Culture medium

DR SHARAD SHARMA

..

y culture medium is a liquid or gel designed to support

the growth of microorganisms , cells or small plants.

y culture media consist of nutrients used to grow micro

organism or cells outside of their natural habitats.

Basic requirements of culture media

y Energy source , (carbon source,nitrogen source) y Mineral salts ( sulphates phosphates and chlorides

of K, Mg nd Ca ) y pH - 7.2 to 7.4 y Accessory growth factors ( e.g , tryptophan for salmonella typhi )

Agar agar : most important constituents of culture media

y Agar- agar: It is used as solidifing agent , y contains long chain poly saccharides , inorganic y y y y

salt and proteins , solidify at 42c Melts at 98c Generally not metabolized by microbes, Obtained from sea weed (new zealand)

Types of culture media

     

Nutrient media Minimal media Selective media Differential media Enriched media Transport media

Nutrient media
 source of amino acids and nitrogen is , beef yeast or extract.
 Undefined medium ( amino acid source contains a

variety of compounds with the exact composition being unknown)  Nutrient media contain all the essential elements that most bacteria need for growth and are nonselective, so they are used for the general cultivation and maintenance of bacteria kept in laboratory culture collections.

Nutrient media

Minimal media
 Contains the minimum nutrients possible for

colony growth, without the presence of amino acids.  It is often used by microbiologists and geneticists to grow wild type microorganisms.  Minimal medium typically contains:  a carbon source (may be glucose or a less energyrich source like succinate)  various salts, which may vary among bacteria species and growing conditions; these generally provide essential elements such as magnesium, nitrogen, phosphorus, and sulfur to allow the bacteria to synthesize protein and nucleic acid  water

Selective media
 used for the growth of only select microorganisms. (e.g, if a microorganism is resistant to a certain antibiotic, such as ampicillin or tetracycline, then that antibiotic can be added to the medium in order to prevent other cells, which do not possess the resistance, from growing).
 Selective growth media are also used in cell culture

to ensure the survival or proliferation of cells with certain properties, such as antibiotic resistance or the ability to synthesize a certain metabolite.

Selective media

Some examples of selective media include:

y Eosin-methylene blue agar (EMB) contains methylene blue

y y y y y y

toxic to Gram-positive bacteria, allowing only growth of Gram negative bacteria Yeast and mold has a low pH, deterring bacterial growth blood agar contains bovine heart blood that becomes transparent in the presence of hemolytic Streptococcus MacConkey agar for Gram-negative bacteria Hektoen enteric agar (HE) which is selective for Gramnegative bacteria mannitol salt agar (MSA) which is selective for Gram-positive bacteria and differential for mannitol xylose lysine desoxyscholate , is selective for Gram-negative bacteria

Differential media
Differential media or indicator media distinguish one microorganism type from another growing on the same media.]  This type of media uses the biochemical characteristics of a microorganism growing in the presence of specific nutrients or indicators (such as neutral red, phenol red, eosin y, or methylene blue).  Examples of differential media include: y eosin methylene blue, which is differential for lactose and sucrose fermentation y MacConkey, which is differential for lactose fermentation y X-gal plates, which are differential for lac operon mutants

Enriched media
 Enriched media contain the nutrients required to

support the growth of a wide variety of organisms, including some of the more fastidious ones.  They are commonly used to harvest as many different types of microbes as are present in the specimen.  Blood agar is an enriched medium in which nutritionally rich whole blood supplements the basic nutrients.  Chocolate agar is enriched with heat-treated blood (40-45C), which turns brown and gives the medium the color for which it is named.

Transport media
y Transport media is used fortemporary storage of

y y y y y

specimens being transported to the laboratory for cultivation. maintains the viability of all organisms in the specimen without altering their concentration. contain only buffers and salt. lack of carbon, nitrogen, and organic growth factors prevents microbial multiplication. transport media used in the isolation of anaerobes must be free of molecular oxygen. Example: Thioglycollate broth for strict anaerobes, Stuart transport medium-a non-nutrient soft agar gel containing a reducing agent to prevent oxidation, Venkat Ramakrishnan medium for v. cholerae

Sterilization of culture media

y Culture media are sterlized at : y Temperature 121c, y Pressure 15 pascals, y Time 15 t0 20 minutes, y media containing heat sensitive substances like

serum , fatty acid ,sugars are sterlized by free steam or filtration.

Storage of culture media

y Individual media is stored in screw capped bottles . y Culture media is preserved at low temperature (in

refrigeratirs or cold rooms)

y Deep freeze refrigerators are used for preservation

of media containing antibiotics and amino acids ( 10 to -400c).

Refrences
y Madigan M, Martinko J (editors). (2005). Brock Biology of y

y y

Microorganisms (11th ed.). Prentice Hall. ISBN 0131443291. Birgit Hadeler, Sirkka Scholz, Ralf Reski (1995) Gelrite and agar differently influence cytokinin-sensitivity of a moss. Journal of Plant Physiology 146, 369-371 Ryan KJ, Ray CG (editors) (2004). Sherris Medical Microbiology (4th ed.). McGraw Hill. ISBN 0838585299. Cooper GM (2000). "Tools of Cell Biology". The cell: a molecular approach. Washington, D.C: ASM Press. ISBN 0-87893-106-6. http://www.ncbi.nlm.nih.gov/books/bv.fcgi?&rid=cooper.section.1 28#148. Washington JA (1996). "Principles of Diagnosis". Baron's Medical Microbiology (Baron S et al., eds.) (4th ed.). Univ of Texas Medical Branch. ISBN 0-9631172-1-1. http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=mmed.section.54 51#5457