Microbial Growth Chapter 6

1
Microbial Growth Chapter 6
2
Microbial Growth

• Requirements for Growth
• Culture Media
• Growth Patterns
• Evaluation/Measurement of Growth
• Lecture 6

3
Physical Requirements for Growth

• Three realms of requirement
• Temperature
• pH
• Osmotic pressure
• For all of the three, different groups of
  microorganisms have optimal ranges.

4
Temperature (Fig. 6.1)
5
Temperature

• Psychrophiles cold-loving microbes
• Mesophiles moderate temperature loving microbes
• Thermophiles heat-loving microbes
• Most species will grow within a limited range
  or _____ Celsius

6
Temperature

• _________ growth temperature lowest temperature
  at which a species will grow
• _________ growth temperature temperature at
  which the species grows best
• _________ growth temperature highest temperature
  at which growth is possible

7
Temperature

• Psychrophiles two groups
• True psychrophiles are native to cold
  environments in the Earths deep oceans and polar
  regions
• Can grow at ____ degrees Celsius, but optimum is
  at 15 degrees C, cant grow above 25 deg. C
• Psychrotrophs can grow at zero C, but optimum is
  20-30 degrees C, and max is 40 degrees C
• _____________ organisms

8
Temperature

• Mesophiles Optimum growth ________C
• For many pathogens, optimum is body temp. 37C
  (clinical incubators are set at this temp.)
• Include most common food spoilage organisms (so
  refrigeration works!)
• Thermophiles optimum 50-60C, many cant grow
  below _________C
• Spores may survive canning, and may grow is cans
  kept at high temp., but not considered a health
  problem

9
Extreme Thermophiles or Hyperthermophiles

• Members of the Archaea
• Optimums of _____C or higher
• Associated with hot springs and volcanic activity
• The record Deep sea vent communities growth
  at _____C (water doesnt boil because of extreme
  pressures)

10
pH

• Most bacteria have a narrow range for optimum
  growth near neutral pH between pH _______________
• Since bacterial fermentation produces acids,
  natural food preservation is accomplished in
  sauerkraut, pickles and cheeses!
• Acidophiles record is a chemoautotrophic form
  that lives in the drainage water from coal mines
  at a pH of ________!
• Molds and yeasts tolerate a wider range and have
  _______ pH optimums

11
Culturing microorganisms and pH

• Species which produce acids as they grow, will
  create hostile conditions for themselves!
• __________ are added to growth media
• Peptones
• Amino acids
• Phosphate salts

12
Osmotic Pressure

• Microorganisms are 80-90 water
• If too much water leaves the cell, shrinkage
  called _____________occurs
• When membrane pulls away from cell wall, growth
  is inhibited
• As a result, high osmotic pressure preserves
  food
• Salted fish
• Honey
• Condensed milk

13
Plasmolysis Fig. 6.4
14
Osmotic Pressure

• Halophiles (salt lovers)
• Extreme halophiles or obligate halophiles need up
  to _______ salt
• Facultative halophiles _______
• Most need a medium nearly all water
• _________________ however can cause rupture of
  some types with weak cell walls

15
Chemical Requirements for Growth

• Carbon
• ______________ get carbon along with their source
  of energy-organic compounds
• Chemo- and ________autotrophs get their carbon
  from Carbon dioxide

16
Chemical Requirements for Growth

• Nitrogen, Phosphorus, Sulfur
• N 14 of dry weight-mainly _________and__________
  __
• S, P 4 of dry weight- proteins and nucleic
  acids, cell membranes and ATP
• Trace elements cofactors/coenzymes- usually
  enough in _________________
• Organic Growth Factors vitamins, amino acids,
  purines, pyrimindines
• Nitrogen
• Decomposed (digested) protein
• Ammonium ions
• Nitrate ions
• Ecologically important relationships..

17
Nitrogen

• Nitrogen fixation photosynthetic _______bacteria
  and others which live symbiotically within the
  root nodules
• of such plants as the ________ family convert
  nitrogen gas to usable forms
• Others groups return the nitrogen to the
  atmosphere. In this way, nitrogen is cycled
  through the _____________.

18
OXYGEN

• __________ Aerobes require oxygen, growth at or
  near surface of liquid medium, have enzymes to
  deal with the toxic forms of oxygen
• ____________ aerobes can continue growth without
  oxygen with fermentation or other electron
  acceptors
• ________ anerobes unable to use oxygen in
  energy manipulating processes and most are HARMED
  by it!

19
HOW OXYGEN causes HARM

• ________ oxygen high energy and extremely
  reactive form of oxygen(present in phagocytes)
• Superoxide free radicals very toxic to cellular
  components-neutralized by ___________
• ______________________ (SOD) converts the
  superoxide into hydrogen peroxide and _____
• Peroxide broken down by _________ (yields water
  and oxygen)and ____________ (yields water only)

20
Other types of response to oxygen

• _______________ anerobes dont use oxygen, but
  tolerate it and have enzyme systems for dealing
  with the toxic forms of oxygen
• ______________ require oxygen, but only in
  concentrations ______ than air. Grow in a narrow
  zone. Sensitive to superoxides and peroxides

21
Summary of Oxygen Relationships Table 6.1
22
Culture Media

• Vocabulary
• Culture medium nutrient material prepared in a
  laboratory
• ___________ microbes introduced inot a culture
  medium
• ________ treated so that no microorganisms are
  present
• _____ solidifying agent derived from marine
  algae, composed of a complex, ____________
  polysaccharide

23
Agar

• Also used a s a thickener in jellies and ice
  cream
• Melts at 100C and remains liquid until 40-45C
  usually held at ________C
• ________ agar in a tube at an angle
• _________ agar in a tube solidified when
  vertical
• ____________ agar in a Petri dish

24
Media

• Chemically defined media
• Exact ____________ composition is known
• Experimental Laboratory work and autotrophs
  (Table 6.2 and 6.3)
• Complex media
• Contains ___________ from yeasts, meat, plants,
  digests of proteins (See Table 6.4)

25
Anerobic Growth

• Reducing agents such as sodium ___________________
  _which combine with oxygen are part of media
• For plates
• Special jar with means to remove ______________
  from around the plates is created and an
  indicator so that removal of oxygen can be
  monitored

26
Special techniques

• Some bacteria are really difficult
• Mycobacterium leprae (leprosy) is cultured is in
  ___________________!!!
• Syphillis spirochete not grown
• Intracellular parasitic forms Rickettsias and
  chlamydias
• Carbon dioxide enrichment for forms that live in
  intestinal and respiratory tract
  _________________________
• Candle jars, chemical packets in bags for plates

27
Candle Jars Fig. 6.7
28
Selective and Differential Media

• For detection of specific microorganisms
• _____________ media
• Examples
• Bismuth sulfite agar-Salmonella typhi
• Sabourauds Dextrose agar-pH 5.6 fungi
• Brilliant Green agar inhibits G, see G-
  Salmonella
• ____________media make it easier to see a
  desired organism

29
Differential Media

• Example Blood agar-
• Streptococcus pyogenes shows a ___________________
  ___ around their colonies where enzymes they have
  secreted have destroyed the red cells

30
Combining Selective and Differential
Qualities Examples

• Mannitol Salt agar
• Contains _______ salt and a pH indicator, plus
  mannitol (a sugar)
• Staph. aureus can tolerate the salt and ferments
  mannitol to form an acid
• MacConkey agar
• Contains bile salts and crystal violet and
  lactose
• Crystal violet and bile salts inhibit __________
• Lactose fermenters show as red or pink colonies
• Non-fermenters show as _____________
• Used to differentiate Salmonella and related
  species

31
Obtaining Pure Cultures

• Visible colonies theoretically derive from a
  single spore or cell or clump or chain of the
  same species
• Colonies must be far enough apart to assure they
  are separate
• _____________________ Is a technique for
  obtaining pure cultures by diluting by streaking
  on a Petri plate
• Only works if organism is present in large enough
  numbers-otherwise an enrichment step must precede
• Final step is to remove bacteria from one colony
  to grow in pure culture

32
Streak Plate for Isolation Fig. 6.10
33
Preserving Cultures

• Deep Freeze -50 to 95C several years
• ________________(freeze drying) -54 to 72 C
  and water is removed under vacuum. Sealed from
  air, bacteria can live for many years in this form

34
Patterns of Bacterial Growth

• Binary fission and budding

35
Exponential Growth Fig. 6.13
36
Phases of Growth Fig. 6.14
37
Direct Measurements of Bacterial Growth

• Plate Counts serial dilution (count 25-250
  colony plates)

38
Pour Plates/Spread Plates Fig. 6.16
39
Most Probable Number (set-up)
40
Most Probable Number (MPN) Fig. 6.18
41
Direct Microscopic Count Fig. 6.19
42
Indirect Methods spectrophotometry
43
Indirect Methods

• Metabolic activity
• Amount of product (say CO2 or and acid) is
  assumed to be in direct relationship with the
  number of cells present.)
• Dry Weight Used for fungi-so that amount of
  growth can be monitored
• Remove from culture medium, filter, dry and weigh
• Bacteria can also be monitored in this way and
  are removed from the growth medium (liquid) by
  centrifugation