Urtica dioica agglutinin (UDA) is a carbohydrate-binding small monomeric protein isolated from stinging nettle rhizomes. It inhibits replication of a broad range of viruses, including coronaviruses, in multiple cell types, with appealing selectivity. In this work, we investigated the potential of UDA as a broad-spectrum antiviral agent against SARS-CoV-2. UDA potently blocks transduction of pseudotyped SARS-CoV-2 in A549.ACE2⁺-TMPRSS2 cells, with IC₅₀ values ranging from 0.32 to 1.22 µM. Furthermore, UDA prevents viral replication of the early Wuhan-Hu-1 strain in Vero E6 cells (IC₅₀ = 225 nM), but also the replication of SARS-CoV-2 variants of concern, including Alpha, Beta and Gamma (IC₅₀ ranging from 115 to 171 nM). In addition, UDA exerts antiviral activity against the latest circulating Delta and Omicron variant in U87.ACE2⁺ cells (IC₅₀ values are 1.6 and 0.9 µM, respectively). Importantly, when tested in Air-Liquid Interface (ALI) primary lung epithelial cell cultures, UDA preserves antiviral activity against SARS-CoV-2 (20A.EU2 variant) in the nanomolar range. Surface plasmon resonance (SPR) studies demonstrated a concentration-dependent binding of UDA to the viral spike protein of SARS-CoV-2, suggesting interference of UDA with cell attachment or subsequent virus entry. Moreover, in additional mechanistic studies with cell-cell fusion assays, UDA inhibited SARS-CoV-2 spike protein-mediated membrane fusion. Finally, pseudotyped SARS-CoV-2 mutants with N-glycosylation deletions in the S2 subunit of the spike protein remained sensitive to the antiviral activity of UDA. In conclusion, our data establish UDA as a potent fusion inhibitor for the current variants of SARS-CoV-2.