Dihydroartemisinin inhibits plasmid transfer in drug-resistant Escherichia coli via limiting energy supply

Conjugative transfer of antibiotic resistance genes(ARGs)by plasmids is an important route for ARG dissemination.An increasing number of antibiotic and nonantibiotic compounds have been reported to aid the spread of ARGs,highlighting potential challenges for controlling this type of horizontal transfer.Development of conjugation inhibitors that block or delay the transfer of ARG-bearing plasmids is a promising strategy to control the propagation of antibiotic resistance.Although such inhibitors are rare,they typically exhibit relatively high toxicity and low efficacy in vivo and their mechanisms of action are inadequately understood.Here,we studied the effects of dihydroartemisinin(DHA),an artemisinin derivative used to treat malaria,on conjugation.DHA inhibited the conjugation of the IncI2 and IncX4 plasmids carrying the mobile colistin resistance gene(mcr-1)by more than 160-fold in vitro in Escherichia coli,and more than two-fold(IncI2 plasmid)in vivo in a mouse model.It also suppressed the transfer of the IncX3 plasmid carrying the carbapenem resistance gene bla_(NDM-5)by more than twofold in vitro.Detection of intracellular adenosine triphosphate(ATP)and proton motive force(PMF),in combination with transcriptomic and metabolomic analyses,revealed that DHA impaired the function of the electron transport chain(ETC)by inhibiting the tricarboxylic acid(TCA)cycle pathway,thereby disrupting PMF and limiting the availability of intracellular ATP for plasmid conjugative transfer.Furthermore,expression levels of genes related to conjugation and pilus generation were significantly down-regulated during DHA exposure,indicating that the transfer apparatus for conjugation may be inhibited.Our findings provide new insights into the control of antibiotic resistance and the potential use of DHA.

Calculation of Mass Concrete Temperature and Creep Stress under the Influence of Local Air Heat Transfer

Temperature-induced cracking during the construction of mass concrete is a significant concern.Numerical simulations of concrete temperature have primarily assumed that the concrete is placed in an open environment.The problem of heat transfer between the air and concrete has been simplified to the concrete’s heat dissipation boundary.However,in the case of tubular concrete structures,where air inlet and outlet are relatively limited,the internal air temperature does not dissipate promptly to the external environment as it rises.To accurately simulate the temperature and creep stress in tubular concrete structures with enclosed air spaces during construction,we establish an air–concrete coupled heat transfer model according to the principles of conjugate heat transfer,and the accuracy of the model is verified through experiments.Furthermore,we conduct a case study to analyze the impact of airflow within the ship lock corridor on concrete temperature and creep stress.The results demonstrate that enhancing airflow within the corridor can significantly reduce the maximum concrete temperature.Compared with cases in which airflow within the corridor is neglected,the maximum concrete temperature and maximum tensile stress can be reduced by 12.5℃ and 0.7 MPa,respectively,under a wind speed of 4 m/s.The results of the traditional calculation method are relatively close to those obtained at a wind speed of 1 m/s.However,the temperature reduction process in the traditional method is faster,and the method yields greater tensile stress values for the corridor location.

质粒介导的利奈唑胺耐药基因optrA在粪肠球菌的水平转移机制

目的研究临床上耐利奈唑胺粪肠球菌(LREf)携带的optrA基因通过质粒介导在粪肠球菌间的水平转移机制。方法收集昆明市某三级甲等医院2022年8月—2023年8月留取的非重复LREf,使用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)仪进行鉴定,VITEK 2 Compact、纸片扩散法、微量肉汤稀释法进行药敏试验,聚合酶链式反应(PCR)检测optrA基因,全基因组测序(WGS)分析LREf的分子生物学特征,以LREf为供体菌、临床来源粪肠球菌为受体菌进行接合实验。结果共收集17株LREf,12株LREf的质粒检测出optrA基因,检测出多种耐药基因及毒力基因。12株LREf菌株以ST16(50.0%)为主要分型。接合试验的24株接合子中有8株接合成功,接合率为33.3%,进一步分析发现,菌株接合前后位于质粒上的optrA基因上下游均存在IS1216E插入序列,且形成IS1216E-fexA-optrA-erm(A)-IS1216E样的转座单元。结论optrA基因可通过同时携带erm(A)、fexA基因的质粒介导在临床粪肠球菌间进行水平转移,而插入序列IS1216E在其水平转移过程中发挥着重要作用。

基于空气幕冷却的涡轮端壁改进冷却结构的高温数值验证

叶栅端壁是航空发动机高压涡轮中流动结构与传热冷却特征最为复杂的区域之一。为提高采用轴向气膜冷却孔布局的原型涡轮端壁的综合冷却性能,本文依据各区域的流场结构和热负荷特性有针对性地进行局部高效气膜冷却强化设计,包括应用端壁通道进口空气幕冷却、扇形气膜孔以及等马赫数线优化布局等方法,使得冷气可以克服横流和二次流的影响,冷气附壁性和覆盖面积大幅提升。本文采用数值模拟方法在某型航空发动机高压涡轮真实进口条件下,验证了端壁改进冷却设计的耦合换热特性。与端壁原型冷却设计相比,在相同冷气消耗量下,端壁改进冷却设计在涡轮真实平均进口温度2150 K下实现了更低且更均匀的金属温度分布,有效消除了原型冷却设计中存在的局部高温热斑,端壁面积平均综合冷却有效度提升了14.2%。叶栅气动分析表明,端壁改进冷却设计还可以降低叶栅出口总压损失,改善叶栅气动性能。冷气流线分布规律表明,空气幕冷却在端壁应用中表现出良好的冷却性能;当空气幕冷气吹风比达到1.85时,可以克服端壁附近涡系的影响而达到尾缘区域形成冷却,并削弱二次流对压力面侧气膜冷却的不利影响;但当空气幕冷气吹风比达到2.12时则会由于过大的射流动量而脱离端壁表面,从而损失部分冷却效果,因此需要合理分配冷气用量。

多孔介质扰流柱对双层壁冷却系统性能影响的数值研究

为了提升双层壁冷却系统性能并强化系统的冷却优势,将双层壁冷却系统的实体扰流柱结构替换为多孔介质结构,采用气热耦合方法数值研究了多孔介质结构对双层壁冷却系统的提升性能,分析并对比了不同吹风比、扰流柱直径与扰流柱高度对多孔介质结构和实体结构的流动及传热特性的影响规律。数值结果表明:对于多孔介质高渗透率结构,双层壁内、外壁面之间的速度和温度分布更加均匀;将实体扰流柱结构替换为多孔介质结构后,双层壁综合冷却有效度明显增大,面平均综合冷却有效度可提升3.08%~5.03%,而压力损失变化不大;随着多孔介质扰流柱直径的增大,综合冷却有效度呈现增大趋势,提升幅度为2.13%~3.03%;扰流柱高度对冷却性能的影响较为微弱。研究结果验证了多孔介质扰流柱在双层壁冷却系统中应用的可行性。

Construction of human eukaryotic expression plasmid vascular endothelial growth factor 165 and its expression in transfected vascular smooth muscles

BACKGROUND: The highly specific vascular endothelialgrowth factor (VEGF) induces the growth of vascular en-dothelial cell. This study was to construct the eukaryoticexpression plasmid of vascular endothelial growth factorl65(VEGF165) and observe its expression in vascular smoothmuscles (VSMCs).METHODS: The primers were designed and synthesizedaccording to the gene sequences of human VEGF165. TheVEGF165 gene was obtained from umbilic artery tissue bythe method of RT-PCR, then it was cloned to eukaryoticexpression plasmid pBudCE4.1 by recombination strategy.The eukaryotic expression plasmid named pBudCE4.1/VEGF165 was identified by restriction enzyme digestion,and was sequenced. The pBudCE4.1/VEGF165 was trans-fected into VSMCs by using lipofection. The VEGF165 ex-pression of mRNA and protein was detected by RT-PCRand Western blot respectively.RESULTS: VEGF165 was shown about 576bp by RT-PCR.Sequencing revealed the amplified VEGF165 gene was iden-tical with that in the GeneBank. Restrictive enzyme (HindBam HI) digestion analysis showed that recombinantexpression plasmid pBudCE4. l/tVEGF165 had been con-structed successfully. The expression of VEGF165 at mRNAand protein levels in the transformed VSMCs had beendemonstrated by RT-PCR and Western blot.CONCLUSIONS: The recombinant eukaryotic expressionplasmid pBudCE4.1/VEGF165 has been successfully con-structed and expressed in transformed VSMCs. The presentstudy has laid a foundation for VEGF165 gene therapy ofvascular stenosis in the transplant organ.

Towards a better understanding of antimicrobial resistance dissemination:what can be learnt from studying model conjugative plasmids?

Bacteria can evolve rapidly by acquiring new traits such as virulence,metabolic properties,and most importantly,antimicrobial resistance,through horizontal gene transfer(HGT).Multidrug resistance in bacteria,especially in Gram-negative organisms,has become a global public health threat often through the spread of mobile genetic elements.Conjugation represents a major form of HGT and involves the transfer of DNA from a donor bacterium to a recipient by direct contact.Conjugative plasmids,a major vehicle for the dissemination of antimicrobial resistance,are selfish elements capable of mediating their own transmission through conjugation.To spread to and survive in a new bacterial host,conjugative plasmids have evolved mechanisms to circumvent both host defense systems and compete with co-resident plasmids.Such mechanisms have mostly been studied in model plasmids such as the F plasmid,rather than in conjugative plasmids that confer antimicrobial resistance(AMR)in important human pathogens.A better understanding of these mechanisms is crucial for predicting the flow of antimicrobial resistance-conferring conjugative plasmids among bacterial populations and guiding the rational design of strategies to halt the spread of antimicrobial resistance.Here,we review mechanisms employed by conjugative plasmids that promote their transmission and establishment in Gram-negative bacteria,by following the life cycle of conjugative plasmids.

Construction of human VEGF165 gene eukaryotic expression plasmid and its effect on proliferation of vascular endothelial cells

After organ transplantation, rapid repair of injured vascular endothelial cell (VEC) is a key to prevent graft chronic dysfunction besides control of immunological rejection. Many studies have confirmed that vascular endothelial growth factor 165 (VEGF165) could accelerate the repair of VEC injury, decrease thrombosis and thrombotic occlusion, and inhibit hyperplasia of the intima. This study was designed to construct eukaryotic expression plasmid pBudCE4.1/VEGF165, and observe its effect on the prolife ration of VEC. METHODS:The VEGF165 gene cloned from human heart tissue by RT-PCR was cloned into eukaryotic expression plasmid pBudCE4.1. The recombinant expression plasmid pBudCE4.1/VEGF165 was identified by restriction enzyme (Hind III and BamH I) digestion analysis, and was sequenced. The pBudCE4.1/VEGF165 was introduced into VEC through lipofection transfection. The VEGF165 mRNA expression by Northern blot and VEGF165 protein expression was detected by immunocytochemical staining. The effect of expression protein on VEC proliferation was detected by flow cytometry. RESULTS:The RT-PCR product of the VEGF165 gene was about 576bp. Sequencing analysis revealed that the sequence of the amplified VEGF165 gene was identical with that in GenBank. Restrictive enzyme digestion analysis showed that recombinant expression plasmid pBudCE4.1/ tVEGF165 had been constructed successfully. The expression of VEGF165 at mRNA and protein levels in the transformed VSMCs had been demonstrated by Northern blot and immunocytochemical staining respectively. The expressed product of VEGF165 could notably accelerate the proliferation of VECs. CONCLUSIONS:pBudCE4.1/VEGF165 is successfully cons- tructed and is expressed in VECs. Expressed VEGF165 can accelerate the VEC proliferation. The present study has laid a foundation for potential use of VEGF165 gene transfection to prevent and treat vascular stenosis in the transplanted organ.

Conjugate Heat Transfer Analysis of an Ultrasonic Molten Metal Treatment System

In piezoceramic ultrasonic devices,the piezoceramic stacks may fail permanently or function improperly if their working temperatures overstep the Curie temperature of the piezoceramic material.While the end of the horn usually serves near the melting point of the molten metal and is enclosed in an airtight chamber,so that it is difficult to experimentally measure the temperature of the transducer and its variation with time,which bring heavy difficulty to the design of the ultrasonic molten metal treatment system.To find a way out,conjugate heat transfer analysis of an ultrasonic molten metal treatment system is performed with coupled fluid and heat transfer finite element method.In modeling of the system,the RNG model and the SIMPLE algorithm are adopted for turbulence and nonlinear coupling between the momentum equation and the energy equation.Forced air cooling as well as natural air cooling is analyzed to compare the difference of temperature evolution.Numerical results show that,after about 350 s of working time,temperatures in the surface of the ceramic stacks in forced air cooling drop about 7 K compared with that in natural cooling.At 240 s,The molten metal surface emits heat radiation with a maximum rate of about 19 036 W/m2,while the heat insulation disc absorbs heat radiation at a maximum rate of about 7922 W/m2,which indicates the effectiveness of heat insulation of the asbestos pad.Transient heat transfer film coefficient and its distribution,which are difficult to be measured experimentally are also obtained through numerical simulation.At 240 s,the heat transfer film coefficient in the surface of the transducer ranges from–17.86 to 20.17 W/（m2？K）.Compared with the trial and error method based on the test,the proposed research provides a more effective way in the design and analysis of the temperature control of the molten metal treatment system.

INCREASED EXPRESSION OF DIRECT GENE TRANSFER INISCHEMIC SKELETAL MUSCLES OF RABBITS

Objective The gene expression of skeletal muscle under ischemic condition by direct gene injectionwas observed in order to find a new gene delivery method to treat chronic arterial occlusion disease. Methods Weestablished the rabbit hindlimb ischemic model and used plasmid PSV-β-gal as a reporter gene. We transferedgene intramuscularly and detected the activity of β-galactosidase by histochemistry method. Results Weobserved that the gene expression of skeletal muscle under ischemic condition was higher than normalmuscle. Conclusion The result demonstrated that the direct gene injection was suitable for the chronic peripheralarterial occlusion disease, and might be a novel gene delivery method for this disease.

Streamline upwind finite element method for conjugate heat transfer problems

This paper presents a combined finite element method for solving conjugate heat transfer problems where heat conduction in a solid is coupled with heat convection in viscous fluid flow. The streamline upwind finite element method is used for the analysis of thermal viscous flow in the fluid region, whereas the analysis of heat conduction in solid region is performed by the Galerkin method. The method uses the three-node triangular element with equal-order interpolation functions for all the variables of the velocity components, the pressure and the temperature. The main advantage of the proposed method is to consistently couple heat transfer along the fluid-solid interface. Three test cases, i.e. conjugate Couette flow problem in parallel plate channel, counter-flow in heat exchanger, and conjugate natural convection in a square cavity with a conducting wall, are selected to evaluate the efficiency of the present method.

Numerical Simulation and Control of Two-Phase Flow with Evaporation in a Vertical Tube Submitted to a Conjugate Heat Transfer

A better understanding of two-phase flows with evaporation allows leading to an optimal design of evaporators. For that purpose, numerical simulations are very useful. In this paper, a numerical study has been carried out in order to model and simulate the combination of a two-phase flow with evaporation in a vertical tube. The VOF （volume-of-fluid） multiphase flow method and a phase-change model for the mass transfer have been used. For an accurate modeling, the effect of axial conduction has been also taken into account using a conjugate heat transfer model. Since thermal oscillations are undesirable as they can lead to the failure of the tube, flow instabilities have also been analyzed, using FFT （fast Fourier transforms）, in order to comprehend their behavior and influence. A control study of the flow instabilities in the tube is also presented. For that purpose tube inlet temperature has been varied using a gain control parameter.

The Expression of the Plasmid DNA Encoding TGF-β_1 in Endothelium after Injection into the Anterior Chamber

The method of gene transfer into corneal endothelium was investigated to provide a foundation for the study of TGF-β1 gene transfer to inhibit corneal graft rejection.Two days after direct injection of p MAM TGF-β1 mediated by liposom e into the anterior cham ber of rabbits,one half of corneas were made into paraffin slides and the endothelial layer was carefully torn from the other half to make a single layer slide of endothelia.By means of im munohistochemical technique, the plasmid p MAM TGF- β1 expression product TGF- β1 in the endothelia was detected.Specific TGF- β1 expression was positive in the endothelia on both the paraffin slide and the single layer slide.The results showed that by direct injection into the anterior cham ber,foreign plasmid DNA could be transferred into the endothelia and its expression was obtained.This may provide a foun- dation for further study on TGF-β1 participating in local induction of corneal imm une tolerance.

CONJUGATE MODEL FOR HEAT AND MASS TRANSFER OF POROUS WALL IN THE HIGH TEMPERATURE GAS FLOW

Heat and mass transfer of a porous permeable wall in a high temperature gas dynamical flow is considered. Numerical simulation is conducted on the ground of the conjugate mathematical model which includes filtration and heat transfer equations in a porous body and boundary layer equations on its surface. Such an approach enables one to take into account complex interaction between heat and mass transfer in the gasdynamical flow and in the structure subjected to this flow. The main attention is given to the impact of the intraporous heat transfer intensity on the transpiration cooling efficiency.

Analysis of Drug Resistance Spectra and Conjugative Plasmid Carrying Rates of P.aeruginosa and Acinetobacter

Antimicrobial susceptibility test was performed on 57 clinical isolates of P. aeruginosa and 36 clinical isolates of Acinetobacter with 11 antimicrobial agents including getamicin, amikacin, ciprofloxacin, ofloxacin, fleroxacin, piperacillin, cefotaxime, cefoperazone/sulbactam, ceftazidime, cefoperazone and doxycycline. Transferable drug resistance plasmid carrying rates of these clinical isolates were also studied. On the basis of the in vitro activities, 52.63%(30/57) of the isolated strains of P. aeruginosa were susceptible to antimicrobial agents selected (except doxycycline), 41.67%(15/36) of the isolated strains of Acinetobacter were susceptible to 11 antimicrobial agents. The sensitivity rate of P.aeruginosa and Acinetobacter to antimicrobial agents selected was 70% or greater to all except doxycycline. Furthermore, the sensitivity rate of P.aeruginosa to amikacin ciprofloxacin, ceftazidime, cefoperazone, cefoperazone/sulbactam, and that of Acinetobacter to cefoperazone/sulbactam, amikacin was more than 90%,among them amikacin, cefoperazone/sulbactam being the most effective. Plasmid analysis showed that 15.79%(9/57) P.aeruginosa strains and 13.89%(5/36) Acinetobacter strains carried plasmid. Conjugative plasmid carrying rates of P. aeruginosa strains and Acinetobacter strains were 7.02%(4/57), 13.89%(5/36), respectively. Conjugative plasmid didn′t play an important role in the formation and dissemination of drug resistance of P. aeruginosa and Acinetobacter.

Accurate reconstruction of the optical parameter distribution in participating medium based on the frequency-domain radiative transfer equation

Reconstructing the distribution of optical parameters in the participating medium based on the frequency-domain radiative transfer equation （FD-RTE） to probe the internal structure of the medium is investigated in the present work. The forward model of FD-RTE is solved via the finite volume method （FVM）. The regularization term formatted by the generalized Gaussian Markov random field model is used in the objective function to overcome the ill-posed nature of the inverse problem. The multi-start conjugate gradient （MCG） method is employed to search the minimum of the objective function and increase the efficiency of convergence. A modified adjoint differentiation technique using the collimated radiative intensity is developed to calculate the gradient of the objective function with respect to the optical parameters. All simulation results show that the proposed reconstruction algorithm based on FD-RTE can obtain the accurate distributions of absorption and scattering coefficients. The reconstructed images of the scattering coefficient have less errors than those of the absorption coefficient, which indicates the former are more suitable to probing the inner structure.

Construction and Expression of Recombinant Plasmid pCD-rbFGF in Osteoblasts

Summary: To construct basic fibroblast growth factor (bFGF) eukaryotic expression vector and to evaluate the possibility of bFGF gene therapy in orthopedic disease, the pCD-rbFGF recombinant plasmid was constructed by cloning rat basic fibroblast growth factor (bFGF) cDNA into an eukaryotic expression vector, pcDNA 3. Rat osteoblasts were transfected with pCD-rbFGF plasmid by lopofectin mediated gene transfer, the transient expression was detected by streptavidin-biotin-enzyme complex (SABC) method. It was observed that the expression of rat bFGF gene was detected 72 h after transfected distinctly. Basic fibroblast growth factor gene therapy is a method of potential for a wide array of orthopedic diseases.

Construction of plasmid containing human blood- clotting factor Ⅷ and observation its expression in Cos-7 cells

Factor Ⅷ deficiency or hemophilia A is X-linkedgenetic disorder in human. General treatment of severehemophilia A consists of adiministration of plasma-derived or recombinant clotting factor concentrates. Ithas caused a series of problems, i. e. viral infection andcost too much to use rF Ⅷ. Nowadays, people havedeveloped the retroviral vector and the adcnoviral

Investigation of convection cooling guide vane with conjugate heat transfer method

This paper studied a certain blade with ten radial cooling holes which employed conjugate heat transfer method. The cooling air entered the cooling channel from the bottom of the blade and went out from the top, it was not ejected into the main flow. This paper used different numerical conditions including different turbulence models,turbulence intensities,thermal conduction coefficients and the influence on fluid property via temperature variation. The temperature distribution and pressure distribution of the blade were compared with experimental data. The results show that the numerical results using different turbulence models are almost identical to experimental data even little deviation occurs at shock wave location. The trends of temperature distribution under different numerical conditions are coincident to experimental data,especially Reynolds stress turbulence model. It can be concluded that anisotropic turbulence models can simulate the transition from laminar to turbulence,and the influence of turbulence intensity on laminar region and transition region is more than that on developed turbulent region.

Theoretical Study of the Isomerization Reaction of 1-3 H Transfer on Formamidine Substituted by Halogen

The ab initio method has been used to study the 1-3 H transfer reaction on formamidine substituted by halogen. The calculation results show that the substituted halogen has two effects on the 1-3 H transfer reaction: decreasing the activation energy and stabilizing the C=N double bond owing to the conjugative effect of p-π-p of products and transition states.