106 research outputs found
Fast emergence of phage-resistant Pseudomonas aeruginosa biofilm cells in response to the pressure exerted by bacteriophage treatment
Antibiotic resistance constitutes currently one of the most serious threats
to the global public health and it urgently requires new and effective
solutions. Bacteriophages are bacterial viruses increasingly recognized
as an attractive alternative to the conventional antibiotic therapies. In the
present study, the efficacy of phages against Pseudomonas aeruginosa
PAO1 biofilm and planktonic cell cultures was evaluated over the course
of 48 hours. Although significant reductions in the number of viable cells
were achieved for both cases, the high adaptation capability of bacteria in
response to the selective pressure caused by phage treatment, resulted
in the inevitable arising of phage-resistant variants. In most cases, those
variants appeared later in planktonic cultures than in biofilms. Given the
interest in further understanding their genetic makeup and possible
mutations accumulated, some were selected for further phenotypic and
genotypic characterization. The complete genomes of five P. aeruginosa
PAO1 phage-resistant variants were sequenced and all revealed to carry
mutations in the galU gene, which is involved in lipopolysaccharide core
biosynthesis, as well as in one pil gene, which is involved in type IV pilus
synthesis. Three of the P. aeruginosa PAO1 variants further revealed
large deletions (> 200 kbp) in their genomes. Overall the results of this
study reveal that the selective pressure caused by phages while targeting
biofilms results in a faster emergence of resistance compared to
planktonic cultures, probably due to the high genetic diversity of cells
within biofilms. Furthermore phage-resistant variants seem to be quite
adapted to the biofilm phenotype
Evaluation of a microarray-hybridization based method applicable for discovery of single nucleotide polymorphisms (SNPs) in the Pseudomonas aeruginosa genome
<p>Abstract</p> <p>Background</p> <p>Whole genome sequencing techniques have added a new dimension to studies on bacterial adaptation, evolution and diversity in chronic infections. By using this powerful approach it was demonstrated that <it>Pseudomonas aeruginosa </it>undergoes intense genetic adaptation processes, crucial in the development of persistent disease. The challenge ahead is to identify universal infection relevant adaptive bacterial traits as potential targets for the development of alternative treatment strategies.</p> <p>Results</p> <p>We developed a microarray-based method applicable for discovery of single nucleotide polymorphisms (SNPs) in <it>P. aeruginosa </it>as an easy and economical alternative to whole genome sequencing. About 50% of all SNPs theoretically covered by the array could be detected in a comparative hybridization of PAO1 and PA14 genomes at high specificity (> 0.996). Variations larger than SNPs were detected at much higher sensitivities, reaching nearly 100% for genetic differences affecting multiple consecutive probe oligonucleotides. The detailed comparison of the <it>in silico </it>alignment with experimental hybridization data lead to the identification of various factors influencing sensitivity and specificity in SNP detection and to the identification of strain specific features such as a large deletion within the PA4684 and PA4685 genes in the Washington Genome Center PAO1.</p> <p>Conclusion</p> <p>The application of the genome array as a tool to identify adaptive mutations, to depict genome organizations, and to identify global regulons by the "ChIP-on-chip" technique will expand our knowledge on <it>P. aeruginosa </it>adaptation, evolution and regulatory mechanisms of persistence on a global scale and thus advance the development of effective therapies to overcome persistent disease.</p
Microbiological findings in early and late implant loss: an observational clinical case-controlled study
Background
Implants are a predictable and well-established treatment method in dentistry. Nevertheless, looking at possible failures of dental implants, early and late loss have to be distinguished. The intent of the study was to report microbiological findings on the surface of implants with severe peri-implantitis, which had to be explanted.
Methods
53 specimens of implants from 48 patients without severe general illnesses have been examined. The groups investigated were implants that had to be removed in the period of osseointegration (early loss, 13 patients with 14 implants) or after the healing period (late loss, 14 patients with 17 implants). The implant losses were compared with two control groups (implants with no bone loss directly after completed osseointegration, two to four months after implant placement (17 patients with 17 implants) and implants with no bone loss and prosthetic restoration for more than three years (5 patients with 5 implants)). Data about the bacteria located in the peri-implant sulcus was collected using amplification and high throughput sequencing of the 16S rRNA gene.
Results
The biofilm composition differed substantially between individuals. Both in early and late implant loss, Fusobacterium nucleatum and Porphyromonas gingivalis were found to be abundant. Late lost implants showed higher bacterial diversity and in addition higher abundances of Treponema, Fretibacterium, Pseudoramibacter and Desulfobulbus, while microbial communities of early loss implants were very heterogeneous and showed no significantly more abundant bacterial taxa.
Conclusions
Specific peri-implant pathogens were found around implants that were lost after a primarily uneventful osseointegration. P. gingivalis and F. nucleatum frequently colonized the implant in early and late losses and could therefore be characteristic for implant loss in general. In general, early lost implants showed also lower microbial diversity than late losses. However, the microbial results were not indicative of the causes of early and late losses
Evolutionary conservation of essential and highly expressed genes in Pseudomonas aeruginosa
<p>Abstract</p> <p>Background</p> <p>The constant increase in development and spread of bacterial resistance to antibiotics poses a serious threat to human health. New sequencing technologies are now on the horizon that will yield massive increases in our capacity for DNA sequencing and will revolutionize the drug discovery process. Since essential genes are promising novel antibiotic targets, the prediction of gene essentiality based on genomic information has become a major focus.</p> <p>Results</p> <p>In this study we demonstrate that pooled sequencing is applicable for the analysis of sequence variations of strain collections with more than 10 individual isolates. Pooled sequencing of 36 clinical <it>Pseudomonas aeruginosa </it>isolates revealed that essential and highly expressed proteins evolve at lower rates, whereas extracellular proteins evolve at higher rates. We furthermore refined the list of experimentally essential <it>P. aeruginosa </it>genes, and identified 980 genes that show no sequence variation at all. Among the conserved nonessential genes we found several that are involved in regulation, motility and virulence, indicating that they represent factors of evolutionary importance for the lifestyle of a successful environmental bacterium and opportunistic pathogen.</p> <p>Conclusion</p> <p>The detailed analysis of a comprehensive set of <it>P. aeruginosa </it>genomes in this study clearly disclosed detailed information of the genomic makeup and revealed a large set of highly conserved genes that play an important role for the lifestyle of this microorganism. Sequencing strain collections enables for a detailed and extensive identification of sequence variations as potential bacterial adaptation processes, e.g., during the development of antibiotic resistance in the clinical setting and thus may be the basis to uncover putative targets for novel treatment strategies.</p
Efficient Bioelectrochemical Conversion of Industrial Wastewater by Specific Strain Isolation and Community Adaptation
The aim of this study was the development of a specifically adapted microbial community for the removal of organic carbon from an industrial wastewater using a bioelectrochemical system. In a first step, ferric iron reducing microorganisms were isolated from the examined industrial wastewater. In a second step, it was tested to what extent these isolates or a cocultivation of the isolates with the exoelectrogenic model organism Geobacter sulfurreducens (G. sulfurreducens) were able to eliminate organic carbon from the wastewater. To establish a stable biofilm on the anode and to analyze the performance of the system, the experiments were conducted first under batch-mode conditions for 21 days. Since the removal of organic carbon was relatively low in the batch system, a similar experiment was conducted under continuous-mode conditions for 65 days, including a slow transition from synthetic medium to industrial wastewater as carbon and electron source and variations in the flow rate of the medium. The overall performance of the system was strongly increased in the continuous- compared to the batch-mode reactor and the highest average current density (1,368 mA/m2) and Coulombic efficiency (54.9%) was measured in the continuous-mode reactor inoculated with the coculture consisting of the new isolates and G. sulfurreducens. The equivalently inoculated batch-mode system produced only 82-fold lower current densities, which were accompanied by 42-fold lower Coulombic efficiencies
Gut Microbiome Composition in Obese and Non-Obese Persons: A Systematic Review and Meta-Analysis
Whether the gut microbiome in obesity is characterized by lower diversity and altered composition at the phylum or genus level may be more accurately investigated using high-throughput sequencing technologies. We conducted a systematic review in PubMed and Embase including 32 cross-sectional studies assessing the gut microbiome composition by high-throughput sequencing in obese and non-obese adults. A significantly lower alpha diversity (Shannon index) in obese versus non-obese adults was observed in nine out of 22 studies, and meta-analysis of seven studies revealed a non-significant mean difference (-0.06, 95% CI -0.24, 0.12, I2 = 81%). At the phylum level, significantly more Firmicutes and fewer Bacteroidetes in obese versus non-obese adults were observed in six out of seventeen, and in four out of eighteen studies, respectively. Meta-analyses of six studies revealed significantly higher Firmicutes (5.50, 95% 0.27, 10.73, I2 = 81%) and non-significantly lower Bacteroidetes (-4.79, 95% CI -10.77, 1.20, I2 = 86%). At the genus level, lower relative proportions of Bifidobacterium and Eggerthella and higher Acidaminococcus, Anaerococcus, Catenibacterium, Dialister, Dorea, Escherichia-Shigella, Eubacterium, Fusobacterium, Megasphera, Prevotella, Roseburia, Streptococcus, and Sutterella were found in obese versus non-obese adults. Although a proportion of studies found lower diversity and differences in gut microbiome composition in obese versus non-obese adults, the observed heterogeneity across studies precludes clear answers
RNASeq Based Transcriptional Profiling of Pseudomonas aeruginosa PA14 after Short- and Long-Term Anoxic Cultivation in Synthetic Cystic Fibrosis Sputum Medium
The opportunistic human pathogen Pseudomonas aeruginosa can thrive under microaerophilic to anaerobic conditions in the lungs of cystic fibrosis patients. RNA based comparative RNA profiling of the clinical isolate PA14 cultured in synthetic cystic fibrosis medium was performed after planktonic growth (OD = 2.0; P), 30 min after shift to anaerobiosis (A-30) and after anaerobic biofilm growth for 96h (B-96) with the aim to reveal differentially regulated functions impacting on sustained anoxic biofilm formation as well as on tolerance towards different antibiotics. Most notably, functions involved in sulfur metabolism were found to be up-regulated in B-96 cells when compared to A-30 cells. Based on the transcriptome studies a set of transposon mutants were screened, which revealed novel functions involved in anoxic biofilm growth.In addition, these studies revealed a decreased and an increased abundance of the oprD and the mexCD-oprJ operon transcripts, respectively, in B-96 cells, which may explain their increased tolerance towards meropenem and to antibiotics that are expelled by the MexCD-OprD efflux pump. The OprI protein has been implicated as a target for cationic antimicrobial peptides, such as SMAP-29. The transcriptome and subsequent Northern-blot analyses showed that the abundance of the oprI transcript encoding the OprI protein is strongly decreased in B-96 cells. However, follow up studies revealed that the susceptibility of a constructed PA14ÎoprI mutant towards SMAP-29 was indistinguishable from the parental wild-type strain, which questions OprI as a target for this antimicrobial peptide in strain PA14
Microbiological findings in early and late implant loss: an observational clinical case-controlled study
Background:
Implants are a predictable and well-established treatment method in dentistry. Nevertheless, looking at possible failures of dental implants, early and late loss have to be distinguished. The intent of the study was to report microbiological findings on the surface of implants with severe peri-implantitis, which had to be explanted.
Methods:
53 specimens of implants from 48 patients without severe general illnesses have been examined. The groups investigated were implants that had to be removed in the period of osseointegration (early loss, 13 patients with 14 implants) or after the healing period (late loss, 14 patients with 17 implants). The implant losses were compared with two control groups (implants with no bone loss directly after completed osseointegration, two to four months after implant placement (17 patients with 17 implants) and implants with no bone loss and prosthetic restoration for more than three years (5 patients with 5 implants)). Data about the bacteria located in the peri-implant sulcus was collected using amplification and high throughput sequencing of the 16S rRNA gene.
Results:
The biofilm composition differed substantially between individuals. Both in early and late implant loss, Fusobacterium nucleatum and Porphyromonas gingivalis were found to be abundant. Late lost implants showed higher bacterial diversity and in addition higher abundances of Treponema, Fretibacterium, Pseudoramibacter and Desulfobulbus, while microbial communities of early loss implants were very heterogeneous and showed no significantly more abundant bacterial taxa.
Conclusions:
Specific peri-implant pathogens were found around implants that were lost after a primarily uneventful osseointegration. P. gingivalis and F. nucleatum frequently colonized the implant in early and late losses and could therefore be characteristic for implant loss in general. In general, early lost implants showed also lower microbial diversity than late losses. However, the microbial results were not indicative of the causes of early and late losses
Does the antidiabetic drug metformin affect embryo development and the health of brown trout (Salmo trutta f. fario)?
Abstract Background Due to the rising number of type 2 diabetes patients, the antidiabetic drug, metformin is currently among those pharmaceuticals with the highest consumption rates worldwide. Via sewage-treatment plants, metformin enters surface waters where it is frequently detected in low concentrations (”g/L). Since possible adverse effects of this substance in aquatic organisms have been insufficiently explored to date, the aim of this study was to investigate the impact of metformin on health and development in brown trout (Salmo trutta f. fario) and its microbiome. Results Brown trout embryos were exposed to 0, 1, 10, 100 and 1000 ”g/L metformin over a period from 48 days post fertilisation (dpf) until 8 weeks post-yolk sac consumption at 7 °C (156 dpf) and 11 °C (143 dpf). Chemical analyses in tissues of exposed fish showed the concentration-dependent presence of metformin in the larvae. Mortality, embryonic development, body length, liver tissue integrity, stress protein levels and swimming behaviour were not influenced. However, compared to the controls, the amount of hepatic glycogen was higher in larvae exposed to metformin, especially in fish exposed to the lowest metformin concentration of 1 ”g/L, which is environmentally relevant. At higher metformin concentrations, the glycogen content in the liver showed a high variability, especially for larvae exposed to 1000 ”g/L metformin. Furthermore, the body weight of fish exposed to 10 and 100 ”g/L metformin at 7 °C and to 1 ”g/L metformin at 11 °C was decreased compared with the respective controls. The results of the microbiome analyses indicated a shift in the bacteria distribution in fish exposed to 1 and 10 ”g/L metformin at 7 °C and to 100 ”g/L metformin at 11 °C, leading to an increase of Proteobacteria and a reduction of Firmicutes and Actinobacteria. Conclusions Overall, weight reduction and the increased glycogen content belong to the described pharmaceutical effects of the drug in humans, but this study showed that they also occur in brown trout larvae. The impact of a shift in the intestinal microbiome caused by metformin on the immune system and vitality of the host organism should be the subject of further research before assessing the environmental relevance of the pharmaceutical
The UBA domain of conjugating enzyme Ubc1/Ube2K facilitates assembly of K48/K63âbranched ubiquitin chains
The assembly of a specific polymeric ubiquitin chain on a target protein is a key event in the regulation of numerous cellular processes. Yet, the mechanisms that govern the selective synthesis of particular polyubiquitin signals remain enigmatic. The homologous ubiquitin-conjugating (E2) enzymes Ubc1 (budding yeast) and Ube2K (mammals) exclusively generate polyubiquitin linked through lysine 48 (K48). Uniquely among E2 enzymes, Ubc1 and Ube2K harbor a ubiquitin-binding UBA domain with unknown function. We found that this UBA domain preferentially interacts with ubiquitin chains linked through lysine 63 (K63). Based on structural modeling, in vitro ubiquitination experiments, and NMR studies, we propose that the UBA domain aligns Ubc1 with K63-linked polyubiquitin and facilitates the selective assembly of K48/K63-branched ubiquitin conjugates. Genetic and proteomics experiments link the activity of the UBA domain, and hence the formation of this unusual ubiquitin chain topology, to the maintenance of cellular proteostasis.Deutsche Forschungsgemeinschaft (DFG)
http://dx.doi.org/10.13039/501100001659MaxâPlanckâGesellschaft (MPG)
http://dx.doi.org/10.13039/501100004189Peer Reviewe
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