The goal of this study was to evaluate the clinical and microbiological factors associated with orthodontic appliances during an episode of gingival inflammation and the impact of appliance removal on periodontal health. This prospective study included 10 patients, aged 12 to 20 years, with clinical signs of gingival inflammation at the final phase of orthodontic treatment (appliance removal). Plaque index, gingival index, and probing depth were evaluated, and microbiological samples were collected from teeth 16, 11, and 26 at 2 times: during the gingival inflammation (baseline) and 30 days after the removal of the appliance and professional prophylaxis. Polymerase chain reaction analysis was used to detect Porphyromonas gingivalis, Bacteroides forsythus, Actinobacillus actinomycetemcomitans, Prevotella intermedia, and Prevotella nigrescens. A statistically significant improvement of the plaque and gingival indexes was seen, as well as a reduction in probing depth (P <.05). Periodontal pathogens were associated with gingival inflammation during orthodontic treatment. The improvement in periodontal health at 30 days was concomitant with a reduction of sites positive for A. actinomycetemcomitans and B. forsythus (P <.05). Periodontal pathogens associated with gingival inflammation during orthodontic treatment can be significantly reduced by orthodontic appliance removal and professional prophylaxis.