Influenza B virus (IBV) is considered a major human pathogen, responsible for seasonal epidemics of acute respiratory illness. Two antigenically distinct IBV HA lineages co-circulate worldwide with little cross-reactivity between each other. Live attenuated influenza virus vaccines (LAIV) have been shown to provide improved cross-protective immune responses compared to inactivated vaccines by eliciting local mucosal immunity and systemic B cell and T cell-mediated memory responses. We have previously shown that incorporation of temperature sensitive (ts) mutations in the PB1 and PB2 subunits along with a modified HA epitope tag in the C-terminus of PB1 resulted in influenza A viruses (IAV) that are safe and effective as modified live attenuated (att) virus vaccines (IAV att). We explored whether analogous mutations in the IBV polymerase subunits would result in a stable virus with an att phenotype. The PB1 subunit of the influenza B/Brisbane/60/2008 strain was used to incorporate ts mutations and a C-terminal HA tag. Such modifications resulted in a B/Bris att strain with ts characteristics in vitro and att phenotype in vivo Vaccination studies in mice showed that a single dose of the B/Bris att candidate stimulated sterilizing immunity against lethal homologous challenge and complete protection against heterologous challenge. These studies show the potential of an alternative LAIV platform for development of IBV vaccines.IMPORTANCE The licensed LAIV vaccine in the United States (FluMist) has suffered a number of issues concerning vaccine effectiveness over the past three seasons (2013-2014, 2014-2015, and 2015-2016). While the reasons for the limited robustness of vaccine-elicited immune response remains controversial, it also highlights the critical importance of continued investment in LAIV development and opens up the opportunity to improve the current strategies in order to develop more efficacious vaccines. Our laboratory has developed an alternative strategy, which incorporates 2 amino acid mutations and a modified HA tag at the C-terminus of PB1, which are sufficient to attenuate the IBV. As a LAIV, this novel strategy provides complete protection against IBV strains. The availability of attenuated IAV and IBV backbones based on contemporary strains offers alternative platforms for the development of LAIVs that may overcome current limitations.