The non-structural (NS) gene of highly pathogenic avian influenza viruses of the H5N1 subtype (HPAI-H5N1) isolated in Baltic Sea area of Sweden in 2006 was studied. The phylogenetic analysis data demonstrated that two distinct sub-lineages of HPAI-H5N1 were circulating during the outbreak in Northern Europe in Spring 2006. Sub-lineage I viruses fell into the same clade as viruses found in Denmark and Germany and formed a sub-clade which also included viruses isolated in the Russian Federation in late 2005. Sub-lineage II viruses formed a sub-clade closely related to European, Middle Eastern and African isolates reported in 2006. Analysis of the inferred amino acid sequences of the NS1 protein showed a deletion of five amino acids at positions 80-84. No viruses represented in this study contained Glu92 in the NS1 and all isolates contained the avian-like ESKV amino acid sequences at the NS1 C-terminal end. Sub-lineage I isolates contained unique substitutions V194I in NS1 and G63E in Nuclear export protein (NEP).