In a multicentre study, influenza A/H1N1/09v 222G/N variants were more frequently detected in patients admitted to the intensive-care unit for invasive mechanical ventilation or extracorporeal membrane oxygenation (10/23; 43.5%) than in patients hospitalized in other units (2/27; 7.4%) and community patients (0/81; 0.0%) (p <0.01). A significantly higher virus load (p 0.02) in the lower vs the upper respiratory tract was observed. Predominance of 222G/N variants in the lower respiratory tract (40% of total virus population) vs the upper respiratory tract (10%) was shown by clonal analysis of haemagglutinin sequences in paired nasal swab and bronchoalveolar lavage samples. The time from illness onset to sampling was significantly longer in patients with severe infection vs community patients (p <0.001). It was concluded that the 222G/N variants showed increased virulence; mutant variants were probably selected in individual patients; and the longer duration of illness might have favoured the emergence of adaptive mutations through multiple replication cycles.