Reduced severity of middle ear infection caused by nontypeable Haemophilus influenzae lacking the hemoglobin/hemoglobin–haptoglobin binding proteins (Hgp) in a chinchilla model of otitis media

Morton, Daniel J.; Bakaletz, Lauren O.; Jurcisek, Joseph A.; VanWagoner, Timothy M.; Seale, Thomas W.; Whitby, Paul W.; Stull, Terrence L.

doi:10.1016/j.micpath.2003.08.007

Cited by 53 publications

(61 citation statements)

References 41 publications

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“…The degree of inflammation was assessed during otoscopic examination using a scoring system, and a qualitative score of 1 to 4 was recorded for each of the following criteria: erythema, tympanic membrane discoloration and opacity, vessel dilation, and fluid accumulation. Changes in middle ear pressure, tympanic width, and tympanic membrane compliance were measured by tympanometry (EarScan, Daytona, FL) as previously described (27). As indicated in the text, the presence of inflammatory mediators in middle ear fluids was compared using RayBiotek protein arrays.…”

Section: Methodsmentioning

confidence: 99%

Phosphorylcholine Decreases Early Inflammation and Promotes the Establishment of Stable Biofilm Communities of Nontypeable Haemophilus influenzae Strain 86-028NP in a Chinchilla Model of Otitis Media

et al. 2007

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Haemophilus influenzae is a gram-negative pleiomorphic bacterium that is a common commensal/mutualist within the human airways (30). Encapsulated H. influenzae strains are overt pathogens causing invasive disease (3) and have largely been contained by a vaccine effective against the predominant capsular serotype b strains (32). In contrast, the so-called nontypeable H. influenzae (NTHi) strains lacking capsular polysaccharides remain predominant in asymptomatic carriage and localized airway infections (14, 29). These infections are mostly opportunistic in nature and include bronchiopneumonia, sinusitis, and otitis media (OM). OM is among the most common pediatric infections, causing an estimated ϳ$5 billion in costs of treatment and parents' missed work days per year (20). OM infections include chronic OM that is difficult to resolve with antibiotic therapy, and it has long been postulated that chronic OM involves the formation of bacterial biofilm communities (5, 35). In support of that hypothesis, biofilms have been visualized in tympanostomy drain tubes removed from patients with OM and on middle ear tissue from experimentally infected chinchillas (7,18,33). More recent evidence shows that NTHi and other bacterial agents are present within biofilms on tissue specimens obtained from patients with chronic and recurrent OM (13).The H. influenzae surface is covered with lipooligosaccharide (LOS) endotoxins that lack a repeating O side chain. Instead, the H. influenzae LOS features a diverse collection of LOS glycoforms that differ in the length, content, and nature of the chemical linkages found in the oligosaccharide portion. These LOS oligosaccharides include structures that are antigenically similar to host cell-surface glycolipids and may also contain the host membrane constituents sialic acid (NeuAc) and phosphorylcholine (PCho) (41). LOS confers resistance to host killing (8,9,37) and is also the primary target of the Toll-like receptor 4 pathway that mediates protection against H. influenzae in the airways (47). It has been established that NTHi strains that express NeuAc-LOS forms comprise a greater proportion of biofilm communities than of planktonic cultures, and that mutations eliminating these forms decrease biofilm formation and bacterial persistence in animal models of OM (4,12,18,43). More recently, we showed that LOS purified from biofilms has decreased potency as an inflammatory agonist, which correlated with an increase in PCho ϩ LOS forms that were present within biofilms (55). In this study, we compared the virulence

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Section: Methodsmentioning

confidence: 99%

Phosphorylcholine Decreases Early Inflammation and Promotes the Establishment of Stable Biofilm Communities of Nontypeable Haemophilus influenzae Strain 86-028NP in a Chinchilla Model of Otitis Media

et al. 2007

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“…H. influenzae strains HI689 and HI1717 were transformed to kanamycin resistance (25 g/ml with plasmid pDJM343 DNA using a modification of the static aerobic method of Gromkova et al (22) as previously described (38). Appropriate chromosomal insertions were confirmed by the size of a PCR product, and hxuC deletion mutants of strains HI689 and HI1717 were identified, phenotypically confirmed by growth characteristics in vitro, and designated strains HI1787 and HI1810, respectively.…”

Section: Methodsmentioning

confidence: 99%

“…Release of heme from methemoglobin could result in the presence of free heme if plasma levels of hemopexin and albumin in 5-day-old infant rats are low or formation of heme-hemopexin and heme-albumin complexes if their concentrations are high. Aerobic growth of H. influenzae in vitro is supported by each of these heme sources by mechanisms other than those involving the Hgps (38,40,63). The hypothesis that plasma from infant rats contains heme sources capable of supporting the growth of strain HI1717 was evaluated first in plate bioassays on hdBHI agar ( Table 2).…”

Section: Vol 74 2006 Role Of H Influenzae Hgp Genes In Invasive DImentioning

confidence: 99%

“…Previously we have shown that the hgpA gene and other iron acquisition genes are expressed in vivo during acute human otitis media infection (68). More recently we showed that deletion of the complement of three hgp genes from a nontypeable strain of H. influenzae attenuated virulence in the chinchilla model of otitis media (38). The present study was undertaken to characterize the contribution of the hgp genes to virulence of H. influenzae in the rat model of invasive disease.…”

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confidence: 90%

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Complex Role of Hemoglobin and Hemoglobin-Haptoglobin Binding Proteins in Haemophilus influenzae Virulence in the Infant Rat Model of Invasive Infection

Seale¹,

Morton²,

Whitby³

et al. 2006

Infect Immun

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Haemophilus influenzae requires an exogenous heme source for aerobic growth in vitro. Hemoglobin or hemoglobin-haptoglobin satisfies this requirement. Heme acquisition from hemoglobin-haptoglobin is mediated by proteins encoded by hgp genes. Both Hgps and additional proteins, including those encoded by the hxu operon, provide independent pathways for hemoglobin utilization. Recently we showed that deletion of the set of three hgp genes from a nontypeable strain (86-028NP) of H. influenzae attenuated virulence in the chinchilla otitis media model of noninvasive disease. The present study was undertaken to investigate the role of the hgp genes in virulence of the wild-type serotype b clinical isolate HI689 in the infant rat model of hematogenous meningitis, an established model of invasive disease requiring aerobic growth. Bacteremia of high titer and long duration (>14 days) and histopathologically confirmed meningitis occurred in >95% of infant rats challenged at 5 days of age with strain HI689. While mutations disrupting either the Hgp-or Hxu-mediated pathway of heme acquisition had no effect on virulence in infant rats, an isogenic mutant deficient for both pathways was unable to sustain bacteremia or produce meningitis. In contrast, mutations disrupting either pathway decreased the limited ability of H. influenzae to initiate and sustain bacteremia in weanling rats. Biochemical and growth studies also indicated that infant rat plasma contains multiple heme sources that change with age. Taken together, these data indicate that both the hgp genes and the hxuC gene are virulence determinants in the rat model of human invasive disease.

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“…The erythromycin resistance marker from pERMR was excised with SmaI and cloned into the unique SnaBI site (internal to hel) of pSG5 to yield pSG17. H. influenzae were transformed to erythromycin resistance with pSG17 as previously described [12]. Correct chromosomal recombinations were confirmed by the molecular size of a PCR product resolved on an agarose gel.…”

Section: Construction Of Hel Mutantsmentioning

confidence: 99%

Lipoprotein e (P4) of Haemophilus influenzae: role in heme utilization and pathogenesis

Morton

Smith

VanWagoner

et al. 2007

Microbes and Infection

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Lipoprotein e (P4) of Haemophilus influenzae is a phosphomonoesterase, encoded by the hel gene, that has been implicated in the acquisition of heme by this fastidious organism. However, lipoprotein e (P4) is also involved in the utilization of NAD and NMN. Some reports have concluded that the reported heme-related growth defect actually reflects a growth defect for NAD. In the current study hel insertion mutants were constructed and a role for e (P4) in heme acquisition was demonstrated independent of its role in NAD or NMN acquisition. In addition a rat model of infection demonstrated a role for e (P4) in the pathogenesis of invasive disease.

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Reduced severity of middle ear infection caused by nontypeable Haemophilus influenzae lacking the hemoglobin/hemoglobin–haptoglobin binding proteins (Hgp) in a chinchilla model of otitis media

Cited by 53 publications

References 41 publications

Phosphorylcholine Decreases Early Inflammation and Promotes the Establishment of Stable Biofilm Communities of Nontypeable Haemophilus influenzae Strain 86-028NP in a Chinchilla Model of Otitis Media

Phosphorylcholine Decreases Early Inflammation and Promotes the Establishment of Stable Biofilm Communities of Nontypeable Haemophilus influenzae Strain 86-028NP in a Chinchilla Model of Otitis Media

Complex Role of Hemoglobin and Hemoglobin-Haptoglobin Binding Proteins in Haemophilus influenzae Virulence in the Infant Rat Model of Invasive Infection

Lipoprotein e (P4) of Haemophilus influenzae: role in heme utilization and pathogenesis

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