Internalization and Trafficking of Nontypeable Haemophilus influenzae in Human Respiratory Epithelial Cells and the Roles of IgA1 Proteases for Optimal Internalization and Persistence

Nontypeable Haemophilus influenzae (NTHI) is a leading cause of opportunistic infections of the respiratory tract in children and adults. Although considered an extracellular pathogen, NTHI has been observed repeatedly within and between cells of the human respiratory tract, and these observations have been correlated to symptomatic infection. These findings are intriguing in light of the observation that NTHI persists in the respiratory tract despite antibiotic therapy and the development of bactericidal antibodies. We hypothesized that intracellular NTHI avoid, escape, or neutralize the endolysosomal pathway and persist within human respiratory epithelial cells, and that human IgA1 proteases are required for optimal internalization and persistence of NTHI. Virtually all strains of NTHI contain the human IgA1 protease gene, iga, and we previously characterized a novel human IgA1 protease gene, igaB, that is associated with disease-causing strains and is homologous to an IgA1 protease of pathogenic Neisseria . Here, we show that NTHI invades human bronchial epithelial cells in vitro in a lipid raft-independent manner, is subsequently trafficked via the endolysosomal pathway, and is killed in lysosomes after variable durations of persistence. IgaA is required for optimal invasion. IgaB appears to play no role in adherence or invasion, but is required for optimal intracellular persistence of NTHI. IgaB cleaves lysosome-associated membrane protein 1 (LAMP1) at pHs characteristic of the plasma membrane, early endosome, late endosome, and lysosome. Collectively, these observations establish that the two IgA1 proteases of NTHI play important but different roles in NTHI invasion and trafficking in respiratory epithelial cells.