The supragingival plaque will be collected with sterile spoon dentin and subgingival plaque will be collected with a sterile paper cone inserted in selected sites for 30 seconds , after drying the surface and careful removal of supragingival plaque . The cones are stored in sterile plastic tube at -80 ° C until analysis. The identification of total bacterial load and microbial periodontal ( Porphyromonas gingivalis , Fusobacterium nucleatum , Prevotella intermedia and Streptpcoccus mitis ) will be performed using the technique of real -time PCR and T - RFLP .