Mutations within the N-terminal domain (NTD) of the spike (S) protein play a pivotal role in the emergence of successful SARS-CoV-2 viral lineages. This study investigates the influence of novel combinations of NTD lineage-defining mutations found in the Alpha, Delta, and Omicron variants on viral success. We performed comparative genomics of more than 10 million public SARS-CoV-2 samples to decipher the transmission success of different NTD markers. Additionally, we characterized the viral phenotype of such markers in a surrogate in vitro system. We found that viruses bearing repaired deletions SDeltaH69/V70 and SDeltaY144 in Alpha background were associated with increased transmission rates. After the emergence of the Omicron BA.1 lineage, Alpha viruses harbouring both repaired deletions still showed increased transmission compared to their BA.1 counterparts. Remarkably, Alpha viruses with the SDeltaH69/V70 repair displayed the highest emergence rate, while those in BA.1 exhibited the lowest. Moreover, repaired deletions were more frequently observed among older individuals infected with Alpha, but not with BA.1. In vitro biological characterization of Omicron BA.1 spike deletion repair patterns revealed substantial differences with Alpha. In BA.1, SDeltaV143/Y145 repair enhanced fusogenicity and susceptibility to neutralization by vaccinated individuals' sera. In contrast, the SDeltaH69/V70 repair did not significantly alter these traits but reduced viral infectivity. Simultaneous repair of both deletions led to lower fusogenicity. These findings highlight the intricate genotype-phenotype landscape of the spike NTD in SARS-CoV-2, which impacts viral biology, transmission efficiency, and susceptibility to neutralization. Overall, this study advances our comprehension of SARS-CoV-2 evolution, carrying implications for public health and future research.