5. Cultivation of M. plutonius

For many experiments, it is imperative to use bacterial cultures in which all cells are genetically identical. Since all cells in a colony develop from one single cell, a single isolated colony of M. plutonius is an excellent source of a genetically pure bacterial stock. In order to propagate any bacterium, it is necessary to provide the appropriate biochemical and biophysical environment to encourage bacterial growth. The biochemical or nutritional environment is provided as a culture medium based on special needs for particular bacteria, and can be used for isolation and maintenance of bacterial cultures. Bacterial culture media can be classified based on consistency. Liquid media are sometimes referred to as “broths” where bacteria grow uniformly, and tend to be used when a large quantity of bacteria have to be grown. Moreover, liquid media can be used to obtain a viable bacterial count i.e. to physically quantify the amount of organism present (see section 9.1.1). Any liquid media can be solidified by the addition of agar (e.g. Oxoid Technical Agar No. 1) at a concentration of 1-3 %. Although M. plutonius can be isolated from honey and diseased brood by cultivation, bacterial culture methods seem to be very insensitive detecting less than 0.2 % of the bacterial cells (Djordjevic et al., 1998; Hornitzky and Smith, 1998).