The objectives of this study were to determine the prevalence of Salmonella, trends, major serovars, and their clusters from fluff samples, in poultry hatcheries in Ontario between 1998 and 2008. Multi-level logistic regression modelling with random effects for hatchery and sampling visit (day on which samples were collected from a hatchery) was used to identify factors [poultry breeder type, year (trend), and season] associated with the prevalence of Salmonella and a cluster detection test was used to identify clusters of common serovars. The period prevalence of Salmonella in fluff samples was 8.7% in broiler-breeders, 3.1% in layer-breeders, 13.2% in turkey-breeders, and 11.9% in other-breeder birds, such as ducks, geese, quail, partridges, and pheasants. There was an overall increasing trend in Salmonella prevalence in broiler-breeders and other-breeder birds, and a decreasing trend in layer-breeders. The 4 most common serovars identified were Salmonella Kentucky, Heidelberg, Enteritidis, and Senftenberg in broiler-breeders; Salmonella Heidelberg, Senftenberg, Braenderup, and Typhimurium in layer-breeders; Salmonella Senftenberg, Heidelberg, Saintpaul, and Montevideo in turkey-breeders; and Salmonella Enteritidis, Thompson, Typhimurium, and Heidelberg in other-breeder birds. Temporal clusters were identified for 12 of 13 serovars examined in broiler-breeders, and 4 of 4 serovars in all other poultry-breeders. The seasonal effects varied by year with the highest probability of Salmonella most often occurring in the summer, followed by the fall season. Variance components suggested that control measures should be directed at the hatchery and the sampling visit levels. Further studies are needed to identify risk factors for Salmonella in broiler-breeder, turkey-breeder, and other-breeder bird hatcheries in order to implement necessary control measures.