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BACKGROUND: Myostatin (Mstn) plays an important role in adipocyte growth, differentiation and metabolism, leading to the development of obesity. OBJECTIVES: We aimed to explore the effect of Mstn on white fat browning in a mouse model of type 2 diabetes mellitus (T2DM). MATERIAL AND METHODS: Twelve wild-type (WT), 12 heterozygous (Mstn(+/-)) and 12 homozygous (Mstn(-/-)) male mice were randomly divided into 6 groups: WT, Mstn(+/-), Mstn(-/-), WT+DM, Mstn(+/-)+DM, and Mstn(-/-)+DM. The first 3 groups were fed normal chow, while the last 3 were fed high-fat diet and administered streptozotocin to generate T2DM. Subsequently, body mass, length, and white and brown fat masses were measured, after which Lee's index, white-brown ratio and fat index were calculated. The serum free fatty acid (FFA) levels were detected using enzyme-linked immunosorbent assay (ELISA). Hematoxylin and eosin (H&E) staining was used to analyze white and brown fat cell morphology. The relative expression levels of peroxisome proliferator-activated receptor-gamma (PPARγ), peroxisome proliferator-activated receptor-gamma coactivator-1 alpha (PGC-1α), uncoupling protein 1 (UCP1), and cluster of differentiation 137 (CD137) protein were determined with western blotting. RESULTS: The Mstn(-/-) group displayed higher levels of PPARγ, PGC-1α and CD137 proteins in white and brown fat compared to the WT and Mstn(+/-) groups, while the expression level of UCP1 protein in the Mstn(-/-) group was higher than in the WT group. The expression levels of PPARγ, PGC-1α, UCP1, and CD137 proteins in the WT+DM group were lower than in the WT group. Moreover, PPARγ, PGC-1α, UCP1, and CD137 proteins were more highly expressed in the Mstn(-/-)+DM group compared to the WT+DM and Mstn(+/-)+DM groups. CONCLUSIONS: The Mstn gene inhibition antagonizes obesity phenotypes, such as white fat accumulation and lipid metabolism derangement caused by T2DM, thus promoting white fat browning.
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Ecological value realization of blue carbon in marine ranching is essential to achieve carbon neutrality. The motivation for conducting the study is to verify whether multi-agent cooperation can promote the ecological value realization of blue carbon in marine ranching. Based on the blue carbon ecological value realization model of marine ranching enterprises, blue carbon demand enterprises, blue carbon trading platforms and government, this paper explores the cooperative governance strategy of marine ranching for each subject using cooperative game and non-cooperative game models. Further, we conduct a comparative analysis to arrive at the optimal strategy. The conclusions are as follows. Multi-agent cooperation is more conducive to the ecological value realization of blue carbon in marine ranching. Compared with non-cooperative governance, the platform's commission and blue carbon price are lower, and the blue carbon output, profit of each market subject, government utility and overall profit are higher in cooperative governance. The strengths of this paper lie in 2 aspects. First, we focuses on the ecological value of blue carbon in marine ranching instead of economic value, providing a new theoretical basis for ecological compensation in marine ranching. Second, we construct a government-led and market-oriented operation of marine ranching's blue carbon ecological value realization mechanism, incorporating blue carbon trading platform and government into the value realization model.
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To grasp the epidemiological trend and drug resistance mechanisms of Clostridioides difficile (C. diff) in Beijing, 302 C. diff isolates were obtained from patients with diarrhea. The sequence types (STs) from mainstream strains were all susceptible to metronidazole, vancomycin, piperacillin/tazobactam, meropenem, and tigecycline but almost resistant to ciprofloxacin and clindamycin. The missense mutation of GyrA/GyrB and RpoB resulted in fluoroquinolone and rifamycin resistance, respectively. Toxigenic strains from clade IV were likely to be missed due to the deficiency of tcdA gene. Four tcdC genotypes were first detected in strains from clade III and IV. The truncating mutation of TcdC disabled its function working as a toxin suppressor. In conclusion, the molecular epidemiology of C. diff in Beijing is different from other regions of China. The antimicrobial resistance patterns and toxin-producing abilities of strains with different STs varied greatly, which suggests that continuous surveillance and control are meaningful and urgent.
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Clostridioides difficile , Infecciones por Clostridium , Humanos , Antibacterianos/farmacología , Epidemiología Molecular/métodos , Clostridioides/genética , Infecciones por Clostridium/epidemiología , Hospitales de Enseñanza , China/epidemiología , Pruebas de Sensibilidad MicrobianaRESUMEN
Clostridioides difficile infection (CDI) is associated with high recurrence rates that have substantial effects on patients' quality of life. To investigate the risk factors and potential mechanisms contributing to recurrent CDI (rCDI), a total of 243 cases were enrolled in this study. The history of omeprazole (OME) medication and ST81 strain infection were considered the two independent risks with the highest odds ratios in rCDI. In the presence of OME, we detected concentration-dependent increases in the MIC values of fluoroquinolone antibiotics against ST81 strains. Mechanically, OME facilitated ST81 strain sporulation and spore germination by blocking the pathway of purine metabolism and also promoted an increase in cell motility and toxin production by turning the flagellar switch to the ON state. In conclusion, OME affects several biological processes during C difficile growth, which have fundamental impacts on the development of rCDI caused by ST81 strains. Programmed OME administration and stringent surveillance of the emerging ST81 genotype are matters of considerable urgency and significance in rCDI prevention.
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Clostridioides difficile , Infecciones por Clostridium , Humanos , Omeprazol/farmacología , Omeprazol/uso terapéutico , Calidad de Vida , Clostridioides difficile/genética , Infecciones por Clostridium/prevención & control , Fluoroquinolonas/farmacología , Recurrencia , Antibacterianos/farmacología , Antibacterianos/uso terapéuticoRESUMEN
At present, the diffusion and trading volume of new energy vehicles (NEVs) account for only a small part of the automobile market. How to effectively promote the production of energy vehicle manufacturers and the purchase of consumers for NEVs has become a key and urgent problem to be solved. Our research builds an evolutionary game model including government, consumers and energy vehicle manufacturers from the perspective of supply chain research, and discusses the participants' evolutionary stability strategy and portfolio stability strategy. In addition, the method of simulated moments is used to assign values to the model data, and MATLAB software is used for simulation verification and analysis. Through analysis, it is found that: (1) The government, energy vehicle manufacturers and consumers influence each other in their strategic choices, but to different degrees. (2) The difference of government subsidy objects will not affect the evolution direction and result of the diffusion game model of NEVs, but will affect the stable speed of strategic behavior and the change direction of strategic choice of consumers and energy vehicle manufacturers. And in the case of moderate subsidies, subsidies to energy vehicle manufacturers can promote the NEV diffusion. (3) Improving the environmental friendliness of NEVs and improving consumers' low-carbon preference for NEVs have about the same effect on the replicative power system. And compared with other key factors, they are the most favorable way to promote NEVs diffusion. At last, suggestions are put forward according to the research conclusions.
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The ultrasonic detectability of buried defects within composite materials is dependent on the anisotropy of the composite material by which the propagation property of acoustic wave in each direction is variably affected. In this study, the characteristics of acoustic waves propagating in different directions for composite materials are explored based on the full matrix capture (FMC) data using an ultrasonic phased array. The elastic constant of multidirectional carbon fiber reinforced plastic (CFRP) laminate is first derived based on the genetic algorithm. The characteristics of transmitted and reflected waves in higher angles are predicted by implementing the Christoffel equation, and the focal law used in post-processing of FMC data can be optimized accordingly. The imaging results of the total focusing method (TFM) using the improved focal law are compared with the results of the conventional TFM. The results suggest that the optimized TFM can effectively characterize the defect by reducing the background noise. Furthermore, since it is impractical to theoretically correct angle-dependent velocity for in situ inspection, a linear extrapolation method based on the experimentally measurable velocity at low angles is proposed to estimate the velocity profile at higher angles. The imaging results using the fast extrapolated velocity profile is then compared with the theoretical, and it has been demonstrated that while the difference between the images using the theoretical focal law and the linearly extrapolated one is barely visible, the later one is overwhelmingly advantageous to be realiszd for engineering practices.
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BackgroundMerkel cell carcinoma (MCC) is an aggressive neuroendocrine (NE) skin cancer caused by severe UV-induced mutations or expression of Merkel cell polyomavirus (MCPyV) large and small T antigens (LT and ST). Despite deep genetic differences between MCPyV-positive and -negative subtypes, current clinical diagnostic markers are indistinguishable, and the expression profile of MCC tumors is, to our knowledge, unexplored.MethodsHere, we leveraged bulk and single-cell RNA-Seq of patient-derived tumor biopsies and cell lines to explore the underlying transcriptional environment of MCC.ResultsStrikingly, MCC samples could be separated into transcriptional subtypes that were independent of MCPyV status. Instead, we observed an inverse correlation between a NE gene signature and the Hippo pathway transcription factors Yes1-associated transcriptional regulator (YAP1) and WW domain-containing transcriptional regulator 1 (WWTR1). This inverse correlation was broadly present at the transcript and protein levels in the tumor biopsies as well as in established and patient-derived cell lines. Mechanistically, expression of YAP1 or WWTR1 in a MCPyV-positive MCC cell line induced cell-cycle arrest at least in part through TEA domain-dependent (TEAD-dependent) transcriptional repression of MCPyV LT.ConclusionThese findings identify what we believe to be a previously unrecognized heterogeneity in NE gene expression within MCC and support a model of YAP1/WWTR1 silencing as essential for the development of MCPyV-positive MCC.FundingUS Public Health Service grants R35CA232128, P01CA203655, and P30CA06516.
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Carcinoma de Células de Merkel , Poliomavirus de Células de Merkel , Infecciones por Polyomavirus , Neoplasias Cutáneas , Infecciones Tumorales por Virus , Humanos , Carcinoma de Células de Merkel/genética , Carcinoma de Células de Merkel/patología , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/patología , Poliomavirus de Células de Merkel/genética , Péptidos y Proteínas de Señalización Intracelular , Línea Celular , Infecciones por Polyomavirus/genética , Infecciones Tumorales por Virus/genética , Proteínas Coactivadoras Transcripcionales con Motivo de Unión a PDZRESUMEN
Cancers avoid immune surveillance through an array of mechanisms, including perturbation of HLA class I antigen presentation. Merkel cell carcinoma (MCC) is an aggressive, HLA-I-low, neuroendocrine carcinoma of the skin often caused by the Merkel cell polyomavirus (MCPyV). Through the characterization of 11 newly generated MCC patient-derived cell lines, we identified transcriptional suppression of several class I antigen presentation genes. To systematically identify regulators of HLA-I loss in MCC, we performed parallel, genome-scale, gain- and loss-of-function screens in a patient-derived MCPyV-positive cell line and identified MYCL and the non-canonical Polycomb repressive complex 1.1 (PRC1.1) as HLA-I repressors. We observed physical interaction of MYCL with the MCPyV small T viral antigen, supporting a mechanism of virally mediated HLA-I suppression. We further identify the PRC1.1 component USP7 as a pharmacologic target to restore HLA-I expression in MCC.
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Carcinoma de Células de Merkel , Poliomavirus de Células de Merkel , Infecciones por Polyomavirus , Neoplasias Cutáneas , Antígenos Virales de Tumores/genética , Antígenos Virales de Tumores/metabolismo , Carcinoma de Células de Merkel/genética , Carcinoma de Células de Merkel/patología , Epigénesis Genética , Humanos , Poliomavirus de Células de Merkel/genética , Poliomavirus de Células de Merkel/metabolismo , Infecciones por Polyomavirus/genética , Neoplasias Cutáneas/patología , Peptidasa Específica de Ubiquitina 7/metabolismoRESUMEN
Merkel cell carcinoma (MCC) is an aggressive malignancy with neuroendocrine (NE) features, limited treatment options, and a lack of druggable targets. There is no reported involvement of the MUC1-C oncogenic protein in MCC progression. We show here that MUC1-C is broadly expressed in MCCs and at higher levels in Merkel cell polyomavirus (MCPyV)-positive (MCCP) relative to MCPyV-negative (MCCN) tumors. Our results further demonstrate that MUC1-C is expressed in MCCP, as well as MCCN, cell lines and regulates common sets of signaling pathways related to RNA synthesis, processing, and transport in both subtypes. Mechanistically, MUC1-C (i) interacts with MYCL, which drives MCC progression, (ii) is necessary for expression of the OCT4, SOX2, KLF4, MYC, and NANOG pluripotency factors, and (iii) induces the NEUROD1, BRN2 and ATOH1 NE lineage dictating transcription factors. We show that MUC1-C is also necessary for MCCP and MCCN cell survival by suppressing DNA replication stress, the p53 pathway, and apoptosis. In concert with these results, targeting MUC1-C genetically and pharmacologically inhibits MCC self-renewal capacity and tumorigenicity. These findings demonstrate that MCCP and MCCN cells are addicted to MUC1-C and identify MUC1-C as a potential target for MCC treatment.
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Carcinoma de Células de Merkel , Poliomavirus de Células de Merkel , Mucina-1 , Neoplasias Cutáneas , Carcinoma de Células de Merkel/tratamiento farmacológico , Carcinoma de Células de Merkel/genética , Carcinoma de Células de Merkel/virología , Humanos , Mucina-1/metabolismo , Transducción de Señal , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/virologíaRESUMEN
Clear evidence supports a causal link between Merkel cell polyomavirus (MCPyV) and the highly aggressive human skin cancer called Merkel cell carcinoma (MCC). Integration of viral DNA into the human genome facilitates continued expression of the MCPyV small tumor (ST) and large tumor (LT) antigens in virus-positive MCCs. In MCC tumors, MCPyV LT is truncated in a manner that renders the virus unable to replicate yet preserves the LXCXE motif that facilitates its binding to and inactivation of the retinoblastoma tumor suppressor protein (pRb). We previously developed a MCPyV transgenic mouse model in which MCC tumor-derived ST and truncated LT expression were targeted to the stratified epithelium of the skin, causing epithelial hyperplasia, increased proliferation, and spontaneous tumorigenesis. We sought to determine if any of these phenotypes required the association between the truncated MCPyV LT and pRb. Mice were generated in which K14-driven MCPyV ST/LT were expressed in the context of a homozygous RbΔLXCXE knock-in allele that attenuates LT-pRb interactions through LT's LXCXE motif. We found that many of the phenotypes including tumorigenesis that develop in the K14-driven MCPyV transgenic mice were dependent upon LT's LXCXE-dependent interaction with pRb. These findings highlight the importance of the MCPyV LT-pRb interaction in an in vivo model for MCPyV-induced tumorigenesis.
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Carcinoma de Células de Merkel , Poliomavirus de Células de Merkel , Infecciones por Polyomavirus , Neoplasias Cutáneas , Infecciones Tumorales por Virus , Animales , Antígenos Transformadores de Poliomavirus/genética , Antígenos Transformadores de Poliomavirus/metabolismo , Antígenos Virales de Tumores/genética , Antígenos Virales de Tumores/metabolismo , Transformación Celular Neoplásica , Hiperplasia/patología , Células de Merkel/metabolismo , Células de Merkel/patología , Poliomavirus de Células de Merkel/genética , Ratones , Neoplasias Cutáneas/patologíaRESUMEN
INTRODUCTION: Bone mineral density (BMD) decreases with age, leading to fractures, decreased mobility, and impaired quality of life. We aimed to determine the effects of brisk walking and exposure to sunlight on BMD and balance in the elderly with osteopenia. MATERIALS AND METHODS: We recruited 81 elderly subjects with osteopenia from January 2019 to March 2019. They were divided into four groups: a daytime-walking group (n = 20), a night-time-walking group (n = 20), a sun-exposure-only group (n = 20), and a control group (n = 21). The subjects walked briskly for 30-60 min three times a week for 24 weeks. The sun-exposure-only group received sunlight for 20-30 min three times a week. All four groups received supplemental calcium. Lumbar L1-L4 BMD, serum 25-hydroxyvitamin D3, timed-up-go-test (TUGT), five-times-sit-stand-test (FTSST), open-eye and closed-eye one-leg-stance-test (OLST) were measured at baseline and 1 day after program completion. RESULTS: The lumbar L1-L4 BMD was higher in all intervention groups (P < 0.05), with the daytime-walking group outperforming the others. There was no significant difference between the night-time-walking and sun-exposure-only groups (P > 0.05). The levels of serum 25-hydroxyvitamin D3 in the daytime-walking and sun-exposure-only groups were higher than those in the night-time-walking and control groups (P < 0.05). The TUGT and FTSST times decreased in all three intervention groups and predominantly so in the daytime-walking group, whereas the open-eye and closed-eye OLST times increased (P < 0.05). CONCLUSION: Brisk walking and sun exposure increase BMD and improve dynamic and static balance in the elderly with osteopenia.
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Densidad Ósea , Enfermedades Óseas Metabólicas , Anciano , Calcifediol , Humanos , Calidad de Vida , Luz Solar , CaminataRESUMEN
Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been used in the field of clinical microbiology since 2010. Compared with the traditional technique of biochemical identification, MALDI-TOF MS has many advantages, including convenience, speed, accuracy, and low cost. The accuracy and speed of identification using MALDI-TOF MS have been increasing with the development of sample preparation, database enrichment, and algorithm optimization. MALDI-TOF MS has shown promising results in identifying cultured colonies and rapidly detecting samples. MALDI-TOF MS has critical research applications for the rapid detection of highly virulent and drug-resistant pathogens. Here we present a scientific review that evaluates the performance of MALDI-TOF MS in identifying clinical pathogenic microorganisms. MALDI-TOF MS is a promising tool in identifying clinical microorganisms, although some aspects still require improvement.
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Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has been accepted as a rapid, accurate, and less labor-intensive method in the identification of microorganisms in clinical laboratories. However, there is limited data on systematic evaluation of its effectiveness in the identification of phylogenetically closely-related yeast species. In this study, we evaluated two commercially available MALDI-TOF systems, Autof MS 1000 and Vitek MS, for the identification of yeasts within closely-related species complexes. A total of 1,228 yeast isolates, representing 14 different species of five species complexes, including 479 of Candida parapsilosis complex, 323 of Candida albicans complex, 95 of Candida glabrata complex, 16 of Candida haemulonii complex (including two Candida auris), and 315 of Cryptococcus neoformans complex, collected under the National China Hospital Invasive Fungal Surveillance Net (CHIF-NET) program, were studied. Autof MS 1000 and Vitek MS systems correctly identified 99.2% and 89.2% of the isolates, with major error rate of 0.4% versus 1.6%, and minor error rate of 0.1% versus 3.5%, respectively. The proportion of isolates accurately identified by Autof MS 1000 and Vitek MS per each yeast complex, respectively, was as follows; C. albicans complex, 99.4% vs 96.3%; C. parapsilosis complex, 99.0% vs 79.1%; C glabrata complex, 98.9% vs 94.7%; C. haemulonii complex, 100% vs 93.8%; and C. neoformans, 99.4% vs 95.2%. Overall, Autof MS 1000 exhibited good capacity in yeast identification while Vitek MS had lower identification accuracy, especially in the identification of less common species within phylogenetically closely-related species complexes.
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Infecciones Fúngicas Invasoras , Candida , China , Humanos , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Espectrometría de Masas en TándemRESUMEN
Merkel cell polyomavirus (MCPyV) causes the majority of human Merkel cell carcinomas (MCC), a rare but highly aggressive form of skin cancer. We recently reported that constitutive expression of MCC tumor-derived MCPyV tumor (T) antigens in the skin of transgenic mice leads to hyperplasia, increased proliferation, and spontaneous epithelial tumor development. We sought to evaluate how the MCPyV T antigens contribute to tumor formation in vivo using a classical, multi-stage model for squamous cell carcinoma development. In this model, two chemical carcinogens, DMBA and TPA, contribute to two distinct phases of carcinogenesis-initiation and promotion, respectively-that are required for tumors to develop. By treating the MCPyV transgenic mice with each chemical carcinogen, we determined how the viral oncogenes contributed to carcinogenesis. We observed that the MCPyV T antigens synergized with the tumor initiator DMBA, but not with the tumor promoter TPA, cause tumors. Therefore, the MCPyV tumor antigens function primarily as tumor promoters, similar to that seen with human papillomavirus (HPV) oncoproteins. These studies provide insight into the role of MCPyV T antigen expression in tumor formation in vivo and contribute to our understanding of how MCPyV may function as a human DNA tumor virus.
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The improvement of detection accuracy and reliability of micro delamination defect is restricted by the rough surface. The flat bottom holes (FBHs) are frequently employed as reference targets to evaluate the sensitivity of ultrasonic testing for internal defects. A roughness-modified analytical model for ultrasonic testing of FBHs is established based on the principle of multi-Gaussian beam and phase-screen approximation. The signal of reference reflector is obtained from two-dimensional ultrasonic simulation model. The amplitude changes of echo signals and noises of FBHs with different diameters and depths under rough surfaces are presented. The analyses results indicate that the root-mean-square (rms) height plays a dominant role in the amplitude change of signals compared with the correlation length. The reflected wave amplitude of FBHs decreases nonlinearly with an increase of roughness whereas the amplitude of noise increases slightly. Subsequently, a method is proposed further combining the noise amplitude acquired from numerical simulation and the echo signal amplitude obtained by the analytical model, which is to predict and estimate the detection accuracy of internal defects under different surface roughness. The parallel experiments are performed on several samples with different roughness to validate the evaluation method.
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BACKGROUND: This study reports the global trends of antimicrobial susceptibility to ceftaroline and ceftazidime-avibactam using data from the Antimicrobial Testing Leadership and Surveillance (ATLAS) program between 2012 and 2016. METHODS: For the 2012-2016 ATLAS program, 205 medical centers located in Africa-Middle East (n = 12), Asia-Pacific (n = 32), Europe (n = 94), Latin America (n = 26), North America (n = 31), and Oceania (n = 10) consecutively collected the clinical isolates. The minimum inhibitory concentrations (MICs) and in vitro susceptibilities to ceftaroline and ceftazidime-avibactam were assessed using the Clinical and Laboratory Standards Institute (CLSI) 2019and European Committee on Antimicrobial Susceptibility Testing (EUCAST) 2019 guidelines. RESULTS: Between 2012 and 2016, 176,345 isolates were collected from around the globe and included in the analysis. Regarding Gram-negative bacteria, ceftazidime-avibactam demonstrated high susceptibility (> 90%) against Enterobacteriaceae and Pseudomonas aeruginosa, with increased antimicrobial activity observed from the addition of avibactam (4 mg/L) to ceftazidime. Regarding Gram-positive bacteria, ceftaroline showed > 90% susceptibility against Staphylococcus aureus, Streptococcus pneumoniae, α-and ß-hemolytic Streptococcus. The antimicrobial susceptibilities to ceftaroline and ceftazidime-avibactam were mostly stable from 2012 to 2016, but the susceptibilities to ceftazidime-avibactam to carbapenem-resistant (CR) Klebsiella pneumonia (88.4-81.6%) and to CR-P. aeruginosa (89.6-72.7%) decreased over time. In terms of regional difference, the susceptibilities of methicillin-resistant S. aureus to ceftaroline in Asia and of CR-K. pneumonia to ceftazidime-avibactam in Asia/Africa-Middle East were lower compared with other regions, while the susceptibility of CR-P. aeruginosa to ceftazidime-avibactam in North America was higher. CONCLUSION: The addition of avibactam improves the activity of ceftazidime against Enterobacteriaceae and P. aeruginosa. The global antimicrobial susceptibilities to ceftaroline and ceftazidime-avibactam were, in general, stable from 2012 to 2016, but a marked reduction in the susceptibilities of specific species and CR-P. aeruginosa to ceftazidime-avibactam was observed.
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Antibacterianos/farmacología , Compuestos de Azabiciclo/farmacología , Bacterias/efectos de los fármacos , Ceftazidima/farmacología , Cefalosporinas/farmacología , Adulto , Anciano , Combinación de Medicamentos , Farmacorresistencia Bacteriana Múltiple , Humanos , Pruebas de Sensibilidad Microbiana , Persona de Mediana Edad , Estudios Retrospectivos , Factores de Tiempo , Adulto Joven , CeftarolinaRESUMEN
PURPOSE: Clostridioides difficile toxin B (TcdB) plays a critical role in C. difficile infection (CDI), a common and costly healthcare-associated disease. The aim of the current study was to explore the intracellular and potent systemic effects of TcdB on human colon epithelial cells utilizing Gene Expression Omnibus and bioinformatic methods. METHODS: Two datasets (GSE63880 and GSE29008) were collected to extract data components of mRNA of TcdB-treated human colon epithelial cells; "limma" package of "R" software was used to screen the differential genes, and "pheatmap" package was applied to construct heat maps for the differential genes; Metascape website was utilized for protein-protein interaction network and Molecular Complex Detection analysis, and Genome Ontology (GO) was used to analyze the selected differential genes. Quantitative real-time PCR (qRT-PCR) and Western blot were performed to validate the expression of hub genes. RESULTS: GO terms involved in DNA replication and cell cycle were identified significantly enriched in TcdB-treated human colon epithelial cells. Moreover, the decreased expression of DNA replication-related genes, MCM complex, and CDC45 in C. difficile (TcdA-/TcdB+)-infected Caco-2 cells were validated via qRT-PCR and Western blot assays. CONCLUSIONS: In conclusion, the integrated analysis of different gene expression datasets allowed us to identify a set of genes and GO terms underlying the mechanisms of CDI induced by TcdB. It would aid in understanding of the molecular mechanisms underlying TcdB-exposed colon epithelial cells and provide the basis for developing diagnosis biomarkers, treatment, and prevention strategies.
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Proteínas Bacterianas/metabolismo , Toxinas Bacterianas/metabolismo , Clostridioides difficile/patogenicidad , Biología Computacional/métodos , Enterocolitis Seudomembranosa/patología , Proteínas Bacterianas/genética , Toxinas Bacterianas/genética , Células CACO-2 , Línea Celular Tumoral , Clostridioides difficile/genética , Enterocolitis Seudomembranosa/microbiología , Células Epiteliales/patología , Expresión Génica/genética , Humanos , Mucosa Intestinal/patología , Análisis por Matrices de Proteínas , Mapas de Interacción de ProteínasRESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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Clostridium difficile infection (CDI) is the leading cause of antibiotic-associated diarrhoea worldwide. In order to gain a better understanding about the molecular epidemiology of C. difficile in Beijing, China, molecular typing, antimicrobial susceptibility testing and drug resistance gene sequencing were performed on 174 strains of C. difficile collected from four large tertiary hospitals in Beijing. In total, 31 sequence types (STs) were identified among the 174 strains. ST81 was found to be the most prevalent (26.4%, 46/174), followed by ST2 (16.7%, 29/174) and ST54 (9.8%, 17/174). All isolates were susceptible to metronidazole and vancomycin. The test strains displayed resistance rates of 97.1%, 44.3% and 44.3% for ciprofloxacin, levofloxacin and moxifloxacin, respectively. ST81 isolates displayed a drug resistance rate of 97.8% for levofloxacin and moxifloxacin, which was significantly higher than ST2 (0%), ST54 (17.6%) and ST42 (0%) isolates (P<0.05). An amino acid mutation (T82I) was identified in GyrA, and the total mutation rate of the C. difficile strains was 40.8% (71/174). The mutation rate of ST81 isolates was 95.7% (44/46). Three amino acid mutations (D426N, S366A and D426V) were identified in GyrB, and the total mutation rate of GyrB was 39.1%. A double-site mutation in GyrB (S366A+D426V) was identified in all ST81 (n=46) isolates. In conclusion, the C. difficile ST81 clone showed a high level of resistance to fluoroquinolones in Beijing, highlighting the need for nationwide surveillance of CDI.