Background Several SARS-CoV-2 variants of concern (VOC) have emerged through 2020 and 2021. There is need for tools to estimate the relative transmissibility of emerging variants of SARS-CoV-2 with respect to circulating strains. Aim We aimed to assess the prevalence of co-circulating VOC in Italy and estimate their relative transmissibility. Methods We conducted two genomic surveillance surveys on 18 February and 18 March 2021 across the whole Italian territory covering 3,243 clinical samples and developed a mathematical model that describes the dynamics of co-circulating strains. Results The Alpha variant was already dominant on 18 February in a majority of regions/autonomous provinces (national prevalence: 54%) and almost completely replaced historical lineages by 18 March (dominant across Italy, national prevalence: 86%). We found a substantial proportion of the Gamma variant on 18 February, almost exclusively in central Italy (prevalence: 19%), which remained similar on 18 March. Nationally, the mean relative transmissibility of Alpha ranged at 1.55–1.57 times the level of historical lineages (95% CrI: 1.45–1.66). The relative transmissibility of Gamma varied according to the assumed degree of cross-protection from infection with other lineages and ranged from 1.12 (95% CrI: 1.03–1.23) with complete immune evasion to 1.39 (95% CrI: 1.26–1.56) for complete cross-protection. Conclusion We assessed the relative advantage of competing viral strains, using a mathematical model assuming different degrees of cross-protection. We found substantial co-circulation of Alpha and Gamma in Italy. Gamma was not able to outcompete Alpha, probably because of its lower transmissibility.