September 14, 2015

• Defined as the nutrient material prepared for the growth of organisms in the lab
– in vivo—growth inside normal biological context
– in vitro—growth outside their biological context
• Inoculum is the microorganisms that are introduced to the media to initiate growth

1. Must contain the necessary nutrients to promote growth
2. Incubated at optimal temperature
3. Sterility

• Broth is liquid media
• Agar can be added to broth to produce solid media
– Complex polysaccharide derived from marine algae
– Liquefies at 100°C and becomes solid at 40°C
– Not metabolized by microbes
– Useful in make petri dishes, slants, and deeps

• There are several different varieties of culture media
– Chemically defined media—exact chemical composition is known
– Chemically undefined media—exact chemical composition is not known
• Complex media contain extracts and digests of yeasts, meat, or plants
– Nutrient broth
– Nutrient agar

Table 6.2 A Chemically Defined Medium for Growing a Typical Chemoheterotroph, Such as Escherichia coli

Table 6.4 Composition of Nutrient Agar, a Complex Medium for the Growth of Heterotrophic Bacteria

• Cultivation of anaerobic bacteria poses a problem to scientists
– Must absorb all the oxygen from medium/environment in order to grow microbes
• Reducing media
– Contain chemicals (sodium thioglycolate or oxyrase) that combine O
2 and removes all available oxygen
– Usually contained in screw cap test tubes or jars
– Media is heated before use to drive off O
2 tube)
(screw cap

Figure 6.6 A jar for cultivating anaerobic bacteria on Petri plates.
Lid with
O-ring gasket
Envelope containing sodium bicarbonate and sodium borohydride
CO
2
Anaerobic indicator
(methylene blue)
Petri plates
Clamp with clamp screw
H
2
Palladium catalyst pellets

Figure 6.7 An anaerobic chamber.
Air lock
Arm ports

• Microbes that require high CO
2
– Camplyobacter spp.
conditions
• CO
2 packet
– Chemical packets are used to generate carbon dioxide within containers
• Candle jar
– Contains a lit candle that depletes the oxygen in an environment and generates carbon dioxide
• Low-oxygen, high-CO
2 conditions resemble the conditions of the intestinal and respiratory tract

• In clinics and public health microbiology, it is necessary to identify specific microbes to distinguish colonies of different microbes
– Identifying the microbes that cause disease
• Clinicians use both selective and differential media to accomplish this task

• Selective media contains compounds that inhibit the growth of certain bacteria while promoting the growth of others
– Eosin-methylene blue (EMB) plates are an example of a selective media
• Inhibits the growth of gram positive bacteria

E. coli colonies on an EMB plate

• Differential Media grows a variety of different organisms. Various organisms produce different products that affects the appearance of the microbe on the media
– Blood Agar Plate

• Erythrocytes are incorporated into nutrient agar medium
• Certain bacteria produce products that lyse Red
Blood Cells
– Alpha-hemolytic- partial lysis
– Beta-hemolytic- complete lysis
– Gamma-hemolytic- no lysis

• Some culture plates are both Selective AND Differential
• Mannitol Salt Agar (MSA)
– High salt concentration (7.5%) only permits
Staphylococcous spp. growth
– Mannitol is a sugar alcohol that is fermented by certain species
– If mannitol is fermented, acidic products are formed.
Indicated by phenol red (yellow is acidic)
• Pathogenic Staph will ferment mannitol
• Non-pathogenic Staph will not ferment mannitol

Figure 6.10 Differential and Selective medium.
Uninoculated
Staphylococcus epidermis
Staphylococcus aureus