Characterization of the Gene Encoding a 26-Kilodalton Protein (OMP26) from Nontypeable Haemophilus influenzae and Immune Responses to the Recombinant Protein

Abstract

A 26-kDa protein (OMP26) isolated and purified from nontypeable Haemophilus influenzae (NTHI) strain 289 has been shown to enhance clearance of infection following pulmonary challenge with NTHI in rats. DNA sequence analysis revealed that it was 99% identical to a gene encoding a cell envelope protein of the H. influenzae Rd strain (TIGR accession no. HI0916). The deduced amino acid sequence revealed a hydrophilic polypeptide rich in basic amino acids. Restriction fragment length polymorphism analysis suggested that the OMP26 gene was relatively conserved among isolates of NTHI. Analysis of the deduced amino acid sequence of the OMP26 gene from 20 different isolates showed that similarity with NTHI-289 ranged from 96.5% (1 isolate) to 99.5% (14 isolates). Two recombinant forms of OMP26, a full length 28-kDa protein (equivalent to preprotein) and a 26-kDa protein lacking a 23-amino-acid leader peptide (equivalent to processed protein), were assessed in immunization studies for the ability to induce an immune response that would be as effective as the native protein in enhancing the clearance of NTHI following pulmonary challenge in rats. Immunization with the recombinant protein that included the leader peptide was more effective in enhancing pulmonary clearance, and it induced a better cell-mediated response and higher titers of systemic and mucosal antibody. This study has characterized a 26-kDa protein from NTHI that shows significant potential as a vaccine candidate.

Bactericidal antibody response against P6, protein D, and OMP26 of nontypeableHaemophilus influenzaeafter acute otitis media in otitis-prone children

“…P6 is antigenically conserved among both nontypeable and type b H. influenzae strains and has been shown to elicit bactericidal antibodies (De Maria et al, 1996;Murphy et al, 1986) and is considered a vaccine candidate against NTHi infections. OMP26 has been evaluated for its potential as a vaccine candidate as it has been shown to enhance pulmonary clearance of NTHi in a rat model (Adhami et al, 1999;Kyd & Cripps, 1998).…”

Effectiveness of engineering the nontypeableHaemophilus influenzaeantigen Omp26 as an S-layer fusion in bacterial ghosts as a mucosal vaccine delivery

“…These results are in accordance with previous findings showing a combination of IPP/IT to be very successful for immunization with purified Omp26. 20,21 Bacterial clearance achieved by delivering Omp26 in bacterial ghosts without the addition of adjuvants is comparable to that seen following IPP/IT immunization with purified Omp26 formulated in incomplete Freund`s adjuvant (IFA). IPP/IT immunization with recombinant bacterial ghosts elicited slightly lower levels of Omp26 specific antibodies than immunization with purified Omp26.…”

“…For the construction of a maltose binding protein (MBP) Omp26 fusion protein, the omp26 gene was amplified by PCR from the vector pQE30-omp26 21 with oligonucleotides ER-Omp26B (5`-ggc gga tcc atg aaa aac atc gca aaa gt-3`) and ER-Omp26H (5`-ggc aag ctt tta ttt ttt ctc ttg agc ttt ttc tga agc-3`) containing BamHI and HindIII restriction enzyme sites, respectively (restriction enzyme sites are underlined). PCR was performed as previously described.…”

“…15,17,18,19 A NTHi outer membrane protein (Omp26) has been shown to be present on all NTHi strains tested and is highly conserved between strains. 20,21 Rats immunized mucosally with Omp26 enhanced the clearance of both homologous and heterologous strains of NTHi post pulmonary challenge and more recently, Omp26 was shown to be an effective immunogen against NTHi otitis media and nasopharyngeal carriage in the chinchilla model. 20,21,22,23 Thus, these animal experiments provide evidence to support the potential of Omp26 as a vaccine candidate.…”

Construction of recombinant S-layer proteins (rSbsA) and their expression in bacterial ghosts – a delivery system for the nontypeableHaemophilus influenzaeantigenOmp26

“…While they have been found to elicit immune responses in animal models, many exhibit significant ‘between strain’ sequence heterogeneity which limits their capacity to protect against infection with heterologous bacterial strains [20,23–25]. A NTHi outer membrane protein (Omp26) has been shown to be present on all NTHi strains tested and is highly conserved between strains [26,27]. Animals immunized mucosally with Omp26 enhanced the clearance of both homologous and heterologous strains of NTHi post pulmonary challenge [26].…”