Helicobacter pylori plays a significant role in gastric disease. However, the presence of this bacterium in the oral cavity remains controversial. The aim of the present study was to detect and quantify H. pylori in 29 different sites of the oral cavity in non-dyspeptic subjects by means of real-time polymerase chain reactions (PCR). Ten subjects without gastric symptoms were studied. Samples from unstimulated saliva, three sites of the tongue, oral mucosa, and 12 sites of both supragingival and subgingival plaque were collected from each subject. DNA was extracted from the oral samples and analysed for the presence of H. pylori by real-time PCR (LightCycler) using JW23/22 primers which targeted the 16S rRNA gene. DNA from H. pylori DSM 4867 was used as a positive control. Amplification efficiency for the LightCycler 2.0 runs ranged from 1.8 to 2.4. Melting curve analysis identified all the positive control capillaries, which contained H. pylori reference DNA, as a single and narrow peak at a melting temperature between 84.5 and 84.9 degrees C. All the negative control capillaries with no template control and the 29 oral samples from each subject showed either no melting peaks or broad melting peaks below 80 degrees C, which were considered as primer dimers. Therefore, H. pylori was not detected from any of the 290 oral samples. As a conclusion, H. pylori seems not to be permanently present in the oral cavity of a non-dyspeptic population.