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Whole Transcriptome Analysis of Acinetobacter baumannii Assessed by RNA-Sequencing Reveals Different mRNA Expression Profiles in Biofilm Compared to Planktonic Cells  [PDF]
Soraya Rumbo-Feal, Manuel J. Gómez, Carmen Gayoso, Laura álvarez-Fraga, María P. Cabral, Ana M. Aransay, Naiara Rodríguez-Ezpeleta, Ane Fullaondo, Jaione Valle, María Tomás, Germán Bou, Margarita Poza
PLOS ONE , 2013, DOI: 10.1371/journal.pone.0072968
Abstract: Acinetobacter baumannii has emerged as a dangerous opportunistic pathogen, with many strains able to form biofilms and thus cause persistent infections. The aim of the present study was to use high-throughput sequencing techniques to establish complete transcriptome profiles of planktonic (free-living) and sessile (biofilm) forms of A. baumannii ATCC 17978 and thereby identify differences in their gene expression patterns. Collections of mRNA from planktonic (both exponential and stationary phase cultures) and sessile (biofilm) cells were sequenced. Six mRNA libraries were prepared following the mRNA-Seq protocols from Illumina. Reads were obtained in a HiScanSQ platform and mapped against the complete genome to describe the complete mRNA transcriptomes of planktonic and sessile cells. The results showed that the gene expression pattern of A. baumannii biofilm cells was distinct from that of planktonic cells, including 1621 genes over-expressed in biofilms relative to stationary phase cells and 55 genes expressed only in biofilms. These differences suggested important changes in amino acid and fatty acid metabolism, motility, active transport, DNA-methylation, iron acquisition, transcriptional regulation, and quorum sensing, among other processes. Disruption or deletion of five of these genes caused a significant decrease in biofilm formation ability in the corresponding mutant strains. Among the genes over-expressed in biofilm cells were those in an operon involved in quorum sensing. One of them, encoding an acyl carrier protein, was shown to be involved in biofilm formation as demonstrated by the significant decrease in biofilm formation by the corresponding knockout strain. The present work serves as a basis for future studies examining the complex network systems that regulate bacterial biofilm formation and maintenance.
Growth of Acinetobacter baumannii in Pellicle Enhanced the Expression of Potential Virulence Factors  [PDF]
Sara Marti, Yassine Nait Chabane, Stéphane Alexandre, Laurent Coquet, Jordi Vila, Thierry Jouenne, Emmanuelle Dé
PLOS ONE , 2011, DOI: 10.1371/journal.pone.0026030
Abstract: Background Interestingly, Acinetobacter baumannii presents an enhanced capacity to form biofilms (also named pellicles) at the air-liquid interface as compared to the other Acinetobacter species. This characteristic questions the contribution of this phenotype to an increased risk of clinical infections by this pathogen. Methodology/Principal Findings By a proteomic approach using 2-D gel electrophoresis-LC-MS/MS mass spectrometry, we compared the membrane protein patterns of A. baumannii 77, a pellicle-forming clinical isolate, grown in planktonic and in sessile modes. We identified 52 proteins with a differential expression, including 32 up-regulated and 20 down-regulated in the pellicle state. Several proteins, differentially expressed during pellicle development, were of particular interest. We determined the over-expression of four siderophore iron uptake systems including the acinetobactin and enterobactin receptors and confirmed that the development of this type of biofilm is promoted by ferric ions. Two over-expressed proteins, CarO and an OprD-homologue, putative carbapenem-resistance associated porins, would be involved in the transport of specific compounds, like ornithine, a biosynthesis precursor of a siderophore from the hydroxamate family. We evidenced the overexpression of a lipase and a transporter of LCFA that may be involved in the recycling of lipids inside the pellicle matrix. Finally, we demonstrated both by proteomic and by AFM studies that this particular type of biofilm required multiple pili systems to maintain this cohesive structure at the air-liquid interface; two of these systems have never been described in A. baumannii. Conclusions/Significance Our study demonstrated that several proteins, overexpressed at a late state of pellicle development, could be potentially involved in virulence processes. Therefore, regarding the number of potential virulence factors that are over-expressed in this growth mode, the pellicle-forming clinical isolates should be kept under survey.
Deciphering the Multifactorial Nature of Acinetobacter baumannii Pathogenicity  [PDF]
Luísa C. S. Antunes, Francesco Imperi, Alessandra Carattoli, Paolo Visca
PLOS ONE , 2011, DOI: 10.1371/journal.pone.0022674
Abstract: Background Acinetobacter baumannii is an emerging bacterial pathogen that causes a broad array of infections, particularly in hospitalized patients. Many studies have focused on the epidemiology and antibiotic resistance of A. baumannii, but little is currently known with respect to its virulence potential. Methodology/Principal Findings The aim of this work was to analyze a number of virulence-related traits of four A. baumannii strains of different origin and clinical impact for which complete genome sequences were available, in order to tentatively identify novel determinants of A. baumannii pathogenicity. Clinical strains showed comparable virulence in the Galleria mellonella model of infection, irrespective of their status as outbreak or sporadic strains, whereas a non-human isolate was avirulent. A combined approach of genomic and phenotypic analyses led to the identification of several virulence factors, including exoproducts with hemolytic, phospholipase, protease and iron-chelating activities, as well as a number of multifactorial phenotypes, such as biofilm formation, surface motility and stress resistance, which were differentially expressed and could play a role in A. baumannii pathogenicity. Conclusion/Significance This work provides evidence of the multifactorial nature of A. baumannii virulence. While A. baumannii clinical isolates could represent a selected population of strains adapted to infect the human host, subpopulations of highly genotypically and phenotypically diverse A. baumannii strains may exist outside the hospital environment, whose relevance and distribution deserve further investigation.
Growth in glucose-based medium and exposure to subinhibitory concentrations of imipenem induce biofilm formation in a multidrug-resistant clinical isolate of Acinetobacter baumannii
Elisabetta Nucleo, Laura Steffanoni, Giulia Fugazza, Roberta Migliavacca, Ernesto Giacobone, Antonella Navarra, Laura Pagani, Paolo Landini
BMC Microbiology , 2009, DOI: 10.1186/1471-2180-9-270
Abstract: Over a six-year period, 73 isolates of A. baumannii were collected from infected patients in two hospitals in Italy. While 69 out of the 73 isolates displayed identical multidrug antibiotic resistance pattern, they were susceptible to carbapenems. Genetic profiles of these 69 isolates, determined by Pulsed Field Gel Electrophoresis (PFGE), indicated that they were genetically related and could be clustered in a specific clone, called SMAL. We tested the ability of the SMAL clone to form biofilm, an important determinant for bacterial colonization of the human host and for persistence in the hospital environment. Biofilm formation by A. baumannii SMAL, measured as surface adhesion to polystyrene, is strongly affected by growth conditions, being impaired in rich growth media such as LB, while being favoured in glucose-based medium. Surface adhesion in glucose-based media is inhibited by treatment with cellulase, suggesting that it depends on production of cellulose or of a chemically related extracellular polysaccharide. Exposure of A. baumannii SMAL to subinhibitory concentrations of imipenem resulted in biofilm stimulation and increased production of iron uptake proteins. Growth in iron-supplemented medium also stimulated surface adhesion, thus suggesting that increased intracellular iron concentrations might act as an environmental signal for biofilm formation in A. baumannii SMAL.Our results indicate that exposure to subinhibitory concentrations of imipenem can stimulate biofilm formation and induce iron uptake in a pathogenic strain of A. baumannii, with potential implications on antibiotic susceptibility and ability to persist in the human host.Bacteria belonging to the genus Acinetobacter, in particular Acinetobacter baumannii and the closely related Acinetobacter 13 TU and gen.sp. 3 (referred to as Acinetobacter baumannii sensu lato), are important opportunistic pathogens in hospital-acquired infections (reviewed in [1]).A. baumannii can cause pneumonia, wound
Fulminating septicemia due to persistent pan-resistant community-acquired metallo-β-lactamase (IMP-1)-positive Acinetobacter baumannii  [cached]
Telang Nilima,Satpute Meenakshi,Dhakephalkar Prashant,Niphadkar Krishna
Indian Journal of Pathology and Microbiology , 2011,
Abstract: Acinetobacter baumannii is considered as an emerging nosocomial pathogen and is renowned for its multi-drug resistance. We report a case of community-acquired pan-resistant A. baumannii caused fulminating septicemia. The treatment failure led to death. The A. baumannii strain isolated from blood, pus, urine and tracheal aspirate was confirmed by 16S r-RNA sequence homology and found positive for metallo-β-lactamase IMP-1, and was found to be a strong biofilm producer. The isolate was only susceptible (moderately) to colistin.
Correlation between biofilm production and multiple drug resistance in imipenem resistant clinical isolates of Acinetobacter baumannii  [cached]
Rao R,Karthika R,Singh S,Shashikala P
Indian Journal of Medical Microbiology , 2008,
Abstract: Purpose: To study the qualitative and quantitative methods for the investigation of biofilm formation and to examine the correlation between biofilm and antibiotic resistance among the clinical isolates of Acinetobacter baumannii . We also verified the association between biofilm and presence of extended spectrum β-lactamases, particularly, bla PER-1 . Methods: A total of 55 isolates were subjected to susceptibility testing by disc diffusion method for 13 clinically relevant antibiotics. Screening for biofilm production was done by both qualitative and quantitative methods through tube and microtitre plate assay respectively. The presence of bla PER-1 was checked by PCR. Results: A. baumannii isolates showed very high resistance (>75%) to imipenem, cephotaxime, amikacin and ciprofloxacin. Only cefoperazone, netillin and norfloxacin were found to be effective agents. Results of microtitre and tube methods were concordant with 34 isolates (62%) showing biofilm formation. Resistance to four antibiotics such as amikacin (82% vs. 17.6%, P < 0.001), cephotaxime (88% vs. 11%, P P < 0.001), ciprofloxacin (70% vs. 29%, P =0.005) and aztreonam (38% vs. 11%, P =0.039) was comparatively higher among biofilm producers than non-biofilm producers. Microtitre assay additionally detected 14 weakly adherent isolates. Only 11 isolates had bla PER-1 gene and among these two were strong biofilm producers, while remaining were weakly adherent isolates. Conclusion: Microtitre plate method was found to be a more sensitive method for biofilm detection. This study demonstrates a high propensity among the clinical isolates of A. baumannii to form biofilm and a significant association of biofilms with multiple drug resistance. Presence of bla PER-1 appears to be more critical for cell adherence than for biofilm formation.
An MFS Transporter-Like ORF from MDR Acinetobacter baumannii AIIMS 7 Is Associated with Adherence and Biofilm Formation on Biotic/Abiotic Surface  [PDF]
Praveen K. Sahu,Pavithra S. Iyer,Madhumita B. Gaikwad,Sheetal C. Talreja,Karishma R. Pardesi,Balu A. Chopade
International Journal of Microbiology , 2012, DOI: 10.1155/2012/490647
Abstract: A major facilitator superfamily (MFS) transporter-like open reading frame (ORF) of 453?bp was identified in a pathogenic strain Acinetobacter baumannii AIIMS 7, and its association with adherence and biofilm formation was investigated. Reverse transcription PCR (RT-PCR) showed differential expression in surface-attached biofilm cells than nonadherent cells. In vitro translation showed synthesis of a 17 kDa protein, further confirmed by cloning and heterologous expression in E. coli DH5 . Up to 2.1-, 3.1-, and 4.1- fold biofilm augmentation was observed on abiotic (polystyrene) and biotic (S. cerevisiae/HeLa) surface, respectively. Scanning electron microscopy (SEM) and gfp-tagged fluorescence microscopy revealed increased adherence to abiotic (glass) and biotic (S. cerevisiae) surface. Extracellular DNA(eDNA) was found significantly during active growth; due to probable involvement of the protein in DNA export, strong sequence homology with MFS transporter proteins, and presence of transmembrane helices. In summary, our findings show that the putative MFS transporter-like ORF (pmt) is associated with adherence, biofilm formation, and probable eDNA release in A. baumannii AIIMS 7. 1. Introduction Multidrug resistant (MDR) Acinetobacter baumannii [1–3] causes a range of infections in hospital environments [4, 5] which are often difficult to treat. Its pathogenesis and resistance are enhanced largely due to formation of biofilms; especially on clinically important surfaces [6]. Acinetobacter is capable of surviving on nutrient-limited abiotic surfaces and stressful environmental conditions by forming biofilms [7]. Biofilm formation is a dynamic process involving the attachment of bacteria to a biotic or abiotic surface followed by growth and maturation [8]. Property of adhesion to surfaces and subsequent formation of biofilm can be conferred by large number of genetic factors in MDR A. baumannii which are still not completely understood. Biofilm development in A. baumannii on abiotic surfaces is mediated by biofilm associated protein-Bap, as shown earlier by Loehfelm et al. [9]. Study by Tomaras and group [10] shows that BfmR, a part of a two-component regulatory system, plays an important role in the morphology of A. baumannii 19606 and the ability to form biofilms on abiotic surfaces. Earlier it was shown that expression of csuAB gene cluster is required for pili formation and attachment to abiotic surfaces, ensuing biofilm formation [11]. Adherence of A. baumannii to polystyrene and respiratory epithelial cells is correlated to antibiotic resistance
Predictors of Mortality in Acinetobacter baumannii Bacteremia  [cached]
Ay?egül Ulu-K?l??,?nder Erg?nül,Aysel Kocagül-?elikba?,Ba?ak Dokuzo?uz
Klimik Journal , 2011,
Abstract: Objective: We aimed to describe the risk factors for fatality among the patients with Acinetobacter baumannii bacteremia. Methods: Patients with a positive blood culture for A. baumannii by BacT/Alert (bioMérieux, Durham, NC) system were selected. Surviving and nonsurviving patients were compared in terms of demographic characteristics, underlying diseases, invasive procedures, and antibiotic resistance of the microorganisms. Therapies were evaluated according to the results of antibiotic susceptibility tests, and impact of inappropriate therapies for A. baumannii bacteremia was evaluated statistically.Results: Of the 40 patients enrolled, 21 (52.5%) died due to A. baumannii bacteremia. Univariate analysis of risk factors for fatality revealed the following parameters to be significant: >50 years of age, an APACHE II score >16, and transfer from one unit to another more than once (p<0.05). Cox regression survival analysis demonstrated that the death rate was 3.5 times higher in the patients older than 50 years of age, and 2.8 times higher in the patients with an APACHE II score >16. Overall imipenem resistance in A. baumannii was 67.5%, and carbapenem resistance among fatal cases was 76%. When the antibiotic sensitivity results were assessed, it was noted that 70% of the patients had been receiving inappropriate therapy. However, the antibiotic therapy received or initiated empirically was not shown to be a significant risk factor for mortality (p>0.05).Conclusions: Knowing the risk factors that indicate mortality in such patients and ensuring early diagnosis are of critical importance with respect to prognosis. Starting appropriate therapy by making the right decisions regarding the severity of acute and underlying conditions will have favorable effects on the management of A. baumannii bacteremia.
Inhibition of Acinetobacter baumannii Biofilm Formation by Methanolic Extract of Nothoscordum bivalve  [PDF]
Hernández-Marín David Alejandro, Mu?oz-Ortega Martín Humberto, Guevara-Lara Fidel, Gúzman-Lucio Marco Antonio, Sánchez-García Eduardo
Advances in Microbiology (AiM) , 2018, DOI: 10.4236/aim.2018.85028
Abstract: Acinetobacter baumannii is one of the most prevalent pathogens in nosocomial infections and has been cause of concern in recent years because, it has presented multiresistance to antibiotics and besides can form biofilms on biotic and abiotic surfaces like tissues and medical devices. Therefore, the search for new alternatives of natural origin to inhibit biofilm formation is being conducted. In the present investigation, sub-lethal concentrations (5.61 mg/mL, 3.74 mg/mL and 1.87 mg/mL) of methanolic extract of Nothoscordum bivalve were evaluated, showing biofilm formation inhibition up to 40.8%, in one nosocomial isolated of A. baumannii by the microtiter biofilm formation assay using crystal violet. On the other hand, the concentrations of 5.61 mg/mL and 3.74 mg/mL, caused an overexpression (up to 15.4 times) in the genes involved in the formation of biofilm (abaI, bap and csuE); due to this, the interaction of the extract with the bacteria was analyzed by scanning electron microscopy (SEM) and cellular damage was observed in the structure and stability of biofilm.
Successful treatment of multidrug resistant Acinetobacter baumannii meningitis
Acinetobacter baumannii, Post-surgical meningitis, High dose meropenem
Journal of Infection in Developing Countries , 2007,
Abstract: Background: Acinetobacter baumannii is a major cause of nosocomial infections in many hospitals and appears to have a propensity for developing multiple antimicrobial resistance rapidly.Cases: We report two cases with post-surgical meningitis due to multidrug resistant A. baumannii which were successfully treated with high-dose intravenous meropenem therapy.Conclusions: Multidrug resistant Acinetobacter spp. in intensive care units are a growing concern. High-dose meropenem is used in the treatment of these infections.
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