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Growth and Multiplication of Bacteria

Growth and Multiplication of Bacteria. Hugh B. Fackrell Sept 1997 Filename: Growth.ppt. Requirements for Growth/Multiplication. ALL required nutrients correct pH temperature salinity, moisture redox potential atmosphere. Growth Liquid vs Solid Media. Liquid: clear >>turbid

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Growth and Multiplication of Bacteria

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  1. Growth and Multiplication of Bacteria Hugh B. Fackrell Sept 1997 Filename: Growth.ppt

  2. Requirements for Growth/Multiplication • ALL required nutrients • correct • pH • temperature • salinity, • moisture • redox potential • atmosphere

  3. GrowthLiquid vs Solid Media • Liquid: clear >>turbid • Solid: individual colonies • each colony derived from a single cell

  4. Growth Event • Absorption of water & nutrients • Catabolism of carbon source • inorganic or organic • Biosynthesis of new cellular components • major energy consumption • Cell enlargement • Cell division ( Binary Fission)

  5. Binary Fission • DNA replication • Plasma membrane invaginate • Cell wall deposited in invaginated space • Cross wall completed • Cells separate

  6. Binary Fission • Light micrograph

  7. Binary Fission

  8. Consequences of Binary Fission • Very large number of cells very fast • Mathematical progressions • arithmetic (1>2>4>6>8>10>12>14>16) • geometric(1>2>4>8>16) • exponential expression (20 > 21 > 22 >23>24) • logarithmic expression(0 >log21>log22>log23>log24

  9. Logarithmic Plots • Can plot very large Range of numbers • Phases of growth demonstrated • Generation time easily calculated

  10. Cell Multiplication • 1 20 0 l • 2 21 log21 ll • 4 22 log22 llll • 8 23 log23 lllllllll • 16 24 log24 lllllllllllllllll

  11. Mathematics of bacterial growth Cells Generation # Log 2 Log10 • 0 1 0 0.000 • 1 2 1 0.301 • 2 4 2 0.602 • 3 8 3 0.903 • 4 16 4 1.204 • 5 32 5 1.505 • 6 64 6 1.806 • 7 128 7 2.107 • 8 256 8 2.408

  12. Growth Data • #Generation #cells Log10 • 1 1 0 • 5 32 1.51 • 10 1,024 3.01 • 15 32,768 4.50 • 20 1,048,576 6.02

  13. Number of cells Log number of cells Time (hours) Growth curves for exponentially increasing population

  14. Stationary phase Death phase Log phase Lag phase Bacterial Growth Curve 1 5 10 Time (hours)

  15. Measurement of Growth Constants • G: Generation Time • K: Mean Growth Rate Constant G := 1/K

  16. G: Generation time Time in minutes or hours for a population of bacteria to double in number

  17. Double # cells Log phase Generation time Calculation of Generation Time Log Number of Bacteria 1 5 10 Time (hours)

  18. Fast Log Number of bacteria Medium Doubling number Time (hours) Slope of Log phase proportional to generation time Slow

  19. K: Mean Growth Rate Constant • K= n/t • K= (log10Nt - log10Nt0)/ 0.301t • N= number of cells • n=: number of generations • t = time (hr or min) • K = 1/slope ( semi log growth plot) • Therefore G = 1/K

  20. Sample calculation for K & G • Population increase from 103 to 109 in 10hrs • K= (log 109 - log 103) / 0.301 x 10 • K= 9-3/3.01 = 2 generations/hours • G = 1/K = 1/2 = 0.5 hr/generation

  21. Factors influencing lag phase • Age of culture inoculum • old culture -> long lag • young culture-> short lag • Size of inoculum • few cells -> long lag • many cells -> short lag • Environment • pH, temp, gases,salinity • sub optimum -> long lag • optimum-> short lag

  22. Thermophile Mesophile Extreme Thermophile Rate of Growth Psychrotroph Pyschrophile Growth Responses: Temperature -10 0 10 20 30 40 50 60 70 80 90 100 Temperature (o C)

  23. Neutrophile Alkalophile Rate of Growth Acidophile Growth Responses: pH 1 2 3 4 5 6 7 8 9 10 11 12 pH

  24. Diauxic Growth • Growth on two carbon sources • Mixed sugars • Each sugar used separately • Glucose ALWAYS used first • Second sugar ONLY used when glucose GONE

  25. Growth [Sugar] Arabinose Glucose Time (hr) Diauxic Growth: 2 carbon sources

  26. Synchronous Growth • Filtration • Smaller cells • all same size • Temperature shock • Hot/cold brings cells to same metabolic state • Starvation • deplete medium of selected nutrient

  27. Number of Cells Synchronousgrowth Asynchronous growth Time (min) Synchronous vs Asynchronous growth

  28. Growth in Limited Nutrients • Limiting concentration of Required nutrient • YIELD • number of cells • Linear increase yield with nutrient conc Yield = Mass of organisms formed Mass of nutrients used

  29. Growth Rate Total Growth [Nutrient] Growth in Limited Nutrients

  30. Applications of Limiting [Nutrient] • Chemostat (continuous culture) • Bio-Assay

  31. Bio-Assay: Procedure • Bacterium: CANNOT synthesize nutrient • Medium: all growth requirements except nutrient to be assayed • Add • equal amounts of medium to each tube • equal numbers of bacteria to each tube • increasing amounts of the nutrient to be assayed • [Unknown] • Incubate • Measure growth (turbidity or viable count)

  32. 0 Growth of known [nutrient] Microbial Growth Growth of unknown 0 0 0 0 [Nutrient] in unknown 0 [Nutrient] mg/ml Bio-Assay • Vitamin B-12 measurement in Green beans • Lactobacillus leichmanni

  33. Chemostat • Description of Instrument • Principle • Steady State • Sample Results • Application

  34. Chemostat: Description of Instrument

  35. Steady State K = D Flow rate D = Vessel volume Chemostat: Principle • Essential nutrient is limited • Growth rate(K) controlled by supply rate of nutrient • Yield controlled by concentration of nutrient • Dilution rate (D): speed of nutrient flow into the culture vessel

  36. Cell density or biomass Measurement Value Generation time Nutrient conc Dilution Rate of Nutrient Chemostat: Sample Results

  37. Chemostat: Applications • Growing large amounts of cells • Industrial production • vaccines • pharmaceuticals • hormones • Long term studies of specific growth phase • Selecting for specific mutants • Aquatic systems

  38. Bacterial Growth in Natural Environments • Natural Environments • Animal Tissues • Soil • Water- freshwater- marine • Plants

  39. Bacterial Growth in Natural Environments • Active • Short bursts of growth & metabolism • usually low rates of growth • Quiescent • Viable cannot culture • Stressed • starvation semi viable

  40. Biofilms: Body • Catheter • Foley: • latex silicone • Intravenous: • polyurethane S. epidermidis • Prostheses • Hip joints • Dental implants • voicebox • Tampons • IUD

  41. Biofilms: Water • Dental lines • Spacecraft • Drinking filters • ALL surfacces Biofilm in gut of a mollusc

  42. Biofilms: Disease • Cystic fibrosis • lung-alveolar surface • Ulcers • Helicobacter jejuni • Dental caries • Streptococccus spp

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