MOLECULAR MICROBIAL ECOLOGY APPROACHES APPLIED TO DIAZOTROPHS ASSOCIATED WITH NON-LEGUMES

Species- and genus-specific oligonucleotide probes, as well as monoclo nal antibodies and polyclonal antisera, were used to study the taxonom ic identity and ecology of diazotrophic bacteria associated with non-l egumes. Species-specific oligonucleotide probes directed against varia ble regions of the ribosomal rRNAs of five Azospirillum spp., Herbaspi rillum seropedicae, Herbaspirillum rubrisubalbicans, Acetobacter diazo trophicus and new Burkholderia spp. isolates, as well as genus-specifi c Azospirillum and Azoarcus probes, have been developed and applied to prove the identification of new isolates. In addition, different appr oaches to identify bacteria on the basis of restriction fragment lengt h polymorphism (RFLP) and PCR-fingerprinting techniques allow the char acterization of the diversity of populations even at strain level. On the other hand, strain-specific monoclonal antibodies directed against different Azospirillum brasilense strains enabled us to localize and quantify inoculant bacteria in different parts of the rhizosphere. In addition, antibodies were used successfully to enrich and (re)isolate specific bacteria from different sources. The application of confocal laser scanning microscopy offers high resolution images of diazotrophi c bacteria specifically labelled with fluorescent probes in plant tiss ues. (C) 1997 Elsevier Science Ltd.