Membrane Pore Protein Research Articles

Predictive modeling and insight into protein fouling in microfiltration and ultrafiltration through one-dimensional convolutional models

Membrane fouling, a critical issue in membrane performance associated with proteins, is often difficult to analyze due to limited knowledge of contamination scenarios. This study uses a one-dimensional convolutional neural network (ODCNN) to examine the complex relationships among membrane properties, protein characteristics, and operational parameters, aiming to elucidate the dynamics of protein-related contamination. To improve the predictive performance of the deep learning model, we propose a novel strategy that learns numerical and categorical features in sequence. Compared with the other seven conventional models, our results reveal that the ODCNN model shows the best performance, which achieves impressive coefficients of determination, reaching 0.833 for flux and 0.723 for protein rejection, indicating the model’s robustness even when dealing with incomplete information on membrane fouling. To mitigate the issue of insufficient data, we applied the Synthetic Minority Over-Sampling Technique to increase the dataset’s diversity, validating this approach with experimental evidence. To improve the interpretability of the model, we used the Shapley Additive exPlanations method to assess the contribution of various features, which revealed that membrane pore size, protein concentration, transmembrane pressure, and crossflow velocity are key factors influencing both flux and protein retention. We also analyzed the causes and assessed the degree of contamination according to the degree of their influence. By leveraging machine learning techniques to analyze and predict membrane filtration performance, particularly concerning protein fouling, this study demonstrates significant potential for practical applications in environments with limited information. This innovative approach is poised to contribute to ongoing advancements in membrane filtration technology and offers a promising pathway for enhancing the understanding and management of membrane fouling.

A comparative study of genotyping and antimicrobial resistance between carbapenem-resistant Klebsiella pneumoniae and Acinetobacter baumannii isolates at a tertiary pediatric hospital in China.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) clinical isolations have rapidly increased in pediatric patients. To investigate a possible health care-associated infections of CRKP in a tertiary pediatric hospital, the circulating clones and carbapenem-resistant pattern between CRKP and carbapenem-resistant Acinetobacter baumannii (CRAB) isolates were compared to classify their epidemiological characteristics. The results will help to identify the epidemic pattern of the CRKP transmission in the hospital. Ninety-six CRKP and forty-eight CRAB isolates were collected in Kunming Children's Hospital from 2019 through 2022. These isolates were genotyped using repetitive extragenic palindromic-PCR (REP-PCR). Carbapenemase phenotypic and genetic characterization were investigated using a disk diffusion test and singleplex PCR, respectively. In addition, these characteristics of the two pathogens were compared. The rates of CRKP and CRAB ranged from 15.8% to 37.0% at the hospital. Forty-nine and sixteen REP genotypes were identified among the 96 and 48 CRKP and CRAB isolates tested, respectively. The CRKP isolates showed more genetic diversity than the CRAB isolates. Of the 96 CRKP isolates, 69 (72%) produced Class B carbapenemases. However, all 48 CRAB isolates produced Class D carbapenemase or extended-spectrum β-lactamases (ESBL) combined with the downregulation of membrane pore proteins. Furthermore, the carbapenemase genes bla KPC, bla IMP, and bla NDM were detected in CRKP isolates. However, CRAB isolates were all positive for the bla VIM, bla OXA-23, and bla OXA-51 genes. These CRKP isolates exhibited different biological and genetic characteristics with dynamic changes, suggesting widespread communities. Continuous epidemiological surveillance and multicenter research should be carried out to strengthen the prevention and control of infections.

Evaluation of immunogenicity of gene-deleted and subunit vaccines constructed against the emerging pseudorabies virus variants

BackgroundPseudorabies (PR) (also called Aujeszky’s disease, AD) is a serious infectious disease affecting pigs and other animals worldwide. The emergence of variant strains of pseudorabies virus (PRV) since 2011 has led to PR outbreaks in China and a vaccine that antigenically more closely matches these PRV variants could represent an added value to control these infections.MethodsThe objective of this study was to develop new live attenuated and subunit vaccines against PRV variant strains. Genomic alterations of vaccine strains were based on the highly virulent SD-2017 mutant strain and gene-deleted strains SD-2017ΔgE/gI and SD-2017ΔgE/gI/TK, which constructed using homologous recombination technology. PRV gB-DCpep (Dendritic cells targeting peptide) and PorB (the outer membrane pore proteins of N. meningitidis) proteins containing gp67 protein secretion signal peptide were expressed using the baculovirus system for the preparation of subunit vaccines. We used experimental animal rabbits to test immunogenicity to evaluate the effect of the newly constructed PR vaccines.ResultsCompared with the PRV-gB subunit vaccine and SD-2017ΔgE/gI inactivated vaccines, rabbits (n = 10) that were intramuscularly vaccinated with SD-2017ΔgE/gI/TK live attenuated vaccine and PRV-gB + PorB subunit vaccine showed significantly higher anti-PRV-specific antibodies as well as neutralizing antibodies and IFN-γ levels in serum. In addition, the SD-2017ΔgE/gI/TK live attenuated vaccine and PRV-gB + PorB subunit vaccine protected (90–100%) rabbits against homologous infection by the PRV variant strain. No obvious pathological damage was observed in these vaccinated rabbits.ConclusionsThe SD-2017ΔgE/gI/TK live attenuated vaccine provided 100% protection against PRV variant challenge. Interestingly, the subunit vaccines with gB protein linked to DCpep and PorB protein as adjuvant may also be a promising and effective PRV variant vaccine candidate.

Understanding Single-Protein Fouling in Micro- and Ultrafiltration Systems via Machine-Learning-Based Models

Protein fouling is a complex mechanism. To enhance understanding on protein fouling of membranes, the target of this study is twofold: (i) to determine the relative influences of parameters via the Random Forest (RF) model and (ii) to evaluate the predictive capability of the Neural Network (NN) model. Membrane pore size is the most dominant influence on fouling followed by transmembrane pressure (TMP), while membrane configuration (i.e., flat-sheet, hollow fiber, or tubular) is the least dominant. The NN model gives modest predictive capability despite inconsistencies and variabilities of the experimental setups and protocols, which invariably affects the important parameters in the database compiled from past publications. The database was divided into microfiltration (MF) and ultrafiltration (UF) subsets based on the membrane pore size values. It was found that the dominant parameters for permeate flux are different, with membrane pore size and protein concentration being dominant for MF and UF, respectively, while TMP is dominant for protein rejection for both cases. For permeate flux, membrane material is the most dominant parameter for the non-BSA database, while membrane pore size remains the most dominant parameter for protein rejection regardless of the protein used. Results show that such data-driven RF and NN models can enhance the understanding on the relative dominance of the parameters on different phenomena and provide adequate prediction of protein fouling, in the absence of any governing equations.

Phenotypic and Genetic Analysis of KPC-49, a KPC-2 Variant Conferring Resistance to Ceftazidime-Avibactam and Maintaining Resistance to Imipenem and Meropenem.

Klebsiella pneumoniae, a gram-negative bacterium, poses a severe hazard to public health, with many bacterial hosts having developed resistance to most antibiotics in clinical use. The goal of this study was to look into the development of resistance to both ceftazidime-avibactam and carbapenems, including imipenem and meropenem, in a K. pneumonia strain expressing a novel K. pneumoniae carbapenemase-2 (KPC-2) variant, referred to as KPC-49. After 1 day of incubation of K1 on agar containing ceftazidime-avibactam (MIC = 16/4 mg/L), a second KPC-producing K. pneumoniae strain (K2) was recovered. Antimicrobial susceptibility assays, cloning assays, and whole genome sequencing were performed to analyse and evaluate antibiotic resistance phenotypes and genotypes. K. pneumoniae strain (K1), that produced KPC-2, was susceptible to ceftazidime-avibactam but resistant to carbapenems. The K2 isolate harboured a novel bla KPC-49 variant, which differs from bla KPC-2 by a single nucleotide (C487A), and results in an arginine-serine substitution at amino acid position 163 (R163S). The mutant K2 strain was resistant to both ceftazidime-avibactam and carbapenems. We demonstrated the ability of KPC-49 to hydrolyse carbapenems, which may be attributed to high KPC-49 expression or presence of an efflux pump and/or absence of membrane pore proteins in K2. Furthermore, blaKPC-like was carried on an IncFII (pHN7A8)/IncR-type plasmid within a TnAs1-orf-orf-orf-orf-orf-orf-ISKpn6-bla KPC-ISKpn27 structure. The bla KPC-like gene was flanked by various insertion sequences and transposon elements, including the Tn3 family transposon, such as TnAs1, TnAs3, IS26, and IS481-ISKpn27. New KPC variants are emerging owing to sustained exposure to antimicrobials and modifications in their amino acid sequences. We demonstrated the drug resistance mechanisms of the new mutant strains through experimental whole genome sequencing combined with bioinformatics analysis. Enhanced understanding of laboratory and clinical features of infections due to K. pneumoniae of the new KPC subtype is key to early and accurate anti-infective therapy.

Light-Activated Assembly of Connexon Nanopores in Synthetic Cells.

During developmental processes and wound healing, activation of living cells occurs with spatiotemporal precision and leads to rapid release of soluble molecular signals, allowing communication and coordination between neighbors. Nonliving systems capable of similar responsive release hold great promise for information transfer in materials and site-specific drug delivery. One nonliving system that offers a tunable platform for programming release is synthetic cells. Encased in a lipid bilayer structure, synthetic cells can be outfitted with molecular conduits that span the bilayer and lead to material exchange. While previous work expressing membrane pore proteins in synthetic cells demonstrated content exchange, user-defined control over release has remained elusive. In mammalian cells, connexon nanopore structures drive content release and have garnered significant interest since they can direct material exchange through intercellular contacts. Here, we focus on connexon nanopores and present activated release of material from synthetic cells in a light-sensitive fashion. To do this, we re-engineer connexon nanopores to assemble after post-translational processing by a protease. By encapsulating proteases in light-sensitive liposomes, we show that assembly of nanopores can be triggered by illumination, resulting in rapid release of molecules encapsulated within synthetic cells. Controlling connexon nanopore activity provides an opportunity for initiating communication with extracellular signals and for transferring molecular agents to the cytoplasm of living cells in a rapid, light-guided manner.

Effects comparison between the secondary nanoplastics released from biodegradable and conventional plastics on the transfer of antibiotic resistance genes between bacteria

Antibiotic resistance genes (ARGs) have caused widespread concern because of their potential harm to environmental safety and human health. As substitutes for conventional plastics, the toxic effects of short-term degradation products of biodegradable plastics (polylactic acid (PLA) and polyhydroxyalkanoates (PHA)) on bacteria and their impact on ARGs transfer were the focus of this study. After 60 days of degradation, more secondary nanoplastics were released from the biodegradable plastics PLA and PHA than that from the conventional plastics polystyrene (PS). All kinds of nanoplastics, no matter released from biodegradable plastics or conventional plastics, had no significant toxicity to bacteria. Nanoplastic particles from biodegradable plastics could significantly increase the transfer efficiency of ARGs. Although the amount of secondary nanoplastics produced by PHA microplastics was much higher than that of PLA, the transfer frequency after exposure to PLA was much higher, which may be due to the agglomeration of PHA nanoplastics caused by plastic instability in solution. After exposure to the 60d PLA nanoplastics, the transfer frequency was the highest, which was approximately 28 times higher than that of control. The biodegradable nanoplastics significantly enhanced the expression of the outer membrane pore protein genes ompA and ompC, which could increase cell membrane permeability. The expression levels of trfAp and trbBp were increased by repressed major global regulatory genes korA, korB, and trbA, which eventually led to an increase in conjugative transfer frequency. This study provides important insights into the evaluation of the environmental and health risks caused by secondary nanoplastics released from biodegradable plastics.

Cefiderocol: a novel siderophore cephalosporin against multi-drug resistant Gram-negative bacilli infections

Antimicrobial resistance is one of the critical public health issues in the world. There is an urgent need to develop effective broad-spectrum antibiotics to treat the infection of multi-drug resistant Gram-negative bacilli. Cefiderocol, developed by the Shionogi Inc. in Japan, is a new type of iron carrier cephalosporin antibiotics, which overcomes the drug resistance of Gram-negative bacilli due to the down-regulation of outer membrane pore protein and the up-regulation of efflux pump, and has good stability to serine- and metallo-carbapenemases. This drug has a broad spectrum and strong antibacterial activity against carbapenem-resistant Enterobacteriaceae (CRE), Pseudomonas aeruginosa, Acinetobacter baumannii, and Stenotrophomonas maltophilia. Cefiderocol can be used to treat complex urinary tract infections (including pyelonephritis), hospital-acquired pneumonia, and ventilator-associated pneumonia. By summarizing the chemical structure, antibacterial mechanism, in vitro antibacterial activity, pharmacokinetics, pharmacodynamics, and clinical treatment of cefiderocol, this review shows the application potential of cefiderocol as a new iron carrier cephalosporin in the treatment of multi-drug resistant Gram-negative bacilli infections.

Fitness cost and compensation mechanism of sulfonamide resistance genes (sul1, sul2, and sul3) in Escherichia coli.

The fitness cost of antibiotic resistance is a crucial factor to determine the evolutionary and transmission success of resistant bacteria. Exploring the fitness cost and compensation mechanism of antibiotic resistance genes (ARGs) in bacteria may effectively reduce the transmission of drug-resistant genes in the environment. Engineered bacteria with the same genetic background that carry sulfonamide resistance gene were generated to explore the fitness cost of sulfonamide resistance gene in Escherichia coli. There were significant differences in the protein expression of the two-component system pathway (fliZ, fliA, fliC and lrhA), folate biosynthesis pathway (sul1, sul2 and sul3), ABC transporter system (ugpC, rbsA and gsiA), and outer membrane pore protein OmpD through the comparative analysis of differential proteins compared to sensitive bacteria. Thus, we could speculate the possible fitness compensation mechanism. Finally, quantitative Real-time PCR (qRT-PCR) was used to verify the functions of some differential proteins at the transcriptional level. The fitness cost and compensatory evolution of antibiotic resistance are an essential part of bacterial evolution.

Identification of c-di-GMP/FleQ-Regulated New Target Genes, Including cyaA, Encoding Adenylate Cyclase, in Pseudomonas putida.

The bacterial second messenger cyclic diguanylate (c-di-GMP) modulates plankton-to-biofilm lifestyle transition of Pseudomonas species through its transcriptional regulatory effector FleQ. FleQ regulates transcription of biofilm- and flagellum-related genes in response to c-di-GMP. Through transcriptomic analysis and FleQ-DNA binding assay, this study identified five new target genes of c-di-GMP/FleQ in P. putida, including PP_0681, PP_0788, PP_4519 (lapE), PP_5222 (cyaA), and PP_5586 Except lapE encoding an outer membrane pore protein and cyaA encoding an adenylate cyclase, the functions of the other three genes encoding hypothetical proteins remain unknown. FleQ and c-di-GMP coordinately inhibit transcription of PP_0788 and cyaA and promote transcription of PP_0681, lapE, and PP_5586 Both in vitro and in vivo assays show that FleQ binds directly to promoters of the five genes. Further analyses confirm that LapE plays a central role of in the secretion of adhesin LapA and that c-di-GMP/FleQ increases lapE transcription, thereby promoting adhesin secretion and biofilm formation. The adenylate cyclase CyaA is responsible for synthesis of another second messenger, cyclic AMP (cAMP). FleQ and c-di-GMP coordinate to decrease the content of cAMP, suggesting that c-di-GMP and FleQ coregulate cAMP by modulating cyaA expression. Overall, this study adds five new members to the c-di-GMP/FleQ-regulated gene family and reveals the role of c-di-GMP/FleQ in LapA secretion and cAMP synthesis regulation in P. putida IMPORTANCE c-di-GMP/FleQ promotes the plankton-to-biofilm lifestyle transition at the transcriptional level via FleQ in Pseudomonas species. Identification of new target genes directly regulated by c-di-GMP/FleQ helps to broaden the knowledge of c-di-GMP/FleQ-mediated transcriptional regulation. Regulation of lapE by c-di-GMP/FleQ guarantees highly efficient LapA secretion and biofilm formation. The mechanism of negative correlation between c-di-GMP and cAMP in both P. putida and P. aeruginosa remains unknown. Our result concerning transcriptional inhibition of cyaA by c-di-GMP/FleQ reveals the mechanism underlying the decrease of cAMP content by c-di-GMP in P. putida.

Promoter engineering for high ectoine production in a lower saline medium by Halomonas hydrothermalis Y2.

For the stress from fermenters, downstream processing equipment, and wastewater treatment to be alleviated, lowering salt-dependence in the ectoine synthesis process is of great significance in the moderately halotolerant Halomonas hydrothermalis Y2. In H. hydrothermalis Y2, the σ70- and σ38-controlled promoters of ectA are predicted to be involved in the osmotic regulation of ectoine synthesis. By substituting the ectA promoter with a promoter P265 that identified in the outer membrane pore protein E of H. hydrothermalis Y2, the salt dependence of ectoine synthesis was significantly decreased. In the 500-ml flask containing various NaCl contents, the engineered strain (p/Y2/△ectD/△doeA) showed a remarkably enhanced ability in ectoine synthesis, especially under lower saline stress. After a 36-h fed-batch fermentation in the 1-l fermenter, p/Y2/△ectD/△doeA synthesized 11.5g ectoine l-1 in the presence of 60g NaCl-1l, with a high 0.32g ectoine l-1 h-1 productivity, a specific productivity of 512.2mg ectoine per g cell dry weight (CDW)-1, and an excretion ratio of 67 % ectoine. As no impaired growth was observed in strain p/Y2/△ectD/△doeA while ectoine synthesis was increased, this promoter engineering strategy provides a practical protocol for lowering the salt-dependence of ectoine synthesis in this moderately halotolerant strain.

Cryo-EM analysis of the SctV cytosolic domain from the enteropathogenic E. coli T3SS injectisome

The bacterial injectisome and flagella both rely on type III secretion systems for their assembly. The syringe-like injectisome creates a continuous channel between the bacterium and the host cell, through which signal-modulating effector proteins are secreted. The inner membrane pore protein SctV controls the hierarchy of substrate selection and may also be involved in energizing secretion. We present the 4.7 Å cryo-EM structure of the SctV cytosolic domain (SctVC) from the enteropathogenic Escherichia coli injectisome. SctVC forms a nonameric ring with primarily electrostatic interactions between its subunits. Molecular dynamics simulations show that monomeric SctVC maintains a closed conformation, in contrast with previous studies on flagellar homologue FlhA. Comparison with substrate-bound homologues suggest that a conformational change would be required to accommodate binding partners.

Construction of a bacterial surface display system based on outer membrane protein F

BackgroundBacterial surface display systems were developed to surface expose heterologous proteins or peptides for different applications, such as peptide libraries screening and live bacterial vaccine design. Various outer membrane proteins, such as outer membrane protein A (OmpA), OmpC and outer membrane pore protein E precursor (PhoE), have been used as carriers for surface display, fused to the proteins or peptides of interest in Gram-negative bacteria. Here, we investigated the utility of constitutively expressed OmpF for the display of foreign immune epitopes on the Escherichia coli cell surface and then compared it with plasmid-induced expression of OmpF and OmpC.ResultsEnhanced expression of OmpF was linked to a mutation in the OmpF promoter sequence. This mutation rendered OmpF an ideal carrier protein for the enriched display of a target of interest on the bacterial surface. To this end, we grafted two peptides, harboring important epitopes of the hepatitis B virus (HBV) S antigen and human papilloma virus (HPV) L2 protein, onto OmpF of E. coli by genome editing. The resultant fused OmpF proteins were constitutively expressed in the edited E. coli and purified by membrane component extraction. The epitope that displayed on the bacterial surface was verified by SDS-PAGE, western blotting, flow cytometry, and immunoelectron microscopy of the intact bacteria. We further compared this constitutive expression with plasmid-induced expression of OmpF and OmpC in bacterial cells using the same methods for verification. We found that plasmid-induced expression is much less efficient than constitutive expression of OmpF from the bacterial genome.ConclusionsEnhanced expression of OmpF in a plasmid-independent manner provides an amenable way to display epitopes on the bacterial surface and sheds light on ways to engineer bacteria for biotechnological applications.

In silico Analysis of Different Signal Peptides for the Excretory Production of Recombinant NS3-GP96 Fusion Protein in Escherichia coli

Escherichia coli is one of the simplest hosts which is widely being used to express heterologous proteins. However, without appropriate signal peptide, this host cannot be applied for secretory proteins. Secretory production of recombinant proteins in E. coli has been an issue of interest because of its diverse advantages including cost and time savings, as well as reduction of endotoxin. NS3 from hepatitis C virus (HCV) was chosen as an antigen for vaccine development against HCV virus infections and it connected to gp96 as an adjuvant for stimulating Toll-like receptors (TLRs) to stimulate cytokines secretion by T cells. It was successfully produced in E. coli without using signal peptide previously. In this study, in order to increase the expression level of recombinant NS3-gp96 fusion protein (rNS3-gp96) in periplasmic space, we selected a series of signal peptides. Therefore, to foretell the best signal peptides for expression of NS3-gp96 recombinant protein in E. coli, 52 signal peptides from gram-negative bacteria were chosen and the most important physicochemical features of them were investigated. Therefore, n, h and c regions and signal peptide probability of them were evaluated by signalP software “version 4.1”, and physicochemical features were assessed by ProtParam and PROSO II tools. Eventually, prsK protein, outer membrane pore protein E (phoE), and fimbrial adapter papK protein were determined as the best candidates for the secretory production of rNS3-gp96 in E. coli in our study (with D score 0.899, 0.806, 0.797, respectively). Although, in the experimental investigation, should be considered other influencing parameters.

Correlation Study of PVDF Membrane Morphology with Protein Adsorption: Quantitative Analysis by FTIR/ATR Technique

Microporous PVDF membranes were used as protein capture matrices in immunoassays. Because the most common labels in immunoassays were detected based on the colour change, an understanding of how protein concentration varies on different PVDF surfaces was needed. Herein, the correlation between the membrane pore size and protein adsorption was systematically investigated. Five different PVDF membrane morphologies were prepared and FTIR/ATR was employed to accurately quantify the surface protein concentration on membranes with small pore sizes. SigmaPlot® was used to find a suitable curve fit for protein adsorption and membrane pore size, with a high correlation coefficient, R2, of 0.9971.

αB‐Crystallin Phosphorylation as a Precursor to Cataractogenesis

PurposeThe molecular mechanism behind age‐related cataract formation is yet to be elucidated, although phosphorylation induced hyperactivity of αB‐crystallin (αB) and its subsequent association with fibre cell membranes has been implicated. This study evaluates the effect phosphorylation of αB at three well characterised sites (S19, S45 and S59) has on its chaperone activity and aims to characterise αB's membrane association with a focus on its role in cataractogenesis.MethodsThe chaperone activity of αB phosphomimics (S→D) were measured via aggregation assays using creatine phosphokinase (CPK) as a substrate. In vivo association of αB phosphoisotypes with two abundant membrane proteins, connexin 46 (cx46) and aquaporin 0 (AQP0) in aged human lenses was visualised using Duolink proximity assays and confocal microscopy.ResultsThe chaperone activity of αB increased with an increase in modification sites. Modification at the S19 and S59 residues activated αB to a greater extent than at S45. In the lens, phosphorylated αB was found to interact with Cx46 and AQP0 at cell membranes.ConclusionsThis study confirms that multiple phosphorylation events of αB cause a cumulative increase in activity, and that membrane association, at least in part, is mediated by interactions with membrane pore proteins. The proposed mechanism of cataractogenesis is that phosphorylation induced increases in substrate affinity of αB may lead to increased association with membrane pore proteins, leading to the obstruction of the diffusion of small molecules into inner regions of the lens, contributing to cataract formation.

Clinical significance of serum anti-microbial antibodies test in inflammatory bowel disease

Objective To investigate the clinical significance of serum anti-Saccharomyces cerevisias antibody （ASCA）,anti-outer membrane porin C （anti-OmpC）,antibody to Pseudomonas fluorescens-associated sequence I2 （anti-I2 ）and antibody to bacterial flagellin （anti-CBirl ）in the diagnosis and treatment of inflammatory bowel disease （IBD）.Methods From 2011 to 2013,87 patients with IBD were enrolled and divided into Crohn′s disease （CD）group （66 cases）and ulcerative colitis （UC）group （21 cases）.A total of 62 age and gender matched healthy individuals were enrolled as the control group. Fasting blood samples （2 mL）of the subjects were collected.The expression of ASCA,anti-OmpC,anti-I2 and anti-Cbirl antibodies was detected with enzyme-linked immunosorbent assay （ELISA）kits.The diagnosis value of each antibody in IBD and the differential diagnostic value of in UC and CD were compared by receiver operating characteristic （ROC）curve.Results The area under the curve （AUC）of ASCA between IBD and the healthy control group,between CD group and UC group was 0.580 and 0.512, respectively;the accuracy in diagnosis was low.The AUC of anti-CBirl between IBD and the healthy control group was 0.617.There was no differential diagnosis significance of the other antibodies.The positive rate of ASCA in IBD group was 62.1 % （54/87）,which was significantly higher than that in the control group （38.7%,24/62）.The positive rates of anti-OmpC and anti-I2 in IBD group was significantly lower than those in the control group and the differences were statistically significant （both P 〈0.05）.No difference was observed in positive rates of serum antibodies among the others groups （all P 〉0.05）.The specificity,sensitivity,positive predictive value （PPV）and negative predictive value （NPV）of ASCA in differential diagnosis of CD and UC was 52.4%,66.7%,81 .48% and 33.33%,respectively.The specificity and sensitivity of anti-OmpC,anti-I2 and anti-CBirl in differential diagnosis of CD and UC was 81 .0% to 100.0% and 9.1 % to 37.9%,respectively.The specificity,sensitivity,PPV and NPV of double-positive ASCA and anti-I2 in the diagnosis of CD was 57.1 %,86.4%,82.6% and 50.0%, respectively.The positive rate of ASCA and anti-I2 in CD group was significantly higher than that in UC group （84.8%（56/66）vs 57.1 % （12/21 ）;χ2 =5 .633,P =0.018 ）.Conclusions Positive ASCA has some significance in the diagnosis of patients with IBD in our country.The detection of anti-I2 can help to diagnose ASCA negative CD.Because of low sensitivity and positive rate,anti-OmpC and anti-CBirl have limited value in the diagnosis of IBD and the differential diagnosis of UC and CD in our country. Key words: Inflammatory bowel diseases; Diagnosis, differential; Antibodies, fungal; Antibodies, bacterial; Anti-Saccharomyces cerevisiae antibodies~ Bacterial outer membrane pore protein C; Pseudomonas fluorescens associated sequence I2

Reciprocal regulation of resistance-nodulation-division efflux systems and the Cpx two-component system in Vibrio cholerae.

The Cpx two-component regulatory system has been shown in Escherichia coli to alleviate stress caused by misfolded cell envelope proteins. The Vibrio cholerae Cpx system was previously found to respond to cues distinct from those in the E. coli system, suggesting that this system fulfills a different physiological role in the cholera pathogen. Here, we used microarrays to identify genes that were regulated by the V. cholerae Cpx system. Our observations suggest that the activation of the V. cholerae Cpx system does not induce expression of genes involved in the mitigation of stress generated by misfolded cell envelope proteins but promotes expression of genes involved in antimicrobial resistance. In particular, activation of the Cpx system induced expression of the genes encoding the VexAB and VexGH resistance-nodulation-division (RND) efflux systems and their cognate outer membrane pore protein TolC. The promoters for these loci contained putative CpxR consensus binding sites, and ectopic cpxR expression activated transcription from the promoters for the RND efflux systems. CpxR was not required for intrinsic antimicrobial resistance, but CpxR activation enhanced resistance to antimicrobial substrates of VexAB and VexGH. Mutations that inactivated VexAB or VexGH efflux activity resulted in the activation of the Cpx response, suggesting that vexAB and vexGH and the cpxP-cpxRA system are reciprocally regulated. We speculate that the reciprocal regulation of the V. cholerae RND efflux systems and the Cpx two-component system is mediated by the intracellular accumulation of an endogenously produced metabolic by-product that is normally extruded from the cell by the RND efflux systems.

Upregulation and lysosomal degradation of AQP4 in rat brains with bacterial meningitis

Brain edema is among the major complications in children with bacterial meningitis. Aquaporins are integral membrane pore proteins that form channels to regulate cellular water content. Aquaporin-4 (AQP4), which is enriched in parts of astrocytic membranes that are apposed to pial or perivascular basal laminae, is the predominant aquaporin in the central nervous system. Dystroglycan is among the proteins that are responsible for the site-specific anchorage of AQP4. To elucidate the role of AQP4 in the development of brain edema induced by meningitis, a model of bacterial meningitis was established by injecting group B β-hemolytic Streptococci into the cerebrospinal fluid of three-week-old rats. The brain water content increased in this model compared with that in the control group. The expression of AQP4 and dystroglycan was examined by Western blot and the degradation route of AQP4 was investigated by double immunofluorescence labeling. Western blot results showed that the expression of AQP4 and dystroglycan in rat brain increased in the meningitis model. Meanwhile, AQP4 was co-localized with the marker of lysosome in this model, indicating that the lysosome is involved in AQP4 degradation.

English

This study aimed to investigate the existence and molecular epidemiological characteristics of  multi-drug resistant Acinetobacter baumannii (MDRAB)-related resistance genes isolated from our hospital. Microdilution and disk diffusion methods were used to detect the antibiotic susceptibility of 46 MDRAB isolates that carried OXA-51 gene. Pulsed-field gel electrophoresis (PFGE) was used to analyze the homologies among the strains. Resistance genes were tested through polymerase chain reaction. Among the 46 MDRABA, a total of nine, isolates were categorized as strain A (19.6%), eight as strain B (17.4%), and four as strain O (8.7%). A total of 41 isolates (89.1%) carried the OXA23 gene, 17 (37.0%) carried the PER gene, and six (13%) carried the IMP gene. Six strains (13%) did not carry the membrane pore protein gene carO. The numbers of isolates that carried a particular gene were as follows: 40 (87.0%), armA; 41 (89.1%), ant(3”)-I; 33 (71.7%), aac(3)-I; 2 (4.3%), aac(3)-II; 1 (2.2%), aac(6’)-II; and 1 (2.2%) and aph(3’)-VI. Most of the isolates (93.5%) carried the qacE∆1 gene. These hospital MDRAB isolates were proven to simultaneously carry a variety of drug resistance genes. Strains A and B were the major epidemic strains of nosocomial MDRAB. Key words: Acinetobacter baumannii, resistance gene, pulsed-field gel electrophoresis, multi-drug resistant.