OBJECTIVE: The aim of the present study was to investigate the reproducibility of bacterial enumeration from subsequent subgingival samples collected from patients with peri-implant mucositis. MATERIAL AND METHODS: Duplicate microbial samples from 222 unique implant sites in 45 adult subjects were collected with paper points and analyzed using the checkerboard DNA-DNA hybridization technique. Whole genomic probes of 74 preselected bacterial species were used. Based on the bacterial scores, Cohen's kappa coefficient was used to calculate the inter-annotator agreement for categorical data. The percentage agreement was considered as "good" when the two samples showed the same score or differed by 1 to the power of 10. RESULTS: Moderate to fair kappa values were displayed for all bacterial species in the test panel (range 0.21-0.58). There were no significant differences between Gram-positive and Gram-negative species. The percentage of good agreement between the first and second samples averaged 74.7 % (n = 74; range 56-83 %), while the proportion of poor agreement ranged from 1 to 19 % for the various strains. CONCLUSION: While an acceptable clinical agreement was obtained in most cases, diverging bacterial scores may appear in subgingival samples collected at the same time point from patients with peri-implant mucositis. CLINICAL RELEVANCE: The broad bulky base of implant crowns may present an obstacle for the collection of reproducible subgingival samples with paper points.