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Bacteria as communities sociomicrobiology

Biofilms Community of microbes embedded in an organic polymer matrix that adheres to a surface" . 3D structure containing 1 or more microbial speciesForms at interfacesSolid/liquid, liquid/air, air/solidSpacial heterogeneityExopolysaccharide glue"Water channelsImportant in human infectionsD

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Bacteria as communities sociomicrobiology

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    1. Bacteria as communities “sociomicrobiology” Biofilms Quorum sensing

    2. Biofilms “Community of microbes embedded in an organic polymer matrix that adheres to a surface” 3D structure containing 1 or more microbial species Forms at interfaces Solid/liquid, liquid/air, air/solid Spacial heterogeneity Exopolysaccharide “glue” Water channels Important in human infections Decreased antibiotic susceptibility Less conspicuous to immune system Targets organisms to favorable niches Protection from physical injury Promote horizontal gene transfer Enhance cell-cell communication

    3. Components

    4. Biofilms Abundant in aquatic ecosystems Targets specialized microbes to specific locales Nutrionally favorable, abundant nutrients Symbiosis Rhizobium/legumes Ruminant digestive tracts: degrade/recycle insol materials Non-hostile environment, Protection from predators Insurance hypothesis Presence of diverse subpopulations increases survival of community as a whole

    5. Bacterial species associated with biofilms P. aeruginosa S. epidermidis S. aureus E. coli Lactobacillus spp. Streptococcal spp. …and many more…..including candida albicans

    6. Biofilms are important in human infections Dental biofilms >500 species; mixed aerobes/anaerobes Caries, gigivitis, periodontitis Pneumonia in CF patients Endocarditis, osteomyelitis Artificial surfaces Catheter-related infections Contact lens infections Prosthetic joints Difficult to eradicate Antibiotic resistant Resist phagocytosis

    8. Technological advances Early steps mimicked on plastic surfaces Flow cells Confocal scanning laser microscopy Allows visualization of hydrated state 3D reconstruction Spatial & temporal heterogeneity GFP labeled bacteria reporter constructs

    9. Multiple steps and gene products required for biofilm formation

    10. Pseudomonas aeruginosa biofilm formation

    12. Initial clonal growth followed by migration

    13. PilA mutants fail to climb the mushroom stalk pilA WT

    15. Biofilms are antiobiotic resistant

    16. Why are they abx resistant? Physical or chemical diffusion barriers Bind to Glycocalyx Slow growth 2° to nutrient deprivation Altered microenvironment pH, reduced O2 Activation of stress response E. coli RpoS mutants cannot make biofilms Biofilm-specific state Induction of efflux pumps Phase variation

    17. Molecular elucidation of a mechanism of abx resistance Screened tranposon library for mutants with increased antibiotic sensitivity when grown as biofilms but not when grown planktonically Secondary screen for nl biofilm formation

    18. Biofilms mutant that is antiobiotic sensitive

    20. NvdB encodes a glucosyl transferase required for cyclic glucan synthesis Mutation in NvdB gene: glucosyltransferase required for synthesis of cyclic b (1,3) glucans (normally found in periplasm) Mutant is defective in periplasmic glycans Binds to tobramycin Gene is expressed during biofilm growth Likely binds other antibiotics?

    21. Self generated diversity produces insurance effects in biofilm communities Short term growth in biofilms generates extensive genetic diversity RecA-dependent

    22. Increases ability of biofilm to withstand external stress WT biofilm-grown bacteria survive exposure to oxidative stress better than RecA mutant

    23. Quorum sensing Intermicrobe communication Signaling (language) system that allows bacteria to sense cell density Inter- vs intra-species Early examples Vibrio harveyi-only produces light at high cell densities Can induce light by adding supernatants from stationary phase cultures Strep pneumonie-competence induced at late-log phase Produces “competence” factor at high cell densities Important in many processes Symbiosis Virulence Antibiotic production Biofilm formation Microbe-host communication Novel drug targets

    24. Why quorum sense?

    25. Quorum sensing in Vibrio harveyi Bacteria live in light organ of fish Need late log-phase bacteria in vitro to get light production Can recapitulate with late log-phase supernatants

    26. Gram negative QS Found in > 70 GNs Players LuxI homolog: acyl homoserine lactone (AHSL) synthetase AHSL: membrane permeable signal Lux R homolog: DNA binding transcription factor that is activated upon binding AHSL

    27. LuxI & LuxR LuxI LuxR Intraspecies communication C-terminus allows Multimerization In absence of HSL, N-terminus binds to C-terminus and inhibits DNA binding Lux box: 20 bp palindrome at -40 Specificity comes from exquisite specificity of HSL binding; LuxR homologs recognize similar DNA sequences (Lux Box)

    28. Gram positive QS

    30. Hybrid QS AI1 is a HSL but signals through 2 CS. Intraspecies AI2: furanosyl Borate diester. LuxS is widespread: interspecies communication?

    32. Combinatorial Diversity Parallel circuit B. subtilis competence Series circuit P. aeruginosa virulence factor production

    33. V. Harveyi QS private vs public conversations multilingual communication

    34. B. subtilis: either/or Signals have opposing effects on response regulator One causes phosphorylation Other causes dephosphorylation Allows choice of 2 mutually exclusive states Competence vs Sporulation

    36. Biofilm maturation QS

    37. Interspecies cell-cell communication Many bacteria encode LuxS and produce AI-2 V. harvyei can respond to cell-free sups from many gram pos and gram neg bacteria Eavesdropping P. aeruginosa responds to AI-2 even though it lacks LuxS High levels of AI-2 in sputum from CF patients Induces QS-dependent virulence factors Eukaryotes may also respond to bacterial QS signals IL-8 The literature is complicated because the presence of host-encoded QS-degrading enzymes has only recently been appreciated

    38. QS in S. aureus

    39. Inhibitory QS: AIP from one strain inhibits agr response of other strains

    40. Quorum quenching Bacterial antagonists S. aureus Proteases Bacillus AiiA destroys AHSLs required for plant pathogen virulence factor production Allows Bacillis to compete with Erwinia Transgenic potato & tobacco plants expressing AiiA are resistant to Erwinia infections Therapeutics? Seaweed Halogenated furanones Inhibit Pseudomonas biofilm formation & Serratia swarming Human airway epithelial cells membrane-associated activity that degrades Pseudomonas C12 HSL but not C4 HSL Synthetic halogenated furanones abrogate pneumonia in mouse model of acute Pseudomonas pneumonia

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