Bacillus thuringiensis (Bt) is a soil-dwelling bacterium of great interest for agronomical research because of its use as biological pesticide. There are some limitations regarding the subspecies classification. Phenotyping and genotyping studies are important to ascertain its variability. The diversity of 40 environmental strains, isolated from different regions in Mexico, was analyzed by ERIC-PCR and the ability of biofilm formation. Thirty-nine different fingerprinting patterns revealed enough data to discriminate among the 40 strains. A total of 24 polymorphic fragments with sizes between 139 and 1,468 bp were amplified. Almost all (95 %) strains showed biofilm formation after 96 h of incubation. At 96 h of incubation the biofilm-forming strains from the CINVESTAV collection showed a more heterogeneous ability as biofilms producers. Results showed a large intra-species genomic variability in Bt. However, some strains could be correlated as they were found within clusters depending on the location of isolation.