Background: Since the first introduction of rabbit hemorrhagic disease virus (RHDV) in 1991, Egyptian authorities started a national vaccination program against the disease. However, RHDV clinical cases are still being reported. Objectives: This study aimed to investigate genetic characterization of recent RHDV isolates from naturally-infected rabbits in Sharkia governorate, Egypt in 2015/2016. Methods: A total of 24 clinical samples were investigated by real-time reverse transcription PCR (RRT-PCR). Hemagglutination analysis of RRT-PCR positive samples were performed, also these positive RRT-PCR samples were purified and directly partial sequenced for C-terminal VP60 gene. Results: Only two samples were positive for partial C-terminal VP60 amplification. The first positive sample was collected from non-vaccinated young rabbit (45-day old) and designed here as SHAH 2015, whereas the second sample was collected from non-vaccinated adult dam (nine-month old) and designed as SHMK2016. Hemagglutination analysis using human red blood cells type O revealed that both isolates were non-hemagglutinating. Sequence analysis revealed that both isolates shared amino acid identity of 98.57 % and 95.71 % with each other and with commonly used vaccinal strain (RHDV-Giza 2006), respectively. Phylogenetic analysis revealed that both isolates clustered with the vaccine strain and isolate UT01 within genotype 6 RHDVa. Molecular analysis of antigenic regions of VP60 (part of E and F regions) revealed neither detection of recombination points nor sites under positive selection. Conclusion: The obtained results indicate that non-hemagglutinating RHDVa G6 variant viruses are currently circulating in Sharkia with the ability to infect both young and adult rabbits that may be due to inadequate application of vaccine and not due to vaccine mismatch. We recommend broad application of currently applied vaccine should include both young and adult rabbits