Acute otitis media (AOM) is the most common reason for antimicrobial treatment in children. Novel methods for primary prevention of AOM are needed. Previously, probiotic lactobacilli have been ineffective in AOM prevention. In contrast to lactobacilli, alpha-hemolytic streptococci belong to the normal core microbiome of the oral cavity and nasopharynx. In this randomized controlled trial, we set out to investigate the microbiological effects of Streptococcus salivarius K12 probiotic product in the saliva and nasopharyngeal microbiome of 121 children attending daycare centres. The study products were used every evening for one month. We obtained saliva and nasopharyngeal samples at 0, 1, and 2 months, and used next-generation sequencing of bacterial 16S gene.