© 2016 Asian Pacific Tropical Medicine Press. Objective: To determine the contamination levels of Nosema ceranae in honeybees and its molecular linkages in different geographical areas of Northern Thailand. Methods: Seventy-eight apiaries in Northern Thailand were chosen at random. The detection was accomplished both by microscopic examination and multiplex PCR. Nosema positive samples were evaluated by PCR sequencing and phylogenetic analysis. Results: Of the samples subjected to microscopic examination, 11.54% were found to be positive for Nosema while 29.49% of the samples evaluated by PCR were found to be positive for the disease. Honeybees from four of the six provinces surveyed in Northern Thailand were positive for Nosema, with the highest prevalence in Chiang Mai Province (48.57%). There was a high diversity of Nosema strains in some locations, while the same strain of pathogen was identified in many locations in Northern Thailand. Conclusions: This is the first report about the contamination levels and distribution pattern of nosemosis in Thailand. The study found the same group of Nosema in different locations, and different groups of Nosema in the same location. This pattern of distribution will be an advantage for disease control in the future.