Characterization of P1-deficient isogenic mutant of Haemophilus influenzae biogroup aegyptius associated with Brazilian purpuric fever

Segada, Lynn M.; Carlone, George M.; Gheesling, Linda L.; Lesse, Alan J.

doi:10.1006/mpat.1999.0334

Cited by 6 publications

(5 citation statements)

References 47 publications

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“…It is not known if H. influenzae OMP P1 is also involved in LCFA transport. In previous studies, OMP P1‐deficient strains were analyzed in animal models of invasive Hib or Hinf ‐aeg disease, where no differences in the growth and pathogenic potential compared with wild‐type bacteria were observed, when these bacteria were injected intraperitoneally (Hanson et al ., ; Munson and Hunt, ; Segada et al ., ). This indicates that OMP P1 is not essential for bacterial survival and growth under such in vivo conditions.…”

Section: Discussionmentioning

confidence: 97%

Outer membrane protein P1 is the CEACAM‐binding adhesin of Haemophilus influenzae

Tchoupa

Lichtenegger

Reidl

et al. 2015

Molecular Microbiology

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SummaryHaemophilus influenzae is a Gram-negative pathogen colonizing the upper respiratory tract mucosa. H. influenzae is one of several human-restricted bacteria, which bind to carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) on the epithelium leading to bacterial uptake by the eukaryotic cells. Adhesion to CEACAMs is thought to be mediated by the H. influenzae outer membrane protein (OMP) P5. However, CEACAMs still bound to H. influenzae lacking OMP P5 expression, and soluble CEACAM receptor ectodomains failed to bind to OMP P5, when heterologously expressed in Escherichia coli. Screening of a panel of H. influenzae OMP mutants revealed that lack of OMP P1 completely abrogated CEACAM binding and supressed CEACAM-mediated engulfment of H. influenzae by epithelial cells. Moreover, ectopic expression of OMP P1 in E. coli was sufficient to induce CEACAM binding and to promote attachment to and internalization into CEACAM-expressing cells. Interestingly, OMP P1 selectively recognizes human CEACAMs, but not homologs from other mammals and this binding preference is preserved upon expression in E. coli. Together, our data identify OMP P1 as the bona fide CEACAM-binding invasin of H. influenzae. This is the first report providing evidence for an involvement of the major OMP P1 of H. influenzae in pathogenesis.

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Section: Discussionmentioning

confidence: 97%

Outer membrane protein P1 is the CEACAM‐binding adhesin of Haemophilus influenzae

Tchoupa

Lichtenegger

Reidl

et al. 2015

Molecular Microbiology

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“…Inactivation of the coding region of genes by insertion of antibiotic resistance markers is a general strategy for the construction of defined mutants (9,24,36,44). The use of suicide vectors is fundamental to this technique, allowing the creation of deletions or insertions in specific genes on chromosomes (24).…”

Section: Discussionmentioning

confidence: 99%

Role of Various Enterotoxins in Aeromonas hydrophila- Induced Gastroenteritis: Generation of Enterotoxin Gene-Deficient Mutants and Evaluation of Their Enterotoxic Activity

2002

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Three enterotoxins from the Aeromonas hydrophila diarrheal isolate SSU have been molecularly characterized in our laboratory. One of these enterotoxins is cytotoxic in nature, whereas the other two are cytotonic enterotoxins, one of them heat labile and the other heat stable. Earlier, by developing an isogenic mutant, we demonstrated the role of a cytotoxic enterotoxin in causing systemic infection in mice. In the present study, we evaluated the role of these three enterotoxins in evoking diarrhea in a murine model by developing various combinations of enterotoxin gene-deficient mutants by marker-exchange mutagenesis. A total of six isogenic mutants were prepared in a cytotoxic enterotoxin gene (act)-positive or -negative background strain of A. hydrophila. We developed two single knockouts with truncation in either the heat-labile (alt) or the heat-stable (ast) cytotonic enterotoxin gene; three double knockouts with truncations of genes encoding (i) alt and ast, (ii) act and alt, and (iii) act and ast genes; and a triple-knockout mutant with truncation in all three genes, act, alt, and ast. The identity of these isogenic mutants developed by double-crossover homologous recombination was confirmed by Southern blot analysis. Northern and Western blot analyses revealed that the expression of different enterotoxin genes in the mutants was correspondingly abrogated. We tested the biological activity of these mutants in a diet-restricted and antibiotic-treated mouse model with a ligated ileal loop assay. Our data indicated that all of these mutants had significantly reduced capacity to evoke fluid secretion compared to that of wild-type A. hydrophila; the triple-knockout mutant failed to induce any detectable level of fluid secretion. The biological activity of selected A. hydrophila mutants was restored after complementation. Taken together, we have established a role for three enterotoxins in A. hydrophila-induced gastroenteritis in a mouse model with the greatest contribution from the cytotoxic enterotoxin Act, followed by the Alt and Ast cytotonic enterotoxins.

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“…Strains of H. influenzae were grown on chocolate agar, in brain heart infusion (BHI) broth supplemented with hemin and isovitalex, or in chemically defined media (CDM) as described previously (53).…”

Section: Methodsmentioning

confidence: 99%

Characterization of igaB , a Second Immunoglobulin A1 Protease Gene in Nontypeable Haemophilus influenzae

Fernaays¹,

Lesse²,

Cai

et al. 2006

Infect Immun

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Nontypeable Haemophilus influenzae is an important respiratory pathogen, causing otitis media in children and lower respiratory tract infection in adults with chronic obstructive pulmonary disease (COPD). Immunoglobulin A1 (IgA1) protease is a well-described protein and potential virulence factor in this organism as well as other respiratory pathogens. IgA1 proteases cleave human IgA1, are involved in invasion, and display immunomodulatory effects. We have identified a second IgA1 protease gene, igaB, in H. influenzae that is present in addition to the previously described IgA1 protease gene, iga. Reverse transcriptase PCR and IgA1 protease assays indicated that the gene is transcribed, expressed, and enzymatically active in H. influenzae. The product of this gene is a type 2 IgA1 protease with homology to the iga gene of Neisseria species. Mutants that were deficient in iga, igaB, and both genes were constructed in H. influenzae strain 11P6H, a strain isolated from a patient with COPD who was experiencing an exacerbation. Analysis of these mutants indicated that igaB is the primary mediator of IgA1 protease activity in this strain. IgA1 protease activity assays on 20 clinical isolates indicated that the igaB gene is associated with increased levels of IgA1 protease activity. Approximately one-third of 297 strains of H. influenzae of diverse clinical and geographic origin contained igaB. Significant differences in the prevalence of igaB were observed among isolates from different sites of isolation (sputum > middle ear > nasopharynx). These data support the hypothesis that the newly discovered igaB gene is a potential virulence factor in nontypeable H. influenzae.Nontypeable Haemophilus influenzae, a gram-negative coccobacillus, is an important respiratory pathogen (36). This organism is the second most common cause of bacterial otitis media in children and the leading cause of recurrent otitis media (24,36). Nontypeable H. influenzae also plays an important role in the course and pathogenesis of chronic obstructive pulmonary disease (COPD). It is the most commonly isolated organism from the lower airways of adults with COPD and is the most common bacterial cause of exacerbations or periodic worsening of the disease (57). Nontypeable H. influenzae is also a contributing etiologic agent in sinusitis in children and adults as well as in adult pneumonia (36).A number of virulence factors have been identified in nontypeable H. influenzae, including (i) several adhesin molecules, which promote attachment to host structures; (ii) lipooligosaccharide, which promotes inflammation at the site of infection; and (iii) proteins involved in iron acquisition (49, 59, 60). Additionally, an enzyme which cleaves immunoglobulin A1 (IgA1) has been identified in H. influenzae (22,32). This capability is advantageous to this organism because IgA is the predominant mucosal immunoglobulin, and most of the IgA in the respiratory mucosa is of the IgA1 subclass (34).IgA1 proteases have been identified in several human mucosal pathogens, ...

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Characterization of P1-deficient isogenic mutant of Haemophilus influenzae biogroup aegyptius associated with Brazilian purpuric fever

Cited by 6 publications

References 47 publications

Outer membrane protein P1 is the CEACAM‐binding adhesin of Haemophilus influenzae

Outer membrane protein P1 is the CEACAM‐binding adhesin of Haemophilus influenzae

Role of Various Enterotoxins in Aeromonas hydrophila- Induced Gastroenteritis: Generation of Enterotoxin Gene-Deficient Mutants and Evaluation of Their Enterotoxic Activity

Characterization of igaB , a Second Immunoglobulin A1 Protease Gene in Nontypeable Haemophilus influenzae

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