Project description:The human-adapted pathogen group A Streptococcus (GAS) utilizes wounds as portals of entry into host tissue, wherein surface adhesins interact with the extracellular matrix, enabling bacterial colonization. The streptococcal collagen-like protein 1 (Scl1) is a major adhesin of GAS that selectively binds to two fibronectin type III (FnIII) repeats within cellular fibronectin, specifically the alternatively spliced extra domains A and B, and the FnIII repeats within tenascin-C. Binding to FnIII repeats was mediated through conserved structural determinants present within the Scl1 globular domain and facilitated GAS adherence and biofilm formation. Isoforms of cellular fibronectin that contain extra domains A and B, as well as tenascin-C, are present for several days in the wound extracellular matrix. Scl1-FnIII binding is therefore an example of GAS adaptation to the host's wound environment. Similarly, cellular fibronectin isoforms and tenascin-C are present in the tumor microenvironment. Consistent with this, FnIII repeats mediate GAS attachment to and enhancement of biofilm formation on matrices deposited by cancer-associated fibroblasts and osteosarcoma cells. These data collectively support the premise for utilization of the Scl1-FnIII interaction as a novel method of anti-neoplastic targeting in the tumor microenvironment.
Project description:Bacteria can form single- and multispecies biofilms exhibiting diverse features based upon the microbial composition of their community and microenvironment. The study of bacterial biofilm development has received great interest in the past 20 years and is motivated by the elegant complexity characteristic of these multicellular communities and their role in infectious diseases. Biofilms can thrive on virtually any surface and can be beneficial or detrimental based upon the community's interplay and the surface. Advances in the understanding of structural and functional variations and the roles that biofilms play in disease and host-pathogen interactions have been addressed through comprehensive literature searches. In this review article, a synopsis of the methodological landscape of biofilm analysis is provided, including an evaluation of the current trends in methodological research. We deem this worthwhile because a keyword-oriented bibliographical search reveals that less than 5% of the biofilm literature is devoted to methodology. In this report, we (i) summarize current methodologies for biofilm characterization, monitoring, and quantification; (ii) discuss advances in the discovery of effective imaging and sensing tools and modalities; (iii) provide an overview of tailored animal models that assess features of biofilm infections; and (iv) make recommendations defining the most appropriate methodological tools for clinical settings.
Project description:The clinical management of bacterial biofilm infections represents an enormous challenge in today's healthcare setting. The NIH estimates that 65% of bacterial infections are biofilm-related, and therapeutic outcomes are positively correlated with early intervention. Currently, there is no reliable imaging technique to detect biofilm infections in vivo, and current clinical protocols for accurate and direct biofilm identification are nonexistent. In orthopedic implant-associated biofilm infections, for example, current detection methods are based on nonspecific X-ray or radiolabeled white blood cell imaging, coupled with peri-prosthetic tissue or fluid samples taken invasively, and must be cultured. This approach is time-consuming and often fails to detect biofilm bacteria due to sampling errors and a lack of sensitivity. The ability to quantify bacterial biofilms by real-time noninvasive imaging is an urgent unmet clinical need that would revolutionize the management and treatment of these devastating types of infections. In the present study, we assembled a collection of fluorescently labeled peptide candidates to specifically explore their biofilm targeting properties. We evaluated these fluorescently labeled peptides using various in vitro assays for their ability to specifically and nondestructively target biofilms produced by model bacterial pathogen Pseudomonas aeruginosa. The lead candidate that emerged, 4Iphf-HN17, demonstrated rapid biofilm labeling kinetics, a lack of bactericidal activity, and biofilm targeting specificity in human cell infection models. In vivo fluorescently labeled 4Iphf-HN17 showed enhanced accumulation in biofilm-infected wounds, thus warranting further study.
Project description:Cationic antimicrobial peptides are multifunctional molecules that have a high potential as therapeutic agents. We have identified a histone H1-derived peptide from the Komodo dragon (Varanus komodoensis), called VK25. Using this peptide as inspiration, we designed a synthetic peptide called DRGN-1. We evaluated the antimicrobial and anti-biofilm activity of both peptides against Pseudomonas aeruginosa and Staphylococcus aureus. DRGN-1, more than VK25, exhibited potent antimicrobial and anti-biofilm activity, and permeabilized bacterial membranes. Wound healing was significantly enhanced by DRGN-1 in both uninfected and mixed biofilm (Pseudomonas aeruginosa and Staphylococcus aureus)-infected murine wounds. In a scratch wound closure assay used to elucidate the wound healing mechanism, the peptide promoted the migration of HEKa keratinocyte cells, which was inhibited by mitomycin C (proliferation inhibitor) and AG1478 (epidermal growth factor receptor inhibitor). DRGN-1 also activated the EGFR-STAT1/3 pathway. Thus, DRGN-1 is a candidate for use as a topical wound treatment. Wound infections are a major concern; made increasingly complicated by the emerging, rapid spread of bacterial resistance. The novel synthetic peptide DRGN-1 (inspired by a peptide identified from Komodo dragon) exhibits pathogen-directed and host-directed activities in promoting the clearance and healing of polymicrobial (Pseudomonas aeruginosa & Staphylococcus aureus) biofilm infected wounds. The effectiveness of this peptide cannot be attributed solely to its ability to act upon the bacteria and disrupt the biofilm, but also reflects the peptide's ability to promsote keratinocyte migration. When applied in a murine model, infected wounds treated with DRGN-1 healed significantly faster than did untreated wounds, or wounds treated with other peptides. The host-directed mechanism of action was determined to be via the EGFR-STAT1/3 pathway. The pathogen-directed mechanism of action was determined to be via anti-biofilm activity and antibacterial activity through membrane permeabilization. This novel peptide may have potential as a future therapeutic for treating infected wounds.
Project description:Pseudomonas aeruginosa is an opportunistic multidrug-resistant pathogen, able to grow in biofilms. It causes life-threatening complications in diseases characterized by the up-regulation of type I interferon (IFN) signaling, such as cancer or viral infections. Since type I IFNs regulate multiple functions of neutrophils, which constitute the first line of anti-bacterial host defense, in this work we aimed to study how interferon-activated neutrophils influence the course of P. aeruginosa infection of the lung. In lungs of infected IFN-sufficient WT mice, significantly elevated bacteria load was observed, accompanied by the prominent lung tissue damage. At the same time IFN-deficient animals seem to be partly resistant to the infection. Lung neutrophils from such IFN-deficient animals release significantly lower amounts of neutrophil extracellular traps (NETs) and reactive oxygen species (ROS), as compared to WT neutrophils. Of note, such IFN-deficient neutrophils show significantly decreased capacity to stimulate biofilm formation by P. aeruginosa. Reduced biofilm production impairs in turn the survival of bacteria in a lung tissue. In line with that, treatment of neutrophils with recombinant IFN-? enhances their NETosis and stimulates biofilm formation by Pseudomonas after co-incubation with such neutrophils. Possibly, bacteria utilizes neutrophil-derived NETs as a scaffold for released biofilms. In agreement with this, in vivo treatment with ROS-scavengers, NETs disruption or usage of the bacterial strains unable to bind DNA, suppress neutrophil-mediated biofilm formation in the lungs. Together, our findings indicate that the excessive activation of neutrophils by type I IFNs leads to their boosted NETosis that in turn triggers biofilm formation by P. aeruginosa and supports its persistence in the infected lung. Targeting these mechanisms could offer a new therapeutic approach to prevent persistent bacterial infections in patients with diseases associated with the up-regulation of type I IFNs.
Project description:Dinoflagellates in the genus Symbiodinium exhibit a variety of life styles, ranging from mutualistic endosymbioses with animal and protist hosts to free-living life styles. In culture, Symbiodinium spp. and naturally associated bacteria are known to form calcifying biofilms that produce so-called symbiolites, i.e., aragonitic microbialites that incorporate Symbiodinium as endolithic cells. In this study, we investigated (i) how algal growth and the combined physiological activity of these bacterial-algal associations affect the physicochemical macroenvironment in culture and the microenvironment within bacterial-algal biofilms, and (ii) how these interactions induce the formation of symbiolites. In batch culture, calcification typically commenced when Symbiodinium spp. growth approached stationary phase and when photosynthetic activity and its influence on pH and the carbonate system of the culture medium had already subsided, indicating that symbiolite formation is not simply a function of photosynthetic activity in the bulk medium. Physical disturbance of bacteria-algal biofilms, via repeated detaching and dispersing of the developing biofilm, generally impeded symbiolite formation, suggesting that the structural integrity of biofilms plays an important role in generating conditions conducive to calcification. Microsensor measurements of pH and O2 revealed a biofilm microenvironment characterized by high photosynthetic rates and by dynamic changes in photosynthesis and respiration with light intensity and culture age. Ca2+ microsensor measurements confirmed the significance of the biofilm microenvironment in inducing calcification, as photosynthesis within the biofilm induced calcification without the influence of batch culture medium and under environmentally relevant flow conditions. Furthermore, first quantitative data on calcification from 26 calcifying cultures enabled a first broad comparison of Symbiodinium-induced bacterial-algal calcification with other calcification processes. Our findings support the idea that symbiolite formation is a typical, photosynthesis-induced, bacterial-algal calcification process that is likely to occur under natural conditions.
Project description:We have used microarray analysis to study the transcriptome of the bacterial pathogen Bordetella bronchiseptica over the course of five time points representing distinct stages of biofilm development. The results suggest that B. bronchiseptica undergoes a coordinately regulated gene expression program similar to a bacterial developmental process. Expression and subsequent production of the genes encoding flagella, a classical Bvg(-) phase phenotype, occurs and is under tight regulatory control during B. bronchiseptica biofilm development. Using mutational analysis, we demonstrate that flagella production at the appropriate stage of biofilm development, i.e. production early subsequently followed by repression, is required for robust biofilm formation and maturation. We also demonstrate that flagella are necessary and enhance the initial cell-surface interactions, thereby providing mechanistic information on the initial stages of biofilm development for B. bronchiseptica. Biofilm formation by B. bronchiseptica involves the production of both Bvg-activated and Bvg-repressed factors followed by the repression of factors that inhibit formation of mature biofilms.
Project description:Biofilm formation is a key virulence factor for a wide range of microorganisms that cause chronic infections. The multifactorial nature of biofilm development and drug tolerance imposes great challenges for the use of conventional antimicrobials and indicates the need for multi-targeted or combinatorial therapies. In this Review, we focus on current therapeutic strategies and those under development that target vital structural and functional traits of microbial biofilms and drug tolerance mechanisms, including the extracellular matrix and dormant cells. We emphasize strategies that are supported by in vivo or ex vivo studies, highlight emerging biofilm-targeting technologies and provide a rationale for multi-targeted therapies aimed at disrupting the complex biofilm microenvironment.
Project description:The development of surface-attached biofilm bacterial communities is considered an important source of nosocomial infections. Recently, bacterial interference via signaling molecules and surface active compounds was shown to antagonize biofilm formation, suggesting that nonantibiotic molecules produced during competitive interactions between bacteria could be used for biofilm reduction. Hence, a better understanding of commensal/pathogen interactions within bacterial community could lead to an improved control of exogenous pathogens. To reveal adhesion or growth-related bacterial interference, we investigated interactions between uropathogenic and commensal Escherichia coli in mixed in vitro biofilms. We demonstrate here that the uropathogenic strain CFT073 and all E. coli expressing group II capsules release into their environment a soluble polysaccharide that induces physicochemical surface alterations, which prevent biofilm formation by a wide range of Gram-positive and Gram-negative bacteria. We show that the treatment of abiotic surfaces with group II capsular polysaccharides drastically reduces both initial adhesion and biofilm development by important nosocomial pathogens. These findings identify capsular polymers as antiadhesion bacterial interference molecules, which may prove to be of significance in the design of new strategies to limit biofilm formation on medical in dwelling devices.