The objective of this cross-sectional clinical study was to analyze the differences in the microbiome in gingival sulci of adult patients in the presence or absence of chronic periodontitis. Patients with or without periodontal disease were included in this cross-sectional study. Subgingival biofilm samples were collected and analyzed by 16S massive pyrosequencing. Functional analyses were also performed. A total of 15 phyla, 154 genera and 351 species were detected globally. Differences between disease and non-disease samples were observed in all taxonomical levels which suggest functional profile changes in the community. It was found that the main species associated with non-disease samples were reduced in disease but not completely suppressed. Analysis of the functional potential of the biofilms revealed a significantly higher activity related to endocytosis and phosphatidylinositol signaling in the disease group but lower cell adhesion molecules. Specific differences between health and disease suggest functional profile changes in the community, although bacteria associated with periodontal disease are also increased in health. Transcriptome studies should be conducted to confirm and deepen metabolic dysfunctions.