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Classificationof Bacteria

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Classificationof Bacteria

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Classificationof Bacteria

  1. 1. Bacterial classification <ul><li>Wall structure </li></ul><ul><ul><li>Gram + </li></ul></ul><ul><ul><ul><li>Staphylococcus , Streptococcus , Clostridium, Bacillus </li></ul></ul></ul><ul><ul><li>Gram - </li></ul></ul><ul><ul><ul><li>Enteric, respiratory and others </li></ul></ul></ul><ul><ul><li>Acid-fast </li></ul></ul><ul><ul><ul><li>Mycobacterium </li></ul></ul></ul><ul><ul><li>Wall-less </li></ul></ul><ul><ul><ul><li>Mycoplasma </li></ul></ul></ul><ul><li>Unusual </li></ul><ul><ul><li>Obligate intracellular </li></ul></ul><ul><ul><ul><li>Rickettsia , Chlamydia, Treponema </li></ul></ul></ul>G+ G- AF WL IC Bacteria
  2. 2. Bacterial classification <ul><li>Cell morphology </li></ul><ul><ul><li>Shapes </li></ul></ul><ul><ul><ul><li>Rod </li></ul></ul></ul><ul><ul><ul><li>Cocci </li></ul></ul></ul><ul><ul><ul><li>Spiral </li></ul></ul></ul><ul><ul><li>Associations </li></ul></ul><ul><ul><ul><li>Individual </li></ul></ul></ul><ul><ul><ul><li>Diplo- </li></ul></ul></ul><ul><ul><ul><li>Staphylo- </li></ul></ul></ul><ul><ul><ul><li>Strepto- </li></ul></ul></ul><ul><ul><ul><li>Filaments </li></ul></ul></ul>G+ G- AF WL IC Bacteria Rod Cocci Rod Cocci Spiral
  3. 3. Gram stains
  4. 4. Gram Stains of Mixtures
  5. 5. Cell shapes
  6. 6. Bacterial classification <ul><li>Growth characteristics </li></ul><ul><ul><li>Oxygen requirement </li></ul></ul><ul><ul><ul><li>Aerobic </li></ul></ul></ul><ul><ul><ul><li>Anaerobic </li></ul></ul></ul><ul><ul><ul><ul><li>Microaerophilic, aerotolerant </li></ul></ul></ul></ul><ul><ul><ul><li>Facultative </li></ul></ul></ul><ul><ul><li>Spore formation </li></ul></ul><ul><ul><li>Intracelular/extracellular </li></ul></ul><ul><ul><li>Fastidious/non-fastidious </li></ul></ul>
  7. 7. Endospore
  8. 8. Classification & Diagnosis <ul><li>Type of colonies </li></ul><ul><ul><li>Appearance </li></ul></ul><ul><ul><ul><li>Color, shape, size and smoothness </li></ul></ul></ul><ul><ul><li>On differential media </li></ul></ul><ul><ul><ul><li>Blood, MacConkey, EMB, Manitol salt agar </li></ul></ul></ul><ul><ul><li>On selective media </li></ul></ul><ul><ul><ul><li>MacConkey, EMB, Manitol salt agar </li></ul></ul></ul>
  9. 9. Colony morphology/color
  10. 10. Bacterial colony morphology
  11. 11. Capsule
  12. 12. MacConkey Agar Plate
  13. 13. Blood Agar Plates and Hemolysis
  14. 14. Mannitol salt agar
  15. 15. Classification & Diagnosis <ul><li>Metabolism </li></ul><ul><ul><li>Utilization of specific substrates </li></ul></ul><ul><ul><ul><li>Lactose ( Sal / Shi / Yer /) - </li></ul></ul></ul><ul><ul><ul><li>Citrate ( E.coli - / Klebsiela + ) </li></ul></ul></ul><ul><ul><li>Production of certain end products </li></ul></ul><ul><ul><ul><li>Fermentation end products </li></ul></ul></ul><ul><ul><ul><ul><li>Acid (acetate, propionic acid, butyric acid etc .) </li></ul></ul></ul></ul><ul><ul><ul><ul><li>Aceton </li></ul></ul></ul></ul><ul><ul><ul><ul><li>Alcohol </li></ul></ul></ul></ul><ul><ul><ul><ul><li>Amine </li></ul></ul></ul></ul><ul><ul><ul><ul><li>H 2 S </li></ul></ul></ul></ul>
  16. 16. Triple Sugar Iron Agar Slants <ul><li>TSI </li></ul><ul><li>Fermentation of glucose, lactose, and/or sucrose </li></ul><ul><li>Reduction of sulfur to hydrogen sulfide </li></ul><ul><li>Gas formation </li></ul><ul><li>Used for Enterobacteriaceae </li></ul>
  17. 17. Classification & Diagnosis <ul><li>Specialized tests </li></ul><ul><ul><li>Immunological </li></ul></ul><ul><ul><ul><li>O-, H- & K-Ag (serotype) </li></ul></ul></ul><ul><ul><ul><li>Precipitation, agglutination </li></ul></ul></ul><ul><ul><li>Specialized enzymes </li></ul></ul><ul><ul><ul><li>Catalase--- Staph + . vs. Strep - . </li></ul></ul></ul><ul><ul><ul><li>Coagulase--- S. aureus + vs. S. epidermidis - </li></ul></ul></ul><ul><ul><ul><li>Oxidase--- Neisseria gonorrhoea + </li></ul></ul></ul><ul><ul><ul><li>Urease--- Proteus + , Helicobacter + </li></ul></ul></ul><ul><ul><li>Antibiogram pattern </li></ul></ul><ul><ul><li>Phage typing </li></ul></ul><ul><ul><li>Fatty acid profile </li></ul></ul>
  18. 18. Urease Test Urea CO 2 + NH 3 NH 4 + + OH - H 2 O
  19. 19. Indole Test
  20. 20. API test strip
  21. 21. Bacteriophage Plaques
  22. 22. Conventional method <ul><li>Depend on ability to culture </li></ul><ul><ul><li>Treponema pallidum </li></ul></ul><ul><li>Slow, esp. for fastidious species </li></ul><ul><ul><li>Mycobacterium spp . </li></ul></ul><ul><li>Not always definitive </li></ul>
  23. 23. Immunological detection ELISA
  24. 24. Instant View tests
  25. 25. Molecular diagnosis <ul><li>Ribotyping </li></ul><ul><li>Restriction fragment length polymorphism (RFLP) </li></ul><ul><li>DNA hybridization </li></ul><ul><li>PCR, RT-PCR and RAPD </li></ul><ul><li>Nucleic acid sequence analysis </li></ul><ul><li>Phage-GFP (TB) </li></ul>
  26. 26. RFLP GGATCC CCTAGG
  27. 27. DNA hybridization
  28. 28. In situ Hybridization
  29. 29. Rate of increase 2 n PCR
  30. 30. Real time PCR Q R 5' 3' 5' 3' 5' 3' 3' 5' Q R 5' 3' 3' 5' Q R Q 5' 3' 3' 5' R Primer Primer Probe Taq DNA Pol
  31. 31. RAPD of P. aeruginosa
  32. 32. DNA and protein chips A B C D
  33. 33. Microarray array mediated pathogen detection
  34. 34. Ab chip mediated pathogen detection  -O157:H7  -Salmonella  -O157:H7  -Salmonella E.coli O157:H7 Salmonella typhimurium
  35. 35. Molecular diagnosis <ul><li>Reduce reliance on culture </li></ul><ul><li>Faster </li></ul><ul><li>High sensitive </li></ul><ul><li>More definitive </li></ul><ul><li>More discriminating </li></ul><ul><li>Techniques adaptable to all pathogens </li></ul><ul><li>Technically demanding </li></ul><ul><li>Relatively expensive </li></ul><ul><li>Can be too sensitive </li></ul><ul><li>Provides no information if results are negative </li></ul>
  36. 36. Bioterrorism <ul><li>Pathogen detection </li></ul><ul><ul><li>Fast and accurate </li></ul></ul><ul><ul><li>Mobile </li></ul></ul><ul><ul><li>Inexpensive </li></ul></ul><ul><li>Source investigation </li></ul>
  37. 37. Biosensor Mediated Pathogen Detection Step 1: Attach Capture Antibody to Fiber Optic Waveguide Step 2: Apply Sample. Target Organism Binds to Waveguide Step 3: Wash Away Other Cells and Debris Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y
  38. 38. Step 1: Attach Capture Antibody to Fiber Optic Waveguide Step 2: Apply Sample. Target Organism Binds to Waveguide Step 3: Wash Away Other Cells and Debris Step 4: Add Specific Detection Antibody Step 5: Turn on Laser. Detection Antibody Fluoresces, Indicating a Positive Sample. Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Biosensor Mediated Pathogen Detection Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y
  39. 39. Biosensors Analyte 2000 TM for laboratory research and development RAPTOR TM automated version for field analysis
  40. 40. Differentiating Staphylococci from Streptococci <ul><li>Gram stain and morphology </li></ul><ul><ul><li>Both Gram + </li></ul></ul><ul><ul><li>Staphylococci : bunched cocci </li></ul></ul><ul><ul><li>Streptococci : chained cocci </li></ul></ul><ul><ul><ul><li>S. pneumoniae : diplococcus </li></ul></ul></ul><ul><li>Enzyme tests </li></ul><ul><ul><li>Staphylococci : catalase + </li></ul></ul><ul><ul><li>Streptococci : catalase - </li></ul></ul><ul><li>Growth </li></ul><ul><ul><li>Staph .: large colonies (non-fastidious), some hemolytic </li></ul></ul><ul><ul><li>Strep .: small colonies (fastidious), many hemolytic (  or  ) </li></ul></ul>
  41. 41. Staphylococci <ul><li>S. aureus : coagulase + </li></ul><ul><li>S. epidermidis : coagulase - </li></ul>
  42. 42. Streptococci
  43. 43. Differentiating the Gram - bacteria <ul><li>Cocci </li></ul><ul><ul><li>Neisseria </li></ul></ul><ul><li>Rods </li></ul><ul><ul><li>Type of disease they cause </li></ul></ul><ul><ul><li>Enteric Gram - rods </li></ul></ul><ul><ul><ul><li>API test </li></ul></ul></ul><ul><li>Curved </li></ul><ul><ul><li>Vibrio , Campylobacter , Helicobacter </li></ul></ul><ul><li>Spiral Gram - organisms </li></ul><ul><ul><li>Spirochetes </li></ul></ul>
  44. 44. Gram negative Straight rods Curved rods Lactose+ Lactose- Citrate+ Citrate- H 2 S+ H 2 S- Klebsiella E.coli Salmonella Shigella Campy blood agar 42 o C+ 25 o C- Campylobacter TCBS agar Yellow Oxidase+ Vibrio
  45. 45. Bacteria Gram+ Cocci Rod Staph. Strep. Non-spore Spore S. a. S. e. S. s. A B Pn Vir Fil Rod A.i. C.d. L. m. +O 2 -O 2 B.a. B.c. C.b. C.t. C.p. C.d. Acid Fast M.t. M.l. N.c. Intra Cellular Rickettsia Coxiella Erlichia Chlamydia Wall Less Mycoplasma Gram- Cocci Rod Spiral Straight Curve +O 2 -O 2 +/-O 2 Other Treponema Borrelia Leptospira Neisseria Moraxella P.a. Enteric Bact. Resp. Zoo GU Bordetella. H. influenzae Legionella Yersinia Pasteurella Brucella Francislla Streptobacillus H. ducreyi Gardnerella Calymmatobacterium Vibrio Campylobacter Helicobacter

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