Multidrug-resistant <i>Haemophilus influenzae</i> cluster of severe asthma from sputum bacteriome–resistome

Background Severe asthma encompasses heterogeneous inflammatory phenotypes and airway bacteriome diversity but the state of its airway resistome remains understudied. We therefore evaluated the link between the airway microbiome and the antibiotic-resistant genes by determining the clusters from a bacteriome–resistome integration from sputum samples of patients with severe asthma. Methods Induced sputum samples from severe asthma (SA; n=96), mild–moderate asthma (MMA; n=23) and healthy controls (HCs; n=23) in the European U-BIOPRED asthma cohort were metagenomically sequenced. Respiratory bacteriome was evaluated by taxonomical and functional classification. The comprehensive antibiotic resistance database was used to determine airway resistome and Similarity Network Fusion to cluster integratively the bacteriome–resistome. Results More multidrug-resistance genes were present in SA compared with MMA and HCs with the hmrM , encoded in Haemophilus influenzae chromosome, being highest. Two of the three defined clusters were dominated by commensals with resistance genes from different classes but different in α- and β-diversities. The third cluster was dominated by multidrug-resistant H. influenzae , with SA characteristics of increased asthma duration, reduced pulmonary macrophages and decreased lung function. It had the highest signature expression of neutrophil activation, NETosis and of interleukin (IL)-5, IL-6, IL-13, IL-17 and IL-33 signalling pathways. These clusters were reproduced in an Asian-Singapore SA cohort including the multidrug-resistant H. influenzae cluster, but with an additional cluster of multidrug-resistant Pseudomonas aeruginosa . Conclusion The demonstration of U-BIOPRED multiresistant H. Influenzae and of Asian-Singapore multiresistant P. aeruginosa clusters highlights the potential importance of antibiotic-resistant genes in driving severe asthma.