Cefotaxime resistance in invasive Haemophilus influenzae isolates in Germany 2016-19: prevalence, epidemiology and relevance of PBP3 substitutions

Background: Haemophilus influenzae can cause invasive infections, in which cefotaxime is among the first-Line antibiotics for treatment. The prevalence of cefotaxime-resistant H. influenzae in Europe is reported to be on a Low Level. Nevertheless, systematic studies with a Large set of invasive isolates are scarce. Objectives: To provide prevalence data for cefotaxime resistance in invasive H. influenzae isolates in Germany 2016-19 and investigate the epidemiological relevance of PBP3 mutations known to elevate the cefotaxime MIC. Methods: Cefotaxime susceptibility of invasive H. influenzae isolates, collected in the national Laboratory surveillance programme, was examined by gradient agar diffusion (GAD) testing. Cefotaxime resistance was determined according to EUCAST guidelines (resistance breakpoint MIC >0.125 mg/L). Therefore, the MIC for all resistant isolates was verified by broth microdilution method (BMD). WGS was performed to investigate the genetic relationship of cefotaxime-resistant isolates and to analyse alterations in the PBP3. An analysis of the geographic distribution of the resistant isolates was performed. Results: From 2016 to 2019, the German National Reference Laboratory for Meningococci and H. influenzae received 2432 invasive H. influenzae isolates from blood and CSF. According to GAD results, 27 strains were resistant to cefotaxime. BMD confirmed the resistance in 22 of these isolates, which equals a prevalence of cefotaxime resistance of 0.90% in invasive H. influenzae in Germany. Among cefotaxime-resistant isolates cgMLST revealed three clusters. PBP3 analysis showed previously described mutations in our strains. In comparison with cefotaxime-susceptible strains, the alterations L389F and Y557H were significantly associated with cefotaxime resistance, but were not present in all resistant strains. Geographic analysis showed that the disease cases with cefotaxime-resistant H. influenzae were evenly spread throughout the population in Germany. Conclusions: Cefotaxime is still well suited for the treatment of invasive H. influenzae infections. Rarely occurring cefotaxime resistance is caused by sporadic mutations. The role of PBP3 mutations needs further investigation.