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Microbial Growth

The growth of a population through an increase in the number of cells in a specific amount of time

1

What is Asexual Reproduction called

Binary Fission

2

Binary Fission steps

1. Parent cell increases in size
2. Chromosome is copied
3. DNA attaches to cell membrane
4. New cell walls form
5. Cell divides into two new cells

3

Budding

Yeast and some bacteria species
a bud forms on parent cell

4

Growth curve

Distinct, sequential phases of growth in a lab with a closed system (agar, broth)

5

Steps of Growth Curve

1. Lag Phase
2. Exponential (Log) phase
3. Stationary Phase
4. Death phase

6

Lag Phase

- preparation for cell division; no increase in population size
Metabolic Activity
-Enzyme and protein synthesis
-Breakdown of substances in environment
-Synthesis of new macromolecules and ribosomes

7

How long is lag phase?

Varies because of bacteria type and environment

8

Exponential (Log) Phase

-Cell numbers increase exponentially (slowly at first then extremely rapid)
-Cells are most virulent at this phase
-Most sensitive to antimicrobial medications
-Limited by nutrients, oxygen, waste accumulation, space

9

Generation time

Time it takes for one cell to divide into two cells (or population to double).
-10 minutes to 24 hours (average 30 minutes)

10

Formula for measuring population

Nt = N0 x 2n
N0= original number of cells in a population
Nt= number of cells in a population at a given time
n= number of divisions in a given amount of time

11

Stationary Phase

Environmental conditions do not favor continues growth
-Death rate = division rate
Cells may produce survival structures (glycocalyx, endospores, cytoplasmic inclusions

12

Abcess

collection of pus surrounded by inflamed tissue

13

What is pus made of

WBCs, tissue debris, prtoeins and bacteria

14

Abscess
(why formed and what happens)

Body's response to bacteria
Nutrients are limited so bacteria go to stationary phase

15

Are antimicrobials effective for abscesses?

No because bacteria is not dividing

16

Death phase

Cells begin to die out
Exponential death at a low rate
ATP reserves depleted
Prolonged decay: some cells survive by using nutrients of dead cells

17

Growth Phases in lab vs. nature

-Cultures pass through all growth phases in lab but not in nature.
- In nature nutrients enter cell's environment at low concentrations.
therefore growth is continuous, steady but at a low rate. Limited by nutrients. Metabolic wastes are removed by other microbes

18

Nutrients r/t position in colony
(growth on agar)

Edges: lots of nutrients = exponential growth
Middle: limited nutrients = stationary growth
Center: depleted nutrients = death

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Growth on agar

On solid media bacteria grows in colonies

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Colony

a distinct mass of cells that originate from a single cell

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Microbes can exist in many environments because they are...

small
easily dispersed
need only small quantities of nutrients
diverse in nutritional requirements

22

Environmental influences on Microbial Growth

moisture, temperature, pH, oxygen, solute concentration, hydrostatic pressure, radiation, nutrients

23

Moisture
(influences on microbial growth)

- Bacteria use diffusion to get nutrients from the environment (so water is needed for diffusion to occur)
-Preserving food through dehydration prevents bacteria from multiplying but may not kill bacteria

24

Temperature
(influences on microbial growth)

-Microbial species have specific temperature ranges in which they can grow

25

Reasons why extreme heat or cold affects growth

very cold - proteins not denatures, microbes not killed just slowed
very hot - proteins denatured, metabolism stops, cells destroyed

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Optimum temperature

Usually a small range where organism has fastest rate of growth

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Psychrophiles

cold loving
5-15 C
Grow in polar and glacial regions

28

Psychrotrophs

"cold feeding"
20 - 30 C
do not cause infection in humans
responsible for spoiling of refrigerated and frozen food (Blood)

29

Mesophiles

middle loving
25 - 45 C
Pathogens are mesophiles
optimum temperature for human pathogens is around 37C

30

Normal body temp in C

37 C

31

Most refrigerators run what temp in C

4 C

32

Thermophiles

heat loving
45 - 70 C
Found in natural hot springs, compost

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Hyperthermophiles

extreme heat loving
70 C and higher
usually Archaea
hydrothermal vents in ocean

34

pH
(influences on microbial growth)

pH of external environment - internal pH is neutral

35

Acidophiles

optimal pH is below 5.5
example: H. Pylori

36

Neutrophiles

optimal pH 6-8
Human pathogenic bacteria optimum pH is 7.3

37

Alkaliphiles

optimal pH above 8.5 (very small group of bacteria
ex. alkaline lakes and soils

38

Bleach

pH of 10
kills most bacteria

39

Hydrostatic pressure
(influences on microbial growth)

-pressure exerted by standing water
-Most bacteria killed by high pressure (autoclave)
- Some extreme microbes need high pressure to keep enzymes from denaturing

40

Radiation
(influences on microbial growth)

Can damage or kill microbes
3 types: Ionizing, UV, microwave

41

Ionizing radiation

removes electrons - destroys DNA, damages membranes
ex. Xrays and Gamma rays
Gram + can survive longer than Gram -

42

Uv radiation

destroys DNA (mutations)
some bacteria have repair enzymes to correct mutations

43

Microwave radiation

kills bacteria with heat only

44

Harmful oxygen-containing molecules

Superoxide ion (o2-)
Peroxides(H2O2)
Damage living tissue (especially membranes and can kill cells -> aging

45

Antioxidants

Enzymes present in organisms to detoxify harmful oxygen containing molecules

46

Examples of antioxidants

Superoxide dismutase (SOD)
Catalase

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Superoxide dismutase

converts O2- to O2 + H2O2

48

Catalase

converts H2O2to O2 and H2O

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Obligate Aerobes

-Have SOD and catalase
- must have O2 for metabolism (aerobic respiration) and growth
- ex. most fungi and protists, some bacteria (Bacillus, Pseudomonas)

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Facultative Anaerobes

- Usually have SOD and catalase
-"flexible" - can survive with or without oxygen
- Grow best with oxygen (aerobic respiration
- Ex. Ecoli, Staphylococcus and Saccharomyces(yeast)

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Microaerophiles

-Only need small amounts of SOD and catalase
- Need small concentrations of O2( 2-10%) for aerobic respiration
- Large amounts of oxygen are inhibitory
- Found in mucous linings of hollow organs
- Ex. H. Pylori

52

Obligate Anaerobes

- Usually lack both SOD and catalase
- Cannot grown if O2 is present
- No aerobic respiration
- Found in deep mud, lakes, oceans, inside animal bodies
-Ex. Clostridium

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Aerotolerant Anaerobes

- May have SOD, but not catalase
- Indifferent to oxygen
- Do not use oxygen (OBLIGATE FERMENTERS)
- Ex. Streptococcus pyogenes

54

How do bacteria try to maintain a hypotonic environment?

They will pump in K+ of produce extra amino acids

55

Halotolerant

- can tolerate moderate concentrations of salt - up to 10% NaCl
ex. staph on the skin

56

Halophiles

- REQUIRE a high level of NaCl
- Marine microbes
- Extreme halophiles are Archaea living salt lakes

57

Osmophiles

grow in high sugar concentrations

58

Element reminds

C. Hopkins CaFe Mg
(C. Hopkins Cafe Mighty good)

59

Carbon

energy source (glucose) and building blocks

60

Nitrogen

amino acids, nucleotides
-some can synthesize all 20 amino acids
- others need amino acids added to culture medium

61

Sulfer

amino acids, coenzymes

62

Phosphorus

ATP, phospholipids, nucleotides

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Vitamins

- organic substance required by an organism, usually a coenzyme
- some bacteria make own vitamins
- others need vitamins added to culture medium
- microbes in human intestine make Vitamin K and some B vitamins = mutualistic symbiosis

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Other elements are often...

cofactors

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Media

liquid or solid material used to grow microbes

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Agar

solidifies culture media at 1.5% concentration
- complex polysaccharide, extracted from seaweed
- melts at 100C and solidifies at 45C
- first used by Koch (Mrs. Hess)
- not degraded by microbes

67

Sterilizing agar

in autoclave
121 C for at least 15 minutes at 15 psi of steam

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Liquid Media (broth)

good for growing large numbers of bacteria in a short time, contains low concentrations of agar (0.5%) for motility test

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Defined (synthetic) media

made of exact amounts of pure chemicals

70

Complex media

- made of some ingredients with variable chemical composition
- can vary from batch to batch
- extracts from beef, yeast, blood...
- example nutrient agar

71

Selective media

- encourages the growth of some microbes and suppresses the growth of others
- for isolation of microbes

72

Differential Media

- contains substances that certain microbes can change in a recognizable way
- observable changes in media due to biochemical reaction from microbe
- ex. color or pH change

73

Indirect measurements

measure property of the mass of cells and then estimate the number of microbes

74

Direct measurement

determine the total number of cells, includes living and/or dead cells, more accurate measurements of numbers of microbes

75

Turbidity

-cloudiness as evidence of growth
- can use spectrophotometer
- measures biomass not number of cells
- cannot detect minor concentrations

76

Metabolic Activity

1. Rate of metabolic products that a culture produces: gases, acids
2. Rate of utilization of a substrate: Oxygen, glucose, ATP
3. Rate of reduction of certain dyes: methylene blue becomes colorless when reduced

77

2 types of Indirect measurement

Turbidity
Metabolic Activity

78

Direct cell counts

1. Direct Microscope count
2. Coulter Counter

79

Direct microscope count

special glass slide with light compound microscope

80

Coulter Counter

- electronic, cells pass through a narrow channel
- rapid, accurate

81

Viable cell count

1. Standard plate count
2. Filtration

82

Standard plate count

count colonies against a grid
- Quebec colony counter
- use serial dilutions to decrease cell numbers before counting

83

Filtration
(viable cell count technique)

fine, sterile filter collects microbes, then placed on agar plate, count colonies, used to concentrate samples

84

Countable plate has how many cells

30 -300
more - too many cells produce too many colonies to count
less - not enough colonies for a valid count