Chapter 6

 

Microbial Growth

-    Increase in number of cells, not cell size

-    Populations

-    Colonies

 

The Requirements for Growth

-    Physical requirements

-    Temperature

-    pH

-    Osmotic pressure

-    Chemical requirements

-    Carbon

-    Nitrogen, sulfur, and phosphorous

-    Trace elements

-    Oxygen

-    Organic growth factor

 

Physical Requirements

-    Temperature

-   Maximum

-   Optimum

-   Minimum

 

-    Psychrotrophs

-   Grow between 0C and 20–30C

-   Cause food spoilage

 

pH

-    Most bacteria grow between pH 6.5 and 7.5

-    Molds and yeasts grow between pH 5 and 6

-    Acidophiles grow in acidic environments

 

Osmotic Pressure

-    Hypertonic environments, or an increase in salt or sugar, cause plasmolysis

-    Extreme or obligate halophiles require high osmotic pressure

-    Facultative halophiles tolerate high osmotic pressure

 

Chemical Requirements

Carbon

-    Structural organic molecules, energy source

-    Chemoheterotrophs use organic carbon sources

-    Autotrophs use CO2

Nitrogen

-    In amino acids and proteins

-    Most bacteria decompose proteins

-    Some bacteria use NH4+ or NO3

-    A few bacteria use N2 in nitrogen fixation

Sulfur

-    In amino acids, thiamine, and biotin

-    Most bacteria decompose proteins

-    Some bacteria use SO42– or H2S

 

Phosphorus

-    In DNA, RNA, ATP, and membranes

-    PO43– is a source of phosphorus

Trace elements

-    Inorganic elements required in small amounts

-    Usually as enzyme cofactors

 

Toxic Oxygen

-    Singlet oxygen: O2 boosted to a higher-energy state

-    Superoxide free radicals: O2

-    Peroxide anion: O22–

-    Hydroxyl radical (OH)

 

Organic Growth Factors

-    Organic compounds obtained from the environment

-    Vitamins, amino acids, purines, and pyrimidines

 

Biofilms

-    Microbial communities

-    Form slime or hydrogels

-    Bacteria attracted by chemicals via quorum sensing

-    Share nutrients

-    Sheltered from harmful factors

 

-    Patients with indwelling catheters received contaminated heparin

-    Bacterial numbers in contaminated heparin were too low to cause infection

-    84–421 days after exposure, patients developed infections

-    Pseudomonas fluorescens was cultured from the catheters

-    What happened?

 

Culture Media

-    Culture medium: Nutrients prepared for microbial growth

-    Sterile: No living microbes

-    Inoculum: Introduction of microbes into medium

-    Culture: Microbes growing in/on culture medium

 

Agar

-    Complex polysaccharide

-    Used as solidifying agent for culture media in Petri plates, slants, and -  deeps

-    Generally not metabolized by microbes

-    Liquefies at 100C

-    Solidifies at ~40C

 

Culture Media

-    Chemically defined media: Exact chemical composition is known

-    Complex media: Extracts and digests of yeasts, meat, or plants

-    Nutrient broth

-    Nutrient agar

 

Anaerobic Culture Methods

-    Reducing media

-    Contain chemicals (thioglycolate or oxyrase) that combine O2

-    Heated to drive off O2

 

Capnophiles

-    Microbes that require high CO2 conditions

-    CO2 packet

Candle jar

 

Biosafety Levels

-    1: No special precautions

-    2: Lab coat, gloves, eye protection

-    3: Biosafety cabinets to prevent airborne transmission

-    4: Sealed, negative pressure

-    Exhaust air is filtered twice

 

Selective Media

-    Suppress unwanted microbes and encourage desired microbes

 

Differential Media

-    Make it easy to distinguish colonies of different microbes.

 

Enrichment Culture

-    Encourages growth of desired microbe

-    Assume a soil sample contains a few phenol-degrading bacteria and thousands of other bacteria

-    Inoculate phenol-containing culture medium with the soil, and incubate

-    Transfer 1 ml to another flask of the phenol medium, and incubate

-    Transfer 1 ml to another flask of the phenol medium, and incubate

-    Only phenol-metabolizing bacteria will be growing

 

Obtaining Pure Cultures

-    A pure culture contains only one species or strain

-    A colony is a population of cells arising from a single cell or spore or from a group of attached cells

-    A colony is often called a colony-forming unit (CFU)

-    The streak plate method is used to isolate pure cultures

 

The Streak Plate Method

 

Preserving Bacterial Cultures

-    Deep-freezing: –50 to –95C

-    Lyophilization (freeze-drying): Frozen (–54 to –72C) and dehydrated in a vacuum

 

Reproduction in Prokaryotes

-    Binary fission

-    Budding

-    Conidiospores (actinomycetes)

-    Fragmentation of filaments

 

Measuring Microbial Growth

-    Generation Time

-    Bacterial Growth Curve

-    Phases of Growth

 

Serial Dilutions

 

Direct Methods

-    Plate counts

-    Filtration

-    MPN

-    Direct microscopic count

 

Indirect Methods

-    Turbidity

-    Metabolic activity

-    Dry weight