BRIEF INTRODUCTION ABOUT CULTURE MEDIAS

1. WELCOME………
Prepared by:
ANSAR AHMAD PARAY
BSC.MLT

2. Culture media
 These are required to isolate the bacteria from the
clinical specimen; following which the appropriate
biochemical tests can be performed to identify the
causative agent.

3. Composition of culture media
 Water: serves as source for hydrogen & oxygen.
 Electrolytes: NaCl or other electrolytes.
 Peptone : It is a source of readily available nitrogen and is
complex mixture of partially digested proteins.
 Buffers : maintain constant PH.
 Indicators : useful in detection of acid or alkali production
by microorganism. e.g. methyl red ,bromocresol purple,
neutral red etc .
 Selective agents : these are special chemicals introduced
in culture media for inhibiting some type of bacteria. E.g.
crystal violet inhibits staphylococcus but not TB .
 Meat extract: usually prepared from beef.

4.  Additive for enrichment : substances like sheep blood,
horse blood etc allow fastidious org. to grow
 AGAR:- it is used for solidifying the media. it is
commercially available in powder form, melts in water after
boiling and jellifies after cooling.
it is also called ‘‘agar-agar” and is prepared from cell
wall of seaweeds (red algae etc).it does not add nutrientive
property to media.
melting point :- 98oc
solidifies at :- 42oc
Concentration of agar:
For solid agar preparation : 1-2%
For semisolid agar : 0.5%
For solid agar to inhibit proteus swarming : 6%

5. Types of culture medias
 Classification
culture medias
Based on growth requirement. Based on
consistency.
1.liquid media (or
broth)
2.semisolid media
3. solid media.
Routine lab. Media Defined or
1.simple/basal media synthetic medias.
2.Enriched media
3.Enrichment media 1. simple synthetic media
4.Selective media 2. complex synthetic media
5.Differential media
6.Transport media
7.Anaerobic media

6. Routine laboratory media
 1. simple/basal media:
contain minimum incridents that support the growth of
non-fastidious bacteria.e.g
Peptone water; contains peptone (1%)+NaCl (0.5%)+water
Nutrient broth; peptone water+ meat extract(1%).{used for study of
bact. Growth curve}
Nutrient agar; nutrient broth+ 2% agar.
Uses of basal media:- used for
Testing the non-fastidous bacteria
serve as base for preparation of any other media

7. ENRICHED MEDIA
When a basal media is added with additional nutrients such as
blood,serum or egg it is called enriched media.
it supports growth of both fastidious and non-fastidious bacteria.
E.g
1.Blood agar: molten nutrient broth+ 5-10% sheep blood…it is most
widely used/.
2.Chocolate agar: it is heated blood agar prepared by adding 5-10%
of sheep blood to molten agar at 70oc so that rbc be lysed
changing color of media to brown.
3.Loefflers serum slope: contains serum and used for isolation of
corynebacterium diptheria.

8.  2.Enrichment broth:-
They are liquid medias added with some inhibitory
substances which selectively allow certain organisms to
grow and inhibit others.
..imp for isolation of contaminated specimen.
E.g , Gram-negative broth for shigella
selenite F broth for shigella
Alkaline peptone water(APW) for vibrio cholera
3.Selective media:-
they contain substances tata accelerate the growth
of required pathogen only and prevent the growth
of other org. E.g,
-- lowenstein-jensen(LJ) media- for M.tuberculosis
-- DCA(deoxycholate citrate agar) for salmonella
and shigella

9.  Transport media:-
They are used for the transport of the clinical specimen
suspected to contain delicate org. or when the delay is
expected transporting the specimen from site of collection
to laboratory.
--- bacteria don’t multiply in this media ,they only remain
viable.
Organism Transport media
streptococcus Pike’s medium
Neisseria Amines medium & Stuart's
medium
Vibrio cholera VR (venkatraman-
ramakrishnan) medium
Cary Blair medium
Shigella , salmonella Cary Blair medium

10.  Differential media:-
these media differentiate between two groups of bacteria
by using an indicator ,which changes color of the colonies
of particular group of bacteria but not he other group. E.g.
MacConkey Agar:- it is a diff. and low selective media
commonly used for isolation of gram negative bact.
-- it differentiate org.into LF or lactose fermenters (produce
pink color colonies e.g. E.coli) and NLF or non-lactose
fermenters (produce colorless colonies e.g shigella)
CLED agar ( cystein lactose electrolyte deficient agar)

12. Anaerobic culture media
 They contain reducing substances (e.g. Na
thioglycollate) which take up oxygen and create
low redox potential and thus permit the growth of
obligate anaerobes such as clostridiun . Examples
are as:-
 Robertson’s cooked meat (RCM) broth:-
it contains chopped meat particles (beef heart) which
provide glutathione (a reducing substance) and
unsaturated fatty acids. It is most widelu used
anaerobic culture medium.
It is also used for stock culture maintenance.

13.  Other anaerobic media include :-
 Thioglycollate broth
 Anaerobic blood agar
 BHIS agar (Brain-heart infusion agar) with
supplements (vitamin K and hemin ) .
 Neomycin blood agar
 Phenyl ether agar

14. Defined or synthetic media
 The media which contain know concentration of
components is called as synthetic media.
They are prepared exclusively from pure chemical
substances in such a way that their composition
i.e. exact quantity of each chemical used is known.
Simple synthetic media:-
They contain a Carbon and energy source such
as gulcose or lactose and an inorganic source of
nitrogen usually in form of ammonium chloride or
sulfate and various inorganic salts in a buffered
aqueous solution.
-- don’t support growth of fastidious bacteria

15.  Complex synthetic media:-
here in addition to simple synthetic media certain
amino acids ,purines ,pyrimidines and other growth
factors are added .
Hence they can also support growth of fastidious
bacteria.

16. Classification based on consistency
 Solid culture media:-
this is used in petri dishes and also in test tubes (slope
cultures )
agar conc. Is 1-2 % usually.
Microorganisms Grow and form colonies after
multiplication. This helps to identify organism.
semisolid cultures :-
this is prepared by adding agar 0.4-0.5 % (W/V) to a fluid
medium. These are mainly used as transport medias and
also for motality testing .
Liquid media/fluid culture media
no agar added
these are used as biochemical testing media ,blood culture
media or enrichment media

17. Cultural characteristics /
Morphology of bacterial colony The appearance of bacterial colony on culture
media is usually characteristic which helps in
bacterial identification. Following features of
colony are studied:-
 Size : in mm e.g. pin head size is characteristic of
staphylococcus and pin point of streptococcus.
 Shape : circular or irregular.
 Surface: Glistening or dull.
 Edge : entire, crenated, lobate or filamentous
 Elevation : flat ,raised,convex, unbonate

18.  Consistency: Mucoid ,friable, firm or butyrous.
 Density: opaque , translucent , transparent.
 Hemolysis on blood agar:
partial or α-hemolysis
complete or β-hemolysis
no Hemolysis (γ-hemolysis)
α prime Hemolysis: half incomplete lysis
surrounded by colonies with a second zone of
complete lysis.
Color of the coloney : coloney may be colored due to
propertied of media used due to pigment production .
MacConkey Agar:. it differentiate org.into LF
or lactose fermenters (produce pink color colonies e.g.
E.coli) and NLF or non-lactose fermenters (produce
colorless colonies e.g shigella).

21. Thanks………………..
BY: Ansar ahmad paray