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Lab 13Bacterial cultivation
Selective /differential
media
Lab activities:
Demo selective and differentiation plates
 Streaking bacteria on differentiation plates

Cultivation

The process of growing microorganisms in
culture by taking bacteria from the
infection site (in vivo or environment) and
grow them in artificial environment in the
laboratory (in vitro).
Growth needs

Fastidious bacteria – relatively complex
growth needs

Non-fastidious bacteria- relatively basic
and straightforward growth needs
Phases of growth media

Broth- Growth of bacteria will change the
liquid from clear to turbid (cloudy).

Solid – Agar plates, slants – Bacterial cells
inoculated on solid media will multiply
enough to be seen by naked eye.
Colony- (clone)

Colony- A bacterial population derived from one
bacterial cell. The cells within the colony have
identical, genus, species, genetic and
phenotypic characteristics.

Pure bacteria - derived from a single colony.

Selection of a pure colony -most important for
bacterial identification
Media classification and function

Enrichment – Used to enhance growth of a
particular pathogen

Supportive - support growth of most non
fastidious bacteria
Media classification and function

Selective - Contain inhibitory agents that are
inhibitory to all organisms except those sought

Differential - Contain factors that allow bacterial
species to manifest certain metabolic
characteristics that distinguish them from other
species.

Media can be both selective and differential
based on the ingredients of the medium.
Blood agar plate (BA)

Nutrient agar with 5% sheep blood

Cultivation of fastidious and non fastidious bacteria.

Differential – Identify hemolysis - Some bacteria secrete
enzymes that lyse red blood cells (hemolysins) such that
a clearing around the colony appears.
b
hemolysis- complete clearing (white hemolysis)
a
hemolysis – incomplete clearing (green hemolysis)
g
hemolysis- no hemolysis
Mannitol Salt Agar (MSA)

Both selective and differential medium.

High salt concentration - inhibits most bacteria.

Selective for Staphylococcus sp.

Differentiate between Staphylococcus sp. by the sugar
mannitol fermentation .

Mannitol fermention produce acids that change the
medium pH.


Peach color- neutral- no fermentation
Bright yellow- Acidic – mannitol fermentation (Staph.
coag. pos.- Staph. aureus)
MacConkey Agar (MAC)

Selective and differential medium.

Selective - Gram positive bacteria are inhibited by the
presence of bile salts and crystal violet inhibitors in the
medium Most of gram negative bacteria will grow.

Differentiate- Between Gram negative bacteria by their
ability to ferment lactose.

Pink colonies- Bacteria that ferment lactose
(precipitation of some salts in media by acid production).

Pale colonies- Non fermenters
Eosine Methylene blue (EMB),

Differentiatial
between lactose
fermenting and non
fermenting enteric
bacteria
Tellurite Glycine Agar (TGA)

Selective- Tellurite glycine
and lithium inhibit most
bacteria

Preferential growth of
Staphyloccoci coagulase
positive (Staphyloccocus
aureus)
Bacteria streaked in lab




Staphylococcus aureus
Staphylococcus epidermidis
Salmonella pullorum,
E.coli
S.aureus
S.pullurum
S.epidermidis
E.coli
EMB and TGA