A randomized clinical trial was conducted to investigate the effect of a 0.5% chlorhexidine (CHX) gel on dental plaque superinfecting microorganisms in mentally handicapped patients. Thirty inmates from the institution "Casas André Luiz" were assigned to either test group (CHX gel, n = 15) or control group (placebo gel, n = 15). The gel was administered over a period of 8 weeks. Supragingival plaque samples were collected at baseline, after gel use (8 weeks) and 16 weeks after baseline. The presence of Gram-negative Enterobacteriaceae, Staphylococcus and yeasts was evaluated. No significant growth of any superinfecting microorganism was observed in the CHX group, when compared to the placebo group. The results indicated that the 0.5% chlorhexidine gel did not produce an undesirable shift in these bacterial populations.