Abstract Background The bronchial microbiome in chronic lung diseases presents an abnormal pattern, but its microbial composition and regional differences in severe asthma have not been sufficiently addressed. The aim of the study was to describe the bacterial community in bronchial mucosa and secretions of patients with severe chronic asthma chronically treated with corticosteroids in addition to usual care according to Global Initiative for Asthma. Bacterial community composition was obtained by 16S rRNA gene amplification and sequencing, and functional capabilities through PICRUSt. Results Thirteen patients with severe asthma were included and provided 11 bronchial biopsies (BB) and 12 bronchial aspirates (BA) suitable for sequence analyses. Bacteroidetes, Firmicutes, Proteobacteria and Actinobacteria showed relative abundances (RAs) over 5% in BB, a cutoff that was reached by Streptococcus and Prevotella at genus level. Legionella genus attained a median RA of 2.7 (interquartile range 1.1–4.7) in BB samples. In BA a higher RA of Fusobacteria was found, when compared with BB [8.7 (5.9–11.4) vs 4.2 (0.8–7.5), p = 0.037], while the RA of Proteobacteria was lower in BA [4.3 (3.7–6.5) vs 17.1 (11.2–33.4), p = 0.005]. RA of the Legionella genus was also significantly lower in BA [0.004 (0.001–0.02) vs. 2.7 (1.1–4.7), p = 0.005]. Beta-diversity analysis confirmed the differences between the microbial communities in BA and BB (R2 = 0.20, p = 0.001, Adonis test), and functional analysis revealed also statistically significant differences between both types of sample on Metabolism, Cellular processes, Human diseases, Organismal systems and Genetic information processing pathways. Conclusions The microbiota in the bronchial mucosa of severe asthma has a specific pattern that is not accurately represented in bronchial secretions, which must be considered a different niche of bacteria growth.