Comparative pangenome analysis of capsulated Haemophilus influenzae serotype f highlights their high genomic stability.

Haemophilus influenzae is an opportunistic pathogen adapted to the human respiratory tract. Non-typeable H. influenzae are highly heterogeneous, but few studies have analysed the genomic variability of capsulated strains. This study aims to examine the genetic diversity of 37 serotype f isolates from the Netherlands, Portugal, and Spain, and to compare all capsulated genomes available on public databases. Serotype f isolates belonged to CC124 and shared few single nucleotide polymorphisms (SNPs) (n = 10,999), but a high core genome (> 80%). Three main clades were identified by the presence of 75, 60 and 41 exclusive genes for each clade, respectively. Multi-locus sequence type analysis of all capsulated genomes revealed a reduced number of clonal complexes associated with each serotype. Pangenome analysis showed a large pool of genes (n = 6360), many of which were accessory genome (n = 5323). Phylogenetic analysis revealed that serotypes a, b, and f had greater diversity. The total number of SNPs in serotype f was significantly lower than in serotypes a, b, and e (p < 0.0001), indicating low variability within the serotype f clonal complexes. Capsulated H. influenzae are genetically homogeneous, with few lineages in each serotype. Serotype f has high genetic stability regardless of time and country of isolation.

Antibody Binding and Complement-Mediated Killing of Invasive Haemophilus influenzae Isolates from Spain, Portugal, and the Netherlands.

Haemophilus influenzae is a Gram-negative bacterium that can be classified into typeable (types a through f) and nontypeable (NTHi) groups. This opportunistic pathogen asymptomatically colonizes the mucosal epithelium of the upper respiratory tract, from where it spreads to other neighboring regions, potentially leading to disease. Infection with NTHi can cause otitis media, sinusitis, conjunctivitis, exacerbations of chronic obstructive pulmonary disease, and pneumonia, but it is increasingly causing invasive disease, including bacteremia and meningitis. Invasive NTHi strains are more resistant to complement-mediated killing. However, the mechanisms of complement resistance have never been studied in large numbers of invasive NTHi strains. In this study, we determined the relationship between binding of IgG or IgM and the bacterial survival in normal human serum for 267 invasive H. influenzae strains from Spain, Portugal, and the Netherlands, of which the majority (200 [75%]) were NTHi. NTHi bacteria opsonized with high levels of IgM had the lowest survival in human serum. IgM binding to the bacterial surface, but not IgG binding, was shown to be associated with complement-mediated killing of NTHi strains. We conclude that evasion of IgM binding by NTHi strains increases survival in blood, thereby potentially contributing to their ability to cause severe invasive diseases.

Identification of polysaccharide capsules among extensively drug-resistant genitourinary Haemophilus parainfluenzae isolates.

The human commensal Haemophilus parainfluenzae is emerging as an opportunistic multidrug-resistant pathogen. The objectives of this work were to characterise a new capsular operon of extensively drug-resistant (XDR) H. parainfluenzae clinical isolates and study their resistance mechanisms using whole-genome sequencing. All strains were resistant to: ß-lactams, via amino acid changes in PBP3 (S385T, I442F, V511A, N526K and V562I); quinolones, by alterations in GyrA (S84F and D88Y) and ParC (S84F and S138T); chloramphenicol, through the presence of catS; macrolides, via the presence of mel and mef(E)-carrying MEGA element; and tetracycline, through the presence of tet(M) and/or tet(B). Phylogenetic analysis revealed high genomic diversity when compared to the H. parainfluenzae genomes available on the NCBI, the isolates from this study being closely related to the Swiss XDR AE-2096513. A full capsular operon showing homology to that of H. influenzae was identified, in accordance with the observation of a capsular structure by TEM. This study describes for the first time a capsular operon in H. parainfluenzae, a major determinant of pathogenicity that may contribute to increased virulence in XDR clinical isolates. Moreover, phylogenetic analysis suggests the possible spread of an XDR-encapsulated strain in Europe.