Low β-Lactamase-Negative Ampicillin-Resistant Haemophilus influenzae Strains Are Best Detected by Testing Amoxicillin Susceptibility by the Broth Microdilution Method

Ampicillin resistance in Haemophilus influenzae due to alterations in penicillin-binding proteins (β-lactamase negative ampicillin resistant [BLNAR]) is acquiring increasing clinical and epidemiological importance. BLNAR strains with low ampicillin MICs (0.5 to 4 μg/ml) represent the majority of this population in Europe and the United States, but separating them from susceptible isolates is challenging. To investigate the best method to identify low-BLNAR strains, we studied the antibiotic susceptibilities of 94 clinical isolates of H. influenzae by microdilution, Etest, and disk diffusion: 25 had no resistance mechanisms (gBLNAS), 34 had mutations in the ftsI gene only (gBLNAR), 20 were β-lactamase producers only (gBLPAR), and 15 showed β-lactamase production and mutations in the ftsI gene (gBLPACR). By current CLSI breakpoints, most gBLNAR isolates were ampicillin susceptible by microdilution (76.5%) or by Etest (88.2%). Most gBLNAR strains (79.4%) were nonsusceptible to amoxicillin (the most widely used community antibiotic in the United States and Europe) when tested by microdilution. By Etest, 15% of β-lactamase-positive isolates were nonresistant to ampicillin or amoxicillin. The poorest agreement between Etest and microdilution results was for the gBLPAR isolates (25% for ampicillin, 15% for amoxicillin, and 10% for cefaclor). Low-strength disks of ampicillin and amoxicillin-clavulanic acid poorly identified low-BLNAR isolates and are not recommended as a screening method. We suggest new amoxicillin breakpoints for BLNAR isolates as follows: susceptible, MIC ≤ 0.5 μg/ml (no resistance mechanisms; pharmacokinetic/pharmacodynamic [PK/PD] data favorable); intermediate, MICs = 1 to 2 μg/ml (resistance mechanisms present but PK/PD data favorable), and resistant, MICs ≥ 4 μg/ml (resistance mechanisms present and PK/PD data unfavorable).