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Amantadine and rimantadine are anti-influenza A drugs that target the M2 protein, located within the viral envelope. Over the last century, the circulating human Influenza A viruses in the UK have included H1N1, H2N2 and H3N2 antigenic subtypes. The main aim of this study was to analyse the national archive of influenza A viruses for amantadine susceptibility as there is limited data on natural resistance over the entire period of circulation of influenza A viruses. Over 2300 influenza A viruses were screened from a period representing all influenza seasons and antigenic variants over the last 40 years in the UK. Phenotypic and genotypic analyses determined the frequency of amantadine-resistance. Enzyme- linked immunoassay and plaque reduction assay determined resistance to be 2.4% and 2.1% for H3N2 and MINI viruses, respectively. No evidence of amantadine-resistance was found in the limited number of H2N2 viruses screened. Amino acid mutations within the M2 protein transmembrane domain were found in 22/48 of resistant viruses. Serine to asparagine substitutions at amino acid position 31 were most frequent, conferring resistance in 9/22 viruses. A large group of phenotypically resistant viruses did not contain any M2 transmembrane mutations; no other mutations within the matrix genes could be attributed to the resistance. The occurrence of resistance over time was sporadic and often occurred in clusters. Some of the clusters were traced to outbreaks of influenza where amantadine had been used to limit the spread of disease. The emergence of natural resistance within the population was low and did not persist through influenza seasons. Analysis of the M gene sequence indicated that the two matrix gene products. M1 and M2, evolved independently of each other. M2 evolved at a faster rate of nucleotide and amino acid change than M1. Twelve influenza A H3N2 viruses were isolated from a persistent infection of an immunocompromised host. An amantadine-resistant virus population emerged following a short course of amantadine treatment and was maintained for eighteen months in the absence of drug pressure. Virus isolates were mixtures of amantadine-sensitive and -resistant variants, which varied over time. Mutations within the haemagglutinin (HA) molecule suggested that the viral population had undergone antigenic drift over the study period. There was no detectable immune response within the host, therefore the drift, or evolution of these viruses had occurred without the selection pressure of a competent immune system. Fixed mutations within antigenically important regions of the HA molecule were found to affect receptor-binding properties of the viruses. The HA molecule had an increased rate of evolution compared to field virus strains that circulated over the same period as the persistent infection. The emergence and progression of amantadine-resistance and viral evolution within the normal and immunocompromised host will be discussed.