Detection of Novel BEST1 Variations in Autosomal Recessive Bestrophinopathy Using Third-generation Sequencing

Objective:Autosomal recessive bestrophinopathy(ARB),a retinal degenerative disease,is characterized by central visual loss,yellowish multifocal diffuse subretinal deposits,and a dramatic decrease in the light peak on electrooculogram.The potential pathogenic mechanism involves mutations in the BEST1 gene,which encodes Ca2+-activated Cl−channels in the retinal pigment epithelium(RPE),resulting in degeneration of RPE and photoreceptor.In this study,the complete clinical characteristics of two Chinese ARB families were summarized.Methods:Pacific Biosciences(PacBio)single-molecule real-time(SMRT)sequencing was performed on the probands to screen for disease-causing gene mutations,and Sanger sequencing was applied to validate variants in the patients and their family members.Results:Two novel mutations,c.202T>C(chr11:61722628,p.Y68H)and c.867+97G>A,in the BEST1 gene were identified in the two Chinese ARB families.The novel missense mutation BEST1 c.202T>C(p.Y68H)resulted in the substitution of tyrosine with histidine in the N-terminal region of transmembrane domain 2 of bestrophin-1.Another novel variant,BEST1 c.867+97G>A(chr11:61725867),located in intron 7,might be considered a regulatory variant that changes allele-specific binding affinity based on motifs of important transcriptional regulators.Conclusion:Our findings represent the first use of third-generation sequencing(TGS)to identify novel BEST1 mutations in patients with ARB,indicating that TGS can be a more accurate and efficient tool for identifying mutations in specific genes.The novel variants identified further broaden the mutation spectrum of BEST1 in the Chinese population.

Recent progress in third-generation low alloy steels developed under M^3 microstructure control

During the past thirty years, two generations of low alloy steels(ferrite/pearlite followed by bainite/martensite) have been developed and widely used in structural applications. The third-generation of low alloy steels is expected to achieve high strength and improved ductility and toughness, while satisfying the new demands for weight reduction, greenness, and safety. This paper reviews recent progress in the development of third-generation low alloy steels with an M^3 microstructure, namely, microstructures with multi-phase, meta-stable austenite, and multi-scale precipitates. The review summarizes the alloy designs and processing routes of microstructure control, and the mechanical properties of the alloys.The stabilization of retained austenite in low alloy steels is especially emphasized. Multi-scale nano-precipitates, including carbides of microalloying elements and Cu-rich precipitates obtained in third-generation low alloy steels, are then introduced. The structure–property relationships of third-generation alloys are also discussed. Finally, the promises and challenges to future applications are explored.

Innovation and development of the third-generation hybrid rice technology

The breeding and large-scale application of hybrid rice contribute significantly to the food supply worldwide.Currently,hybrid seed production uses cytoplasmic male sterile(CMS)lines or photoperiod/thermo-sensitive genic male sterile(PTGMS)lines as female parent.Despite huge successes,both systems have intrinsic problems.CMS systems are mainly restricted by the narrow restorer resources that make it difficult to breed superior hybrids,while PTGMS systems are limited by conditional sterility of the male sterile lines that makes the propagation of both PTGMS seeds and hybrid seeds vulnerable to unpredictable climate changes.Recessive nuclear male sterile(NMS)lines insensitive to environmental conditions are widely distributed and are ideal for hybrid rice breeding and production,but the lack of effective ways to propagate the pure NMS lines in a large scale renders it impossible to use them for hybrid rice production.The development of"the third-generation hybrid rice technology"enables efficient propagation of the pure NMS lines in commercial scale.This paper discusses the establishment of"the thirdgeneration hybrid rice technology"and further innovations.This new technology breaks the limitations of CMS and PTGMS systems and will bring a big leap forward in hybrid rice production.

Applications of sediment sudden deposition model based on the third-generation numerical model for shallow water wave

The existing numerical models for nearshore waves are briefly introduced, and the third-generation numerical model for shallow water wave, which makes use of the most advanced productions of wave research and has been adapted well to be used in the environment of seacoast, lake and estuary area, is particularly discussed. The applied model realizes the significant wave height distribution at different wind directions. To integrate the model into the coastal area sediment, sudden deposition mechanism, the distribution of average silt content and the change of sediment sudden deposition thickness over time in the nearshore area are simulated. The academic productions can give some theoretical guidance to the applications of sediment sudden deposition mechanism for stormy waves in the coastal area. And the advancing directions of sediment sudden deposition model are prospected.

The Influences of American Deep Image on the Third-Generation Poets in China

The Third-Generation Poetry of China （namely Post-misty Poetry too） initiated with the introduction of Western modernist poetry, especially sorts of American Post-modernist poetry schools into China. ＂The relation between American poetry and Chinese poetry has a long history, which lies in the influences on the creation of the Third-Generation poets. This influence is probably unprecedented in its depth and breadth.＂ ＂Irrational association＂ and ＂leaping images＂ proposed by American Deep Image poets influenced by Freudian and Jungian unconscious perception gained an extraordinary appreciation among the Third-Generation poets who were in pursuit constantly of the experiments on poetic form and language. This paper mainly discusses the influences of American Deep Image on the Third-Generation poets of China through a case study of WANG Yin and CHEN Dongdong＇s poems.

Development of the Mechanical System on a Third-Generation Industrial Computed Tomography Scanner in Brazil

The development of measurement geometry for medical X-ray computed tomography （CT） scanners was carried out from the first to the fourth-generation. This concept has also been applied for imaging of industrial processes such as pipe flows or for improving design, operation, optimization and troubleshooting. Nowadays, gamma CT permits to visualize failure equipment points in three-dimensional analysis and in sections of chemical and petrochemical industries. The aim of this work is the development of the mechanical system on a third-generation industrial CT scanner to analyze laboratorial process columns which perform highly efficient separation, turning the ^6oCo, ^75Se, ^137Cs and/or ^192Ir sealed gamma-ray source（s） and the NaI（Tl） multidetector array. It also has a translation movement along the column axis to obtain as many slices of the process flow as needed. The mechanical assembly for this third-generation industrial CT scanner is comprised by strength and rigidity structural frame in stainless and carbon steels, rotating table, source shield and collimator with pneumatic exposure system, spur gear system, translator, rotary stage, drives and stepper motors. The use of suitable spur gears has given a good repeatability and high accuracy in the degree of veracity. The data acquisition boards, mechanical control interfaces, software for movement control and image reconstruction were specially development. A multiphase phantom capable to be setting with solid, liquid and gas was testing. The scanner was setting for 90 views and 19 projections for each detector totalizing 11,970 projections. Experiments to determine the linear attenuation coefficients of the phantom were carried out which applied the Lambert-Beer principle. Results showed that it was possible to distinguish between the phases even the polymethylmethacrylate and the water have very similar density and linear attenuation coefficients. It was established that the newly developed third-generation fan-beam arrangement gamma scanner unit has a good spatial resolution acceptable given the size of the used phantom in this study. The tomografic reconstruction algorithm in used 60 ~ 60 pixels images was the Alternative Minimization （AM） technique and was implemented in MATLAB and VB platforms. The mechanical system presented a good performance in terms of strength, rigidity, accuracy and repeatability with great potential to be used for education or program which dedicated to training chemical and petrochemical industry professionals and for industrial process optimization in Brazil.

Analysis of Polymer Impurities in Cephalosporin Antibiotics

[Objectives]To establish a HPLC-MS method for the determination of polymer impurities in cefathiamidine and its preparations.[Methods]Kromasil 100-5 C_(18) column(4.6 mm ×250 mm,5μm)was used for analysis;mobile phase ammonium acetate solution(pH 6.30)-acetonitrile,gradient elution;volumetric flow rate 1.0 mL/min;column temperature 40℃;multi-reaction monitoring mode was used for analysis,and positive ion scanning was chosen as the electrospray ion source.[Results]The resolution between impurities and main peaks under this method was greater than 1.5,and 8 known impurities and 2 polymer impurities could be completely separated and distinguish-ed.It was inferred that the molecular ion peak[M+H]^(+):m/z727.1874,m/z 785.1937 was the possible polymer impurity of this product.[Conclusions]A method for the analysis of polymer impurities in cefathiamidine and its preparations was formed,which could achieve the purpose of simultaneous analysis of small molecule impurities and polymer impurities,and could better control the content of single impurities in the polymer,providing a reliable inspection basis for strict control of cefathiamidine quality.

Historical Cohort Study of the Efficacy and Safety of Piperacillin/Tazobactam Versus Fourth-Generation Cephalosporins for Empirical Treatment of Febrile Neutropenia in Patients with Hematological Malignancies

We retrospectively evaluated the efficacy and safety of the combination drug piperacillin/tazobactam (PIPC/TAZ) in comparison with those of fourth-generation cephalosporins (4th Cephs) as initial empirical treatment in hematological malignancies patients with febrile neutropenia (FN). Among 200 patients assessed in this study, 49 had received PIPC/TAZ and 151 4th Cephs. Patient background characteristics were comparable between the two treatment groups. The overall efficacy rate in those receiving 4th Cephs and PIPC/TAZ was 57.0% (86/151 patients) and 59.2% (29/49 patients), respectively, with no significant difference detected between the two treatment regimens (P = 0.78). Treat-ment did not need to be discontinued or interrupted due to development of adverse drug reactions in any of the patients. Therefore in this study the efficacy and safety of PIPC/TAZ as initial antimicrobial treatment for FN in patients with hematological malignancies were not inferior to those of 4th Cephs. Based on the preliminary data obtained in this study, we propose to conduct a multicenter, prospective, controlled study to compare PIPC/TAZ versus CFPM given as empirical antimicrobial treatment against FN in patients with hematological malignancies.

Selective Association between Cephalosporin, Quinolone, Macrolide, and Penicillin Antibiotic Consumption and Childhood Obesity in Europe

The accelerated weight gain in productive animals as a result of feeding antibiotic enriched fodder has been well known for decades. The better energy harvest is the result of modified gut microbiota as a consequence of applied antibiotics. Similar mechanisms might result obesity in humans as well. Objectives: Finding associations between global antibiotic consumption of different classes in EU countries and obesity data in adults and children prove that antibiotics might play a significant role in the development of obesity “epidemics” and related illnesses. Methods: Antibiotic consumption data were compared with obesity figures in adults and children in European countries and statistically analyzed for significance. Results: Significant correlation was found between the average yearly consumption of cephalosporins (p = 0.007), quinolones (p = 0.031), macrolides (p = 0.000083) and childhood obesity data, but no significant association was observed with the average penicillin consumption. No association was observed between adult obesity and any of the antibiotic classes studied. Conclusions: Our results support the hypothesis that different types of antibiotics might influence the development of obesity among children, and this finding can serve as a unified explanation for the development of obesity “epidemics”, similarly to the obesity and gut flora alteration-related diseases (type 2 diabetes mellitus, autism, etc.).

Nano-Coupling of Cephalosporin Antibiotics with Fe<SUB>3</SUB>O<SUB>4</SUB>Nanoparticles: Trojan Horse Approach in Antimicrobial Chemotherapy of Infections Caused by <i>Klebsiella spp</i>.

In the present study we had an aim to develop the methods of functionalizing the surface of magnetite nanoparticles with cefotaxime and ceftriaxone antibiotics. The quantitative analysis of the nanostructured cephalosporins was determined by Atom Absorbance Spectroscopy (AAS) and based on the Lambert-Beer law. The engineered nanostructures were tested on gram-negative microorganisms Klebsiella spp., of Enterobacteriaceae, and gram-positive bacteria Staphylococcus aureus, each having multi-drug resistance properties.

Spectophotometric method for determination of certain cephalosporins using 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl)

A simple, accurate and precise spectrophotometric method has been proposed for the determination of eleven cephalosporins, namely;cefaclor monohydrate, cefadroxil monohydrate, cefalexin anhydrous, cefradine anhydrous, cefotaxime sodium, cefoperazone sodium, ceftriaxone sodium, ceftazidime penthydrate, cefazolin sodium, cefixime and cefpodoxime pro- xetil in bulk drug and in pharmaceutical formulations. The method depends on hydrolysis of the studied drugs using 0.5M NaOH at 100&#176;C and subsequent reaction of the formed sulfide ions with NBD-Cl (4-chloro-7-nitrobenzo-2-oxa-1, 3-diazole) to form a yellow-colored chromogen measured at 390 nm. Different variables affecting the reaction (e.g. NaOH concentration, hydrolysis time, NBD-Cl concentration and diluting solvent) were studied and optimized. Under the optimum conditions, linear relationships with good correlation coefficients (0.9990- 0.9999) were found in the range of 5-160 μg mL-1 for all studied drugs. The limits of assay detection and quantitiation ranged from 0.289 to 5.867 and from 0.878 to 17.778 μg mL-1;respectively. The accuracy and precision of the proposed method were satisfactory. The method was successfully applied for analysis of the studied drugs in their pharmaceutical formulations and the recovery percentages ranged from 96.6 to 103.5%.

Kinetic spectrophotometric determination of certain cephalosporins using iodate/iodide mixture

A simple, precise and accurate kinetic spectro-photometric method for determination of ce-fradine anhydrous, cefaclor monohydrate, ce-fadroxil monohydrate, cefalexin anhydrous and cefixime in bulk and in pharmaceutical formula-tions has been developed. The method based on a kinetic investigation of the reaction of the free carboxylic acid group of the drug with a mixture of potassium iodate and potassium iodide at room temperature to form yellow coloured triiodide ions. The reaction was followed up spectrophotometrically by measuring the increase in absorbance at 352 nm as a function of time. The initial rate, fixed time, variable time and rate-constant methods were adopted for constructing the calibration curves but fixed time method has been found to be more applicable. The analytical performance of the method, in terms of accuracy and precision, was statistically validated;the results were satisfactory. The method has been successfully applied to the determination of the studied drugs in commercial pharmaceutical formulations. Statistical comparison of the results with a well established reported method showed excellent ag- reement and proved that there is no significant difference in the accuracy and precision.

Theoretical Insights Elucidate Novel Active Phosphonate Esters—Cephalosporin Antibiotics’ Intermediate

Theoretical insights elucidate a series of active phosphonate esters application in preparation of Cephalosporin antibiotics’ intermediate. The B3LYP/6-311+G(d,p) method was employed to obtain the stable equilibrium geometries including comparing to the AE-active ester. It was found that the Ethyl-aminothiazoly Loximate (AT) molecule fragment is almost planar sheet, but it is almost perpendicular to the plane of phosphoryl ester. Moreover, the calculated Mulliken atomic charge distribution and frontier molecular orbital analysis of these esters showed that the amino N atom connected to the Thiazole ring of the AT had the maximum negative charge, which suggested that this area had high molecular activity. The value of ΔEL-H was energy gap between EHOMO and ELUMO and indicated that compound 6a had high reaction activity. The theory calculation results can explain the reaction mechanism well and predict that the novel active phosphonate ester has a hopeful application prospect in preparation of Cephalosporin antibiotics’ intermediate.

Synthesis and Antibacterial Activity of New Cephalosporin Compounds

Five new cephalosporin compounds were designed and synthesized, and the antibacterial activities were evaluated by the standard serial 2-fold agar dilution method in vitro. The results showed that the activities of the compounds Ia and lb against ESBL E. coli and K. pneumoniae are comparable to those of Cefepime.

Variable selection in near infrared spectroscopy for quantitative models of homologous analogs of cephalosporins

Two universal spectral ranges(4550-4100 cm^(-1) and 6190-5510 cm^(-1))for construction of quantitative models of homologous analogs of cephalosporins were proposed by evaluating theperformance of five spectral ranges and their combinations,using three data sets of cephalos-porins for injection,ie.,cefuroxime sodium,cetriaxone sodium and cefoperazone sodium.Subsequently,the proposed ranges were validated by using eight calibration sets of otherhomologous analogs of cephalosporins for injection,namely cefmenoxime hydrochloride,ceftezole sodium,cefmetazole,cefoxitin sodium,cefotaxime sodium,cefradine,cephazolin sodium and ceftizoxime sodium.All the constructed quantitative models for the eight kinds of cephalosporinsusing these universal ranges could fulill the requirements for quick quantification.After that,competitive adaptive reweighted sampling(CARS)algorithm and infrared(IR)-near infrared(NIR)two-dimensional(2D)correlation spectral analysis were used to determine the scientific basis of these two spectral ranges as the universal regions for the construction of quantitativemodels of cephalosporins.The CAR.S algorithm demonstrated that the ranges of 4550-4100 cm^(-1) and 6190-5510 cm^(-1) included some key wavenumbers which could be attributed to content changes of cephalosporins.The IR-NIR 2D spectral analysis showed that certain wavenumbersin these two regions have strong correlations to the structures of those cephalosporins that wereeasy to degrade.

Detection and Identification of Gonococci Resistance to Cephalosporin and Determination the Most Effective Empirical Treatment for Gonococcal Urethritis in Male Human in Egypt

Introduction: Gonorrhoeae and antimicrobial resistance AMR of gonococci is a major health problem today, because emerged resistance to last line empirical treatment for gonorrhoeae cephalosporins in many countries is predictable to be untreatable disease in near future. WHO GASP, WHO GLASS and WHO’s global action plan on AMR recommends to expand nationally and internationally to collect data to monitor AMR of gonococci for public health policies. Objective: Our aim is to detect resistance of gonococci to Cepha- losporins and determine the most effective empirical treatment for un-com- plicated gonococcal urethritis in males in Egypt. Methods: We depended in our methodology on selected gonococci from male urethral discharge specimens on Thyer Martien medium;collected 33 isolates during three years from 2017 to 2020;used antibiotics with MIC according to international standards and measuring IZD according to antimicrobial susceptibility testing reference ranges in international standards. Results: By statistical studies, resistance to cephalosporins was as follows: Cephradine 97%, Cefaclor 87.9%, Cefoxitin 97%, Ceftriaxone 90.9% and 42.4% to Cefepime, that shows hetero-genecity in resistance inside cephalosporin group;while resistance to Macrolides group represented by Azithromycin and Tetracyclins group represented by Doxycycline was as follows: Azithromycin 39.4%, Doxycycilne 27.3%;finally fluoroquinolones, the most effective group, resistance, was as: Levofloxacin 15.2%, Ciprofloxacin 15.2% and Ofloxacin 24.2%. Conclusion: The most effective empirical treatment for uncomplicated gonococcal urethritis in males in EGYPT is Fluoroquinolone;especially Levofloxacin ranks first susceptibility as 78.8% and 15.2% resistance followed by Ciprofloxacin susceptibility as 69.7% and 15.2% resistance, finally Ofloxacin susceptibility as 66.7% and 24.2% resistance;for Ceftriaxone not more recommended in EGYPT as empirical treatment for uncomplicated gonococcal urethritis, it is susceptibility as 6.1% and 90.9% resistance;in addition, we can use combination therapy of Fluoroquinolones with Azithromycin or Doxycycline, whose susceptibility is 30.3% for Azithromycin and 42.4% for Doxycycycline, while resistance is 39.4% for Azithromycin and 27.3% for Doxycycline. It is worth noting that only Cefepime in Cephalosporins group represents 42.4% susceptibility and 42.4% resistance;in addition to the Carbapenems group, it represents as 42.4% susceptibility for Imipenem and 45.5% resistance, then 42.2% susceptibility for Meropenem and 48.5% resistance, which can play role in combination therapy.

Studies on Semisynthesis and Anibacterial Activity of 7 β-(5-Methyl-1-Aryl-1H-1,2,3-Triazoly-4-Carboxamido) Cephalosporins

Nine new derivatives of 7 β - (5- methyl- 1- aryl- 1H-1, 2, 3- triazoly- 4- carboxamido) cephalosporins were synthesized by acylction of 7 β -amino group of 7-ACA, 7-ADCA and 7-ACT with 5 -methsyl- 1 -aryl- 1 H- 1,2,3-triazoly-4-formyl chloride. The structure of the compounds were confirmed by elementray analysis IR, HNMR and FAB-MS. Some of them showed significant antibacterial activity

Computational Calculations of Molecular Properties and Molecular Docking of New and Reference Cephalosporins on Penicillin Binding Proteins and Various β-Lactamases

An approach of using molinspiration calculations and molecular docking on PBPs （penicillin-binding proteins） and certain β-lactamases is employed to predict the molecular properties, bioactivity and resistance of newer and reference cephalosporins. The previously synthesized cephalosporins 1-8 and reference cephalosporins were subjected to extensive evaluations by calculating the molecular properties, drug-likeness scores on the bases of Lipinski＇s rule and bioactivity prediction using the method of molinspiration web-based software. The TPSA （topological polar surface area）, OH-NH interactions, n-violation and the molinspiration Log partition coefficient （miLogP） values were also calculated. The investigated cephalosporins were subjected to molecular docking study on PBPs （lpyy） and on β-lactamases produced by S. aureus, K. pneumonia, E. coil and P. auroginosa using 1-click-docking website. Molecular properties of 1-8 recorded higher ＂FPSA than cephalexin and were lower than the reference cephalosporins and do not fulfill the requirements for Lipinski＇s rule. Bioactivities of 1-8 were predicted to be less and their docking scores on PBPs were comparable to those of the reference cephalosporins, particularly ceftobiprole. The references recorded various docking scores on the above β-lactamases and as expected, cefiobiprole recorded the lowest scores on all β-lactarnases. Cephalosporins 1-8 recorded various docking scores on β-lactamases. Molecular docking studies on PBPs and β-lactamases are considered as very useful, reliable and practical approach for predicting the bioactivity scores and to afford some information about the stability and selectivity of the newly proposed cephalosporins against β-lactamases of certain pathogenic microbes, such as P. auroginosa and MRSA, by recording the relative docking scores in comparison with those of reference cephalosporins.

第三代头孢菌素治疗社区获得性自发性细菌性腹膜炎效果预测模型的临床价值

目的探讨第三代头孢菌素(3^(rd)GC)治疗社区获得性自发性细菌性腹膜炎(CASBP)效果预测模型的临床应用价值。方法前瞻性选取2021年1月—2022年6月南昌市第九医院新入院的肝硬化伴CASBP患者50例,随机分为优化治疗组(n=25)与传统治疗组(n=25),前者基于效果预测模型采用头孢他啶或亚胺培南初治,后者均采用头孢他啶初治,此后依据初治效果调整抗生素,比较两组的初治有效率、第5天治愈率及30天病死率。计量资料两组间比较采用成组t检验或MannWhitney U检验。计数资料两组间比较采用χ^(2)检验或Fisher精确概率法。结果所有患者均完成研究。优化治疗组初治有效率明显高于传统治疗组(88.0%vs 60.0%,χ^(2)=5.094,P=0.024),两组第5天治愈率相当(80.0%vs 56.6%,χ^(2)=3.309,P=0.069)。均使用头孢他定初治的患者中,优化治疗组初治有效率亦明显高于传统治疗组(88.9%vs 60.0%,χ^(2)=4.341,P=0.037),两组第5天治愈率相当(83.3%vs 56.0%,χ^(2)=2.425,P=0.119)。两组患者的30天病死率分别为8.0%和20.0%,差异无统计学意义(χ^(2)=0.664,P=0.415)。所有入组患者初治有效与第5天治愈关联明显(OR=9.643,95%CI:2.292~40.564),第5天治愈与患者30天死亡关联明显(OR=0.138,95%CI:0.023~0.813)。结论该疗效预测模型有助于临床医生筛选3rdGC治疗的优势患者,提高3rdGC经验性治疗CASBP的初治疗效。