Influenza remains a significant global public health concern. Live-attenuated influenza vaccines (LAIVs) are recognized as effective interventions for influenza prevention. Currently, two types of LAIVs are licensed for human use: one developed through cold-adapted viral gene mutation and the other through the deletion of the viral NS1 gene. However, the similarities and differences in these two LAIVs’ efficacy, safety, and immune responses have not been thoroughly studied. This study constructed a gene-deficient live-attenuated vaccine strain, CA4-dNS1, and a gene locus-mutated attenuated vaccine strain, CA4-cold, to compare their in vivo and in vitro replication capacity, broad-spectrum protective efficacy, safety, and immunogenicity. The results showed that both LAIVs provide comparable broad-spectrum protection against lethal H1N1 and H5N1 influenza challenges in mice and induce similar humoral and mucosal immune responses. Notably, the CA4-cold vaccine strain induces superior influenza memory T-cell responses, while the CA4-dNS1 vaccine strain demonstrates greater safety. These findings underscore the importance of gene modification in LAIVs in striking a balance between their safety and efficacy. The NS1 gene-deficient CA4-dNS1 strain may offer a more advantageous approach for developing next-generation LAIVs and other intranasal influenza virus vectored vaccines due to enhanced safety.