An experimental approach for identifying microorganisms involved in specified functions: utilisation for understanding a nematode suppressive soil

An experimental approach was developed for identifying microorganisms involved in specified functions such as pathogen suppressiveness. In this approach, it was postulated that the microorganisms involved in pathogen suppressiveness could be discovered by identifying those organisms whose populations positively correlate with high levels of suppressiveness. The approach has three phases. The first phase is to identify bacterial and fungal rRNA genes ( rDNA) from soils possessing various levels of suppressiveness. Ribosomal DNA sequences that are more abundant in the highly suppressive soils than in the less suppressive soils are considered candidate sequences. A method termed oligonucleotide fingerprinting of rRNA genes (OFRG) is used to obtain extensive analysis of microbial community composition. The second phase of this experimental approach is to verify the results obtained from phase one using quantitative PCR. Here, selective PCR primers for each of the candidate rDNA sequences are designed. These primers are then used to determine the relative amounts of the candidate sequences in soils possessing various levels of suppressiveness produced by several different methods such as mixing various quantities of suppressive and fumigation-induced non-suppressive soil, biocidal treatments and temperature treatments. In phase three, the organisms that consistently correlate with suppressiveness are isolated and amended to non-suppressive soils to assess their abilities to produce suppressiveness. The utility of this experimental approach was demonstrated by using it to identify microorganisms involved in suppressiveness against the plant-parasitic nematode, Heterodera schachtii. This general experimental approach should also be useful for the identifying microorganisms involved in functions other then pathogen suppressiveness.