Molecular characterisation of a meropenem-resistant Haemophilus influenzae

Background: Haemophilus influenzae β-lactam resistance is mediated by production of β-lactamase and mutations in penicillin-binding protein 3 (PBP3, encoded by ftsI). Only one of the 646 patient-unique H. influenzae isolated in our laboratory between 2012–July 2022 was meropenem (amoxicillin, co-amoxiclav and ceftriaxone) resistant. Methods: Identification and susceptibility of this meropenem-resistant H. influenzae (MRHI) was confirmed by disk diffusion and E-test (bioMérieux) (CLSI M100). WGS was performed on GridION (Nanopore) and reference mapped with Geneious Prime 2022.2. PCR and Sanger sequencing was used to confirm results of WGS. Results: The MRHI isolate was β-lactamase negative and non-typeable. ftsI subgroup IIc1 mutations were present, signifying previously described low level PBP3 alterations that should not solely account for carbapenem resistance. Group VII2 acrR (repressor of AcrAB efflux pump) substitutions were observed but without the reported carbapenem resistance associated frameshift mutation. There were also multiple substitutions and insertions in ompP2 (outer membrane protein P2) porin, associated with carbapenem resistance.2 The 100% concordance between Sanger sequencing and Nanopore increases our confidence for application of this technology for future investigations. Conclusion: Meropenem resistance in this isolate is likely multifactorial, with PBP3 alterations and decreased influx as contributing factors. References 1. Dabernat H, et al. Diversity of β-lactam resistance-conferring amino acid substitutions in penicillin-binding protein 3 of Haemophilus influenzae. Antimicrob Agents Chemother 2002; 46: 2208–2218. 2. Cherkaoui A, et al. Imipenem heteroresistance in nontypeable Haemophilus influenzae is linked to a combination of altered PBP3, slow drug influx and direct efflux regulation. Clin Microbiol Infect 2017; 118: e9–118.e19