The microflora of 25 wheat sourdoughs from the Apulia region, Southern Italy, was characterized. The sourdoughs were mainly produced from Triticum durum wheat. The number of lactic acid bacteria and yeasts ranged from ca. log 7.5 to log 9.3 colony forming units (cfu)/g and from log 5.5 to log 8.4 cfu/g, respectively. About 38% of the 317 isolates of lactic acid bacteria were identified by conventional physiological and biochemical tests. Phenotypic identification was confirmed by 16S rDNA and 16S/23S rRNA spacer region PCR. Overall, 30% of the isolates were identified as Lactobacillus sanfranciscensis, 20% as Lb. alimentarius, 14% as Lb. brevis, 12% as Leuconostoc citreum, 7% as Lb. plantarum, 6% as Lactococcus lactis subsp. lactis, 4% as Lb. fermentum and Lb. acidophilus, 2% as Weissella confusa and 1% as Lb. delbrueckii subsp. delbrueckii. Some of these species have not been previously isolated from sourdoughs. Since bakers yeast is widely used in sourdough production, Saccharomyces cerevisiae was largely found. The phenotypical relationships within the main lactic acid bacteria identified were established by using cluster analysis. A microbial map of the 25 sourdoughs was plotted showing characteristic associations among lactic acid bacteria and differences in the lactic acid bacteria species which were mainly due to the species of wheat flour, use of bakers yeast and type of bread.