04 BIO+210+FQ+2014+Ch+6+Microbial+Nutrition+and+Growth

8/10/2019 04 BIO+210+FQ+2014+Ch+6+Microbial+Nutrition+and+Growth

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Ch. 6. Microbial Nutrition & Growth

Learning Objectives: Chemical and energy requirements

(autotrophs, heterotrophs, chemotrophs, phototrophs)

Factors affecting growth (oxygen, temperature, pH, water availability)

Culturing microbes and different growth media

(selective, differential, anaerobic, etc.)

Assessing microbial growth (calculating growth, growth curve, methods to measure growth)


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Two groups of organisms based on source of carbon

Autotrophs: use inorganic source of carbon ( CO2 )Heterotrophs: catabolize reduced organic

molecules as source of carbon

Two groups of organisms based on use of chemicalsor light as source of energy

Chemotrophs:acquire energy from redox reactionsusing inorganic and organic compounds

Phototrophs: use light as their energy source


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Major factors influencing growth

Oxygen

Temperature

pH

Water availability


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Oxygen requirements

Obligate aerobes:Bacteria must grow withoxygen (essential for growth)

Obligate anaerobes:Bacteria must growwithout oxygen (forms of oxygen are toxic to their

cell structure)


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Environmental factors that influence microbes: e.g. gas needs

Oxygen can transform into toxic products:

Singlet oxygen (1O2), superoxide ion (O2-), peroxides (H2O2), andhydroxyls (OH-) can destroy cells

Most cells have enzymes to capture and neutralize these toxic

products

O2-+ O2

-+ 2H+ Superoxide dismutase H2O2+ O2

H2O2+ H2O2 Catalase 2H2O + O2

Aerobic, facultative anaerobic organisms have these enzymes,

but anaerobic organisms do not have these enzymes


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Oxygen requirements

Aerobesundergo aerobic respiration

Microaerophilesaerobes that require oxygenlevels from 2 to 10%

Facultative anaerobescan performfermentation or anaerobic respiration or aerobic

respiration

Aerotolerant anaerobesdo not useaerobic metabolism but have some enzymes that

detoxify oxygens poisonous forms

Anaerobesdo not use aerobic metabolism


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Temperature

Each microbe has its own

optimum temperature forgrowth

Too high a

temperaturedenatures proteins

and cell membranes

become too fluid

Too low a temperature

results in rigid and

fragile membranes


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Temperature

Psychrophiles

-5C to 15C

Arctic and Antarctic regions

Psychrotrophs

20C to 30C

Mesophiles25C to 45C

Thermophiles

45C to 70C

Hot springs

Hyperthermophiles

70C to 110C

Usually members ofArchaea

Hydrothermal vents


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Thermophilic bacteria


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An example of a psychrophilic alga


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pH

Neutrophiles: bacteria and protozoa that grow best in a narrow

range around neutral pH (6.5-7.5)

Acidophiles: bacteria and fungi that grow best in acidic

habitats,

Alkalinophiles: live in alkaline soils and water up to pH 11.5


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All microorganisms require water for growth

Facultative halophiles: can tolerate high

salt environments

Obligate halophiles: bacteria that must havehigh salt for cell growth (up to 30% salt)

E.g. Halobacterium (a member of Domain Archaea)

Water availability


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Physical effects of water

Water exerts pressure in proportion to its depth

Barophiles:organisms that live under extreme pressure


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Culturing (growing) microbes..

Inoculum refers to the cells that you introduce into

medium (broth or solid)

Pure culture: population of cells derived from single cell

All cells genetically identical

Pure culture obtained using aseptic technique

Cells grown on culture media

Can be broth (liquid) or solid form (agar plate/slant)


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Isolation of cells in a mixed culture of bacteria:

Colony: formed from a single cell that undergoes

many cell divisions

Agar plate: agar is the solid medium required for

growth of the colony; agar comes from red algae


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Obtaining Pure Culture

Streak-plate method obtaining isolated colonies

Object is to reduce number of cells being spread

Each successive spread decreases number of cells per

streak


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Pour plate method of obtaining isolated colonies


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Culture Media

Types of media

Defined media

Complex media

Selective media

Differential media

Anaerobic media


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Defined (synthetic) media: exact chemical composition is

known and each batch is chemically identical


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Blood agar is also a differential complex medium


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Selective medium:selecting growth ofsome microbes andinhibiting growth ofother microbes.

Bl d d diff ti l di


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Beta-hemolysisAlpha-hemolysis

No hemolysis

(gamma-hemolysis)

Blood agarused as a differential medium

Streptococcus pyogenes

Streptococcus pneumoniae

Enterococcus faecalis

M C k l ti d diff ti l di


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MacConkey agar as a selective and differential medium

An anaerobic culture system


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An anaerobic culture system

Clamp

Airtight lid

Envelopecontainingchemicals to

release CO2and H2

Palladium pelletsto catalyze reaction

removing O2

Methylene blue(anaerobicindicator)


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Bacterial cells divide by binary fission

Doubling time/Generation time: time required for parent cell to

divide and produce two daughter cells

Principles of Bacterial Growth

The growth curve in a bacterial culture


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Lag phase: growth lags; cells adjusting, not multiplying at max. rate

Log phase: exponential growth, max. rate of cell divisionStationary phase: cells stop growing/grow slowly; metabolic rate

declines; see depletion of nutrients, buildup of wastes

Death phase: number of viable cells decreases, rate depends on

species

The growth curve in a bacterial culture

P i i l f B i l G h


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Growth can be calculated

Example we have a culture of 3 cells in original

population

assume 20 minute generation time

after 2 hours of incubation the populationcontains:

3 x 2nwhere n is the number of generations

3 x 26

= 3 x 64 = 192 cells

Principles of Bacterial Growth

Formula: initial # of cells x 2n = # cells after growth


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Measuring Microbial Growth

Viable plate counts

Direct cell counts

Membrane filtration

Turbidity method

Estimating microbial population size viable plate count method


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Estimating microbial population size- viable plate count method

Detecting Bacterial Growth


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Detecting Bacterial Growth

Viable plate counts

Measures viable cells growing on solid culture media

Count based on assumption that one cell gives rise toone colony

Number of colonies = number of cells in sample

Ideal number to count

Between 30 and 300 colonies


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Direct cell count

does not distinguishbetween living anddead cells

bacterial cell numberis measured in aknown volume in ahemocytometer

Membrane filtration used to estimate microbial population


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Membrane filtration used to estimate microbial population


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- measures with

spectrophotometer

measures light

transmitted through

sample

Measurement is

inversely

proportional to cell

concentration

Limitation

Must have high number

of cells

Turbidity Method

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04 BIO+210+FQ+2014+Ch+6+Microbial+Nutrition+and+Growth

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