Barnard Health Care

Growth media for the cultivation of bacteria

A synthetic growth medium may be defined, that is, its exact chemical composition is known, or undefined. A defined growth medium may have few or many constituents, depending on the nutritional requirements of the organism in question. Examples of each are given in Table 4.3. An undefined or complex medium may have a variable composition due to the inclusion of a component such as blood, yeast extract or tap water (Table 4.4). Peptones are also commonly found in complex media; these are the products of partially digesting protein sources such as beef or casein. The exact composition of a complex medium is neither known nor critically important. A medium of this type would generally be chosen for the cultivation of fastidious bacteria such as Neisseria gonorrhoeae (the causative agent of gonorrhoea); it is easier and less expensive to supply the many nutrients required by such an organism in this form rather than supplying them all individually. Bacteria whose specific nutrient requirements are not known are also grown on complex media.

Whilst media such as nutrient agar are used to support the growth of a wide range of organisms, others are specifically designed for the isolation and identification of particular types. Selective media such as bismuth sulphite medium preferentially support the growth of particular bacteria. The bismuth ion inhibits the growth of Gram-positive organisms as well as many Gram-negative types; this medium is used for the isolation of the

A defined medium is one whose precise chemical composition is known.

An undefined or complex medium is one whose precise chemical composition is not known.

A fastidious organism is unable to synthesise a range of nutrients and therefore has complex requirements in culture.

A selective medium is one that favours the growth of a particular organism or group of organisms, often by suppressing the growth of others.

MICROBIAL NUTRITION AND CULTIVATION Table 4.3 Defined growth media

(a) Medium for Acidithiobacillus ferrooxidans

FeSO4.7H2O 40 g

(NH4)2SO4 2g

KH2PO4 0.5 g

MgSO4.7H2O 0.5 g

KCl 0.1g

Ca(NO3)2 0.01g

Distilled H2O (pH 3.0) to 1 litre

(b) Medium for Leuconostoc mesenteroides

100 mg 100 mg 50 mg 200 mg 40 mg 100 mg 250 mg 10 mg 10 mg 10 mg 10 mg 1 mg 1 mg 0.5 mg 0.5 mg 0.5 mg 0.3 mg 0.3 mg 0.1 mg 1 Mg 10 Mg

Distilled H2O to 1 litre

(b) Medium for Leuconostoc mesenteroides

Glucose	25 g	Phenylalanine
Sodium acetate	20 g	Proline
NH4Cl	3g	Serine
KH2PO4	0.6 g	Threonine
K2HPO4	0.6 g	Tryptophan
NaCl	3g	Tyrosine
MgSO4 • 7H2O	0.2 g	Valine
MnSO4 • 4H2O	20 mg	Adenine
FeSO4.7H2O	10 mg	Cytosine
Alanine	200 mg	Guanine
Arginine	242 mg	Uracil
Aspartic acid	100 mg	Nicotinic acid
Asparagine	400 mg	Pyridoxine
Cysteine	50 mg	Riboflavin
Glutamic acid	300 mg	Thiamine
Glycine	100 mg	Ca pantothenate
Histidine	62 mg	Pyridoxamine
Isoleucine	250 mg	Pyridoxal
Leucine	250 mg	p-Aminobenzoic acid
Lysine	250 mg	Biotin
Methionine	100 mg	Folic acid

Distilled H2O to 1 litre

Examples of defined (synthetic) media for (a) the iron-oxidising bacterium Acidithiobacillus ferrooxidans and (b) the lactic acid bacterium Leuconostoc mesenteroides. Note how L. mesenteroides must be provided with numerous amino acids, nucleotides and vitamins as well as glucose as a carbon source, whereas A. ferrooxidans requires only mineral salts, including reduced iron to act as an energy source.

Table 4.4 Composition of an undefined growth medium

Calf brain infusion 200 g

Beef heart infusion 250 g

Proteose peptone 10 g

Glucose 2 g

NaCl 5 g

Na2HPO4 2.5 g

Brain heart infusion broth contains three undefined components. It is used for the culture of a wide variety of fastidious species, both bacterial and fungal.

A differential medium allows colonies of a particular organism to be differentiated from others growing in the same culture.

pathogenic bacterium Salmonella typhi, one of the few organisms that can tolerate the bismuth. Specific media called differential media can be used to distinguish between organisms whose growth they support, usually by means of a coloured indicator. MacConkey agar contains lactose and a pH indicator, allowing the differentiation between lactose fermenters (red colonies) and non-lactose fermenters (white/pale pink colonies). Many media act both selectively and differentially; MacConkey agar, for example, also contains bile salts and the dye crystal violet, both of which serve to inhibit the growth of unwanted Gram-positive bacteria. Mannitol salt agar is also both selective and differential. The high (7.5 per cent) salt content suppresses growth of most bacteria, whilst a combination of mannitol and an indicator permits the detection of mannitol fermenters in a similar fashion to that just described. Sometimes, it is desirable to isolate an organism that is present in small numbers in a large mixed population (e.g. faeces or soil). Enrichment media provide conditions that selectively encourage the growth of these organisms; the use of blood agar in the isolation of streptococci provides an example of such a medium. Blood agar can act as a differential medium, in allowing the user to distinguish between haemolytic and non-haemolytic bacteria (see Chapter 7).

If we are to culture microorganisms successfully in the laboratory, we must provide appropriate physical conditions as well as providing an appropriate nutrient medium. In the next chapter, we shall examine how physical factors such as pH and temperature influence the growth of microorganisms, and describe how these conditions are provided in the laboratory.

An enrichment culture uses a selective medium to encourage the growth of an organism present in low numbers.

Was this article helpful?

0 0