q-Orbitrap HRMS for the first time. Two major anthocyanins (delphinidin 3-O-sambubioside and cyanidin 3-O-sambubioside) and six kinds of flavonol glycosides (most are quercetin-based) were identified and quantified. The coloration of the purple okra pod mainly arises from the accumulation of both delphinidin 3-O-sambubioside and cyanidin 3-O-sambubioside in most of purple varieties (Hong Yu, Bowling Red and Burgundy), except Jing Orange. The significant differences in the compositions and contents of anthocyanins are responsible for the pod color ranging from brick-red to purplish-red among the various okra cultivars. Furthermore, four representative okra cultivars exhibiting obvious differences in anthocyanin accumulation were further analyzed with transcriptome and more than 4000 conserved differentially expressed genes were identified across the three compared groups (B vs. BR, B vs. HY and B vs. JO). Based on the comprehensive analysis of transcriptomic data, it was indicated that MBW complex consisting of AeMYB114, AeTT8, and AeTTG1 and other transcriptional factors coordinately regulate the accumulation of anthocyanins via the transcriptional regulation of structural genes. Moreover, four independent working models explaining the diversities of anthocyanin pigmentation in okra pods were also proposed. Altogether, these results improved our understanding on anthocyanin accumulation in okra pods, and provided strong supports for the development of okra pod as a functional food in the future.

The present study aimed to discuss the effects and relative mechanisms of NEDD4 E3 ubiquitin protein ligase (NEDD4) in cutaneous melanoma (CMM) occurrence and development. Clinical cancer and adjacent normal tissues samples were collected to analyze pathological changes and protein expression of NEDD4. Moreover, small interfering (si)RNA was used to knockdown NEDD4 expression in SK-MEL-2 and Malme-3M cells. Cellular proliferation, apoptosis, invasiveness and migration were examined using colony formation, flow cytometric, Transwell and wound-healing assays, respectively. In addition, the relative mRNA and protein expression levels of NEDD4, notch receptor 1 (Notch1) and PTEN were evaluated via reverse transcription-quantitative (RT-q) PCR and western blotting. It was found that NEDD4 mRNA and protein expression were significantly upregulated (both P

BackgroundEpigenetic changes of lung adenocarcinoma (LUAD) have been reported to be a relevant factor in tumorigenesis and cancer progression. However, the molecular mechanisms responsible for DNA methylation patterns in the tumor immune-infiltrating microenvironment and in cancer immunotherapy remain unclear.MethodsWe conducted a global analysis of the DNA methylation modification pattern (DMP) and immune cell-infiltrating characteristics of LUAD patients based on 21 DNA methylation regulators. A DNA methylation score (DMS) system was constructed to quantify the DMP model in each patient and estimate their potential benefit from immunotherapy.ResultsTwo DNA methylation modification patterns able to distinctly characterize the immune microenvironment characterization were identified among 513 LUAD samples. A lower DMS, characterized by increased CTLA-4/PD-1/L1 gene expression, greater methylation modifications, and tumor mutation burden, characterized a noninflamed phenotype with worse survival. A higher DMS, characterized by decreased methylation modification, a greater stromal-relevant response, and immune hyperactivation, characterized an inflamed phenotype with better prognosis. Moreover, a lower DMS indicated an increased mutation load and exhibited a poor immunotherapeutic response in the anti-CTLA-4/PD-1/PD-L1 cohort.ConclusionEvaluating the DNA methylation modification pattern of LUAD patients could enhance our understanding of the features of tumor microenvironment characterization and may promote more favorable immunotherapy strategies.

The phenotypic plasticity of Schwann cells (SCs) has contributed to the regenerative potential of the peripheral nervous system (PNS), but also pathological processes. This double-sided effect has led to an increasing attention to the role of extracellular vesicles (EVs) or exosomes in SCs to examine the intercellular communication between SCs and their surroundings. Here, we first describe the current knowledge of SC and EV biology, which forms the basis for the updates on advances in SC-derived exosomes research. We seek to explore in-depth the exosome-mediated molecular mechanisms involved in the regulation of SCs and their microenvironment. This review concludes with potential applications of SC-derived exosomes as delivery vehicles for therapeutics and biomarkers. The goal of this review is to emphasize the crucial role of SC-derived exosomes in the functional integration of the PNS, highlighting an emerging area in which there is much to explore and re-explore.

Obesity and its related metabolic diseases have become great public health threats worldwide. Although accumulated evidence suggests that circRNA is a new type of non-coding RNAs regulating various physiological and pathological processes, little attention has been paid to the expression profiles and functions of circRNAs in white adipose tissue. In this study, 3,771 circRNAs were detected in three stages of white adipogenesis (preadipocyte, differentiating preadipocyte, and mature adipocyte) by RNA-seq. Experimental validation suggested that the RNA-seq results are highly reliable. We found that nearly 10% of genes which expressed linear RNAs in adipocytes could also generate circRNAs. In addition, 40% of them produced multiple circRNA isoforms. We performed correlation analysis and found that a great deal of circRNAs (nearly 50%) and their parental genes were highly correlated in expression levels. A total of 41 differential expression circRNAs (DECs) were detected during adipogenesis and an extremely high ratio of them (80%) were correlated with their parental genes, indicating these circRNAs may potentially play roles in regulating the expression of their parental genes. KEGG enrichment and GO annotation of the parental genes suggesting that the DECs may participate in several adipogenesis-related pathways. Following rigorous selection, we found that many up-regulated circRNAs contain multiple miRNAs binding sites, such as miR17, miR-30c, and miR-130, indicating they may potentially facilitate their regulatory functions by acting as miRNA sponges. These results suggest that plenty of circRNAs are expressed in white adipogenesis and the DECs may serve as new candidates for future adipogenesis regulation.

Objective: To analyze the correlation between serum uric acid, prealbumin levels, lactate dehydrogenase (LDH), and the severity of COVID-19.Methods: The data from 135 patients with COVID-19 was collected, and the patients were divided into a non-severe group (110 cases) and a severe group (25 cases), according to the severity of illness. Sixty cases with normal physical examinations over the same period and 17 cases diagnosed with other viral pneumonia in the past five years were selected as the control group to analyze the correlation between the detection index and the severity of COVID-19.Results: Serum albumin and prealbumin in the severe group were significantly lower than those in the non-severe group (p < 0.01); serum uric acid in the severe group was lower than that in the non-severe group (p < 0.05). LDH and C-reaction protein (CRP) in the severe group were higher than those in non-severe group (p < 0.01); the levels of albumin, prealbumin, serum uric acid, and LDH in the severe group were significantly different from those in healthy control group (p < 0.01) and the levels of prealbumin, serum uric acid, LDH, and CRP in the severe group were significantly different from those in the other viral pneumonia group (p < 0.01). Serum albumin and prealbumin were positively correlated with the oxygenation index (p < 0.001), while LDH was negatively correlated with oxygenation index (p < 0.001).Conclusion: Serum albumin, prealbumin, the oxygenation index, and LDH are risk factors of COVID-19.

Mutations in the plant homeodomain-like finger protein 6 (PHF6) gene are strongly associated with acute myeloid (AML) and T-cell acute lymphoblastic leukemia (T-ALL). In this study, we demonstrated that PHF6 can bind to H3K9me3 and H3K27me1 on the nucleolar chromatin and recruit histone methyltransferase SUV39H1 to the rDNA locus. The deletion of PHF6 caused a decrease in the recruitment of SUV39H1 to rDNA gene loci, resulting in a reduction in the level of H3K9me3 and the promotion of rDNA transcription. The knockdown of either SUV39H1 or PHF6 significantly attenuated the effects of increase in H3K9me3 and suppressed the transcription of rDNA induced by the overexpression of the other interacting partner, thereby establishing an interdependent relationship between PHF6 and SUV39H1 in their control of rRNA transcription. The PHF6 clinical mutants significantly impaired the ability to bind and recruit SUV39H1 to the rDNA loci, resulting in an increase in rDNA transcription activity, the proliferation of in vitro leukemia cells, and the growth of in vivo mouse xenografts. Importantly, significantly elevated levels of pre-rRNA were observed in clinical AML patients who possessed a mutated version of PHF6. The specific rDNA transcription inhibitor CX5461 significantly reduced the resistance of U937 AML cells deficient in PHF6 to cytarabine, the drug that is most commonly used to treat AML. Collectively, we revealed a novel molecular mechanism by which PHF6 recruits methyltransferase SUV39H1 to the nucleolar region in leukemia and provided a potential therapeutic target for PHF6-mutant leukemia.

There is an urgent need for developing new immunosuppressive agents due to the toxicity of long‐term use of broad immunosuppressive agents after organ transplantation. Comprehensive sample analysis revealed dysregulation of FGL1/LAG‐3 and PD‐L1/PD‐1 immune checkpoints in allogeneic heart transplantation mice and clinical kidney transplant patients. In order to enhance these two immunosuppressive signal axes, a bioengineering strategy is developed to simultaneously display FGL1/PD‐L1 (FP) on the surface of small extracellular vesicles (sEVs). Among various cell sources, FP sEVs derived from mesenchymal stem cells (MSCs) not only enriches FGL1/PD‐L1 expression but also maintain the immunomodulatory properties of unmodified MSC sEVs. Next, it is confirmed that FGL1 and PD‐L1 on sEVs are specifically bound to their receptors, LAG‐3 and PD‐1 on target cells. Importantly, FP sEVs significantly inhibite T cell activation and proliferation in vitro and a heart allograft model. Furthermore, FP sEVs encapsulated with low‐dose FK506 (FP sEVs@FK506) exert stronger effects on inhibiting T cell proliferation, reducing CD8+ T cell density and cytokine production in the spleens and heart grafts, inducing regulatory T cells in lymph nodes, and extending graft survival. Taken together, dual‐targeting sEVs have the potential to boost the immune inhibitory signalings in synergy and slow down transplant rejection., MSC‐derived FGL1/PD‐L1 sEVs encapsulated in FK506 exhibited a strong ability to inhibit T cell activation and proliferation, and also induced Tregs in organ recipient mice. An experimental basis for a novel intervention strategy is provided that leverages the function of target delivery sEVs to synergistically enhance two immunosuppression axes and reestablish immune tolerance, ultimately promoting organ acceptance.

Metagenomic sequence data from defined mock communities is crucial for the assessment of sequencing platform performance and downstream analyses, including assembly, binning and taxonomic assignment. We report a comparison of shotgun metagenome sequencing and assembly metrics of a defined microbial mock community using the Oxford Nanopore Technologies (ONT) MinION, PacBio and Illumina sequencing platforms. Our synthetic microbial community BMock12 consists of 12 bacterial strains with genome sizes spanning 3.2–7.2 Mbp, 40–73% GC content, and 1.5–7.3% repeats. Size selection of both PacBio and ONT sequencing libraries prior to sequencing was essential to yield comparable relative abundances of organisms among all sequencing technologies. While the Illumina-based metagenome assembly yielded good coverage with few misassemblies, contiguity was greatly improved by both, Illumina + ONT and Illumina + PacBio hybrid assemblies but increased misassemblies, most notably in genomes with high sequence similarity to each other. Our resulting datasets allow evaluation and benchmarking of bioinformatics software on Illumina, PacBio and ONT platforms in parallel., Measurement(s)metagenomic data • sequence_assemblyTechnology Type(s)ONT MinION • Illumina sequencing • PacBio RS IIFactor Type(s)sequencing platformSample Characteristic - OrganismBacteriaSample Characteristic - Environmentmock community Machine-accessible metadata file describing the reported data: 10.6084/m9.figshare.10260740

Banana (Musa spp.) is an important tropical crop. Banana industry is under biotic and abiotic stresses such as Fusarium wilt, typhoon, cold stress. Genetic engineering offers a powerful strategy to create germplasm of banana with enhanced resistance. The safety of genetically modified organisms has become a bottleneck restricting the popularization and application of genetically modified technology. In this study, a candidate promoter, LEAFY (LFY) for expression and flower initiation in Arabidopsis, was cloned and constructed to ‘Gene-deletor’ vector. Histochemical β-glucuronidase (GUS) staining results showed that the ‘Gene-deletor’ vector driven by LFY promoter could lead to 88.5% excision efficiency from Arabidopsis seeds based on more than 200 T3 progeny examined per event. GUS staining was found to be partially negative in transgenic bananas, however, polymerase chain reaction could still detect the presence of large fragments of the vector. These results suggest that although LFY promoter could not completely drive the ‘Gene-deletor’ vector to achieve the effect of complete elimination of exogenous gene in bananas, its efficiency of eliminating exogenous gene laid a theoretical foundation for cloning banana fruit-specific promoters, that is, ‘non-transgenic’ GM bananas. These results suggest that LFY promoter could not completely drive the ‘Gene-deletor’ vector to achieve the effect of complete elimination of exogenous gene in bananas. Nevertheless, a certain effect of exogenous gene elimination laid a theoretical foundation for the next step of screening banana fruit-specific promoters, removing all exogenous genes from banana fruits, and solving the food safety problem of genetically modified bananas.

It is generally believed that exchange of secondary metabolite biosynthetic gene clusters (BGCs) among closely related bacteria is an important driver of BGC evolution and diversification. Applying this idea may help researchers efficiently connect many BGCs to their products and characterize the products' roles in various environments. However, existing genetic tools support only a small fraction of these efforts. Here, we present the development of chassis-independent recombinase-assisted genome engineering (CRAGE), which enables single-step integration of large, complex BGC constructs directly into the chromosomes of diverse bacteria with high accuracy and efficiency. To demonstrate the efficacy of CRAGE, we expressed three known and six previously identified but experimentally elusive non-ribosomal peptide synthetase (NRPS) and NRPS-polyketide synthase (PKS) hybrid BGCs from Photorhabdus luminescens in 25 diverse γ-Proteobacteria species. Successful activation of six BGCs identified 22 products for which diversity and yield were greater when the BGCs were expressed in strains closely related to the native strain than when they were expressed in either native or more distantly related strains. Activation of these BGCs demonstrates the feasibility of exploiting their underlying catalytic activity and plasticity, and provides evidence that systematic approaches based on CRAGE will be useful for discovering and identifying previously uncharacterized metabolites.

Eleven noteworthy liverworts are reported from Brunei, Indonesia (Seram), and Malaysia (Sabah). Cololejeunea koponenii and Allorgella zantenii previously endemic to New Guinea are newly reported for Indonesia. The poorly known monospecific genus (Cephalolejeunea) is new to Brunei. Cololejeunea streimannii previously known only from Papua New Guinea is newly found in Borneo. Cheilolejeunea obcordata is discovered for the first time except for the type locality. Porella geheebii is new to Seram. Oil bodies of Cephalolejeunea parvilobula, Cheilolejeunea obcordata, Cololejeunea koponenii, C. streimannii, Leptolejeunea spinistipula, and Pictolejeunea mizutanii are reported for the first time. The photographs of Allorgella zantenii, Cephalolejeunea parvilobula, Cheilolejeunea obcordata, Cololejeunea koponenii, Drepanolejeunea blumei., and Porella geheebii are provided.

Purpose The role of pathogenic Escherichia coli colonization in asymptomatic pregnant women is not well understood. The purpose of this work was to determine the prevalence, antimicrobial susceptibility, and neonatal outcomes of pathogenic E. coli colonization in pregnant women. Patients and methods A total of 137 women from southern Taiwan with singleton pregnancies were enrolled between March 2016 and June 2017. The women were prospectively screened for E. coli colonization in the rectovaginal region during prenatal examination. The exclusion criteria are twin pregnancy of the mother and major anomaly of the neonate. All E. coli isolates were identified as either pathogenic or commensal strains, and their susceptibility to various antimicrobials was investigated. Clinical data of the infants were retrieved from their medical records. Results Results showed that 35.8% of asymptomatic pregnant women had pathogenic E. coli colonization in the rectovaginal region. Neonates born to such mothers showed significant morbidities, including hospitalization (OR= 3.74, 95% CI= 1.18~11.87), hyperbilirubinemia (OR= 2.81, 95% CI= 1.24~6.38), and gastrointestinal symptoms (OR= 5.53, 95% CI= 1.39~21.94). Maternal colonization with pathogenic E. coli at rectoanal site was a risk factor for neonatal hyperbilirubinemia after Benjamini-Hochberg (BH) adjustment (52% vs 24%, adjusted P= 0.048). Conclusion The prevalence of pathogenic E. coli colonization in Taiwanese asymptomatic pregnant women was high, and the neonates born to colonized mothers exhibited potential neonatal morbidities. Larger studies are necessary to confirm these findings.

Graphical abstract, Highlights • Emergence of carbapenem-resistant S. algae is a severe problem. • Re-sensitization of S. algae to carbapenem by CRISPR/Cas9 genome editing. • The blaOXA-55-like gene is essential for carbapenem resistance in S. algae. • One-plasmid genome editing system for CRISPR/Cas9 genome editing in S. algae. • CRISPR/Cas9 genome editing is a promising approach to validate the gene function., Antibiotic resistance in pathogens is a growing threat to human health. Of particular concern is resistance to carbapenem, which is an antimicrobial agent listed as critically important by the World Health Organization. With the global spread of carbapenem-resistant organisms, there is an urgent need for new treatment options. Shewanella algae is an emerging pathogen found in marine environments throughout the world that has increasing resistance to carbapenem. The organism is also a possible antibiotic resistance reservoir in humans and in its natural habitat. The development of CRISPR/Cas9-based methods has enabled precise genetic manipulation. A number of attempts have been made to knock out resistance genes in various organisms. The study used a single plasmid containing CRISPR/Cas9 and recE/recT recombinase to reverse an antibiotic-resistant phenotype in S. algae and showed blaOXA-55-like gene is essential for the carbapenem resistance. This result demonstrates a potential validation strategy for functional genome annotation in S. algae.

The neurotoxin β-N-methylamino-l-alanine replaces l-serine in proteins and produces Alzheimer-like pathology. In proteoglycans, e.g. glypican-1, this should preclude substitution with heparan sulfate chains. Reduced release of heparan sulfate should increase β-secretase activity and processing of amyloid precursor protein. Cultured cells were treated with β-N-methylamino-l-alanine during the growth-phase and the effect on heparan sulfate substitution and amyloid precursor protein processing was evaluated using antibodies specific for heparan sulfate, the N- and C-termini of the C-terminal fragment of β-cleaved amyloid precursor protein, and amyloid beta followed by immunofluorescence microscopy, flow cytometry or SDS-PAGE. Mouse fibroblasts, N2a neuroblastoma cells and human neural stem cells released less heparan sulfate when grown in the presence of β-N-methylamino-l-alanine. Cells expressing a recombinant, anchor-less glypican-1 secreted heparan sulfate-deficient glypican-1. There was increased processing of amyloid precursor protein in N2a cells when grown in the presence of the neurotoxin. The degradation products accumulated in cytoplasmic clusters. Secretion of amyloid beta increased approx. 3-fold. Human neural stem cells also developed cytoplasmic clusters containing degradation products of amyloid precursor protein. When non-dividing mouse N2a cells or cortical neurons were exposed to β-N-methylamino-l-alanine there was no effect on heparan sulfate substitution in glypican-1 or on amyloid precursor protein processing.

Shewanella xiamenensis is an emerging pathogen causing intra-abdominal infection and intestinal colonization. Epidemiologic clues suggest its role as a potential food-borne zoonotic agent. To date, four genome sequences of S. xiamenensis have been made publicly available. All of them were isolated from water samples. In this study, we characterized the genome of a S. xiamenensis strain isolated from a giant grouper in Taiwan. The genome of S. xiamenensis ZYW1 is 4,827,717 bp in length and encodes 4,239 open reading frames. Its genomic sequence shares high homology with other S. xiamenensis strains. blaOXA-416 was identified. This is the first detection of S. xiamenensis in Taiwan. These genomic data and analyses contribute to our understanding of S. xiamenensis and may help to elucidate disease-causing mechanisms in future studies.

Ethnopharmacological relevance Er-Zhi-Wan (EZW), a famous traditional Chinese formulation, is used to prevent, or to treat, various liver and kidney diseases for its actions of replenishing liver and kidney. However, the mechanisms of treating Liver-kidney Yin deficiency syndrome (LKYDS) of EZW have not been comprehensively investigated. Aim of the study In this study, a broad range metabolomics strategy coupled with network analysis was established to investigate possible mechanisms of EZW in treating LKYDS. Materials and method The rat models of LKYDS were established using the mixture of thyroxine and reserpine, and the changes of biochemical indices in serum and histopathology were detected to explore the effects of EZW. Next, a broad range metabolomics strategy based on RPLC-Q-TOF/MS and HILIC-Q-TOF/MS has been developed to find the possible significant metabolites in the serum and urine of LKYDS rats. Then, network analysis was applied to visualize the relationships between identified serum and urine metabolites and in detail to find hub metabolites, which might be responsible for the effect of EZW on rats of LKYDS. Furthermore, the shortest path of “disease gene-pathway protein-metabolite” was built to investigate the possible intervention path of EZW from the systematic perspective. Results Five hub metabolites, namely, arachidonic acid, L -arginine, testosterone, taurine and oxoglutaric acid, were screened out and could be adjusted to recover by EZW. After that, the shortest path starting from disease genes and ending in metabolites were identified and disclosed, and the genes of aging such as CAV1 and ACO1 were selected to explain the pathological mechanism of LKYDS. Conclusion Broad range metabolomics coupled with network analysis could provide another perspective on systematically investigating the molecular mechanism of EZW in treating LKYDS at metabolomics level. In addition, EZW might prevent the pathological process of LKYDS through regulating the disturbed metabolic pathway and the aging genes such as CAV1 and ACO1, which may be potential targets for EZW in the treatment of LKYDS.

Phytochemical investigation of the roots of Glehnia littoralis Fr. Schmidt. ex Miq. led to the isolation of 16 known compounds, including three β-carboline alkaloids (1–3), four phenylpropanoids (4–7), five phenolic acids (8–12), three polyacetylenes (13–15) and one fatty acid (16). The structures of these compounds were elucidated on the basis of spectral analysis and comparison with those reported in literatures. To the best of knowledge, the report of the first β-carboline alkaloid in the Umbelliferae family. Additionally, compounds 1–5, 9, 10 and 16 have not been reported from any species in Umbelliferae family, compounds 7, 8 and 12 were isolated from the genus Glehnia for the first time and could be of the chemotaxinomic significance and serve as valuable chemotaxonomic makers for G. littoralis. The chemotaxonomic significance of the isolated compounds was summarised.

Shewanella algae is an aquaculture pathogen and an emerging human pathogen. There are increasing reports of carbapenem resistance S. algae from different hosts. However, little is known about the carbapenem-resistant S. algae in aquaculture and the genetic basis of these resistant bacteria in the special niche remains largely unknown. With the progress of sequencing technologies, it became possible to detect the multiple molecular mechanisms leading to antimicrobial resistance using whole genome sequencing data. In this study, we conduct a genome-scale investigation of S. algae SYT4, a carbapenem-resistant bacterium from cultured Asian hard clam in Taiwan. Whole-genome sequencing of Shewanella algae SYT4 was performed using Illumina MiSeq platform. The genome size of Shewanella algae SYT4 is 4,838,024 bp in length. The DNA G + C content is 53.09%. It encodes 4207 proteins and 104 RNA genes. The phylogenetic relationships of S. algae SYT4 and related Shewanella strains were determined using average nucleotide identity and 16S rRNA gene sequence. Candidate genes for antimicrobial resistance and virulence were identified. S. algae SYT4 carries blaOXA-55 and multiple genes encoding efflux pumps. In addition, genes related to chemotaxis, biofilm formation, and stress response were identified, suggesting potential pathogenic mechanism. We demonstrate that whole-genome sequencing is capable to decipher the resistance determinants of carbapenem-resistant bacterium associated with aquaculture. The panoply of antibiotics resistance genes found suggests the organisms can act as a reservoir of antimicrobial drug resistance determinants in seafood, which is an issue of considerable concern.

As an oil crop with good taste and profuse nutrition, peanut ( Arachis hypogaea L.) is grown worldwide, mainly for edible seeds. Black peanuts attract more attention for their appealing color and health-promoting anthocyanins. Here, two cyanidin-based anthocyanins and four quercetin-based flavonols were separated and identified from skins of two black cultivars (Zi Yu and Zi Guan) by HPLC-ESI-Q-TOF-MS. To study the anthocyanin accumulation, libraries constructed from the mRNA of skins of Zi Yu and white cultivar (Bai Yu) were sequenced, and 4042 differentially expressed genes were identified. Gene ontology and KEGG pathway analysis underlined the importance of the high expression of flavonoid biosynthetic and regulatory genes in seed skin of Zi Yu. Furthermore, expression profiles of these genes were analyzed carefully in four representative peanut cultivars. Altogether, these results strongly indicate that the up-regulation of transcriptional activators (AhMYB1, AhMYB2, and AhTT8) accounts for the skin-specific accumulation of anthocyanins in black peanut.

Human bocavirus 1 (HBoV1), an autonomous parvovirus, is a helper virus supporting replication of wild-type adeno-associated virus 2 (AAV2). In this study, we compared the helper functions from HBoV1 with those from adenovirus (Ad) for the production of recombinant AAV (rAAV) vector in HEK293 cells. We demonstrated that triple plasmids transfection of (1) a cloned HBoV1 helper minigenome (pBocaHelper) that expresses HBoV1 genes NP1, NS2, and BocaSR, (2) pAAV transfer plasmid, and (3) pAAVRepCap supports rAAV production in HEK293 cells. Despite a production yield of 1–2 log lower than that using pAdHelper (expressing Ad genes E2A, E4, and VA), rAAV vector produced using pBocaHelper transduced cells as efficiently as that produced using pAdHelper. The low vector production is largely due to the inefficient expression of the AAV Rep52 and capsid proteins, as well as reduced rAAV genome replication. When the AAV capsid proteins and Rep52 were ectopically expressed under strong promoters, the enhanced protein expression significantly improved the rAAV production using pBocaHelper, approaching a level of 50%–70% of that produced using pAdHelper. Through further dissection of the helper functions from pAdHelper in a five-plasmid transfection system, we found that the addition of the Ad E2A gene to the above HBoV1 helper system significantly increased rAAV DNA replication, which increased the rAAV vector production to a level of 3–7 times higher than that using pAdHelper. We finally combined HBoV1 NP1 and NS2 genes with Ad helper genes to create a novel dual helper plasmid (pABHelper) for rAAV vector production in the conventional three-plasmid transfection system. The pABHelper facilitated rAAV production at a yield ∼2 times higher than that using the pAdHelper. Keywords: rAAV, HBoV1, helper gene, DNA replication, gene expression

Magnetic nanoparticles with novel core-shell structure were prepared for immunoglobulin (IgG) separation, in which thiophilic property of sulfone groups and protein resistance of poly(ethylene glycol) (PEG) moieties were integrated. The step-wise surface reactions on the nanoparticles were characterized by 1H nuclear magnetic resonance (NMR) and surface zeta potential measurements. With human IgG and bovine serum albumin (BSA) as model proteins, the effects of PEG chain length, conjugation group, solution pH and salt concentration on IgG selectivity were investigated using static adsorption experiments. The experiment results showed that mPEG2000-NH2 modified magnetic nanoparticles had an adsorption capacity of 132.8 mg g−1 and selectivity of 32.5 towards IgG under the condition of pH 7.45 and 0.15 M NaCl. In complex biological fluids, the PEG modified magnetic nanoparticles could separate IgG from fetal calf serum and Omalizumab from cell culture supernatant with purities of 96% and 99%, respectively. Moreover, the binding affinities of the proposed core-shell structure towards IgG from four animal species (human, bovine, rabbit and goat) were quantified by bio-layer interferometer (BLI). The results showed that the selectivity of this structure towards IgG varied from traditional Protein A method, suggesting its potentials in rapid separation and purification of IgG with low affinity towards Protein A.

Parvovirus B19 (B19V) infection causes diseases in humans ranging from the mild erythema infectiosum to severe hematological disorders. The unique region of the minor structural protein VP1 (VP1u) of 227 amino acids harbors strong neutralizing epitopes which elicit dominant immune responses in patients. Recent studies have shown that the VP1u selectively binds to and enters B19V permissive cells through an unknown cellular proteinaceous receptor. In the present study, we demonstrated that purified recombinant VP1u effectively inhibits B19V infection of ex vivo expanded primary human erythroid progenitors. Furthermore, we identified the amino acid sequence 5-68 of the VP1 (VP1u5-68aa) is sufficient to confer the inhibition of B19V infection at a level similar to that of the full-length VP1u. In silico structure prediction suggests that the VP1u5-68aa contains three α-helices. Importantly, we found that the inhibition capability of the minimal domain VP1u5-68aa is independent of its dimerization but is likely dependent on the structure of the three predicated α-helices. As VP1u5-68aa outcompetes the full-length VP1u in entering cells, we believe that VP1u5-68aa functions as a receptor-binding ligand during virus entry. Finally, we determined the effective inhibition potency of VP1u5-68aa in B19V infection of human erythroid progenitors, which has a half maximal effective concentration (EC50) of 67 nM, suggesting an anti-viral peptide candidate to combat B19V infection.IMPORTANCEHuman parvovirus B19 infection causes severe hematological disorders, including transient aplastic crisis, pure red cell aplasia, and hydrops fetalis. A productive B19 infection is highly restricted to human erythroid progenitors in human bone marrow and fetal liver. In the current study, we identified that the N-terminal 5-68 amino acids domain of the minor viral capsid protein VP1 enters ex vivo expanded human erythroid progenitors, which is nearly 5 times more efficient than the full-length VP1 unique region (1-227aa). Importantly, purified recombinant 5-68aa of the VP1 has a high efficiency in inhibition of parvovirus B19 infection of human erythroid progenitors, which has a half maximal effective concentration (EC50) of 67 nM and a low cytotoxicity. The N-terminal 5-68 amino acids holds the potential as an effective antiviral of parvovirus B19 caused hematological disorders, as well as a carrier to deliver proteins to human erythroid progenitors.

Background The prevalence of clinical multidrug-resistant (MDR) Pseudomonas aeruginosa has been increasing rapidly worldwide over the years and responsible for a wide range of acute and chronic infections with high mortalities. Although hundreds of complete genomes of clinical P. aeruginosa isolates have been sequenced, only a few complete genomes of mucoid strains are available, limiting a comprehensive understanding of this important group of opportunistic pathogens. Herein, the complete genome of a clinically isolated mucoid strain P. aeruginosa JNQH-PA57 was sequenced and assembled using Illumina and Oxford nanopore sequencing technologies. Genomic features, phylogenetic relationships, and comparative genomics of this pathogen were comprehensively analyzed using various bioinformatics tools. A series of phenotypic and molecular-genetic tests were conducted to investigate the mechanisms of carbapenem resistance in this strain. Results Several genomic features of MDR P. aeruginosa JNQH-PA57 were identified based on the whole-genome sequencing. We found that the accessory genome of JNQH-PA57 including several prophages, genomic islands, as well as a PAPI-1 family integrative and conjugative element (ICE), mainly contributed to the larger genome of this strain (6,747,067 bp) compared to other popular P. aeruginosa strains (with an average genome size of 6,445,223 bp) listed in Pseudomonas Genome Database. Colony morphology analysis and biofilm crystal staining assay respectively demonstrated an enhanced alginate production and a thicker biofilm formation capability of JNQH-PA57. A deleted mutation at nt 424 presented in mucA gene, resulted in the upregulated expression of a sigma-factor AlgU and a GDP mannose dehydrogenase AlgD, which might explain the mucoid phenotype of this strain. As for the carbapenem resistance mechanisms, our results revealed that the interplay between impaired OprD porin, chromosomal β-lactamase OXA-488 expression, MexAB-OprM and MexXY-OprM efflux pumps overexpression, synergistically with the alginates-overproducing protective biofilm, conferred the high carbapenem resistance to P. aeruginosa JNQH-PA57. Conclusion Based on the genome analysis, we could demonstrate that the upregulated expression of algU and algD, which due to the truncation variant of MucA, might account for the mucoid phenotype of JNQH-PA57. Moreover, the resistance to carbapenem in P. aeruginosa JNQH-PA57 is multifactorial. The dataset presented in this study provided an essential genetic basis for the comprehensive cognition of the physiology, pathogenicity, and carbapenem resistance mechanisms of this clinical mucoid strain.

Background: Current neurocognitive models of language function have been primarily built from evidence regarding object naming, and their hypothesized white-matter circuit mechanisms tend to be coarse grained. Methods: In this cross-sectional, observational study, we used novel correlational tractography to assess the white-matter circuit mechanism behind verb retrieval, measured through action picture-naming performance in adults with chronic aphasia. Results: The analysis identified tracts implicated in current neurocognitive dual-stream models of language function, including the left inferior fronto-occipital fasciculus, inferior longitudinal fasciculus, and arcuate fasciculus. However, the majority of tracts associated with verb retrieval were not ones included in dual-stream models of language function. Instead, they were projection pathways that connect frontal and parietal cortices to subcortical regions associated with motor functions, including the left corticothalamic pathway, frontopontine tract, parietopontine tract, corticostriatal pathway, and corticospinal tract. Conclusions: These results highlight that corticosubcortical projection pathways implicated in motor functions may be importantly related to language function. This finding is consistent with grounded accounts of cognition and may furthermore inform neurocognitive models. IMPACT STATEMENT: This study suggests that in addition to traditional dual-stream language fiber tracts, the integrity of projection pathways that connect frontal and parietal cortices to subcortical motor regions may be critically associated with verb-retrieval impairments in adults with aphasia. This finding challenges neurological models of language function.