Evaluation of prokaryotic diversity by restrictase digestion of 16S rDNA directly amplified from hypersaline environments

Abstract

A novel molecular strategy to study microbiological diversity is described. This method is based on the restriction digestion of a population of 16S rDNA sequences directly amplified from an environmental sample. Digested fragments separated by polyacrylamide electrophoresis generate characteristic profile data for estimation of diversity and overall similarities between the organisms of different environments. The methodology has been applied to a set of five ponds in a multi-pond solar saltern covering the salinity gradient from about twice that of seawater (6.4%) to NaCl precipitation (30.8%). Bacterial (eubacterial) diversity estimated from the complexity of the banding pattern obtained by restriction of the amplicons from the different ponds decreased with increasing salinity, while for Archaea (archaebacteria) the reverse was true, i.e. the higher the salinity the higher the number of bands. The similarities in taxonomic composition of the prokaryotic populations present in those ponds were evaluated from the number of restriction bands shared by the different samples. The relationships found among the different environments were independent of the enzyme used for digestion and were consistent with previous descriptions obtained by the study of isolates from the different environments. The technique appears to be promising as a rapid method for microbial biodiversity fingerprinting, useful to compare several environments and detect major shifts in species composition of the microbial population.