Margo Diricks (Borstel / DE), Sabine Petersen (Borstel / DE), Lennart Bartels (Borstel / DE), Thien-Tri Lam (Würzburg / DE), Heike Claus (Würzburg / DE), Maria Paula Bajanca-Lavado (Lisbon / PT), Susanne Hauswaldt (Lübeck / DE), Ricardo Stolze (Borstel / DE), Omar Jiménez Vázquez (Borstel / DE), Christian Utpatel (Borstel / DE), Stefan Niemann (Borstel / DE), Jan Rupp (Lübeck / DE), Inken Wohlers (Borstel / DE), Matthias Merker (Borstel / DE)
Background:
Haemophilus influenzae, an opportunistic bacterial pathogen, can lead to severe respiratory tract infections, septicemia, and meningitis. The emergence of β-lactamase-negative ampicillin-resistant (BLNAR) strains and difficulties in correlating genotypic (gBLNAR) and phenotypic resistance complicate empirical treatment and patient management.
Methods:
We conducted a systematic meta-analysis of previously identified H. influenzae gBLNAR groups characterized by distinct substitution patterns in penicillin-binding protein 3 (PBP3), and analyzed their associations with ampicillin and cefotaxime resistance. In addition, we used phylogenomics and a genome-wide association study (GWAS) to identify novel mutations implicated in resistance in a public global cohort (n=555), and a new clinical cohort from three European centers (n=322), respectively.
Results:
Isolates characterized as gBLNAR groups II and III were associated with ampicillin resistance (p<0.03), while only group III+/III-like+ isolates were associated with cefotaxime resistance (p<0.03). However, group II isolates had low specificities (≤ 71%) to rule in resistance against ampicillin, challenging the currently endorsed clinical breakpoint for its ability to distinguish between gBLNAR and susceptible wildtype isolates. Known and newly identified PBP3 substitutions often occur step-wise and show patterns of positive selection, and convergent evolution in the two independent cohorts. Furthermore, we discovered new genes and novel combinations of PBP3 substitutions associated with ampicillin resistance.
Conclusion:
This study reveals new insights on molecular ampicillin resistance determinants in β-lactamase-negative H. influenzae isolates. We emphasize an artifact with the currently applied clinical breakpoints for ampicillin. Further investigations, including treatment outcome data, are crucial to understand the impact of bacterial genome variations on susceptibility to cefotaxime and other cephalosporins.