Immunogenicity of Nontypeable Haemophilus influenzae Outer Membrane Vesicles and Protective Ability in the Chinchilla Model of Otitis Media

Abstract

Outer membrane vesicles (OMVs) produced by Gram-negative bacteria are enriched in several outer membrane components, including major and minor outer membrane proteins and lipooligosaccharide. We assessed the functional activity of nontypeable Haemophilus influenzae (NTHi) OMV-specific antisera and the protective ability of NTHi OMVs as vaccine antigens in the chinchilla otitis media model. OMVs were purified from three HMW1/HMW2-expressing NTHi strains, two of which were also engineered to overexpress Hia proteins. OMV-specific antisera raised in guinea pigs were assessed for their ability to mediate killing of representative NTHi in an opsonophagocytic assay. The three OMV-specific antisera mediated killing of 18 of 65, 24 of 65, and 30 of 65 unrelated HMW1/HMW2-expressing NTHi strains. Overall, they mediated killing of 39 of 65 HMW1/HMW2-expressing strains. The two Hia-expressing OMV-specific antisera mediated killing of 17 of 25 and 14 of 25 unrelated Hia-expressing NTHi strains. Overall, they mediated killing of 20 of 25 Hiaexpressing strains. OMVs from prototype NTHi strain 12 were used to immunize chinchillas and the course of middle ear infection was monitored following intrabullar challenge with the homologous strain. All control animals developed culturepositive otitis media, as did two of three HMW1/HMW2-immunized animals. All OMVimmunized animals, with or without supplemental HMW1/HMW2 immunization, were completely protected against otitis media. NTHi OMVs are the first immunogens examined in this model that provided complete protection with sterile immunity after NTHi strain 12 challenge. These data suggest that NTHi OMVs hold significant potential as components of protective NTHi vaccines, possibly in combination with HMW1/HMW2 proteins. KEYWORDS Haemophilus influenzae, outer membrane vesicles, vaccines Nontypeable Haemophilus influenzae (NTHi) organisms are small Gram-negative bacteria that colonize the upper respiratory tract of humans beginning at a very early age (1). Although these organisms are normally commensals, when host defenses are compromised by underlying medical conditions such as malnutrition, immunodeficiency, chronic lung disease, or acute viral infection, disease caused by NTHi may develop (2, 3). Among children in the developed world, NTHi infections are currently responsible for an estimated 40 to 50% of the cases of acute otitis media and an even higher percentage of cases of chronic and recurrent disease (4, 5). Among adults, particularly among patients with chronic obstructive pulmonary disease, NTHi infections are a major cause of illness, especially during the acute exacerbations that often characterize this disease (6). A vaccine capable of preventing disease caused by these organisms would offer substantial benefit to the adult and pediatric populations alike.NTHi vaccine development efforts are ongoing in many laboratories. A large number

Panel 8: Vaccines and immunology

“…A monoclonal antibody against ketodeoxyoctanoate (KDO) was bactericidal against NTHi in vitro and KDO decoration of lipooligosaccharide within NTHi biofilms from the chinchilla middle ear was shown ex vivo [55]. Immunization of chinchillas with outer membrane vesicles isolated from HMW1/ HMW2-and Hia-expressing NTHi prevented experimental OM [56]. Therapeutic immunization with the outer membrane protein (OMP) P5-and Type IV pilus-directed antigen, chimV4, delivered by band-aid, resolved NTHi-induced OM in a chinchilla model [57] and additionally prevented OM in a viral-bacterial model of experimental disease [58].…”

The Contribution of Membrane Vesicles to Bacterial Pathogenicity in Cystic Fibrosis Infections and Healthcare Associated Pneumonia

“…Furthermore, active immunization of mice with P. aeruginosa MVs resulted in mice that were protected from P. aeruginosa infections (Zhang et al, 2018). Also NTHi, K. pneumoniae and S. aureus MVs are potential vaccine candidates, as was demonstrated in several studies (Lee et al, 2015;Roier et al, 2015;Winter and Barenkamp, 2017;Askarian et al, 2018).…”

A Bacterial Epigenetic Switch in Non-typeable Haemophilus influenzae Modifies Host Immune Response During Otitis Media

“…Future work is necessary to fully investigate the mechanisms by which macrophages differentially recognized and kill each of these populations. Many proteins and saccharides on the outer surface of NTHi are known to affect opsonization and host immune response and may be regulated by ModA2 during disease (Duell et al, 2016;Winter and Barenkamp, 2017;Li et al, 2018). The potential ability of the modA2 ON subpopulation to avoid recognition and/or suppress activation by macrophages could result in increased survival and a subsequent shift toward a predominantly modA2 ON population, such as the one observed within middle ear fluids during experimental disease (Atack et al, 2015a).…”