Identification of Single Nucleotide Polymorphisms Associated with Hyperproduction of Alpha-Toxin in Staphylococcus aureus

The virulence factor α-toxin (hla) is needed by Staphylococcus aureus in order to cause infections in both animals and humans. Although the complicated regulation of hla expression has been well studied in human S. aureus isolates, the mechanisms of of hla regulation in bovine S. aureus isolates remain undefined. In this study, we found that many bovine S. aureus isolates, including the RF122 strain, generate dramatic amounts of α-toxin in vitro compared with human clinical S. aureus isolates, including MRSA WCUH29 and MRSA USA300. To elucidate potential regulatory mechanisms, we analyzed the hla promoter regions and identified predominant single nucleotide polymorphisms (SNPs) at positions −376, −483, and −484 from the start codon in α-toxin hyper-producing isolates. Using site-directed mutagenesis and hla promoter-gfp-luxABCDE dual reporter approaches, we demonstrated that the SNPs contribute to the differential control of hla expression among bovine and human S. aureus isolates. Using a DNA affinity assay, gel-shift assays and a null mutant, we identified and revealed that an hla positive regulator, SarZ, contributes to the involvement of the SNPs in mediating hla expression. In addition, we found that the bovine S. aureus isolate RF122 exhibits higher transcription levels of hla positive regulators, including agrA, saeR, arlR and sarZ, but a lower expression level of hla repressor rot compared to the human S. aureus isolate WCUH29. Our results indicate α-toxin hyperproduction in bovine S. aureus is a multifactorial process, influenced at both the genomic and transcriptional levels. Moreover, the identification of predominant SNPs in the hla promoter region may provide a novel method for genotyping the S. aureus isolates

Inclusion of 2d transition metal dichalcogenides in perovskite inks and their influence on solar cell performance

Funding Information: Acknowledgments: V.N. wishes to thank the support from the ERC 3D2DPrint CoG Grant. The authors gratefully acknowledge the project Best4U—“Tecnologia per celle solari bifacciali ad alta Efficienza a 4 terminali per utility scale”. The authors are grateful to the “Progetto Tecnopolo per la Medicina di precisione, Deliberazione della Giunta Regionale n. 2117 del 21 November 2018”.Organic–inorganic hybrid perovskite materials have raised great interest in recent years due to their excellent optoelectronic properties, which promise stunning improvements in photovoltaic technologies. Moreover, two-dimensional layered materials such as graphene, its derivatives, and transition metal dichalcogenides have been extensively investigated for a wide range of electronic and optoelectronic applications and have recently shown a synergistic effect in combination with hybrid perovskite materials. Here, we report on the inclusion of liquid-phase exfoliated molybdenum disulfide nanosheets into different perovskite precursor solutions, exploring their influence on final device performance. We compared the effect of such additives upon the growth of diverse perovskites, namely CH3NH3PbI3 (MAPbI3 ) and triple-cation with mixed halides Csx (MA0.17FA0.83 )(1−x)Pb (I0.83Br0.17 )3 perovskite. We show how for the referential MAPbI3 materials the addition of the MoS2 additive leads to the formation of larger, highly crystalline grains, which result in a remarkable 15% relative improvement in power conversion efficiency. On the other hand, for the mixed cation– halide perovskite no improvements were observed, confirming that the nucleation process for the two materials is differently influenced by the presence of MoS2 .publishersversionpublishe

Coupling of Cell Surface Biotinylation and SILAC-Based Quantitative Proteomics Identified Myoferlin as a Potential Therapeutic Target for Nasopharyngeal Carcinoma Metastasis

Distant metastasis is a major cause of treatment failure in nasopharyngeal carcinoma (NPC) patients. Cell surface proteins represent attractive targets for cancer diagnosis or therapy. However, the cell surface proteins associated with NPC metastasis are poorly understood. To identify potential therapeutic targets for NPC metastasis, we isolated cell surface proteins from two isogenic NPC cell lines, 6-10B (low metastatic) and 5-8F (highly metastatic), through cell surface biotinylation. Stable isotope labeling by amino acids in cell culture (SILAC) based proteomics was applied to comprehensively characterize the cell surface proteins related with the metastatic phenotype. We identified 294 differentially expressed cell surface proteins, including the most upregulated protein myoferlin (MYOF), two receptor tyrosine kinases(RTKs) epidermal growth factor receptor (EGFR) and ephrin type-A receptor 2 (EPHA2) and several integrin family molecules. These differentially expressed proteins are enriched in multiple biological pathways such as the FAK-PI3K-mTOR pathway, focal adhesions, and integrin-mediated cell adhesion. The knockdown of MYOF effectively suppresses the proliferation, migration and invasion of NPC cells. Immunohistochemistry analysis also showed that MYOF is associated with NPC metastasis. We experimentally confirmed, for the first time, that MYOF can interact with EGFR and EPHA2. Moreover, MYOF knockdown could influence not only EGFR activity and its downstream epithelial–mesenchymal transition (EMT), but also EPHA2 ligand-independent activity. These findings suggest that MYOF might be an attractive potential therapeutic target that has double effects of simultaneously influencing EGFR and EPHA2 signaling pathway. In conclusion, this is the first study to profile the cell surface proteins associated with NPC metastasis and provide valuable resource for future researches

SARS-CoV Infection in a Restaurant from Palm Civet

Contact with food animals was associated with SARS-CoV infection in the People’s Republic of China

The C-Terminal Domain of the Novel Essential Protein Gcp Is Critical for Interaction with Another Essential Protein YeaZ of Staphylococcus aureus

Previous studies have demonstrated that the novel protein Gcp is essential for the viability of various bacterial species including Staphylococcus aureus; however, the reason why it is required for bacterial growth remains unclear. In order to explore the potential mechanisms of this essentiality, we performed RT-PCR analysis and revealed that the gcp gene (sa1854) was co-transcribed with sa1855, yeaZ (sa1856) and sa1857 genes, indicating these genes are located in the same operon. Furthermore, we demonstrated that Gcp interacts with YeaZ using a yeast two-hybrid (Y2H) system and in vitro pull down assays. To characterize the Gcp-YeaZ interaction, we performed alanine scanning mutagenesis on the residues of C-terminal segment of Gcp. We found that the mutations of the C-terminal Y317-F322 region abolished the interaction of Gcp and YeaZ, and the mutations of the D324-N329 and S332-Y336 regions alleviated Gcp binding to YeaZ. More importantly, we demonstrated that these key regions of Gcp are also necessary for the bacterial survival since these mutated Gcp could not complement the depletion of endogenous Gcp. Taken together, our data suggest that the interaction of Gcp and YeaZ may contribute to the essentiality of Gcp for S. aureus survival. Our findings provide new insights into the potential mechanisms and biological functions of this novel essential protein

Purification and Characterization of a Novel Chlorpyrifos Hydrolase from Cladosporium cladosporioides Hu-01

Chlorpyrifos is of great environmental concern due to its widespread use in the past several decades and its potential toxic effects on human health. Thus, the degradation study of chlorpyrifos has become increasing important in recent years. A fungus capable of using chlorpyrifos as the sole carbon source was isolated from organophosphate-contaminated soil and characterized as Cladosporium cladosporioides Hu-01 (collection number: CCTCC M 20711). A novel chlorpyrifos hydrolase from cell extract was purified 35.6-fold to apparent homogeneity with 38.5% overall recovery by ammoniumsulfate precipitation, gel filtration chromatography and anion-exchange chromatography. It is a monomeric structure with a molecular mass of 38.3 kDa. The pI value was estimated to be 5.2. The optimal pH and temperature of the purified enzyme were 6.5 and 40°C, respectively. No cofactors were required for the chlorpyrifos-hydrolysis activity. The enzyme was strongly inhibited by Hg2+, Fe3+, DTT, β-mercaptoethanol and SDS, whereas slight inhibitory effects (5–10% inhibition) were observed in the presence of Mn2+, Zn2+, Cu2+, Mg2+, and EDTA. The purified enzyme hydrolyzed various organophosphorus insecticides with P-O and P-S bond. Chlorpyrifos was the preferred substrate. The Km and Vmax values of the enzyme for chlorpyrifos were 6.7974 μM and 2.6473 μmol·min−1, respectively. Both NH2-terminal sequencing and matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometer (MALDI-TOF-MS) identified an amino acid sequence MEPDGELSALTQGANS, which shared no similarity with any reported organophosphate-hydrolyzing enzymes. These results suggested that the purified enzyme was a novel hydrolase and might conceivably be developed to fulfill the practical requirements to enable its use in situ for detoxification of chlorpyrifos. Finally, this is the first described chlorpyrifos hydrolase from fungus

Broadening of Neutralization Activity to Directly Block a Dominant Antibody-Driven SARS-Coronavirus Evolution Pathway

Phylogenetic analyses have provided strong evidence that amino acid changes in spike (S) protein of animal and human SARS coronaviruses (SARS-CoVs) during and between two zoonotic transfers (2002/03 and 2003/04) are the result of positive selection. While several studies support that some amino acid changes between animal and human viruses are the result of inter-species adaptation, the role of neutralizing antibodies (nAbs) in driving SARS-CoV evolution, particularly during intra-species transmission, is unknown. A detailed examination of SARS-CoV infected animal and human convalescent sera could provide evidence of nAb pressure which, if found, may lead to strategies to effectively block virus evolution pathways by broadening the activity of nAbs. Here we show, by focusing on a dominant neutralization epitope, that contemporaneous- and cross-strain nAb responses against SARS-CoV spike protein exist during natural infection. In vitro immune pressure on this epitope using 2002/03 strain-specific nAb 80R recapitulated a dominant escape mutation that was present in all 2003/04 animal and human viruses. Strategies to block this nAb escape/naturally occurring evolution pathway by generating broad nAbs (BnAbs) with activity against 80R escape mutants and both 2002/03 and 2003/04 strains were explored. Structure-based amino acid changes in an activation-induced cytidine deaminase (AID) “hot spot” in a light chain CDR (complementarity determining region) alone, introduced through shuffling of naturally occurring non-immune human VL chain repertoire or by targeted mutagenesis, were successful in generating these BnAbs. These results demonstrate that nAb-mediated immune pressure is likely a driving force for positive selection during intra-species transmission of SARS-CoV. Somatic hypermutation (SHM) of a single VL CDR can markedly broaden the activity of a strain-specific nAb. The strategies investigated in this study, in particular the use of structural information in combination of chain-shuffling as well as hot-spot CDR mutagenesis, can be exploited to broaden neutralization activity, to improve anti-viral nAb therapies, and directly manipulate virus evolution

The Authoritarian Environmentalism in China : - “air policy” implementation research in Jing-Jin-Ji region as case studies

The concept of “Environmental authoritarian” becomes the focus of discussion in the academic field of environmental politics since it has limited observations. In the recent years, the severe air pollute issue is considered as a threat by both citizens and authorities in China. Along with the more stricter environmental protection laws being issued, even the measures of the factory closings and coal ban seems to become acceptable as the population  have suffered in the heavy smog for years. Does this led to the rise of “environmental authoritarian” in China?  Taking this as a heuristic point, I observe the positions of various stakeholders in the environmental governance as well as present the findings from research on the implementation and enforcement of air pollution controls measures in Jing-Jin-Ji region. I offer a critical examination of “environmental authoritarian”, especially the levels of public participation at the policy process. After to do the research, the key funding is that the concept of “Environmental authoritarian” is too simplified, it cannot cover the complex and detailed environmental governance at local empirical cases

Two-dimensional Materials for Energy Storage and Harvesting

Energy storage and harvesting have become a global concern due to the ever-increasing demand for energy and upcoming depletion of natural resources. As a family of Van der Waals layered crystals, metal chalcogenides (MCs) and transition metal chalcogenides (TMDs) possess unique electronic and photonic properties, enabling high-performance electronic devices for broad applications. However, the potential application of MCs and TMDs in energy devices remains relatively unexplored, partially due to their challenges in exfoliation as well as poor electrical conductivity. This PhD thesis mainly discussed the preparation and energy application of 2D MCs and TMDs materials. The liquid phase exfoliation (LPE) method was demonstrated as a viable method to give large-scale 2D MCs and TMDs nanosheets at low cost, enabling efficient solution processing of thin films, composites and devices. The resultant 2D MCs and TMDs nanosheets were utilized to fabricate high-performance lithium-ion batteries (LiBs) and solar cells (SCs). Four different 2D MCs (i.e., GaS, gallium (II) selenide [GaSe], gallium (II) telluride [GaTe] and indium (II) selenide [InSe]) were percolated in single-wall carbon nanotube (SWCNT) networks to obtain the conductive and flexible anode for LiBs. Based on the general comparison of them, we found the 2D InSe-SWCNT composite anode exhibits a superior electrochemical performance (including high capacity and excellent rate capability). Importantly, the capacity per InSe of 2D InSe-SWCNT composite anode increases over prolonged cycling up to 1224 mAh g-1 from 520 mAh g-1 (after 254 cycles, at 500 mA g-1), which is believed to largely relate with the reversible alloy reaction, as confirmed by the operando X-ray diffraction results. By combining the density functional theory (DFT) calculations and post-cycling scanning electron microscope (SEM) images, we reveal that the in situ formed In particles gradually reduce their domain size, forming nanoclusters upon cycling which are capable to accommodate four Li+ instead of one per atomic In, and leading to extra capacity as increasing the cycling numbers. Such a new "nanocluster-alloying" Li storage mechanism may inspire new architectures or methods to prepare anode materials for excellent performance LiBs. Besides, 2D TMDs (e.g. MoS2, MoSe2, etc.) were tried as 1) absorber, 2) hole transport layer (HTL), and 3) buffer layer of solar cells. Through this study, we found that 2D TMDs obtained via LPE method is not suitable to be an active layer of solar cells due to the limitation of reducing the 2D TMDs nanosheets thickness. However, they have potential to be used as a HTL or a buffer layer of perovskite solar cells (PSCs) to reduce the cost and improve the stability of the device. There is still a large space to improve the PV performance of solar cells with 2D TMDs nanosheets, and this practical built the foundation for further research