Isolation of Pure Cultures

Staff should always work beside a lighted bunsen burner. This is the most convenient method of sterilising materials and also provides an up-draught which stops dust falling onto media and materials. 

A second method of isolating micro-organisms used less frequently is serial dilutions. The sample is diluted repeatedly in a sterile diluent until there is theoretically only one organism per tube. This is then grown to give a pure culture although the purity should be checked by plating out as described previously. 

In the case of an organism such as Bacillus, considerable purification can be achieved by boiling the sample, which would kill all organisms except the resistant spores of the genus Bacillus. 

---

The interactions between y-triazines and microorganisms have been studied over nearly 50 years and new research has led to important discoveries. The isolation of pure cultures that are able to modify or completely mineralize y-triazines has led to the discovery of new genes and enzymes that are involved in the degradation and mineralization of y-triazines by soil bacteria. Studies carried out in soils with a history of repeated y-triazine applications indicate that rapid degradation and mineralization of atrazine developed in various soils (Barriuso and Houot, 1996 Bradley et al, 1997 Pussemier et al, 1997). 

A third step in the study of the microbiology of enhanced degradation involved isolation of pure cultures of insecticide-... 

Janssen PH, PS Yates, BE Grinton, PM Taylor, M Sait (2002) Improved cultivability of soil bacteria and isolation in pure culture of novel members of the divisions Acidobacteria, Actinobacteria, Proteobac-teria, and Verrucomicrobia. Appl Environ Microbiol 68 2391-2396. 

The plaque procedure also permits the isolation of pure virus strains, since if a plaque has arisen from one virus particle, all the virus particles in this plaque are probably genetically identical. Some of the particles from this plaque can be picked and inoculated into a fresh bacterial culture to establish a pure virus line. The development of the plaque assay technique was as important for the advance of virology as was Koch s development of solid media for bacteriology. 

Comparison of evolved 14CO from four sterile and non-sterile soils 60 days after treatment with MSMA-l2tC showed that from 1.7 to 10.0% of the MSMA-14C was degraded in non-sterile soil, as compared with 0.7% in steam-sterilized controls. Four soil micro-organisms isolated in pure culture degraded... 

Tenske, A, Sigalevich, P, Cohen, Y, Muyzer, G. (1996). Molecular identification of bacteria from co-culture by denaturing gradient gel electrophoresis of 16S ribosomal DNA fragments as a tool of isolation in pure cultures. Appl Environ. Microbiol, 62, 4210-4215. 

The hemicellulases of rumen micro-organisms have been isolated from pure culture isolates of (a) bacteria, for example, Bacillus firmus,148 Bacteroides amylogenes,38 Butyrivibrio fibrosolvens,38,40 and Ruminococcus flavefaciens41 and (b) protozoa, for example, Epidinium ecaudatum,43-44 an Entodinium sp.,44,216 Eudiplodinium medium,48 Eremoplastron bovis,44,45 and Polyplastron mul-tivesiculatum.47 They have also been found in cell-free preparations of mixed bacteria,40,42,215 protozoa,43 and mixtures of both organisms.42,217,218,254,255... 

---