A total of 42 Newcastle disease virus (NDV) strains were isolated from clinical samples obtained from chickens during field outbreaks among vaccinated broiler chicken's farms in the Upper Egypt in 2011 - 2012. The samples were inoculated into embrayonated chicken eggs and the positive samples for NDV were detected by hemagglutinination (HA) and confirmed by hemagglutination inhibition (HI) tests and reverse transcriptation polymerase chain reaction (RT-PCR). The pathogenicity of the isolates was estimated biologically by intracerebral pathogenicity index (ICPI) test and genetically by sequencing the partial fusion (F) genes. The ICPI indicated that 40 isolates were > 1.5, which characterized velogenic strains, the remaining was mesogenic (ICPI = 1.4). The sequencing analysis of the nucleotides and deduced amino acids, 8 isolates representing the isolates from different localities, revealed that these isolates possess the amino acids motif 112-R-R-Q-K- R-F-117 of the velogenic nature and a diversity of 19 –22 and 19-21 % in nucleotides and amino acids, respectively, from vaccine strains (LaSota, B1, Ulster/67 and I-2) and Texas/GB. Interesting, that these isolates clustered into two subgroups related to the isolates from Jordan, Israel (2011) and China but were distant from vaccine, Egyptian (2005, 2006 and 2010), Sudan, Saudi Arabia and Israel (2003) strains by phylogenetic analysis. In conclusion, a velogenic NDV prove to circulate among chickens in southern of Egypt although intensive vaccination programs were conducted. These isolates were genetically away from the vaccine strains and virulent to chickens. The necessity needs to continuously monitor the virus and evaluate the vaccination programs efficiency.