Transcript Lecture 2 Microbial Nutrition

Microbial Nutrition
Gary Andersen
Reference Chapter 2,5,6, 7, 8, 9 Microbiology by
Tortora, Funke & Case
Metabolism
Chemical
Substances
Cell
Growth
What do microbes eat?
Composition and Nutrition of
Cells
• 96% of cells are composed of HCNOPS.
• Macronutrients are nutrients required in
larger quantities. (carbohydrates, proteins,
fats and other CHO molecules)
• Micronutrients are nutrients required in
trace amounts. (Mn, Zn, Cu, Ni)
Where are the Elements Used?
Sources of Nutrients
• Carbon – CO2 in air and rocks organisms
• Nitrogen – N2 in air, NO2, NO3, NH4 in soil and
water  organisms (NH3)
• Oxygen – O2 in air, inorganic salts SO4, PO4, NO3,
H2O
• Hydrogen – Water, Organic compounds in
organisms
• Phosphorus – Rocks and minerals  organisms
• Sulfur – Rocks and minerals  organisms
Microbe Nutritional Types
• Autotrophs – “Self feeder”
– Photoautotrophs – Photosynthetic (energy from
light)
– Chemoautotrophs – Energy from simple
inorganic chemicals
• Methanogens – Metabolize H2 and CO2 into CH4
and H2O
• Heterotrophs – “Other feeder”
– Chemoheterotroph – Obtain carbon and energy
from organic compounds. CnH2nOn + O2 
CO2 + H2O + ATP (Adenosine tri-phosphate)
• Saprobe – Free living organisms that feed on dead
organisms
• Parasite – Derive nutrients from the tissues of hosts.
Microbial Clean-Up: The 1989 Exxon Valdez oil spill left
great quantities of pooled oil on sites in the Gulf of Alaska,
such as on Green Island
Microbial Clean-Up: Bioremediation in 1989, by the
application of nutrients (nitrogen and phosphorus) to the
shoreline accelerated the bacterial biodegradation of the oil
into carbon dioxide and water
Microbial Clean-Up: In 1991, the area was surveyed
and found to be mostly cleared of oil, with no further
treatment recommended
How do Microbes Eat?
Transport Mechanisms
• Passive Transport
– Diffusion – Movement of molecules from a high
concentration to a low concentration.
– Facilitated Diffusion- diffusion assisted by
conformational change in a protein molecule.
– Osmosis – Diffusion of water through a semipermeable
membrane
• Active Transport – Moving particles against the
diffusion gradient using membrane proteins and
expending energy.
– Endocytosis – Engulfing with cell membrane and
forming a vacuole.
• Phagocytosis – Engulfing of cells or particles by the cell
membrane
• Pinocytosis – Engulfing of liquids by the cell membrane
Passive Transport
• Osmosis animation:
http://www.tvdsb.on.ca
/westmin/science/sbi3a
1/Cells/Osmosis.htm
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Comparing Solutions
• Hypertonic Solution – Higher solute
concentration. Cells in hypertonic solutions lose
water and the cell membrane shrinks away from
the cell wall. (Salt on a slug)
• Hypotonic Solution – Lower solute concentration.
Cells in hypotonic solutions take on water and
swell. (Prune wrinkles of skin)
• Isotonic Solutions – Solutions that have reached
an equilibrium with a cell or another solution. The
concentration of solute is equal and the diffusion
of water proceeds at equal rates.
(See page 93 of text for what happens to a cell in hypertonic
and hypotonic solutions.)
Animations of Passive and
Active Transport
• http://programs.northlandcollege.edu/biology/Bio
logy1111/animations/transport1.html
• http://highered.mcgrawhill.com/olc/dl/120068/bio02.swf
How do microbes metabolize
nutrients?
Fermentation and Respiration
Enzymes
• Provide a surface on which reactions take place
• Active site: the area on the enzyme surface where the enzyme
forms a loose association with the substrate
• Substrate: the substance on which the enzyme acts
• Enzyme-substrate complex: formed when the substrate
molecule collides with the active site of its enzyme
• Enzymes generally have a high degree of specificity
• Endoenzymes (intracellular)/exoenzymes (extracellular)
The Action of Enzymes on
Substrates to Yield Products
Each substrate binds to an active site, producing
an enzyme-substrate complex. The enzyme helps
a chemical reaction occur, and one or more
products are formed
Competitive Regulation and
Inhibition of Enzymes
Noncompetitive (allosteric)
inhibition of enzymes
http://highered.mcgraw-hill.com/olc/dl/120070/bio10.swf
Factors Influencing Enzymes
• Temperature
• pH
• Concentration of substrate, product, and
enzyme
Relationship between temperature
and enzyme activity
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Microbes and Environmental
Factors
• Temperature
• pH
• Oxygen
• Pressure
Extremophiles – Organisms that can survive under
extreme environmental conditions. An interesting
source of chemical products.
Interesting Website on Extremophiles and Chemical
Products: http://www.mediscover.net/Extremophiles.cfm
Temperature
Classification
• Psychrophile
• Mesophile
• Thermophile
Temperature
• Thermophiles –
organisms that grow
at >45 degrees C.
• Pyrococcus fumarii
is an example of a
thermophile that can
survive at 113 C.
Thermal pool
Grand Prismatic Spring
pH
• Acidophiles – grow at
low pH levels. (1-2)
• Alkalinophiles – live
at high pH levels. (910)
14 Alkaline
13
12
11
10
9
8
7 Neutral
6
5
4
3
2
1
0 Acidic
Pressure
• Barophiles –
organisms that grow at
elevated pressure (31000 x air pressure).
(Found in ocean
depths often in
thermal vents)
Presence of Oxygen
• Aerobe – Organism able to use O2 in metabolism.
• Anaerobe – Organism unable to use O2 in metabolism.
Obligate aerobes - oxygen mandatory
Obligate anaerobes - oxygen toxic
Facultative anaerobes – Aerobe that can also live without O2
Microaerophiles - low oxygen levels required.
Aerotolerant - anaerobic metabolism, oxygen not toxic
Microbial Processing of Oxygen
• Step 1: O2- + O2- + 2H+  H2O2 + O2
(Catalyzing
enzyme is Superoxide dismutase)
• Step 2: H2O2 + H2O2  2H2O + O2
Catalase)
(Catalyzing enzyme is
Living without Oxygen….Glucose Fermentation Pathways
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Glucose Fermentation
Net and practical results
•
•
Cells get chemical energy (ATP)
Fermentation products are natural
waste products useful to humans:
1.
2.
3.
4.
Fermented beverages
Bread
Cheese
Yogurt
Using fermentation metabolism to identify microbes: A
positive (yellow) mannitol-fermentation test. This test
distinguishes the pathogenic Staphylococcus aureus
(MSA)
Test
Mannitol
Salts
Agar
Comparing Aerobic Respiration with
Anaerobic Fermentation
Fermentation
• Fermentation yields small amount of ATP (2)
• Partial oxidation of carbon atoms (6 C  3 C)
Respiration
• Substrate molecules are completely oxidized to
C02 (6 C  1 C)
• Far higher yield of ATP (36)
• The Krebs Cycle and Electron Transport Chain
Final Electron Acceptors: Aerobic respiration,
anaerobic respiration, and fermentation have
different final electron acceptors
Microbe Growth
• Binary or
transverse
fission
• Generation or
Doubling Time
– the time
required for
parent cell to
form two new
daughter cells.
Microbe Growth
• Lag – new cells
require adjustment
and enlargement.
The cells are not
multiplying
rapidly.
• Log or exponential
– maximum rate of
growth
Microbe Growth 2
• Stationary –
death and
multiplication
balance out.
Depleted
nutrients and
waste buildup.
• Death – limiting
factors intensify.
May last a long
time.
Calculating Growth of Cells
Nf = (Ni)2n
Nf = Final population
Ni = Initial population
2n = # cells in generation
n = generation number
Use the table in the handout from the Talaro Appendix A-2 to
calculate the number of cells in the generation.
Measuring Growth Serial Dilutions, Plate
Counts and Turbidity
Measuring growth
turbidity
plate counts
Calculation of the number of bacteria per milliliter of culture
using serial dilution
Pour plate: made by
first adding 1.0ml of
diluted culture to 9ml of
molten agar
Spread plate: made by
adding 0.1ml of diluted
culture to surface of
solid medium
Counting colonies using a bacterial colony counter
Bacterial colonies viewed through the magnifying
glass against a colony-counting grid
Countable number of colonies
(30 to 300 per plate)
Which of these plates would be the
correct one to count? Why?
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The Petroff-Hausser Counting Chamber
Turbidity, or a cloudy appearance, is an indicator of
bacterial growth in urine in the tube on the left
A Spectrophotometer: This instrument can be used to
measure bacterial growth by determining the degree of
light transmission through the culture
The Streak Plate Method uses agar plates to
prepare pure cultures
A Streak Plate of Serratia marcescens. Note the greatly
reduced numbers of growth /colonies in each successive region
Types of Culture Media
• Natural Media: In nature, many species of
microorganisms grow together in oceans, lakes, and soil
and on living or dead organic matter
• Synthetic medium: A medium prepared in the
laboratory from material of precise or reasonably welldefined composition
• Complex medium: contains reasonably familiar
material but varies slightly in chemical composition
from batch to batch (e.g. peptone, a product of enzyme
digestion of proteins)
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Commonly Used Media
• Yeast Extract
• Casein Hydrolysate
• Serum
• Blood agar
• Chocolate agar
Selective, Differential, and Enrichment Media
• Selective medium: encourages growth of some
organisms but suppresses growth of others
(e.g. antibiotics)
• Differential medium: contains a constituent that
causes an observable change (e.g. MacConkey agar)
• Enrichment medium: contains special nutrients that
allow growth of a particular organism that might
not otherwise be present in sufficient numbers to
allow it to be isolated and identified
Three species of Candida can be differentiated in mixed
culture when grown on CHROMagar Candida plates
Identification of urinary tract pathogens with
differential media (CHROMagar)
Ecological Associations
Microbial Associations
Symbiotic
Mutualism
Commensalism
Non-Symbiotic
Parasitism
Synergism
Antagonism
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Symbiosis (Mutualism)
• Obligatory
• Both organisms
benefit.
• Examples: algae +
fungus = lichen,
termites and
trychonympha (a
protist)
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Symbiosis (Commensalism)
• One organism
benefits and the
other is not
harmed.
• Examples: Nonpathogenic
bacteria on our
skin; satellitism
between bacteria
colonies.
Symbiosis (Parasitism)
• One organism
benefits and the other
is harmed.
• Examples: Pathogenic
organisms on their
host. Plasmodium
vivax a protozoan
parasite causing
malaria.
Non Symbiotic (Synergism)
• Free living organisms.
• Both benefit
• The relationship is
optional.
• Examples: Shared
metabolism; nitrogen
fixing bacteria in the
soil and plants
Substance A
Microorganism 1
Substance B
Microorganism 2
Substance C
Microorganism 3
End Product used by all three
microorganisms
Non Symbiotic (Antagonism)
• Free living organisms
• Organisms compete
for resources.
• One organism secretes
a substance toxic to
the other.
• Example: Ruminal
cellulose digesting
bacteria and fungi
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End of Microbial Nutrition
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