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Microbiology

Microbiology. Brock 13 th edition: chapters 1, 16. Why study Microbiology ?. Evolution of life Microbial evolution Examples for universal importance of bacteria in biology, environment and health. a. Evolution of life. Mammals. Humans. Vascular plants. Origin of Earth (4.6 bya).

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Microbiology

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  1. Microbiology Brock 13th edition: chapters 1, 16 Why study Microbiology ? Evolution of life Microbial evolution Examples for universal importance of bacteria in biology, environment and health

  2. a. Evolution of life Mammals Humans Vascularplants Origin of Earth(4.6 bya) Shellyinvertebrates Figure 1.6a Present  20% O2 1bya Origin ofcellular life 4bya O2 Anoxygenicphototrophicbacteria Algaldiversity Mic only rob ms for ial life AnoxicEarth 3bya 2bya Earthis slowlyoxygenated Moderneukaryotes Origin ofcyanobacteria

  3. Evolution of life EarlyBacteria EarlyArchaea Figure 16.4 Mound:precipitates of clay,metal sulfides, silica,and carbonates Ocean water(20°C, containingmetals, CO2 andPO42) Flow of substancesup through mound Nitrogenbases Aminoacids Sugars Ocean crust Nutrients in hothydrothermal vent water

  4. Evolution of life Figure 16.4 6. Dispersal (other habitats) 5. diversification, interaction (0.3 to 0.5 billion years) Time 4. LUCA(last universal common ancestor) 3. proteolipid membrane 2. “RNA world” 1. Prebiotic chemistry

  5. LUCA’senergymetabolism e--acceptor e--donor S + H2  H2S ΔG0’ = -20,6 kJ Figure 16.7 PrimitiveATPase Out Primitivehydrogenase pyrite H+ gradient Cytoplasmicmembrane In S0 reductase

  6. LUCA’s C-metabolism S + H2  H2S ΔG0’ = -20,6 kJ 4-4.3 x109 years BC CO2 fixation  organic compounds (i.e. acetate) accumulate Chemoorganotrophic bacteria “metabolic diversification”

  7. Metabolicdiversification S + H2  H2S ΔG0’ = -20,6 kJ 4-4.3 x109 years BC CO2 fixation  organic compounds accumulate Archaea Bacteria methanogenesis 3.7 x109 years BC H2, CO2 acetate 4 H2 + CO2 CH4 + 2H2O H3CCOO-+ H2O CH4 + HCO3-

  8. Bacteria: evolvephototrophy S + H2  H2S ΔG0’ = -20,6 kJ 4-4.3 x109 years BC CO2 fixation  organic compounds accumulate Archaea Bacteria methanogenesis 3.7 x109 years BC H2, CO2 acetate 4 H2 + CO2 CH4 + 2H2O H3CCOO-+ H2O CH4 + HCO3- CO2 fixation Anaerobic phototrophy (H2S S) 3.3 x109 years BC CO2 fixation Oxygen generating phototrophy(H2O O2) 2.7 x109 years BC

  9. Eon BYA Organisms,events O2level Metabolichighlights O2 toxicity New metabolic pathways: - sulfate reduction - nitrification - chemolithotrophy - O2 respiration (grow fast) 0 Extinction of the dinosaurs Cambrian Phanaerozoic 0.5 Early animals Precambrian Multicellulareukaryotes 20% 1.0 Figure 16.6 and 8 10% Proterozoic 1.5 First eukaryoteswith organelles First: Fe-oxidation 1% 2.0 Endosymbiosis? Ozone shield Great oxidation event 0.1% Aerobic respiration 2.5 Oxygenic photosynthesis Cyanobacteria (2H2O  O2 4H) 3.0 Sulfate reductionFe3 reduction Archaean Precambrian Fe3+ sediments Anoxygenic photosynthesis Purple and green bacteria 3.5 Anoxic Acetogenesis Bacteria/Archaeadivergence Methanogenesis 4.0 First cellular life; LUCA Hadean Formation ofcrust and oceans Sterile Earth 4.5 Formation of Earth

  10. Endosymbionttheoryof Eukaryoteevolution b) Hydrogen hypothesis a) From nucleated Archaeon Figure 16.9 Bacteria Eukarya Bacteria Eukarya Archaea Archaea Plants Animals Plants Animals nucleus formed Archaeon with nucleus cyanobacterium cyanobacterium Engulfment of aH2-producing cellof Bacteria by aH2-consuming cellof Archaea Ancestor ofmitochondrion (Bacteria)

  11. three take home messages Bacteriaaretheancient form of life. All otherorganismsevolvedfromthis. LUCA existedprobably 4.3 Bio yearsago. All forms of lifehad extreme effects on theirenvironment... and mediateddramaticchange Intenseinteractions. All organismshaveinteractedwitheachother (directlyorindirectly). E.g. Eukaryoteshavealwaysbeeninteractingintenselywithbacteriathroughouttheirevolution. The evolution and function of onecannotbeunderstood in theabsence of theother.

  12. b.Bacteriaand Archaeaevolution Very high rate of evolution!! Haploid genomes Rapid growth Large populations

  13. Bacteria/Archaeaevolution Wild-type cellPigment mutantsLightDark 15 Rhodobactercapsulatus Mutantslost inlight Figure 16.10 10 Bacteriochlorophyll a/ml of culture Mutantselectedin dark Cell populations Pigment mutants 5 Wildtype 1 3 4 2 5 15 5 20 10 Subculture number

  14. Phylogeneticanalysis of Bacteria, Archaea Small ribosomal subunit RNA sequence: “long distance” relationships Ribosomal RNA genes • 16S (bacteria), 18S (Eukaryotes) • Ubiquitous and essential • Ancient • Easy RNA isolation • Conserved and variable regions • Sufficiently long 16S rRNA Variable…….conserved Base position in 16S RNA gene

  15. Phylogeneticanalysis of Bacteria, Archaea conserved protein-coding genes: “long distance” and strain differentiation EF-Tu (protein biosynth.) Hsp60 aatRNAsynthetases …

  16. Phylogeneticanalysis via 16S rDNA Isolate DNA 16 S gene Amplify 16Sgene by PCR Nonidentities Figure 16.12, 13 Run on agarosegel; check forcorrect size Beforealignment Species 1 Kilo-bases 4 2 3 5 9 1 3.0– 2.0– Species 2 1.5– 1.0– Gaps 0.5– Species 1 Sequence Afteralignment 15 Species 2 A C G G T Align sequences;generate tree Distinctspecies Ancestralcell Distinctspecies

  17. Display phylogeneticrelationship Cladistics = grouping by common features (absent in more distant relatives) Parsimony = assumes least number of steps Rooted trees Figure 16.14 node Unrooted tree Defines unique paths of evolution Employs “outgroup” Relative relationships

  18. Universal phylogenetictree 3 “domains” PROKARYOTES EUKARYOTES Figure 16.16 Archaea Eukarya Bacteria 2 major phyla > 80 phyla > 10 Mio species?? 8 Mio species ? Animals (7.7 Mio species) Slimemolds Entamoebae Green nonsulfurbacteria Euryarchaeota Fungi (0.6 Mio species) Methanosarcina Mitochondrion Methano-bacterium Extremehalophiles Plants (0.3 Mio spec.) Crenarchaeota Gram-positivebacteria Thermoproteus Proteobacteria Ciliates Methano-coccus Pyrodictium Thermoplasma Chloroplast Thermococcus Cyanobacteria Flagellates Flavobacteria MarineCrenarchaeota Pyrolobus Methanopyrus Trichomonads Thermotoga Thermodesulfobacterium Microsporidia Diplomonads(Giardia) Aquifex Extensive genetic exchange?? LUCA

  19. c. Current topics of interest Methods: for analyzing microorganisms Philosophy: the bacterial species problem health: effects of the microbiota health: what is a pathogen? How to kill bacteria? environment: metabolic effects on C, N, P, S… cycles Industry/safety: genetic engineering

  20. AnalyzingBacteria and Archea Physiology: << 5 % of all bacteriahavebeencultured phenotype (motility, morphology, metabolism…) FISH(Fluorecencein situ hybridization): DNA-oligo binding rRNA Bacillus Yeast Universal probe eukaryal probe DNA sequencing (fast evolvingfield!!): - 16S “community sequencing” - “metagenome sequencing”  predict genes/metabolism  predict physiology

  21. Costs of DNA sequencing

  22. DNA sequencingformicrobialcommunityanalysis Phylogenetic snapshotof most members of the community 1. Identification of novelphylotypes 2. Microbialcommunity Extract totalcommunity DNA DNA Figure 22.16 Communitysampling approach Environmentalgenomics approach Restriction digest total DNA andthen shotgun sequence, ORsequence directly (withoutcloning) using a “next generation”DNA sequencer Amplify single gene,for example, geneencoding 16S rRNA Sequence andgenerate tree Assembly andannotation Outcomes Partialgenomes Single-gene phylogenetic tree Total gene pool of the community 1. Identification of all gene categories 2. Discovery of new genes 3. Linking of genes to phylotypes

  23. Animal evolution Example 1: The gut microbiotaRole in health and diseases

  24. Example 1: The gut microbiotaRole in health and diseases Animal evolution =>> A long history of co-evolution

  25. MAMP signaling/innate immunity Microbiota enzyme/function Example 1: The gut microbiotaRole in health and diseases Health Energy: acetate, butyrate Innate defense: priming Vitamins: K, B12, C, niacin, panthotenic acid, biotin, folic acid GALT: maturation Mucosa: maturation Colonization resistance Disease Inflammatory bowel disease: stimulus Th17 immune responses: stimulus Host metabolism: stimulus cancer: stimulus ?

  26. Example 1: The gut microbiotaRole in health and diseases Dethlefsen, 2008, Nature 448, pp. 811ff

  27. Example 2: The “bacterial species problem” Plants/animals: cross fertile offspring Bacteria, Archaea: ??

  28. Example 2: The “bacterial species problem”

  29. The “bacterial species problem” proteobacteriales Phylogenetic tree 181 genomes Phylogenomic tree ≥5 genes exchangedby«horizontal genetransfer» Dagan et al., 2008, PNAS 105, pp. 10039 ff.

  30. The “bacterial species problem” Streptococcus Fraser et al., 2009, Science 323, pp. 741ff

  31. The “bacterial species problem” Bacteria, Archaea: - no sexual cycle - “long distance” gene exchange phylogenetic species concept  niche occupation: “ecotype” Multigene Tree 16S rRNA Gene Tree Photobacterium damselae FS-2.1 FS-4.2 Photobacteriumphosphoreum 50 changes FS-3.1 Photobacterium leiognathi FS-5.1 FS-2.2 Photobacterium mandapamensis ATCC 11040T FS-5.2 Photobacterium angustum ATCC 51761 Photobacterium phosphoreum NCIMB 13476 Photobacteriumiliopiscarium NCIMB 13478 NCIMB 13481 Photobacterium iliopiscarium ATCC 51760T Photobacterium kishitanii chubb.1.1 ckamo.3.1 canat.1.2 hstri.1.1 Photobacteriumkishitanii calba.1.1 BAA-1194T apros.2.1 ckamo.1.1 vlong.3.1

  32. The “bacterial species problem”  ecotypes One microbial habitat Ecotype II Ecotype I Ecotype III Cell containingan adaptivemutation Figure 16.25 Periodic selection Adaptive mutantsurvives. OriginalEcotype III wild-typecells out competed Populationof mutantEcotype III Repeat processmany times New speciesof Ecotype III

  33. Classification: traditional approach Taxonomic systems: Bergey’s Manual of Systematic Bacteriology The Prokaryotes International Committee on Systematics of Prokaryotes

  34. Example 3: global Carbon-cycle , a greenhouse gas CH4 CO2 Figure 24.1 Humanactivities Respiration Animals andmicroorganisms Landplants Aquaticplants andphyto-plankton CO2 Aquaticanimals Biological pump CO2 Fossilfuels Humus Death andmineralization CH4 CH4 Soil formation Earth’s crust Rock formation Atmosphere 0.003

  35. Example 3: global Carbon-cycle (CH2O)n Organic matter Figure 24.2 Oxygenic photosynthesis Respiration Chemolithotrophy Methanotrophy Oxic CO2 Anoxic Methanogenesis Acetogenesis Anaerobicrespirationandfermentation Syntrophassisted Anoxygenicphotosynthesis Organic matter (CH2O)n

  36. Example 3: anaerobic methane oxidation Marine sediments Methanotrophic Archaea(ANME-types) Sulfate-reducing Bacteria Figure 14.28 Organiccompounds

  37. c. Current topics of interest Methods: for analyzing microorganisms Philosophy: the bacterial species problem health: effects of the microbiota health: what is a pathogen? How to kill bacteria? environment: metabolic effects on C, N, P, S… cycles Industry/safety: genetic engineering

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