I. Microbial Growth

1. Bacterial Division

• Binary fission

2. Generation Time

1 à 2

2 à 4

• Exponential
• Cell number = 2ⁿ n = # of doublings (generations)
• Generation time = time req. for bact. pop. to double

3. Growth Curve

• cultivation in liquid medium
• closed system

a. lag phase

• no immediate × in cell #
• new components synth.
• varies in length

b. log phase

• growing & ÷ max. rate
• population doubling - logarithmic
• rate of growth constant
• pop. most uniform
• industrial fermentations

c. stationary phase

• death = growth
• total # live cells constant
• why?

Nutrient ×

O₂ × (aerobes)

waste ×

• antibiotic production

d. death phase

• living cells ×
• logarithmic decline
• endospores

In a local farm field bacteria are degrading the herbicide 2,4 D. You collect some of this soil and inoculate a broth culture in the lab to which you have added 2,4 D. Your bacteria are alive and well in the culture, but for 6 weeks you do not see any increase in cell numbers. In which part of the growth curve are your bacteria?

B. Measurement of Microbial Growth

1. Cell Numbers

a. Direct counts

• advantages - quick, cheap, cell size & morph
• disadvantages - pop. needs to be large, can’t tell living from dead

b. Plate counts

• diluted sample dispersed over solid agar surface
• each microbe à colony
• original # viable cells can be back calculated
• Ex. dilute sample 1 ml into 100 ml (a 1/100 dilution), count 150 colonies

# colonies x inverse of dilution = # original/ml

150 x 100 = 15,000 CFU/ml

• advantages - simple, sensitive
• disadvantages - have to know how to culture the microbe

Modification - counts from membrane filters

2. Cell mass

a. dry weight - time consuming

b. light scatter - spectrophotometer

In what ways do we preserve food?

1. Physical

a. Temperature

i. impt. pts

• cell temp same as environ temp
• enzyme-catalyzed rxns are temp sensitive
• temp speed up chem rxns - double every 10ºC

ii. cardinal temps

• minimum - membranes gel, transport too slow
• optimum - enz. rxns max. rate
• maximum - proteins & nucleic acids denature, cytoplasmic membrane collapses

iii. temp. classification

• Psychrophile - cold-loving (optimum: 15ºC)

Ex. Chlamydomonas nivalis (-36, 0, 4)

• Psychrotroph - cold tolerant (capable of growth 0-30ºC)

Ex. Bacillus cereus, Listeria monocytogenes

• Mesophile - middle temp-loving (opt: 10-50ºC)

Ex. Most pathogens

• Thermophiles - heat-loving (opt: 50-60ºC)

Ex. Thermoplasma

• Hyperthermophiles - extreme-heat (opt: above 80ºC)

Ex. Pyrolobus fumarii (90, 106, 113)

b. pH

i. impt. pts

• internal pH of cell is neutral
• each species has range and opt.
• protozoa & most bact - neutral

fungi & algae - slightly acidic

ii. effects of wrong pH

• disrupts membranes
• inhibits transport
• inactivates enz

iii. pH classification

• Neutrophiles: 5.5 - 8 (most)
• Acidophiles: 0 - 5.5

habitats - ore mines, stomach

some molds, some bact.

Ex. Helobacterium pylori

uses for humans - low pH chem rxns

• Alkalophiles: 8.5 - 11.5

habitats - soda lakes

uses - laundry detergent

c. Water

i. impt. pts

• water is essential to all life (an impt. solvent)

ii. effects of water imbalance

• hypotonic (outside cell solute conc. low & H₂O high) - H₂O enters cell, cell bursts

solute - 1 subst. dissolved in another

most proks, algae, and fungi have cell walls

• hypertonic (outside cell solute conc. high & H₂O low) - H₂O leaves cell, plasma membrane shrinks, metabolic inactivity, DNA disordered

iii. Adapted to hypertonic environs

• Halotolerant - can tolerate higher salt

habitat - skin

Ex. Staphylococcus aureus

• Halophiles - req. NaCl (1-15%)

habitat - sewater

• Extreme halophiles - 15-30% NaCl

habitat - Dead Sea, Great Salt Lake

Ex. Halobacterium (Arch)

• Osmophiles - high sugar (maple sugar, jam)

• Xerophiles - very dry (cereal, dried fruit)

d. Oxygen

i. evolution of Earth's atmosphere

4.8 billion -------------> no O₂ (reducing)

2.25 billion------------>oxygenic photosynthesis

2 billion --------------->1% O₂

today ----------------->20% O₂

ii. why is O₂ bad?

O₂ - accepts e- and becomes reduced to H₂O

O₂ + e- à O₂-

O₂- + e- + 2H⁺ à H₂O₂

H₂O₂ + e- + H⁺ à H₂O + OH

superoxide dismutase (SOD)

2O₂ + 2H⁺ ----------------------> O₂ + H₂O₂

catalase

H₂O₂ --------------------------->2H₂O + O₂

iv. Oxygen tolerance classification

• Anaerobe - does not req.O₂ for growth, will not survive exposure, neither cat nor SOD
• Aerotolerant anaerobe - does not req.O₂ for growth, will survive exposure, has SOD only
• Facultative anaerobe - can grow with or without O₂, both cat and SOD
• Microaerophile - need O₂ but less than is present in air
• Aerobe - req. O₂ for growth, both cat and SOD

II. Control of Microbial Growth

 

A. Terminology

1. sterilization - destroy all viable cells, spores, viruses

2. disinfection - remove pathogens on inanimate surfaces

3. antiseptic - kill pathogens on living tissue

4. sanitize - lower # of pathogens

B. How do we kill microbes?

1. Nonspecific

a. Physical methods heat

b. Chemical methods

i. phenols - denature proteins, disrupt membranes

ii. alcohols - denature proteins, dissolve lipid membranes

iii. halogens - oxidation of cellular material Ex. chlorine, iodine

2. Specific - Antibiotics (refer to CH 20)

 Antibiotic - natural substance prod. by 1 microbe that inhibits growth of another

a. How were antibiotics discovered?

Fleming (1928) Penicillium notatum

b. How do antibiotics work?

Bactericidal - kill

Bacteriostatic - inhibit

Selective toxicity - no harm to host

c. Cellular Target Sites 

i. cell wall

• prevent synth of new peptidoglycan
• works only on log phase cells
• Ex. Penicillin - penicillin-resistant bact. Enz - Ex. Staphylococcus aureus

ii. plasma membrane

• alter permeability
• euks - sterols, proks no sterols
• Ex. nystatin - antifungal

• interfere with enz.
• Ex. rifampin - TB

iv. proteins

target 70S ribosome

greater toxicity - why?

Ex. Tetracycline, chloramphenicol, streptomycin, erthromycin

3. Antibiotic Resistance

a. history

• 1969 - government cuts funding
• 1980 - drug companies cut research
• 1980s - antibiotic resistance gains foothold
• 1990s - rapid á in resistance

b. how did we get into this predicament?

The facts

• Manufacture greater than 50 million pounds/yr
• 150 million courses prescribed/yr
• 17 million for upper respiratory tract infections
• 16 million for bronchitis
• 23 million for ear infections

Clinical Case Example

Day 1 - 32-year-old woman - spiking intermittent fevers

• CBC shows white blood cells very high

Day 2 - GNR recovered from blood - empirical selection of IV antibiotic

Day 3 - Lab confirms E. coli; susceptibility test shows susceptibility to several antibiotics

Day 4 - still spiking fevers - switch antibiotics

• E. coli still recovered from blood - now resistant to all antibiotics but 1

Day 8 - surgeon called back in

Day 19 - dies of multiple organ failure due to overwhelming

• coli infection.

What can be done?

Can we undo resistance?

We can try to slow development

How can we do that?

III. Microbial Nutrition

A. Equation for Cell Growth

MONOMERS + INFORMATION + ENERGY

à POLYMERS à MACROMOLECULES

à NEW CELL

• monomers - from medium
• information = DNA
• energy - from biochem. rxns

B. Nutritional Patterns Among Microbes

CHNOPS

1. phototrophic autotroph
2. phototrophic heterotroph
3. chemolithotrophic autotroph
4. chemoorganotrophic heterotroph

 

C. Adaptations to nutrient limitations

Copiotroph - á nutrients

Oligotroph - â nutrients

1. synth. more carrier proteins

2. use different nutrients

3. adjust metabolic rate based on least plentiful nutrient

Additional Information