The aim of this study was to characterize the main periodontal bacterial species in Down syndrome (DS) pa-tients with and without periodontitis. Method: This cross-sectional study involved 75 DS patients, 45 with and 30 without periodontitis. Informed consent, health and dental questionnaires and periodontitis diagnosis were performed. PCR and LAMP assays were performed on subgingival dental plaque sample. Results: Tannerella forsythia was the most frequent bacteria detected in the group with and without periodontitis (95.5 and 63.3%) followed by Treponema denticola (88.8 and 50%) and Porphyromonas gingivalis (53.3 and 25% respectively). There were statistical differences between groups (p<0.05). Pg fimA type I was the most frequent Porphyromonas gingivalis genotype. Two different sets o primers (Aa-F/Aa-R and ltx3/ltx4) were used to detect Aggregatibacter actinomycetemcomitans and different frequencies were obtained, (68% and 14.6% respectively), they had a weak correlation (Cohen Kappa=0.16). After sequencing of PCR products, ltx3/ltx4 showed more specificity. JP2 clone of A. actinomycetemcomitans was not detected in any sample. Conclusions: The composition of oral biofilm is fundamental for the development of periodontal disease independently of immunological alterations associated with DS. The frequency of detection of A. actinomycetemcomitans reported in the literature has a wide range, because the primers and probes applied.