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Micro 1611

Micro 1611. Exam 1. FYI. myco means fungi, waxy. Name shapes & sizes of bacteria. Coccus Round Bacillus Rod Coccobacillus Short rod Vibrio Curved Spirillum Wavy Spirochete Helical Pleomorphic Lacking distinct shape. Smallest of all living organisms 0.5-2.0 micrometer

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Micro 1611

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  1. Micro 1611 Exam 1

  2. FYI • myco • means fungi, waxy

  3. Name shapes & sizes of bacteria • Coccus • Round • Bacillus • Rod • Coccobacillus • Short rod • Vibrio • Curved • Spirillum • Wavy • Spirochete • Helical • Pleomorphic • Lacking distinct shape • Smallest of all living organisms • 0.5-2.0 micrometer • Large surface-to-volume ratio • Essential ? • Due to lack of cell organelles • Compartmentalization • Reliance of diffusion

  4. Bacterial Arrangements • Dividing in 1 plane: • Pair? • Diplo • Chain? • Strepto • Dividing in 2+ planes: • Grapes? • Staphylo • Packets of 4? • Tetrads • Packets of 8? • sarcina

  5. What is NOT found in bacterial membranes? • Sterols • Why? • Bacteria have external wall to provide rigidity & support • Exceptions? • Mycoplasma – do NOT have a cell wall • Mycoplasma pneumonia • Walking pneumonia • Adheres to epithelium by slipping in between cilia due to lack of rigid cell wall • Resistant to penicillin, cephalosporin, vancomycin– those that interfere w/cell wall synthesis Gram (-) stain poorly since lack cell wall (per Murray text) Neither Grm (+) or (-) ------ per Clinical Micro made simple text

  6. Major functions of Cell Membrane • Selective permeability & transport of solutes • Electron transport & oxy phosphorylation • Secretion of proteins • ie, Hydrolytic enzymes • Housing enzymes & carrier molecules that function in synthesis of DNA, cell wall polymers & membrane lipids • Housing receptors & other proteins of chemotactic & sensory transduction systems

  7. Bacterial Cell Wall • semi-rigid, outside cell membrane • functions: • maintenance of cell shape • prevention of osmotic lysis • environmental protection • anchor for external structures

  8. Bacterial Cell Wall • What’s the major part of bacterial cell walls? • Peptidoglycan (ranging from 1-2 sheets to 40+) • backbone of repeating…… • NAG’s • N-acetylglucosamine • NAM’s • N-acetylmuramic acid • crosslinked with…….. • Tetrapeptides

  9. Clinical Applications • Ways to interfere or prevent synthesis of peptidoglycan……… • Cell Wall Inhibitors (4) • penicillin • vancomycin • bacitracin • cephalosporin • Lysozyme • human secretion • degrades glycan backbone of peptidoglycan

  10. Lab Stuff • Wet Mounts are used to….. • view living organisms • especially useful in: • identifying clue cells (BV) – (Gardnerellavaginalis on a pap smear) • Trichomonas(parasite) • yeast & other fungi FYI Clue cells are vaginal squamous epithelial cells coated with the anaerobic gram-variable coccobacilliGardnerellavaginalis and other anaerobic bacteria causing bacterial vaginosis. Clue cells were first described by Gardner and Dukes in 1955 and were so named as these cells give an important "clue" to the diagnosis of bacterial vaginosis. A clue cell can be detected on simple wet mount of vaginal secretions.

  11. Lab Stuff • Gram Stain • two most common clinical reasons you ‘d do one • presumptive gonorrhea diagnosis • guy w/urethral discharge • gram-negdiplococci would be seen • pneumonic patient • sputum sample to identify etiologic agent • samples come from….. • urethral discharge, sputum, pus, female exudate, urine, joint fluid, CSF • NOT • blood, throat swabs, stool • stool – only if fecal leukocytes (WBC’s) indicative of Campylobacteriosis or invasive amebiasis • results directly relate to pathogenicity, thus choice of antibiotic

  12. Gram Stain • Differential Stain based on……. • Bacterial Cell Envelope • Gram (+) stain • BLUE/Purple (I’m positively BLUE over you!) • primary stain becomes trapped • Gram (–) stain • Red (No RED commies) • thin layer bound to fatty component which means it can be washed off with alcohol-acetone solution

  13. Gram Stain Quiz Time…………. • What happens when you stain Candida (yeast)? • appears large coccal-shaped, Gram (+), • much larger than Staph or Strep • What group of bacteria doesn’t stain well? • Mycobacteria • why? • waxy mycolic acid component of the cell wall • best stain with? • Acid Fast technique • ie Mycobacterium smegmatis

  14. 2 categories for Bacterial Cell Walls

  15. The Classics Mycobacteriaweakly GM(+)/Acid-Fast SpirochetesGM(-) but too small Mycoplasmalack cell wall, so neither Exceptions:

  16. What makes Gram (+) Special All 3 are Virulence Factors – enhance ability to wreck havoc • Wall teichoic acids • covalent links to peptidoglycan layers, (-) charge • contain Glycerophosphate or Ribitol Phosphate • Lipoteichoic acids • covalently linked, (-) charge • contain Glycerophosphate or Ribitol Phosphate • anchored to cell membrane Both function as Adhesins & can initiate Endotoxin-like activities w/peptidoglycan causing host to release inflammatory cytokines • Mycolic acid layer • long-chain fa’s covalently linked to peptidoglycans via arabinogalactan • waxy coat enables it to……… • resist desiccation (drying out), some antibiotics, phagocytosis

  17. Bonus ?’s • Name some Gram (+) that contain mycolic acid layer: • Mycobacterium tuberculosis • M. leprae • Nocardiaasteroides • Do you use a Gram Stain on these bugs? • No due to lipids, so use Acid Fast – detects mycolic acids • Why are Acid-Fast organisms such a hassle? • resistant to antibiotics • grow slowly • thus need Combination therapy for long time Think: TB, Leprosy & Asteroids

  18. What makes Gram (–) Special • 2 phospholipidbilayers • Outer membrane • Periplasmic space

  19. What makes Gram (–) Special • 2 phospholipidbilayers • Outer membrane • Consists of……. • LPS • Porins • Lipoprotein • Periplasmic space

  20. What makes Gram (–) Special • 2 phospholipidbilayers • Outer membrane • Consists of……. • LPS • consists of 3 main parts….. • O antigen • Core polysaccharide • Lipid A • Porins • Lipoprotein • Periplasmic space

  21. What makes Gram (–) Special • 2 phospholipidbilayers • Outer membrane • Consists of……. • LPS • consists of 3 main parts….. • O antigen (Immunogenic portion of LPS) • most external – use it as an adherence • long, linear repeating units of carbs (Adhesions) • highly variable, useful in ID • E coli 0157H7 (Jack in the Box strain) • Core polysaccharide • just links the 2 • Lipid A (Endotoxic area) • embedded in outer layer of outer membrane • heat-stable, B cell mitogen, induces prod of Cytokines & other inflammatory mediators • toxin activity when released can result in? • GRAM (-) BACTERIAL SEPSIS --- SPEPTIC SHOCK • Porins • Lipoprotein • Periplasmic space

  22. What makes Gram (–) Special • 2 phospholipidbilayers • Outer membrane • LPS can….. • directly activate complement • alternative pathway • bind to serum LBP (LPS binding protein), then to membrane-bound LPS receptor (CD14) on Macrophage • cytokine production & phagocytosis • bind to secreted CD14 (sCD14),then to membrane-bound receptors on endothelial & epithelial cells • Porins • Lipoprotein • Periplasmic space What is the End Result? Sepsis

  23. What makes Gram (–) Special • 2 phospholipidbilayers • Outer membrane • LPS • Porins --- what are they? • Trimeric proteins form lg non-specific channels for passage of H2O & small hydrophilic (charged) molecules • Lipoprotein • abundant protein in Grm(-) • functions? • stabilize outer membrane • anchor it to peptidoglycan layer • Periplasmic space

  24. What makes Gram (–) Special • 2 phospholipidbilayers • Outer membrane • Periplasmic space • gel filled space containing? • hydrolytic enzymes • ie: collagenases, B-lactamases, hyaluronidases • may degrade antibiotics, VIRULENCE contributor • some GRM(-) bacteria concentrate them leading to antibiotic resistance • important to run susceptibility test • located between? • cellular membrane & outer membrane

  25. Session 4 Lecture Notes

  26. External Bacterial Structures • Flagellum • Pili • Glycocalyx

  27. External Bacterial Structures • Flagellum • 3 components • Filament • self-assembling helical arrangmt of flagellin protein subunits w/hollow core • source of H antigen (typing bacterial isolates) • E coli 0157H7 • Hook • attaches filament to cell surface, links to Basal body • Basal Body • anchors flagellum in cell wall/membrane • slightly diff in Gm+ and Gm- ‘s • Pili • Glycocalyx

  28. External Bacterial Structures • Flagellum • location & #’s vary • flagellum at 1 end • Monotrichous • 2+ flagellum at 1 end • Lophotrichous • 1+ flagellum at both ends • Amphitrichous • flagella around entire perimeter • Peritrichous • Pili • Glycocalyx What if its non-motile? Atrichous

  29. External Bacterial Structures • Flagellum • function? • motility • directed motility is known as….. • Chemotaxis • which doesn’t always involve flagella • Pili • Glycocalyx

  30. External Bacterial Structures • Flagellum • Pili --- what are they? • proteinaceous • hair-like structures w/adhesion tips • promote adherence to other bacteria or host cells • 2 types: • common pili = fimbriae • tips contain lectins (bind to specific sugars) • arranged? • Peritrichously • function? • adhesion & twitching motility • sex pili = conjugation pili • Glycocalyx Strains of E. coli have several types of fimbriae that aid in colonization of mucosa -- virulence

  31. External Bacterial Structures • Flagellum • Pili --- what are they? • proteinaceous • hair-like structures w/adhesion tips • promote adherence to other bacteria or host cells • 2 types: • common pili = fimbriae • sex pili = conjugation pili • aka F pili, conjugation pili • function? • adhering 2 cells together • allows for exchange of genetic material (donor to recipient) • BACTERIAL CONJUGATION • Glycocalyx Genetic exchange enables transfer of resistance.

  32. External Bacterial Structures • Flagellum • Pili • Glycocalyx • K (or Vi) antigen • polysaccharide material • often contains glycoproteins • surrounds bacterial cells – 2 types • capsule • rigid, uniform and closely surrounds cell • slime layer • loose, non-uniform, more diffusely surrounds cell Not necessary for survival, but can contribute to increased virulence

  33. External Bacterial Structures Streptococcus mutans its polysaccharide capsule adheres to tooth enamel • Flagellum • Pili • Glycocalyx • functions? • protects from desiccation (drying out) • barrier to toxic hydrophobic molecules • inhibits phagocytosis • too slippery, hard for receptor-ligand interaction • decreases neutrophil killing of engulfed bacteria • decreases complement-mediated cell lysis • hard for antibodies to stick • induces abscess formation • promotes adherence to cells/surfaces • ie, teeth Streptococcus pneumoniae only pathogenic w/capsule Quellung Reaction test anticapsular antibodies bind to capsule & make it swell Virulent – has capsule Avirulent – no capsule

  34. Internal Bacterial Structures • Nucleoid region • Ribosomes • Inclusions • Spores

  35. Internal Bacterial Structures • Nucleoid region • contains? • DNA, RNA, proteins • lacks? • No histones, no introns in DNA, NO Nuclear Membrane • Ribosomes • Inclusions • Spores

  36. Internal Bacterial Structures • Nucleoid region • single, circular? • Chromosome • extrachromosomal DNA called? • Plasmids -- non-essential material • can be acquired thru various genetic exchange mechs • encode for drug resistance &/or toxin production • Virulence Factors • Ribosomes • Inclusions • Spores

  37. Internal Bacterial Structures • Nucleoid region • Ribosomes • consist of? • rRna and protein • Different from us in subunits? Which means? • 70S = 30S and 50S (1 smaller subunit & 1 larger) • We are 80S (2 subunits the same size) • Transcription & translation are coupled due to? • lack of compartmentalization so…… • protein synthesis much faster in bacteria than Eukaryotes • Inclusions • Spores Protein Synthesis Inhibitors can target 70S ribosomes and NOT affect 80S in us Selective Toxicity

  38. Internal Bacterial Structures • Nucleoid region • Ribosomes • Inclusions • collective term referring to? • Granules, vesicles, vacuoles • are? • bounded, thin/nonunit membrane • either via protein or lipid monolayer • function? • storage depots for excess carbon, energy, phosphate • Examples (3): • Volutin or metachromatic granules: polymers of inorganic phosphate • Corynebacteriumdiphtheriae– used to ID it • Glycogen granules: polymers of glycogen • PHB granules: chains of B-hydroxybutyric acid • Spores

  39. Internal Bacterial Structures • Nucleoid region • Ribosomes • Inclusions • Spores • endospores, produced by? • Bacillus • Clostridium • some non-pathogens • What does not produce them? • NO Gram (-) produce true endospores

  40. Internal Bacterial Structures Not many pathogens produce spores so helpful in ID Bacillus cereus (anthrax) – central endospore Bacillus subtilis– subterminal spore Clostridium tetani(Tetanus) – terminal endospore • Nucleoid region • Ribosomes • Inclusions • Spores • function? • harsh conditions, convert from vegetative state to dormant state/spore • spore formation & germination NOT REPO function, but survival • contains? • one copy of chromosome • low concentrations of essential proteins & ribosomes • high concentration of calcium bound to dipicolinic acid • lends to increased resistance of spores to increased temps & UV light thru stabilization of protein structure • Keratin coat • impervious to many chemicals, contributes to increased environmental resistance • small acid-soluble spore proteins (SASPs) that protect from UV & desiccation

  41. Let’s Review

  42. Session 5 Lecture Notes

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