Cloning, expression, purification and functional analysis of a specific multi-epitope protein from multi drug resistance Acinetobacter baumannii.
Ontology highlight
ABSTRACT:
SUBMITTER: Davoudi Z
PROVIDER: S-EPMC8629821 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Introduction:Multidrug-resistance in Acinetobacter baumannii has emerged as a serious problem to public health. There is still a significant gap in the understanding of the multidrug-resistance and the genome diversity evolutionary process of A. baumannii in China, especially in the central and western regions. Methods:Ten A. baumannii strains were collected from three hospitals in Chongqing, China. Whole-genome re-sequencing was used to obtain differences in genomic levels among strains. The diversity were determined by multi-locus sequence typing method, and investigate the genetic relationship between the ten strains and others by phylogenetic analysis. Comparative analysis focused on resistance genes related to insertions and deletions (InDels) and single-nucleotide polymorphisms (SNPs) was performed. Results:The overall G+C% content was 39.05%~39.43%, the average sequencing depth was 273.95~428.99, and the alignment ratio of the sequencing data was 92.93%~99.27%. A total of 42 InDels and 11,387 SNPs were detected in the coding sequence region of the isolates. Phylogenetic tree shows that the 10 A. baumannii isolates were divided into four relative groups, and there exist the possibility of cross-regional spread pattern. A total number of 19 drug resistance genes had been found in each strain, and efflux pump-related genes accounted for the most. Only AacA4 underwent a change in InDel. Six types of drug resistance genes were found in the SNPs resistance gene-related loci, among which gene ANT(3'')-II and QacE mutations were found in each strain. Conclusion:In this study, the main mechanism of A. baumannii multi-drug resistance is due to the multi-drug efflux pump related genes. The point mutations at the SNPs sites of the six types of resistance genes are the main differences in A. baumannii between Chongqing and the eastern coastal areas of China.
| S-EPMC6750166 | biostudies-literature
Project description:Acinetobacter baumannii is one of the most successful pathogens causing nosocomial infections and has significantly multidrug-resistant. So far, there are no certain treatments to protect against infection with A. baumannii, therefore an effective A. baumannii vaccine needed. The purpose of this study was to predict antigenic epitopes of CarO protein for designing the A. baumannii vaccine using immunoinformatics analysis. CarO protein is one of the most important factors in the resistance against the antibiotic Carbapenem. In this study, T and B-cell epitopes of CarO protein were predicted and screened based on the antigenicity, toxicity, allergenicity features. The epitopes were linked by suitable linkers. Four different adjuvants were attached to the vaccine constructs which among them, vaccine construct 3 was chosen to predict the secondary and the 3D structure of the vaccine. The refinement process was performed to improve the quality of the 3D model structure; the validation process is performed using the Ramachandran plot and ProSA z-score. The designed vaccine's binding affinity to six various HLA molecules and TLR 2 and TLR4 were evaluated by molecular docking. Finally, in silico gene cloning was performed in the pET28a (+) vector. The findings suggest that the vaccine may be a promising vaccine to prevent A. baumannii infection.
| S-EPMC8397861 | biostudies-literature
Project description:In this study, we aimed to examine the relationships between antibiotic resistance, biofilm formation, and biofilm-specific resistance in clinical isolates of Acinetobacter baumannii. The tested 272 isolates were collected from several hospitals in China during 2010-2013. Biofilm-forming capacities were evaluated using the crystal violet staining method. Antibiotic resistance/susceptibility profiles to 21 antibiotics were assessed using VITEK 2 system, broth microdilution method or the Kirby-Bauer disc diffusion method. The minimum inhibitory concentration (MIC) and minimum biofilm eradication concentration (MBEC) to cefotaxime, imipenem, and ciprofloxacin were evaluated using micro dilution assays. Genetic relatedness of the isolates was also analyzed by pulsed-field gel electrophoresis (PFGE) and plasmid profile. Among all the 272 isolates, 31 were multidrug-resistant (MDR), and 166 were extensively drug-resistant (XDR). PFGE typing revealed 167 pattern types and 103 clusters with a similarity of 80%. MDR and XDR isolates built up the main prevalent genotypes. Most of the non-MDR isolates were distributed in a scattered pattern. Additionally, 249 isolates exhibited biofilm formation, among which 63 were stronger biofilm formers than type strain ATCC19606. Population that exhibited more robust biofilm formation likely contained larger proportion of non-MDR isolates. Isolates with higher level of resistance tended to form weaker biofilms. The MBECs for cefotaxime, imipenem, and ciprofloxacin showed a positive correlation with corresponding MICs, while the enhancement in resistance occurred independent of the quantity of biofilm biomass produced. Results from this study imply that biofilm acts as a mechanism for bacteria to get a better survival, especially in isolates with resistance level not high enough. Moreover, even though biofilms formed by isolates with high level of resistance are always weak, they could still provide similar level of protection for the isolates. Further explorations genetically would improve our understanding of these processes and provide novel insights in the therapeutics and prevention against A. baumannii biofilm-related infections.
| S-EPMC4828443 | biostudies-literature
Project description:Transcriptomic comparison of a multidrug resistant Acinetobacter baumannii strain and a susceptible reference strain.
2009-11-20 | E-MEXP-2254 | biostudies-arrayexpress
Project description:Nosocomial infections with Acinetobacter baumannii are a global problem in intensive care units with high mortality rates. Increasing resistance to first- and second-line antibiotics has forced the use of colistin as last-resort treatment, and increasing development of colistin resistance in A. baumannii has been reported. We evaluated the transcriptional regulator PmrA as potential drug target to restore colistin efficacy in A. baumannii Deletion of pmrA restored colistin susceptibility in 10 of the 12 extensively drug-resistant A. baumannii clinical isolates studied, indicating the importance of PmrA in the drug resistance phenotype. However, two strains remained highly resistant, indicating that PmrA-mediated overexpression of the phosphoethanolamine (PetN) transferase PmrC is not the exclusive colistin resistance mechanism in A. baumannii A detailed genetic characterization revealed a new colistin resistance mechanism mediated by genetic integration of the insertion element ISAbaI upstream of the PmrC homolog EptA (93% identity), leading to its overexpression. We found that eptA was ubiquitously present in clinical strains belonging to the international clone 2, and ISAbaI integration upstream of eptA was required to mediate the colistin-resistant phenotype. In addition, we found a duplicated ISAbaI-eptA cassette in one isolate, indicating that this colistin resistance determinant may be embedded in a mobile genetic element. Our data disprove PmrA as a drug target for adjuvant therapy but highlight the importance of PetN transferase-mediated colistin resistance in clinical strains. We suggest that direct targeting of the homologous PetN transferases PmrC/EptA may have the potential to overcome colistin resistance in A. baumanniiIMPORTANCE The discovery of antibiotics revolutionized modern medicine and enabled us to cure previously deadly bacterial infections. However, a progressive increase in antibiotic resistance rates is a major and global threat for our health care system. Colistin represents one of our last-resort antibiotics that is still active against most Gram-negative bacterial pathogens, but increasing resistance is reported worldwide, in particular due to the plasmid-encoded protein MCR-1 present in pathogens such as Escherichia coli and Klebsiella pneumoniae Here, we showed that colistin resistance in A. baumannii, a top-priority pathogen causing deadly nosocomial infections, is mediated through different avenues that result in increased activity of homologous phosphoethanolamine (PetN) transferases. Considering that MCR-1 is also a PetN transferase, our findings indicate that PetN transferases might be the Achilles heel of superbugs and that direct targeting of them may have the potential to preserve the activity of polymyxin antibiotics.
| S-EPMC6635527 | biostudies-literature
Project description:ObjectiveAcinetobacter baumannii has become an important problem because of the high drug resistance rate. The aim of this study was to assess the antimicrobial resistance profile and explore the role of membrane porin in imipenem resistance of A baumannii.MethodsA total of 63 isolates of imipenem-resistant A baumannii (IRAB) and 21 of imipenem-sensitive A baumannii (ISAB) were collected. Susceptibility testing to 16 kinds of antimicrobial agents was conducted by K-B method. PCR technique was used to detect carO and oprD genes, and sequencing was performed to compare the sequence between IRAB and ISAB. Three-dimensional structure model of CarO protein was established.ResultsWhile ISAB isolates presented sensitive to most classes of antibiotics, isolates of IRAB displayed much higher resistance rate except tigecycline (3.2%), cefoperazone/sulbactam (28.6%), and minocycline (30.2%). All 84 isolates were observed carrying both carO and oprD genes. Further sequencing revealed important mutations of carO gene existed in IRAB in comparison with ISAB. Meanwhile, significant differences in three-dimensional structure of carO protein molecule were also found between IRAB and ISAB.ConclusionsThe drug resistance profile of IRAB is increasingly severe in clinical settings. Mutation of CarO was identified as one of the molecular mechanisms involved in imipenem resistance in A baumannii.
| S-EPMC6805298 | biostudies-literature
Project description:With the growing emergence of pan-drug-resistant Acinetobacter baumannii (PDR-Ab) strains in clinical, new strategies for the treatment of PDR-Ab infections are urgently needed. Egg yolk immunoglobulin (IgY) as a convenient and inexpensive antibody has been widely applied to the therapy of infectious diseases. The aim of this study was to produce IgY specific to PDR-Ab and investigate its antibacterial effects in vitro and in vivo. IgYs specific to two PDR-Ab strains were produced by immunizing hens with formaldehyde inactivated PDR-Ab cells and isolated from yolks with a purity of 90% by water dilution, salt precipitations and ultrafiltration. IgYs showed high titers when subjected to an ELISA and inhibited the growth of PDR-Ab in a dose-dependent manner in liquid medium. Scanning electron microscopy assay showed structural modification and aggregation of PDR-Ab treated with specific IgYs. Freshly cultured PDR-Ab cells were nasally inhaled in BALB/c mice to induce acute pneumonia. The infected mice were intraperitoneally injected with specific IgYs using cefoperazone/sulbactam and dexamethasone as positive controls. The IgYs specific to PDR-Ab lowered the mortality of mice with PDR-Ab-induced acute pneumonia, decreased the level of TNF-? and IL-1? in serum and reduced inflammation in lung tissue. Specific IgY has the potential to be used as a new therapeutic approach for the treatment of A. baumannii-induced infections.
| S-EPMC7126593 | biostudies-literature
Project description:Acinetobacter baumannii is one of the most important opportunistic pathogens that causes serious health care associated complications in critically ill patients. In the current study we report on the diversity of the clinical multi-drug resistant (MDR) A. baumannii in Kuwait by molecular characterization. One hundred A. baumannii were isolated from one of the largest governmental hospitals in Kuwait. Following the identification of the isolates by molecular methods, the amplified bla OXA-51-like gene product of one isolate (KO-12) recovered from blood showed the insertion of the ISAba19 at position 379 in bla OXA-78. Of the 33 MDR isolates, 28 (85%) contained bla OXA-23, 2 (6%) bla OXA-24 and 6 (18%) bla PER-1 gene. We did not detect bla OXA-58, bla VIM, bla IMP, bla GES, bla VEB, and bla NDM genes in any of the tested isolates. In three bla PER-1 positive isolates the genetic environment of bla PER-1 consisted of two copies of ISPa12 (tnpiA1) surrounding the bla PER-1 gene on a highly stable plasmid of ca. 140-kb. Multilocus-sequence typing (MLST) analysis of the 33 A. baumannii isolates identified 20 different STs, of which six (ST-607, ST-608, ST-609, ST-610, ST-611, and ST-612) were novel. Emerging STs such as ST15 (identified for the first time in the Middle East), ST78 and ST25 were also detected. The predominant clonal complex was CC2. Pulsed-field gel electrophoresis and MLST defined the MDR isolates as multi-clonal with diverse lineages. Our results lead us to believe that A. baumannii is diverse in clonal origins and/or is undergoing clonal expansion continuously while multiple lineages of MDR A. baumannii circulate in hospital ward simultaneously.
| S-EPMC4513246 | biostudies-literature
Project description:These data are part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/
2015-08-04 | E-ERAD-375 | biostudies-arrayexpress
Project description:Acinetobacter baumannii has emerged worldwide as a dominant pathogen in a broad range of severe infections, raising an acute need for efficient antibacterials. This is the first report on the resistome and virulome of 33 extended drug-resistant and carbapenem-resistant A. baumannii (XDR CRAB) strains isolated from hospitalized and ambulatory patients in Bucharest, Romania. A total of 33 isolates were collected and analyzed using phenotypic antibiotic susceptibility and conjugation assays, PCR, whole-genome sequencing (WGS), pulsed-field gel electrophoresis (PFGE) and MultiLocus Sequence Typing (MLST). All isolates were extensively drug-resistant (XDR), being susceptible only to colistin. The carbapenem resistance was attributed by PCR mainly to blaOXA-24 and blaOXA-23 genes. PFGE followed by MLST analysis demonstrated the presence of nine pulsotypes and six sequence types. WGS of seven XDR CRAB isolates from healthcare-associated infections demonstrated the high diversity of resistance genes repertoire, as well as of mobile genetic elements, carrying ARGs for aminoglycosides, sulphonamides and macrolides. Our data will facilitate the understanding of resistance, virulence and transmission features of XDR AB isolates from Romanian patients and might be able to contribute to the implementation of appropriate infection control measures and to develop new molecules with innovative mechanisms of action, able to fight effectively against these bugs, for limiting the spread and decreasing the infection rate and mortality.
| S-EPMC8225882 | biostudies-literature