Abstract: H9N2 influenza viruses have been circulating in Iran since 1998 but a systematic investigation of H9N2 in Iran has not been undertaken since 2002. The present study aimed to analyze and evolution of H9N2 influenza viruses in Iran from 2008 to 2009. To determine the genetic relationship of Iranian viruses, the PB2 genes from five isolates of H9N2 viruses isolated from commercial chickens in Iran during 2008-2009 were amplified and sequenced. The Iranian isolates did not exhibit insertions or deletions within (PB2) gene as compare with their prototype A/turkey/winconsin/66 but rather numerous point mutations were registered. The PB2 gene of Iranian viruses showed great genetic diversity and shared a high level of similarity with PB2 genes from H7 subtype rather than with established H9N2 Eurasian lineages. The results of this study indicated that during recent years the Iranian H9N2 viruses have undergone extensive genetic reassortment which led to the generation of H9N2 viruses with novel genotypes.