THE COMMON COMPONENTS OF CULTURE MEDIA FOR MEDICAL MICROBIOLOGY
An examination of the formulae of culture media commonly shows a few key nutrient ingredients, and several other components added for the selection and/or identification of separate groups or species of bacteria.
All life is dependent on the presence of water and all the nutrients from which micro-organisms synthesize cell material and obtain energy must be dissolved in water.
Simple chemical analysis of bacterial cells reveals 11 macro elements – C, H, O, N, S, P, K, Na, Ca, Mg, Fe – and many trace elements that are often found as impurities in the macro elements, e.g. Mn, Mb, Zn, Cu, Co, Ni, V, B, Se, Si, W.
This simple analysis is not very helpful in the construction of specific culture media and it is better to divide the constituents into one or more functional parts.
Protein hydrolysates (peptones) infusions or extracts: these nutrients often contain sufficient energy-rich molecules and trace elements to become the sole ingredient necessary for the growth of common medical bacteria.
This is usually glucose but other easily used carbohydrates may be substituted. Peptones can also provide carbon energy, especially those derived from plants.
Some more fastidious organisms appear to require heat-labile supplements to stimulate growth, e.g. blood, serum and vitamin complexes.
Further work has shown that, with Bordetella, Neisseria, Campylobacter and Legionella species, the essential role of blood is to act as a protective agent against toxic oxygen radicals rather than act as a supplementary nutrient.
Soluble sodium or magnesium phosphate, acetate or citrate salts are commonly added to culture media containing carbohydrates, to maintain pH stability when fermentative organisms are growing.
These buffer salts, however, can chelate essential metals, especially Fe2+ ions (Munro, 1968).
Some growth factors are toxic (e.g. fatty acids) and must be added in a form or composition (e.g. with albumin or as a detergent) to permit growth and survival.
Mineral salts and metals
Supplements of salts and metals are normally restricted to synthetic (defined) culture media.
Undefined media should provide an adequate supply unless extra is required to overcome a chelating substance in the formulation or an excess of NaCl is added to select halophilic organisms.
Toxic chemicals, antibiotics and inhibitory dyes have been used separately or in combination in culture media. It is essential that the selective agents are at the correct strength for the particular medium formulation and the selected organism.
There is an element of compromise for all selective media, i.e. not all unwanted organisms will be inhibited and not all desired organisms will grow (Miller and Banwart, 1965; Bridson, 1978).
The active concentration of an antibacterial agent (e.g. Hg2+, antibiotic) may be influenced by the components of the medium (e.g. by albumin).
Dyes such as phenol red, neutral red and bromocresol purple are added to culture media to indicate changes in pH value, during and after growth.
Some dyes are colourless (e.g. triphenyltetrazolium chloride) and act as electron acceptors, and their reduced forms (e.g. triphenylformazan) are pigmented (red) and insoluble, resulting in dye deposition around colonies in agar medium or red colour in broths as an indication of growth.
Fermentative carbohydrates are normally added to the formulation.
The dyes used can be toxic to sensitive or stressed cells and this fact needs to be kept in mind, if growth is absent.
Agar is the most common gelling agent used in culture media because it has natural advantages over gelatin and alginate.
It is not an inert constituent of culture media; it can contribute metals, minerals, sulphate and pyruvate.
Agar can also bind water and inhibit growth of organisms.