Aims SARS‐CoV‐2 RNA has been recovered from different sites in the human body, including the mouth. The present study aimed to investigate the presence of SARS‐CoV‐2 RNA in the dental biofilm of symptomatic patients who tested positive in nasopharyngeal and oropharyngeal (NASO/ORO) samples. Materials & Methods An observational clinical study of individuals with flu‐like symptoms was conducted between July and September 2020. Dental biofilm (BIO) samples were collected and analysed using real‐time quantitative polymerase chain reaction (RT‐qPCR) to determine the virus's presence. Results Seventy participants (40 ± 9.8 years of age, 71.4% female) tested positive for SARS‐CoV‐2 RNA in NASO/ORO samples and were included in the study. Among them, 13 tested positive in BIO samples (18.6%; 95% CI: [9.5, 27.7]). The median and interquartile range of cycle quantification (Cq) for NASO/ORO and BIO samples were 15.9 [6.9] and 35.9 [4.0] (p = .001), respectively. BIO‐positive participants showed a higher virus load in NASO/ORO samples (p = .012) than those testing negative (Cq = 20.4 [6.1]). Conclusions Dental biofilms from symptomatic COVID‐19 patients harbour SARS‐CoV‐2 RNA and might be a potential reservoir with an essential role in COVID‐19 transmission.