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International Journal of Molecular... Mar 2020Biofilms are a multicellular way of life, where bacterial cells are close together and embedded in a hydrated macromolecular matrix which offers a number of advantages...
Biofilms are a multicellular way of life, where bacterial cells are close together and embedded in a hydrated macromolecular matrix which offers a number of advantages to the cells. Extracellular polysaccharides play an important role in matrix setup and maintenance. A water-insoluble polysaccharide was isolated and purified from the biofilm produced by strain H111, a cystic fibrosis pathogen. Its composition and glycosidic linkages were determined using Gas-Liquid Chromatography-Mass Spectrometry (GLC-MS) on appropriate carbohydrate derivatives while its complete structure was unraveled by 1D and 2D NMR spectroscopy in deuterated sodium hydroxide (NaOD) aqueous solutions. All the collected data demonstrated the following repeating unit for the water-insoluble . biofilm polysaccharide: [3)-α-d-Gal-(1→3)-α-d-Glc-(1→3)-α-d-Gal-(1→3)-α-d-Man-(1→] Molecular modelling was used, coupled with NMR Nuclear Overhauser Effect (NOE) data, to obtain information about local structural motifs which could give hints about the polysaccharide insolubility. Both modelling and NMR data pointed at restricted dynamics of local conformations which were ascribed to the presence of inter-residue hydrogen bonds and to steric restrictions. In addition, the good correlation between NOE data and calculated interatomic distances by molecular dynamics simulations validated potential energy functions used for calculations.
Topics: Biofilms; Burkholderia cenocepacia; Glycosides; Hydrophobic and Hydrophilic Interactions; Polysaccharides, Bacterial; Solubility
PubMed: 32131450
DOI: 10.3390/ijms21051702 -
Applied and Environmental Microbiology May 2017The complex (Bcc) consists of 20 closely related Gram-negative bacterial species that are significant pathogens for persons with cystic fibrosis (CF). Some Bcc strains...
The complex (Bcc) consists of 20 closely related Gram-negative bacterial species that are significant pathogens for persons with cystic fibrosis (CF). Some Bcc strains are highly transmissible and resistant to multiple antibiotics, making infection difficult to treat. A tailocin (phage tail-like bacteriocin), designated BceTMilo, with a broad host range against members of the Bcc, was identified in strain BC0425. Sixty-eight percent of Bcc representing 10 species and 90% of non-Bcc strains tested were sensitive to BceTMilo. BceTMilo also showed killing activity against PAO1 and derivatives. Liquid chromatography-mass spectrometry analysis of the major BceTMilo proteins was used to identify a 23-kb tailocin locus in a draft BC0425 genome. The BceTMilo locus was syntenic and highly similar to a 24.6-kb region on chromosome 1 of J2315 (BCAL0081 to BCAL0107). A close relationship and synteny were observed between BceTMilo and phage KL3 and, by extension, with paradigm temperate myophage P2. Deletion mutants in the gene cluster encoding enzymes for biosynthesis of lipopolysaccharide (LPS) in the indicator strain K56-2 conferred resistance to BceTMilo. Analysis of the defined mutants in LPS biosynthetic genes indicated that an α-d-glucose residue in the core oligosaccharide is the receptor for BceTMilo. BceTMilo, presented in this study, is a broad-host-range tailocin active against spp. As such, BceTMilo and related or modified tailocins have potential as bactericidal therapeutic agents against plant- and human-pathogenic .
Topics: Anti-Bacterial Agents; Bacteriocins; Burkholderia cenocepacia; Burkholderia cepacia complex; Genome, Bacterial; Genome, Viral; Host Specificity; Humans; Mass Spectrometry; Molecular Structure; Pseudomonas aeruginosa
PubMed: 28258146
DOI: 10.1128/AEM.03414-16 -
Applied and Environmental Microbiology Feb 2022It has been demonstrated that quorum sensing (QS) is widely employed by bacterial cells to coordinately regulate various group behaviors. Diffusible signal factor... (Review)
Review
It has been demonstrated that quorum sensing (QS) is widely employed by bacterial cells to coordinately regulate various group behaviors. Diffusible signal factor (DSF)-type signals have emerged as a growing family of conserved cell-cell communication signals. In addition to the DSF signal initially identified in Xanthomonas campestris pv. campestris, iffusible ignal actor (BDSF) (-2-dodecenoic acid) has been recognized as a conserved DSF-type signal with specific characteristics in both signal perception and transduction from DSF signals. Here, we review the history and current progress of the research on this type of signal, especially focusing on its biosynthesis, signaling pathways, and biological functions. We also discuss and explore the huge potential of targeting this kind of QS system as a new therapeutic strategy to control bacterial infections and diseases.
Topics: Bacterial Proteins; Burkholderia; Burkholderia cenocepacia; Fatty Acids, Monounsaturated; Gene Expression Regulation, Bacterial; Quorum Sensing; Suppressor Factors, Immunologic
PubMed: 34985987
DOI: 10.1128/aem.02342-21 -
Journal of Bacteriology Oct 2023Antibiotic resistance in bacteria is a growing global concern and has spurred increasing efforts to find alternative therapeutics, such as the use of bacterial viruses,...
Antibiotic resistance in bacteria is a growing global concern and has spurred increasing efforts to find alternative therapeutics, such as the use of bacterial viruses, or bacteriophages. One promising approach is to use phages that not only kill pathogenic bacteria but also select phage-resistant survivors that are newly sensitized to traditional antibiotics, in a process called "phage steering." Members of the bacterial genus , which includes various human pathogens, are highly resistant to most antimicrobial agents, including serum immune components, antimicrobial peptides, and polymixin-class antibiotics. However, the application of phages in combination with certain antibiotics can produce synergistic effects that more effectively kill pathogenic bacteria. Herein, we demonstrate that serum resistance is due to intact lipopolysaccharide (LPS) and membranes, and phage-induced resistance altering LPS structure can enhance bacterial sensitivity not only to immune components in serum but also to membrane-associated antibiotics such as colistin. IMPORTANCE Bacteria frequently encounter selection pressure from both antibiotics and lytic phages, but little is known about the interactions between antibiotics and phages. This study provides new insights into the evolutionary trade-offs between phage resistance and antibiotic sensitivity. The creation of phage resistance through changes in membrane structure or lipopolysaccharide composition can simultaneously be a major cause of antibiotic sensitivity. Our results provide evidence of synergistic therapeutic efficacy in phage-antibiotic interactions and have implications for the future clinical use of phage steering in phage therapy applications.
Topics: Humans; Bacteriophages; Burkholderia cenocepacia; Anti-Bacterial Agents; Lipopolysaccharides; Virulence
PubMed: 37791751
DOI: 10.1128/jb.00196-23 -
Journal of Clinical Microbiology Nov 2021The Burkholderia cepacia complex (BCC) is known for causing serious lung infections in people with cystic fibrosis (CF). These infections can require lung...
The Burkholderia cepacia complex (BCC) is known for causing serious lung infections in people with cystic fibrosis (CF). These infections can require lung transplantation, eligibility for which may be guided by antimicrobial susceptibility testing (AST). While the Clinical and Laboratory Standards Institute recommends AST for BCC, the European Committee on Antimicrobial Susceptibility Testing (EUCAST) does not, due to poor method performance and correlation with clinical outcomes. Furthermore, limited data exist on the performance of automated AST methods for BCC. To address these issues, reproducibility and accuracy were evaluated for disk diffusion (DD), broth microdilution (BMD), and MicroScan WalkAway using 50 B. cenocepacia and 50 isolates collected from people with CF. The following drugs were evaluated in triplicate: chloramphenicol (CAM), ceftazidime (CAZ), meropenem (MEM), trimethoprim-sulfamethoxazole (TMP-SMX), minocycline (MIN), levofloxacin (LVX), ciprofloxacin (CIP), and piperacillin-tazobactam (PIP-TAZ). BMD reproducibility was ≥ 95% for MEM and MIN only, and MicroScan WalkAway reproducibility was similar to BMD. DD reproducibility was < 90% for all drugs tested when a 3 mm cut-off was applied. When comparing the accuracy of DD to BMD, only MEM met all acceptance criteria. TMP-SMX and LVX had high minor errors, CAZ had unacceptable very major errors (VME), and MIN, PIP-TAZ, and CIP had both unacceptable minor errors and VMEs. For MicroScan WalkAway, no drugs met acceptance criteria. Analyses also showed that errors were not attributed to one species. In general, our data agree with EUCAST recommendations.
Topics: Anti-Bacterial Agents; Burkholderia; Burkholderia Infections; Burkholderia cenocepacia; Burkholderia cepacia complex; Cystic Fibrosis; Humans; Microbial Sensitivity Tests; Reproducibility of Results
PubMed: 34524889
DOI: 10.1128/JCM.01447-21 -
Frontiers in Microbiology 2020During chronic respiratory infections of cystic fibrosis (CF) patients, bacteria adaptively evolve in response to the nutritional and immune environment as well as...
During chronic respiratory infections of cystic fibrosis (CF) patients, bacteria adaptively evolve in response to the nutritional and immune environment as well as influence other infecting microbes. The present study was designed to gain insights into the genetic mechanisms underlying adaptation and diversification by the two most prevalent pathogenic species of the complex (Bcc), and Herein, we study the evolution of both of these species during coinfection of a CF patient for 4.4 years using genome sequences of 9 and 11 This co-infection spanned at least 3 years following initial infection by and ultimately ended in the patient's death by cepacia syndrome. Both species acquired several mutations with accumulation rates of 2.08 () and 2.27 () SNPs/year. Many of the mutated genes are associated with oxidative stress response, transition metal metabolism, defense mechanisms against antibiotics, and other metabolic alterations consistent with the idea that positive selection might be driven by the action of the host immune system, antibiotic therapy and low oxygen and iron concentrations. Two orthologous genes shared by and were found to be under strong selection and accumulated mutations associated with lineage diversification. One gene encodes a nucleotide sugar dehydratase involved in lipopolysaccharide O-antigen (OAg) biosynthesis (). The other gene encodes a putative two-component regulatory sensor kinase protein required to sense and adapt to oxidative- and heavy metal- inducing stresses. This study contributes to understanding of shared and species-specific evolutionary patterns of . and . evolving in the same CF lung environment.
PubMed: 33101250
DOI: 10.3389/fmicb.2020.574626 -
Microbiology (Reading, England) Mar 2013Burkholderia cenocepacia is an opportunistic pathogen that primarily infects cystic fibrosis patients. Previously we have reported that mutations in shvR, a LysR-type...
Burkholderia cenocepacia is an opportunistic pathogen that primarily infects cystic fibrosis patients. Previously we have reported that mutations in shvR, a LysR-type transcriptional regulator, and ShvR-regulated genes BCAS0208 and BCAS0201 (designated afcE and afcF, respectively) affect colony morphotype, biofilm and pellicle formation and virulence in B. cenocepacia. In this study we investigated the role of afcE and afcF in influencing lipid-metabolism-associated phenotypes. As previously reported for K56-2ΔshvR, the Δ2afcE and afcF : : lux mutants had no antifungal activity against Fusarium and Rhizoctonia solani, suggesting that these genes are involved in synthesis of a membrane-associated antifungal lipopeptide. Strains Δ2afcE and afcF : : lux had reduced swarming motility and altered cell membrane morphology, both of which were restored to wild-type levels upon providing these genes in trans. Both K56-2ΔshvR and Δ2afcE showed increased uptake of the hydrophobic fluorescent probe N-phenylnaphthylamine (NPN), indicating altered outer membrane properties. Total lipid profiles determined by TLC revealed distinct differences in cellular lipid compositions of K56-2ΔshvR, Δ2afcE and afcF : : lux compared with K56-2. Taken together, these results indicate that afcE and afcF are involved in metabolic pathway(s) influencing lipid profiles and affect both cell surface and antifungal properties of B. cenocepacia.
Topics: Antifungal Agents; Burkholderia cenocepacia; Cell Membrane; Chromatography, Thin Layer; Fusarium; Gene Deletion; Gene Expression Regulation, Bacterial; Genetic Complementation Test; Humans; Lipid Metabolism; Lipids; Lipopeptides; Locomotion; Phenotype; Rhizoctonia; Transcription Factors
PubMed: 23306671
DOI: 10.1099/mic.0.064683-0 -
Frontiers in Microbiology 2018H111 is an opportunistic pathogen associated with chronic lung infections in cystic fibrosis patients. Biofilm formation, motility and virulence of are regulated by...
H111 is an opportunistic pathogen associated with chronic lung infections in cystic fibrosis patients. Biofilm formation, motility and virulence of are regulated by the second messenger cyclic di-guanosine monophosphate (c-di-GMP). In the present study, we analyzed the role of all 25 putative c-di-GMP metabolizing proteins of H111 with respect to motility, colony morphology, pellicle formation, biofilm formation, and virulence. We found that RpfR is a key regulator of c-di-GMP signaling in , affecting a broad spectrum of phenotypes under various environmental conditions. In addition, we identified Bcal2449 as a regulator of virulence in larvae. While Bcal2449 consists of protein domains that may catalyze both c-di-GMP synthesis and degradation, only the latter was essential for larvae killing, suggesting that a decreased c-di-GMP level mediated by the Bcal2449 protein is required for virulence of . Finally, our work suggests that some individual proteins play a role in regulating exclusively motility (CdpA), biofilm formation (Bcam1160) or both (Bcam2836).
PubMed: 30687272
DOI: 10.3389/fmicb.2018.03286 -
Microbiology (Reading, England) Sep 2009Burkholderia cenocepacia is a member of the B. cepacia complex (Bcc), a group of opportunistic bacteria that infect the airways of patients with cystic fibrosis (CF) and... (Review)
Review
Burkholderia cenocepacia is a member of the B. cepacia complex (Bcc), a group of opportunistic bacteria that infect the airways of patients with cystic fibrosis (CF) and are extraordinarily resistant to almost all clinically useful antibiotics. Infections in CF patients with Bcc bacteria generally lead to a more rapid decline in lung function, and in some cases to the 'cepacia syndrome', a virtually deadly exacerbation of the lung infection with systemic manifestations. These characteristics of Bcc bacteria contribute to higher morbidity and mortality in infected CF patients. In the last 10 years considerable progress has been made in understanding the interactions between Bcc bacteria and mammalian host cells. Bcc isolates can survive either intracellularly within eukaryotic cells or extracellularly in host tissues. They survive within phagocytes and respiratory epithelial cells, and they have the ability to breach the respiratory epithelium layer. Survival and persistence of Bcc bacteria within host cells and tissues are believed to play a key role in pulmonary infection and to contribute to the persistent inflammation observed in patients with CF. This review summarizes recent findings concerning the interaction between Bcc bacteria and epithelial and phagocytic cells.
Topics: Animals; Burkholderia Infections; Burkholderia cepacia complex; Cystic Fibrosis; Epithelial Cells; Host-Pathogen Interactions; Humans; Phagocytes; Respiratory System; Virulence
PubMed: 19542002
DOI: 10.1099/mic.0.031344-0 -
Frontiers in Cellular and Infection... 2022Cystic fibrosis (CF) human and mouse macrophages are defective in their ability to clear bacteria such as . The autophagy process in CF (F508del) macrophages is halted,...
Cystic fibrosis (CF) human and mouse macrophages are defective in their ability to clear bacteria such as . The autophagy process in CF (F508del) macrophages is halted, and the underlying mechanism remains unclear. Furthermore, the role of CFTR in maintaining the acidification of endosomal and lysosomal compartments in CF cells has been a subject of debate. Using 3D reconstruction of z-stack confocal images, we show that CFTR is recruited to LC3-labeled autophagosomes harboring Using several complementary approaches, we report that CF macrophages display defective lysosomal acidification and degradative function for cargos destined to autophagosomes, whereas non-autophagosomal cargos are effectively degraded within acidic compartments. Notably, treatment of CF macrophages with CFTR modulators (tezacaftor/ivacaftor) improved the autophagy flux, lysosomal acidification and function, and bacterial clearance. In addition, CFTR modulators improved CFTR function as demonstrated by patch-clamp. In conclusion, CFTR regulates the acidification of a specific subset of lysosomes that specifically fuse with autophagosomes. Therefore, our study describes a new biological location and function for CFTR in autophago-lysosomes and clarifies the long-standing discrepancies in the field.
Topics: Animals; Burkholderia cenocepacia; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Hydrogen-Ion Concentration; Lysosomes; Macrophages; Mice
PubMed: 35252032
DOI: 10.3389/fcimb.2022.819554