The aim of the present work was to evaluate the effects of two soil management systems so called sustainable treatment (ST) and conventional treatment (CT) on the composition and on genetic, functional, and metabolic diversity of soil microbial communities in a Mediterranean olive orchard. The ST system included no-tillage, integrated chemical fertilization, and organic matter inputs from drip irrigation, spontaneous cover crops, and pruning material. Microbial analyses were carried out by an integrated approach of culture-dependent (microbial cultures and Biolog) and culture-independent methods (denaturing gradient gel electrophoresis [DGGE]). After 7 years of treatment, average olive yield was 8.4 and 3.1 t ha−1 year−1 in ST and CT, respectively. Conventional treatment had a significantly higher number of total culturable bacteria and actinomycetes compared with ST, whereas fungi were significantly lower. In ST, the number of ammonifying bacteria, proteolytic bacteria, and Azotobacter in the wetted areas under the drippers (ST-WET) was significantly higher than along interrows (ST-INTER). The DGGE analysis of microbial 16S/18S rDNA showed differences between ST and CT, whereas 16S/18S rRNA DGGE bands of ST-WET clustered differently from those of CT and ST-INTER. Some Biolog metabolic indexes were significantly different between ST and CT. The results revealed qualitative and quantitative changes of soil microbial communities in response to sustainable agricultural practices that stimulate soil microorganism activity and improve olive yield and fruit quality.