Herpesviruses account for most of the known virus-encoded miRNAs. Herpesvirus of turkey (HVT), a non-pathogenic avian herpesvirus used as an avian vaccine and viral vector, encodes 28 mature miRNAs. This included HVT-miR-H14-3p that showed almost identical sequence to gga-miR-221, suggesting that it is pirated from the avian host. Although the functional homolog between the two miRNAs has been proposed based on the sequence similarity, the direct experimental evidence is still lacking. In this report, we provide the evidence for the first time that HVT-miR-H14-3p is indeed a gga-miR-221 homolog through modulating the expression of p27Kip1, a known target of miR-221 by binding to its 3’UTR. We also created an HVT-miR-H14-3p deletion virus and show that this miRNA is not essential for in vitro replication.