fH-dependent complement evasion by disease-causing meningococcal strains with absent fHbp genes or frameshift mutations

Abstract

Meningococci bind human fH to down-regulate complement, which enhances survival of the bacteria in serum. A major fH ligand is the vaccine candidate, factor H-binding protein (fHbp). Although fHbp has been considered an essential meningococcal virulence factor, rarely, invasive isolates with absent fHbp genes or frameshift mutations have been identified. In previous studies fH binding to these isolates was not detected. The aim of the present study was to investigate fH binding and complement evasion by invasive meningococcal serogroup B clinical isolates with absent fHbp genes or frameshift mutations. Four of the seven isolates tested bound human fH by flow cytometry and survived in IgG-depleted human serum. In all four, fH binding was decreased after inactivating the gene encoding NspA. Binding of fH to fHbp and NspA is specific for human fH. To investigate fH-dependent evasion of host defenses, human fH transgenic infant rats, or control littermates negative for human fH, were challenged IP with 103 to 104 CFU of two of the isolates with no detectable fH binding by flow cytometry. At six hours, bacteremia caused by both strains was higher in human fH transgenic rats than in control rats (P<0.002). In conclusion, six of the seven isolates had evidence of fH binding and/or human fH-dependent complement evasion in transgenic rats. In four, NspA was as an alternative fH ligand. fHbp vaccination may select for mutants that do not require fHbp for complement evasion. Inclusion of additional target antigens in vaccines containing fHbp may delay emergence of these mutants.

Binding of Complement Factor H to PorB3 and NspA Enhances Resistance of Neisseria meningitidis to Anti-Factor H Binding Protein Bactericidal Activity

“…Since FH downregulates the alternative pathway, the ability of anti-FHbp antibodies to block FH binding can be critical for eliciting anti-FHbp bacteriolysis (25,27,39,40). We investigated whether resistance to anti-FHbp bactericidal activity might be explained by binding of FH to ligands other than FHbp (16).…”

“…FHbp-knockout (KO), neisserial surface protein A (NspA)-KO, and FHbp/NspA double-KO mutants were generated as previously described (16,17). In brief, the FHbp gene was replaced by an erythromycin resistance cassette carried on pBS⌬gna1870erm (5).…”

“…In brief, the FHbp gene was replaced by an erythromycin resistance cassette carried on pBS⌬gna1870erm (5). The genes encoding NspA were inactivated by transforming the wild-type or isogenic FHbp-KO mutants with a plasmid that contained a truncated nspA gene interrupted by a gene that conferred spectinomycin resistance (16). The resulting nonfunctional NspA genes were confirmed by sequencing of DNA obtained by PCR and by loss of binding of the antiNspA monoclonal antibody (MAb) 14C7 (18), as measured by flow cytometry.…”

Serum bactericidal antibody assays – The role of complement in infection and immunity

“…The meningococcus takes advantage of its ability to bind fH in order to shield itself from the action of the innate immune response. The meningococcus harbours at least three distinct proteins on its surface to exploit fH: factor H-binding protein (fHbp), Neisserial surface protein A (NspA) [8,10,18] and PorB [19,20]. With these proteins, the organism can bind human fH specifically -the organism becomes surrounded by fH which keeps complement C3b in check, thereby down-regulating complement activation by the human immune system.…”

Vaccines for prevention of group B meningococcal disease: Not your father's vaccines

“…Invasive meningococcal strains that lack porA or fHbp have been described [72,77] and, as mentioned above, nadA is absent from a substantial proportion of meningococcal strains. The ability to cause invasive disease despite absence of FHbp is preserved because of alternative mechanisms by which N. meningitidis can bind to human factor H [78,79]. Disruption of the nhbA gene by insertion sequence IS1301 in a Brazilian invasive group C isolate, which also lacked the nadA gene, has been identified [80].…”