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Background: Dental plaque, also known as dental biofilm, is a pale-yellow sticky film that is generated on teeth due to the activity of biofilm-forming bacteria. This dental health issue can frequently occur, especially in individuals with low tooth hygiene. Objectives: The current study was performed to understand the bacterial profile of dental plaque from adults and children in Al-Diwaniyah Province, Iraq. Materials and methods: Sixty dental-plaque samples were collected from 30 adults (18-56 years old) and 30 children (6-14 years old). Regular techniques (cultivation, colony morphology, and VITEK 2) were applied to identify the bacterial members in each sample. Results: The results uncovered that Gram-positive bacteria were dominant, in which Streptococcusspp was highly prevalent. Lactobacillusspp showed a second-place prevalence, while Staphylococcusspp unveiled less existence of Gram-positive isolates in the dental-plaque samples of all individual ages. For the Gram-negative bacteria, Enterobacterspp, Enterococcus spp, and Klebsiellaaerogeneswere the most frequent bacterial isolates of the dental biofilm of all participant ages. Conclusion: The study informs the dominant presence of Streptococcusspp, Lactobacillusspp, and Staphylococcusspp in the dental-plaque samples of adults and children. This profile might indicate that the oral cavity is a suitable habitat for those bacteria to form the dental biofilm.