Culture media are used to support the growth of microorganisms. They can be liquid, semisolid, or solid and contain various nutrients like water, electrolytes, peptone, agar, meat extract, and blood or serum. Culture media are classified based on consistency and purpose of use. Common types include basal media, enriched media, selective media, differential media, transport media, and anaerobic and blood culture media. Automated systems now allow continuous monitoring of blood cultures for faster detection of microbial growth compared to conventional methods.

1. CULTURE MEDIA
Liquid or solid substance - contains
nutrients to support the growth, and
survival of microorganisms.
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2. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
Constituents of Culture Media
Constituent Explanation
Water and Electrolytes Sodium chloride
Peptone Mixture of partially digested proteins - obtained from various sources - heart muscle, casein
or fibrin, or soya.
Agar Used for solidifying the culture media
Source: Prepared from - cell wall of seaweeds and available commercially in powder form
Preparation: Agar powder is dissolved in water and subjected to sterilization by autoclave.
When the temperature of the molten agar comes down to 45°C, it is poured into the Petri
dishes and then allowed to set for 20 minutes.
Concentration:
Solid agar preparation - 1-2%
Semisolid agar- 0.5%
Solid agar to inhibit Proteus swarming- 6%

3. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
Constituents of Culture Media (Cont..)
Constituent Explanation
Meat extract Highly concentrated meat stock, usually made from beef.
Yeast extract (Prepared from Baker’s yeast) and malt extract (contains maltose)
Blood and serum Important components of enriched media; provide extra nutrition to fastidious
bacteria. 5–10% of sheep blood is used. Alternatively, horse, ox or human blood
can also be used.

4. Types of Culture Media
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Based on consistency, culture media are
grouped into:
▰ Liquid (or broth) media
▰ semisolid media
▰ solid media

6. Types of Culture Media
Based on the growth detection,
culture media are classified as:

7. Conventional culture media:
Prepared from nutrients - aqueous extract of meat, peptone.
 Simple/basal media
 Enriched media
 Enrichment broth
 Selective media
 Differential media
 Transport media
 Anaerobic media.

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Essentials of Medical Microbiology
Contain minimum ingredients that support the growth of non-fastidious
bacteria.
▰ Peptone water: Contains peptone (1%) + NaCl (0.5%) + water
▰ Nutrient broth: Peptone water + meat extract (1%).
▰ Nutrient agar: Nutrient broth + 2% agar
▰ Semisolid medium: Concentration of agar - reduced to 0.2–0.5 %.

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Essentials of Medical Microbiology

10. Uses of Basal Media
▰ Testing the non-fastidiousness of bacteria.
▰ Serve as base for the preparation of many other media.
▰ Nutrient broth - used for studying the bacterial growth curve.
▰ Nutrient agar - preferred medium for:
 Performing the biochemical tests, such as oxidase, catalase,
etc.
 Study the colony morphology
 Pigment demonstration. 10

11. Enriched Media
▰ Basal medium added with additional nutrients - blood,
serum or egg
▰ In addition to non-fastidious organisms, support the
growth of fastidious nutritionally exacting bacteria.
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12. Enriched Media (Cont..)
Blood agar –
▰ Prepared by adding 5-10% of sheep blood to the
molten nutrient agar at 450C.
▰ Tests the hemolytic property of the bacteria,
which may be either- i) partial or α (green)
hemolysis and ii) complete or β hemolysis
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13. Enriched Media (Cont..)
Chocolate agar:
▰ Heated blood agar, prepared by adding 5 -10% of sheep
blood to the molten nutrient agar at 70°C.
▰ More nutritious than blood agar - supports certain
highly fastidious bacteria - Haemophilus influenzae that
does not grow on blood agar.
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14. Enriched Media (Cont..)
Loeffler’s serum slope: Contains serum - used for isolation of
Corynebacterium diphtheriae
Blood culture media: Used for isolating microorganisms from
blood - available as conventional or automated blood culture
media. 14

15. Enrichment Broth
▰ Liquid media added with inhibitory agents which selectively
allow certain organism to grow and inhibit others.
▰ Important for isolation of pathogens from clinical specimens
which also contain normal flora (e.g. stool and sputum
specimen).
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16. Enrichment Broth (Cont..)
Examples :
▰ Tetrathionate broth—Used for Salmonella Typhi
▰ Gram-negative broth—Used for Shigella
▰ Selenite F broth—Used for Shigella
▰ Alkaline peptone water (APW)—Used for Vibrio cholerae.
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17. Selective Media
Solid media containing inhibitory
substances - inhibit the normal flora
present in the specimen and allow
the pathogens to grow.
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Media Used for isolation of
Lowenstein Jensen (LJ)
medium
Mycobacterium
tuberculosis
Thiosulphate Citrate Bile
salt Sucrose (TCBS)
Vibrio species
DCA (Deoxycholate
Citrate Agar)
Salmonella and Shigella
from stool
XLD (Xylose Lysine
Deoxycholate) agar
Salmonella and Shigella
from stool
Potassium tellurite agar
(PTA)
Corynebacterium
diphtheriae

18. Selective Media (Cont..)
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Lowenstein–Jensen medium TCBS agar

19. Selective Media (Cont..)
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DCA (Deoxycholate Citrate Agar) XLD (Xylose Lysine Deoxycholate) agar

20. Transport Media
▰ Used for the transport of the clinical specimens suspected to
contain delicate organism or when delay is expected while
transporting the specimens from the site of collection to the
laboratory
▰ Bacteria do not multiply in the transport media - they only remain
viable.
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21. Transport Media (Cont..)
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Organism Transport media
Neisseria Amies medium, Stuart’s medium
Vibrio cholerae VR (Venkatraman-Ramakrishnan) medium
Autoclaved sea water
Cary Blair medium
Shigella, Salmonella Buffered glycerol saline
Cary Blair medium

22. Differential Media
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Differential media Features
MacConkey agar  Differential and low selective medium - used for the isolation of enteric
GNB.
 Differentiates organisms into LF (pink colonies, e.g. Escherichia coli)
and NLF or (colorless colonies, e.g. Shigella).
 Composition- Peptone, lactose, agar, neutral red (indicator) and
taurocholate

23. Differential Media (Cont..)
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Differential media Features
CLED agar (cysteine lactose
electrolyte-deficient agar)
 Capable of differentiating between LF and NLF.
 Used as an alternative to combination of blood agar and
MacConkey agar, for the processing of urine specimens

24. Differential Media (Cont..)
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MacConkey agar CLED agar

25. Anaerobic Culture Media
▰ Contain reducing substances which take up oxygen and
create lower redox potential - permit the growth of obligate
anaerobes, such as Clostridium.
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Media Features
Robertson’s cooked meat (RCM) broth  Contains chopped meat particles (beef heart), which
provide glutathione (a sulfhydryl group containing
reducing substance) and unsaturated fatty acids.
 Widely used
 Used for maintenance of stock cultures.

26. Anaerobic Culture Media (Cont..)
Other anaerobic media include:
▰ Thioglycollate broth
▰ Anaerobic blood agar
▰ BHIS agar (Brain-heart infusion agar) with supplements (vitamin K and
hemin)
▰ Neomycin blood agar
▰ Egg yolk agar
▰ Phenyl ethyl agar
▰ Bacteroides bile esculin agar (BBE agar).
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27. Blood Culture Media
▰ Recovery of bacteria from blood is difficult - they are usually present in
lesser quantity and many of the blood pathogens are fastidious.
▰ Therefore, enriched media - used for isolating microorganisms from
blood.
▰ Available either as conventional or automated media
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28. Conventional Blood Culture
Media
Two types:
▰ Monophasic medium: Contains brain–heart
infusion (BHI) broth.
▰ Biphasic medium: Liquid phase containing BHI
broth and a solid agar slope made up of BHI agar
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29. Disadvantages of conventional
media
Subcultures are made manually - performed less-frequently (once a
day) as it is cumbersome.
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Broth Disadvantages
Monophasic BHI broth  Subcultures are made, periodically for 1 week.
 Higher risk of contamination - due to opening of the cap of the bottle every
time when subcultures are made.
Biphasic BHI broth  Subcultures - made just by tilting the bottles so that the broth runs over the
agar slope.
 Lower risk of contamination as it obviates the opening of the cap of the
bottle

30. Automated Blood Culture
Techniques
▰ Continuous automated monitoring: Blood culture bottles -
periodically monitored for the microbial growth - every 10 minutes
by the instrument.
▰ Composition: Tryptic soy broth and/or brain heart infusion broth +
polymeric resin beads - adsorb and neutralize the antimicrobials
present in blood specimen.
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31. Automated Blood Culture Techniques
(Cont..)
▰ Specimens: Used for culture of blood, bone marrow and sterile
body fluids.
▰ More sensitive: Higher yield of positive cultures from clinical
specimens
▰ Rapid: Less time than conventional methods
▰ Less labor intensive: fully-automated 31

32. Automated Systems
BacT/ALERT 3D:
▰ Principle - colorimetric detection of
growth.
▰ When bacteria multiply - produce CO2 -
increases the pH - changes the color of
a blue-green sensor present at the
bottom of the bottle to yellow 32

33. Automated Systems (Cont..)
BacT/ALERT VIRTUO (bioMerieux):
Advanced form of BacT/ALERT – has
several advantages, such as:
▰ Automatic loading and unloading of
bottles,
▰ Faster detection of growth,
▰ Determine volume of blood present in
bottle
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34. Automated Systems (Cont..)
BACTEC (BD Diagnostics)
▰ Principle - fluorometric detection of growth.
▰ Uses an oxygen-sensitive fluorescent dye present in the
medium.
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