culture media

1. Culture media
Dr Avinnanda Biswas

2. Content
Definition
Uses of culture media
Basic composition of culture media
Types of culture media
Based on physical state
1. solid medium
2. semi solid medium
3. liquidmedium
Based on ingredients
1. Simple or basal medium.
2. Complexmedium.
3. Synthetic or defined medium.
4. Semisynthetic medium.
5. Special medium
Media preparation

3. Definition
🞄The food material or substances required for growing
microorganisms in vitro (outside the body) is called culture
medium

4. Uses of culture
media
🞄It is important to grow microorganisms outside the body for the
following purposes:
🞄1. to identify the cause of infection from the clinical sample, so
that proper treatment can be given.
🞄2. to study the characteristics or properties of microorganisms.
🞄3. to prepare biological products like vaccines, toxoids,
antigens…etc.

5. Basic
ingredient of
culture
media
WATER- source of hydrogen and oxygen.
Pepton
- Complex mixture of digested protein
- It contains proteoses, aminoacids, polypeptides, phosphates, minerals
and accessory growth factor like nicotinic acid and riboflavin
Electrolytes : sodium chloride

6. AGAR
- It is prepared for using solid media.
- Obtained from sea weeds.
- Long chain polysaccharides.
- Also contains varying amounts of inorganic salts and small
quantities of a protein – like substance.
- Hydrolysed at high temperature at high acid or alkaline ph.
- Melts at 98ºC and usually sets at 42ºC depending on agar
concentration.
- Approximately 2% agar is used to prepare solid media.
Blood or serum
-These are used for enriching culture media

7. Types of
culture media
🞄I.Classification based on physical state
a) solid medium
b) semi solid medium
c) liquid medium
🞄II.Classification based on the ingredients
a) simple medium
b) complex medium
c) synthetic or defined medium
d) Special media

8. Based on
physical state
Solid medium
agar is the most commonly used solidifying agent
🞄 Melts at 98 °C & sets at 42 °C

9. 🞄 Semi-solid media
Such media are soft and are useful in
demonstrating bacterial motility and separating
motile from nonmotile strains
🞄 Liquid media
are sometimes referred as “ broth “.
bacteria grow uniformly producing general
turbidity e.g. Nutrient broth

10. Based on the
ingredients
🞄Simple media
eg: Nutrient broth, glucose broth,Nutrient agar
- NB consists of peptone, meat extract, NaCl
- NB + 2% agar = Nutrient agar
- Simplest and routinely employed method
🞄Complex media
such as blood agar, it has ingredients that exact components are
difficult to estimate.

11. Synthetic or defined media
•specially prepared media from pure chemical substances for
research purpose and composition of every component is well
known
• egg: peptone water – 1% peptone + 0.5% NaCl in water.

12. Special media
- Enriched media
- Enrichmentmedia
- Selective media
- Differential media
- Indicator media
- Transport media
- Anaerobic media

13. Special media
Enriched media
- Substances like blood, serum, egg are added to the simple
medium.
- Used to grow bacteria that are exacting in their nutritional needs.
Blood agar Chocolate agar

14. Blood agar
contains mammalian blood(usually sheep or horse)
typically at a concentration of 5-10%,
used to isolate fastidious organisms and
detect haemolysis
Chocolate agar
contain red blood cells that have been lysed by
slowly heating to 80 c .and it used for growing
fastidious bacteria;
such as Haemophilus influenzae

15. Enrichment
media
🞄Has stimulating effect on bacteria to be grown or inhibits its
competitors
🞄Results in absolute increase in number of wanted bacteria related
to other bacteria
🞄Useful for culture of faeces where non-pathogenic being
overgrown by pathogenic. Eg Salmonella being overgrown by E.
coli
🞄Eg.Tetrathionate broth and alkaline peptone water

16. Selective
media
Isolates particular bacteria
The inhibitory substance is added to a solid media to inhibit
commensal or contaminating bacteria such as :
- Antibiotics, Dyes,Chemicals
Examples
🞄Deoxycholate citrate agar (DCA) : for enteric bacilli ( Salmonella,
Shigella)
🞄Bile salt agar : for vibrio cholerae

17. Eosin methylene blue
- selective for gram negative bacteria
-The dye methylene blue in the medium
inhibit the growth of gram positive bacteria.
Campylobacter agar
- Is used for isolation ofCampylobacter jejuni from
faecal or rectal swab.
-Contain Bacteriological charcoal ,Cefoperazone
andAmphotericin B.

18. Lowenstein –Jenson medium
- is solid medium used for Mycobacterium tuberculosis.
- contain penicillin, nalidixic acid and malachite green to inhibit
growth of gram positive and gram negative bacteria, in order to
limit growth to Mycobacteria species only

19. Differential
media
- are designed in such a way that different bacteria can be
recognized on the basis of their colony colour.
- Dyes and metabolic substrates are incorporated so that those
bacteria that utilize them appear as differently coloured colonies.
🞄Examples:
- MacConkey agar
-TCBS agar
- XLD agar

20. MacConkey medium
- Distinguish between lactose fermenters
& non lactose fermenters.
- Lactose fermenters – Pink colonies
- Non lactose fermenters – colourless colonies
Xylose Lysine DeoxycholateAgar(XLD)
- Used forSalmonella and
Shigella species.

21. Thiosulfate-citrate-bile salts-sucrose agar(TCBS)
- highly selective for the isolation ofV. cholerae andV.
parahaemolyticus

22. Indicator
media
🞄Contains an indicator which changes color when a bacterium
grows in them.
🞄Salmonella typhi grow as black colonies on Wilson and blair
medium containing sulphite
🞄 MacConkey’s agar : pink colonies in presence of neutral red
indicator

23. Transport
media
- Media used for transporting the samples.
- Delicate organisms may not survive the time taken for
transporting the specimen without a transport media.
• Eg –Stuart’s medium : reducing agent (prevents oxidation),
charcoal (neutralise bacterial inhibitor), used for gonococci
- Buffered glycerol saline : enteric bacilli

24. Sugar media
🞄 Contains 1% of sugar in peptone water along with an indicator
(Andrade’s indicator-0.005%)
🞄 a small tube(durham’s tube) is kept inverted in the larger sugar
tube to detect gas production

25. Anaerobic
media
🞄These media are used to grow anaerobic organisms.
- Eg: Robertson’s cooked meat medium.
Thioglycolate broth medium

26. Dehydrated
media
🞄Simply reconstituted in distilled water and sterilised before use
🞄With dehydrated media, the process of media making has become
simpler and its quality more uniform.

27. Culture
methods
🞄Streak culture
🞄Lawn culture

28. Streak culture
It is the most common inoculation method ; used for the inoculation of the
specimens on to the solid media . It is also used for obtaining individual isolated
colonies from a mixed culture of bacteria .

29. Lawn culture
🞄Employed in antibiotic sensitivity testing and in bacteriophage
typing
🞄Lawn culture is obtained by flooding the surface of the plate with
a liquid culture or suspension of bacterium
🞄Culture plate is kept for minute and the excess material is poured
off
🞄Alternatively, the culture plate may be inoculated by a sterile swab
soaked in liquid bacterial culture
🞄Plate is then incubated at 37C overnight to obtain bacterial
colonies

30. Thank you!