Lecture 6 011808

Lecture 6 011808 - MMG 461: MOLECULAR PATHOGENESIS Lecture...

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Unformatted text preview: MMG 461: MOLECULAR PATHOGENESIS Lecture 6: The infectious cycle II Bacterial Communities: biofilms and cell-cell communication CGArvidson January 18, 2008 BIOFILMS – why? • Strength in numbers • Resistance to antimicrobials • Competition with other microbes • Protection from physiochemical changes • Protection from host defenses • Metabolic cooperativity (communities) • Genetic exchange? Biofilm formation V. cholerae Davey and O’Toole 2000 MMBR 64:847 Pseudomonas aeruginosa biofilm WT mutant Stalks formed by clonal growth (non-motile mutant), caps formed by the (motile) WT, which migrated up the stalks of the mutant. Klausen, Aaes-Jorgensen, Molin, and Tolker-Nielsen. 2003. Mol. Microbiol. 50:61-68 Biofilm Structure •Water channels •Nutrient gradients •Differing gene expression profiles •Some selection for heritable changes Confocal microscopy GFP-V. cholerae biofilm Yildiz and Schoolnik 1999 PNAS 96:4028 What happens to “old” biofilms? q q Dispersal/release of bacteria Breakdown of matrix – Directed degradation q Killing of some organisms in biofilm – Phage mediated – Cannibalism? q Temporal gene expression – Motility functions q q Flagella - up Pili - down Parsek and Fuqua JBac 2004 186:4427-4440 Dispersal of bacteria from a biofilm Hall-Stoodley and Stoodley TIM 2005 13:7-10 Biofilms and disease Davey and O’Toole 2000 MMBR 64:847 •P. aeruginosa in CF lung •V. cholerae •Periodontal disease – usually mixed communities MULTI-SPECIES BIOFILMS q Cooperativity – often, the combined metabolic activity of the community is greater than the sum of that of the individuals that make up the community q Typically, the interaction is of mutual benefit to all in the community Nutritional mutualism Palmer et al., 2001 Infect Immun 69:5794 Strep. oralis and Actinomyces naeslundii together can live in conditions where neither can live alone q Bradshaw et al, 1994 Microbiol 140:3407 – Metabolic cooperation between species in the use of mucin as a source of carbon, nitrogen and energy. q Ecology of a Microbial Community Davey and O'toole. MMBR 2000 64:847 MULTI-SPECIES BIOFILMS q All members of the community usually increase the production of EPS → aids in adherence to the subtratum as well as between microbes in the community q Oral biofilms – one of the best characterized multi-species biofilms – composed of hundreds of different species of microbes – Includes many pathogens not considered oral pathogens, – Source of infection? Colonization of other organs? Formation of multi-species biofilms q Attachment of “primary” colonizers – Adhesin-receptor (or some component of biotic/abiotic surface = substratum) q Growth of 1° species into microcolonies – Progression into young/early biofilm q Secondary (late) colonizers attach to 1° colonizers – Coaggregation, inter-bacterial adhesion Formation of a multi-species biofilm Rickard et al, 2003 TIMS 11:94 ORAL BIOFILMS - Plaque q Many different pairings have been demonstrated between 1° and 2° colonizers – Pairings are highly specific q q q 1° can often coaggregate with each other 2° do not typically coaggregate with each other Bridge organism – can coaggregate with 1° and 2° colonizers – Essential to have these for the formation of a complex biofilm; w/o a bridge, 2° colonizers do not attach q Colonization sequence is very important ORAL BIOFILM Cultivable organisms ~30-80% Strep. (sanguis, oralis, mitis) ~30% Actinomyces (naeslundii) Fusobacterium nucleatum - bridge Rickard et al, 2003 TIMS 11:94 Control of biofilm formation q Increased production of EPS – Density dependent q quorum-sensing – Growth rate dependent q q Usually in stationary phase Formation influenced by environment – Osmolarity, pH, C-source, q Other conditions of the biofilm can affect events – Dental microbes under strong selective pressure to remain tightly adhered/aggregated CO-AGGREGATION (interbacterial adhesion) q Lectin-like interactions Protein adhesin binding to a CHO receptor q Streptococcal wall polysaccharides (RPS) – – – q Different receptor motifs can mediate specificity Ag distinct Recognized by protein adhesins on other bacteria Actinomyces fimbriae – Also adhesins that bind to some RPS UNICELLULAR PARADIGM q q q Bacteria are asocial The primary goal of an individual bacterium is to make two bacteria Each individual in a culture (balanced growth conditions) is identical to every other – Genetics depends on it q Observations in 3 groups of bacteria contradicted – Myxobacteria – Luminescent marine vibrios – Gram-positive cocci q density-dependent accumulation of an extracellular factor (a “hormone-like” product) activated competence – Tomasz, Nature (1965) Cell-cell communication q Strength in numbers – enables bacteria to co-ordinate their behaviour q language-signaling molecules – – – q q HSLs peptides Metabolic intermediates accumulation enables a single cell to sense bacterial cell density Usually, but not always, species specific Myxococcus xanthus fruiting body formation • Truly social behavior • soluble (A-signal) and cellcontact (C-signal) signals coordinate developmental gene expression to build fruiting bodies, filled with spores Xlnt review: D. Kaiser. (2004) Ann. Rev. Microbiol. 58:75 Kuner and Kaiser (1982) J. Bacteriol. 151:458 QUORUM SENSING q First described for the squid light organ symbiont, Vibrio fischeri – cultures produced light only when large numbers of bacteria were present – initiated not by the removal of an inhibitor but by the accumulation of an activator molecule or "autoinducer“ (Nealson, K.H., Platt, T. and Hastings, J.W. (1970) J. Bacteriol. 104, 313-322. ) – sense their cell density by monitoring the autoinducer concentration - bacterial “pheromone” deKievit and Iglewski, 2000 Infec Immun 68:4839 Ahmer, B. M. M. Mol Microbiol. 2004 52:933 HSL quorum sensing systems luminescence Vibrio fischeri Virulence genes Pseudomonas aeruginosa Protease, siderophores Burkholderia cepacia xtra cell. Proteases, swarming Serratia liquifaciens Dispersal from aggregates Rhodobacter sphaeroides Antibiotic production Erwinia carotovora Biofilms, xtra cell. proteases Aeromonas species Conjugation Agrobacterium tumefaciens motility Yersinia species nodulation Rhizobium species Quorum sensing network in P. aeruginosa deKievit and Iglewski, 2000 Infec Immun 68:4839 SdiA of Enterobacteriaceae q LuxR homolog – Responds to HSLs q q Orthologs found in Salmonella, E. coli, Klebsiella None have AI-1 synthase – Make no HSLs q Respond to HSLs made by other organisms – Mechanism to detect mixed microbial communities q Control motility, cell division, virulence Other signaling molecules q AI-2 – – q species non-specific Made by many gram-negative and gram-positives AI-3 – made by EPEC, EHEC – Structure unknown q CAI-1 – 3-(s)-hydroxy-4-tridecanone – V. cholerae-specific q Indole – E. coli, but not Salmonella Other signaling molecules Vibrio Salmonella AI-2: an interconverting family of extracellular signal molecules. Other signaling molecules q AI-2 – species non-specific – Made by many gram-negative and gram-positives q AI-3 – made by EPEC, EHEC – Structure unknown q CAI-1 – 3-(s)-hydroxy-4-tridecanone → – V. cholerae-specific q Indole – E. coli, but not Salmonella → Quorum sensing in V. harveyi AI-1 LuxP (4-hydroxy C4 HSL) HK HK LuxN LuxQ PO4 PO4 RR RR PO4 LuxLM AI-2 PO4 HK LuxU LuxS PO4 RR LuxO luxCDABE Adapted from Bassler, BL 1999 Curr Opin Micro Quorum-quenching Wang YJ, and JR Leadbetter. (2005) Appl Environ Microbiol. 71:1291 Rapid acyl-homoserine lactone quorum signal biodegradation in diverse soils. AHL-degradation enzymes have recently been identified in a range of living organisms, including bacteria and eukaryotes. Expression of these enzymes in AHL-dependent pathogens and transgenic plants efficiently quenches the microbial QS signaling and blocks pathogenic infections. Gram-positive signaling q Peptides q Thiolactones Gram-positive (peptide) QS systems virulence genes Staphylococcus aureus Transformation competence, sporulation Bacillus subtilis Transformation competence Streptococcus pneumoniae conjugal plasmid transfer Enterococcus faecalis bacteriocin production lactic acid bacteria Agr regulatory system of S. aureus AIP AgrC AgrA P AgrA P agrB agrD P3 hld agrC RNAIII AIP target genes agrA Other Microbial Communities and Disease q Microbiota shift diseases q Polymicrobial diseases KOCH’S POSTULATES 1. The organism must always be found in the diseased animals but not in healthy ones. 2. The organism must be isolated from diseased animals and grown in pure culture away from the animal. 3. The organism isolated in pure culture must initiate and reproduce the disease when reinoculated into susceptible animals. 4. The organism should be reisolated from experimentally infected animals. Really only works if a single (micro)organism is the causative agent of a specific disease. Microbiota shift diseases q Pathogen usually present, but in numbers too low to cause disease q Some event shifts the population such that the “bad bug” takes over Example: Antibiotic-associated diarrhea Antibiotics, especially broad-spectrum types, can kill off the normal flora to such an extent that the imbalance results in disease. POLYMICROBIAL DISEASE – Presence of one generates niche for another – Presence of one predisposes host to colonization by another – Two or more together cause disease BACTERIAL VAGINOSIS – Shift from lots of lactobacilli to predominantly anaerobes q Esp. Gardnerella, Mycoplasma – Associated with Increased risk of pre-term delivery q Increased susceptibility to STDs q – GC, Chlamydia, HIV, ...
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This note was uploaded on 03/19/2008 for the course MMG 461 taught by Professor 3 during the Spring '08 term at Western Michigan.

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