The activity of garenoxacin was assessed against 412 Streptococcus pneumoniae isolates (49.3% from the adult population). Overall penicillin, erythromycin, and ciprofloxacin (MIC, greater than or equal to4 mug/ml) resistance was 51.7, 35.4, and 1.5%, respectively. For all isolates, the garenoxacin MIC was less than or equal to1 mug/ml. Amino acid replacements in GryA (Ser81 -->Phe or Tyr), ParC (Ser79 -->Phe or Tyr; Asp83 --> Gly; Lys137-->Asn), and ParE (Ile460-->Val; Asp435-->Asn), alone or in combination, were ascribed to the reduced garenoxacin susceptibility (MIC range, 0.5 to 1 mug/ml) found in four isolates. The low impact of these mutations on garenoxacin activity envisages the possible coverage of S. pneumoniae populations resistant to preexisting quinolones.