Infectious bronchitis (IB) is one of the avian diseases with the greatest impact on poultry farming worldwide. In Brazil, strain BR-I (GI-11) is the most prevalent in poultry flocks. The present study aimed to develop a seminested RT-PCR assay specific for the diagnosis of BR-I IBV in Brazilian samples, targeting subunit 1 of the S gene. The detection limit of this assay was 10 copies of the IBV genome. In this study, 62.24% of 572 organ pools from the 5 regions of Brazil tested positive in a 3′UTR screening, and 84.83% were typed as BR-I IBV. BR-I was detected in the respiratory, digestive and urogenital tracts in pooled samples from all Brazilian geographical regions and in all the breeding systems analyzed. Specificity and sensitivity tests as well as phylogenetic analysis successfully confirmed the expected clustering of the sequences detected by this assay with the BR-I (GI-11) group. The nested PCR described in this study represents a suitable and valuable tool in the diagnosis, epidemiology, monitoring and vaccination decisions of IBV.