culture media

2. o For identification of bacteria and diagnosis of disease
o To know the morphology and colony characteristics of bacteria
Eg. Cocci in bunch – Staphylococcus aureus
o For study of bacterial population rather then one bacteria
Why we need culture media?

3.  Media for cultivation of microorganism contain the
substance necessary to support the growth of mocrobs
 The growth of bacteria is dependent on an adequate
supply of suitable nutrient material
 Culture media provide
Water, Carbon,
Energy, Nitrogen,
Trace element Growth factor

4.  The more commonly used ingredients of bacteriological
media are
Meat extract (Source of CHO),
Peptone (Protein and Amino Acid)
Salt
Agar is generally used as a solidifying agent.
(Source-Bluish green Algae)
Gelatin and Silica gel also use for solidify

5. Preparation of Media:-
There are two ways of preparing bacterial culture media viz.
From basic Ingredients.
(a) Mixing the ingredients.
When preparing solid media,
first add the salts and other soluble ingredients like peptone,
beef extracts etc.
 Then adjusting pH, agar is added.

6. (b) Adjustment of pH
Pathogenic bacteria grow best at the pH range of 7.2 to 7.4.
 To make medium alkaline N/10 NaOH and to make it acidic
N/10 HCL is added.
1. pH indicator paper:
2. Colorimetric method:.
3. Electrometric method:

7. (C) Filtering of medium :
(D) Tubing of media :
(e) Plugging of tubes / flakes :
(f) Sterilization :

8. Types of culture media
I. Based on their consistency
a) solid medium
b) liquid medium (Broth medium)
c) semi solid medium
II. Based on the constituents/ ingredients
a) simple medium
b) complex medium
c) synthetic or defined medium
d) Special media

9. Special media
 Enriched media
 Enrichment media
 Selective media
 Indicator media
 Differential media
 Sugar media
 Transport media
 Media for biochemical reactions
III.Based on Oxygen requirement
- Aerobic media
- Anaerobic media

10. Basic Nutrient Media (Simple Media)
 They contain only basic nutrients and there is no addition of
any selective or enriched ingredients.
1) Peptone water:
 Composition: Peptone 10g
Sodium chloride 5g
Distilled water 1 lit
pH 7.6
 Sterilization: In Autoclave

11. 1) Peptone water:
 Composition: Peptone 10g
Sodium chloride 5g
Distilled water 1 lit
pH 7.6
 Sterilization: In Autoclave (15 lb/sq. inch, 127 C
for 20 minutes)
 Use: To prepare nutrient broth and nutrient agar, for
Indole test, basal medium for preparation of bacterial
culture.

12. 2) Nutrient broth:
 Composition: Add 10 gm of Meat extract in 1 lit. of
peptone water. pH: 7.6

13. 3) Nutrient agar:
 Composition:
 Add 20-25 gm of agar in 1 lit of nutrient broth. pH.7.6
(2 to 2.5% Agar use)
 Sterilization: In Autoclave.

14. 4) Robertson’s cooked meat medium (RCM):
Composition:
 Small muscle pieces of bullock heart up to 1” of test tube
and add 10 ml of nutrient broth.
 Sterilization: In Autoclave.
 Use: For cultivation of anaerobic bacteria.
e.g. Clostridia.

15. 5) Dorset’s egg medium:
 Composition: Fresh egg contents 75 ml
Nutrient broth 25 ml
2 % malachite green 1.25 ml
 Sterilization: In Inspissator.
 Use: For cultivation of mycobacteria.

16. B. Enriched media:
 It is a medium which is able to enhance the growth of particular
organism without inhibiting the growth of other organism.
Blood, serum etc. are use in medium for better or enhance
growth of microorganism.
e.g. Blood agar, serum agar, serum glucose agar, Glucose broth.
Blood agar:
 It is prepared by adding 5-10% sterile defibrinated blood in
melted nutrient agar at 55 C temperature or below.
Use:
 For growing pathogenic organism like Pasturella multocida,
streptococcus spp. and Corynabacterium pyogenes and also for
observing haemolysis.

17. (Enriched Media)

18. C. Differential Media:
 Media containing some ingredient which allows to
distinguish between different group of bacteria
Ex:- Mac Conkey agar:
Use:
 differentiate between lactose fermenting (E.coli) and non
lactose fermenting (Salmonella) organism

19. (Mac Conkey for E.coli)

20. D) Selective media:
The media contain components that select for
the growth of specific organism and inhibiting
the growth of other organism
Brilliant green agar:
 Use: For selective isolation of Salmonella other than
Salmonella typhi.
Eosin Methylene Blue agar (EMB):
 Use: For selective isolation of E.coli.

21. (Eosin Methylene Blue Agar)

22. E) Enrichment Media
 These are normally liquid media which facilitate better or
enhance growth of required organism and inhibit the growth
of other type organism.
Tetrathionate broth and selenite broth for isolation of
Salmonella.

23. F) Media for enhancement of pigmentation
 Potato slopes, mannitol yeast extract agar.
G) Media for studying breakdown of carbohydrate
 Sugar media, Glucose Phosphate Peptone water (GPPW)

24. H) Miscellaneous Media
 Citrate agar: to observe the citrate utilization.
 Urea agar: for hydrolysis of urea.
 Gelatin agar: to study hydrolysis of gelatin.

25. I) Media for Fungus:
Sabouraud’s Dextrose Agar (SDA):
 Composition: Peptone 1g
Dextrose or Maltose 4g
Agar 2g
Distilled water 100ml
pH 5.4
 Sterilization: In autoclave.
 Use: The pH of the medium is acidic and is suitable for
cultivation of yeast and fungi.

26.  Muller Hinton Agar= Antibiotic Sensitivity test

27. I) Special Media:
1) Edward’s medium:
 Sterilization: In Autoclave.
 Use: For selective and rapid isolation of Streptococcus
agalactiae and other Streptococcus associated with
bovine mastitis.
2) Potato dextrose Agar:
 Sterilization: In Autoclave.
 Use: For isolation of yeast and mould.

28. 3) Simmens Citrate Agar:
 Sterilization: In Autoclave.
 Use: Recommended for differential faecal coli member
arranged group of the basis of citrate utilization.
4) Basal Medium:
 Sterilization: In Autoclave.
 Use: Determinate the Gram–ve organism, on the basis of
fermentative and oxidative metabolism and
carbohydrates.

29. 5) Plate count Agar:
 Sterilization: In Autoclave.
 Use: For determine plate count if in milk of dairy
products by pour plate technique.
6) Tryptose phosphate broth:
 Sterilization: In Autoclave.
 Use: For cultivation of fastidial bacteria.

30. 7) Mannitol Salt Agar base:
 Sterilization: In Autoclave.
 Use: For selective isolation of staphylococci.
8) Egg Yolk Agar:
 Sterilization: In Autoclave.
 Use: Isolation of Clostridial organisms.
9) PPLO broth Agar:
 Use: For isolation and multiplication of mycoplasma.

31. Solid media – contains 2% agar
 Colony morphology, pigmentation, hemolysis can be
appreciated.
 Eg: Nutrient agar, Blood agar
Liquid media – no agar.
 For inoculum preparation, Blood culture, for the
isolation of pathogens from a mixture.
 Eg: Nutrient broth
Semi solid medium – 0.5% agar.
 Eg: Motility medium

33. Simple media / basal media
- Eg: NB, NA
- NB consists of peptone, meat extract, NaCl,
- NB + 2% agar = Nutrient agar

34. Complex media
 Media other than basal media.
 They have added ingredients.
 Provide special nutrients
Synthetic or defined media
 Media prepared from pure chemical substances and its
exact composition is known
 Eg: peptone water – 1% peptone + 0.5% NaCl in water

35. Enriched media
 Substances like blood, serum, egg are added to the
basal medium.
 Used to grow bacteria that are exacting in their
nutritional needs.
 Eg: Blood agar, Chocolate agar

36. Blood agar Chocolate agar

37. Enrichment media
 Liquid media used to isolate pathogens
from a mixed culture.
 Media is incorporated with inhibitory
substances to suppress the unwanted
organism.
 Eg:
 Selenite F Broth – for the isolation of
Salmonella, Shigella
 Alkaline Peptone Water – for Vibrio
cholerae

38. Selective media
 The inhibitory substance is added to a solid media.
Eg:
 Mac Conkey’s medium for gram negative bacteria
 TCBS – for V.cholerae
 LJ medium – M.tuberculosis
 Wilson and Blair medium – S.typhi
 Potassium tellurite medium – Diphtheria bacilli

39. TCBS
Mac Conkey’s medium

40. Potassium Tellurite media LJ media

41. Sr. no Organism Selective media
1 Staphylococcus spp. •Mannitol salt agar,
•Baired parker media
2 Streptococcus spp. •Sodium azide,
•Crystal violet blood agar
3 Bacillus anthracic •Polymyxin lysozome EDTA
thallium acetate
4 Clostridium spp. •Robertson’s cooked meat medium
5 Corynebacterium spp. •Potassium tellurite blood agar
6 Actinomyces spp. •Brain heart infusion agar (BHIA)
7 Mycobacterium laprae •Foot pad of mice and nine
branded armadilla

42. Sr no Organism Selective media
8 Mycobacterium spp. •L.J. medium
(Lowenstain jensen)
•Doset egg medium
•Stone brink’s medium
9 E.coli •EMB agar
10 Salmonella spp. •Deoxycholate citrate agar
11 Shigell spp. •Deoxycholate citrate agar
12 Listeria •Potassium tellurite in tryptose agar
13 Brucellosis •Trypticase soya agar
•Farrle’s medium
14 Haemophillus spp. •Levinthols agar medium
•Chocalate agar medium

43. Sr no. Organism Selective media
15 pseudomonas •KCN broth medium
16 Campylobacter •Thiol agar
17 Leptospira •Stuart’s medium
•EMJH medium
•Semberg’s medium
•Dinger’s medium
18 Borrelia spp. Egg embryo media
19 Morexella Horse blood agar