Background: Polyomaviruses (PyV) are small, circular, non-enveloped viruses with a diameter of about 40–50 nm, and have a double-stranded DNA molecule descended from the Polyomaviridae family. 32 PyV species are described, more than 10 of which are reported to infect humans (HPyVs). Aims of study: to identify the sequence of the complete genome of the WU polyomavirus strain to study phylogenetic analysis and determine variation in the WUPyV sequence. Methods: 195 samples were collected to identify the viral DNA by real-time and conventional PCR, respectively. One purified viral DNA sample of positive cases was sent for NGS to identify the WGS. Results: The WUPyV sequence is registered in the NCBI GenBank under the accession number LC771087. The current study showed that the WUPyV sequence has a length of 5229 bp. The WUPyV sequence was compared to reference sequences to find any mutations that occurred. By comparing the sequences, it was found that there were many mutations, some of which were silent, and the others caused a change in the amino acid. Ten reference strains from NCBI were used to create a phylogenetic tree for the studied clinical sample. The present results revealed that the strain LC771087.1 WU Polyomavirus showed great similarity with one sequence, EU711054.1 WU Polyomavirus strain WU/ Wuerzburg/01/03. Conclusion: The current study concluded that WU polyomaviruses used in the multiple sequence alignment with our strain have the same length 5229 bp, with 99.5% similarity in DNA sequences and 99.2% identity of amino acid sequences.