The minimum inhibitory concentration (MIC) of five antibiotics and the presence of resistance genes was determined in 163 Haemophilus influenzae isolates collected over 13 years (1987-2000) in four two-yearly sampling periods from patients with respiratory tract infections. The prevalence of beta-lactamase-negative ampicillin-susceptible strains was approximately 80% over the sampling period although fewer strains (65.9%) were recovered in the period 1995-1997. TEM-1 type beta-lactamase-producing strains were less frequent starting at 15.6% and declining to 2.2% in the final sampling period. Low-beta-lactamase-negative ampicillin-resistant (BLNAR) strains were uncommon in 1987-1989 (2.2%), peaked to 19.5% in 1995-1997, but fell back to 11.1% by 2000. Fully BLNAR strains were not detected until the last sampling period (6.7%). The MICs of ampicillin, levofloxacin, cefditoren and ceftriaxone remained stable but there was an eight-fold increase in the MIC of cefdinir over the sampling period. Pulsed-field gel electrophoresis of DNA digests showed that three representative BLNAR strains were genetically distinct and 11 DNA profiles were identified among 17 low-BLNAR strains. These data suggest that the number of genetically altered BLNAR and low-BLNAR strains are increasing in Japan.