Objective: To assess the pathogenicity and to categorize the high pathogenic avian influenze (HPAI) H5N1 of four isolates from different outbreaks during 2007. Methods: Primary assays of pathogenicity test were used such as mean death time (MDT), intracerebral pathogenicity index (ICPI) and intravenous pathogenicity index. Results: The MDT of isolate 1, 2, 3 and 4 was 35 h, 36 h, 38 h and 35 h respectively in ten-day old chicken embryonated eggs. The ICPI in one day old chick for all isolates were more than 0.7 and the intravenous pathogenicity indexes in six-week old specific pathogen free chicken for all isolates were more than 0.7 and 1.2. Conlusions: It is the first report of pathogenicity assessment of H5N1 virus circulating in Pakistan