Ch. 8— Maintaining Microbial Diversity • 209 



once obtained, are kept as pure cultures. Micro- 

 organisms have been isolated to study their in- 

 j terrelationships and the way the dynamics be- 

 ' tween populations influence the entire biologi- 

 cal food chain. Some collections represent 

 sampling of specific taxonomic classes of 

 micro-organisms of economic or agricultural 

 importance. The actinomycete collection of the 

 Battelle-Kettering Laboratory (Yellow Springs, 

 OH] and the many collections of varying sizes 

 of Rhizohium, the nitrogen-fixing bacteria of 

 legumes, are examples of goal-directed collec- 

 tions. The pathogenic characteristics of a 

 micro-organism, as in the case of a disease- 

 producing virus, can also merit spending funds, 

 time, and expertise to isolate it. 



Isolated pure cultures of micro-organisms are 

 necessary for detailed study (7). For some, such 

 as the fungi, a sterile culture of spores on a spe- 

 cially prepared medium may be all that is nec- 

 essary to obtain a pure culture. Nutritional re- 

 quirements for various fungi can, however, be 

 specific and difficult to determine. Most bac- 

 teria must be cultured on a variety of media 

 that will stimulate growth of possible contami- 

 nants, from which pure culture can then be ob- 

 tained. Viruses are frequently isolated from in- 

 fected cells or tissues by centrifugation or 

 filtration techniques that separate them from 

 other cellular components. For micro-organisms 

 that consist only of a small piece of genetic ma- 

 terial, such as viroids, the newly developed 

 technologies for isolating, multiplying, and 

 characterizing DNA and RNA have been impor- 

 tant. The critical determination that an isolated 

 micro-organism is pure can be a lengthy proc- 

 ess of repeated culture or separation under 

 varying conditions and can be a research prob- 

 lem in itself (7). 



Studies of microbial ecology and microbial 

 diversity are limited by the inability of scien- 

 tists at the present time to isolate many micro- 

 organisms (19). Identification, for example, gen- 

 erally requires growth in pure culture to allow 

 for nutritional and physiological testing. It is 

 not currently possible to acquire a knowledge 

 of the total microbial diversity in any one envi- 

 ronment in a readily definable time period be- 

 cause of this inability to isolate, culture, and 



characterize every (or even most) micro-orga- 

 nism present. Thus, sampling of diversity is 

 limited to those micro-organisms for which iso- 

 lation and culture technologies are available. 



Microbial Identification 



Identification of isolated micro-organisms 

 can be a lengthy and complex task (for details 

 of the principles and procedures, see ref. 15). 

 Preliminary identification involves standard 

 staining procedures and microscopic examina- 

 tion. Analysis of the results of these initial ex- 

 aminations requires extensive knowledge of 

 micro-organisms and the general characteris- 

 tics of various taxonomic groups. Information 

 regarding the source of the isolate can also play 

 an important role at this stage. 



Following preliminary identification, the 

 micro-organism is subjected to more detailed 

 analysis, frequently consisting of examination 

 of growth characteristics on varying substrates 

 and under varying environmental conditions. 

 These tests establish specific physiological 

 characteristics that aid identification. The gen- 

 eral protocol is to work with a pure culture and, 

 using selected tests, narrow the range of possi- 

 bilities. Once identified, the isolate is then com- 

 pared to a reference sample using selected diag- 

 nostic tests (24). 



Biochemical analysis of proteins and DNA, 

 as described for plants (see ch. 7), has been used 

 for identification of many strains of micro- 

 organisms (7). Although these technologies rou- 

 tinely identify the micro-organisms used in re- 

 search laboratories, they are not generally ap- 

 plied in offsite collections. As the field of 

 genetic engineering has developed, however, 

 the capacity to study, compare, and identify the 

 genomes of micro-organisms has improved 

 (10,31). Such techniques could greatly enhance 

 assessment of diversity and facilitate identifi- 

 cation of micro-organisms in offsite collections. 



Storage off Micro-Organisms 



The purpose of preserving micro-organisms 

 is to maintain a strain for an indefinite period 

 in a viable state. The continuous culture of a 



