The two most commonly used wine microorganisms, Saccharomyces cerevisiae yeast and Oenococcus oeni bacteria, are responsible for completion of alcoholic and malolactic fermentation (MLF), respectively. For successful co-inoculation, S. cerevisiae and O. oeni must be able to complete fermentation; however, this relies on compatibility between yeast and bacterial strains. For the first time, quantitative trait loci (QTL) analysis was used to elucidate whether S. cerevisiae genetic makeup can play a role in the ability of O. oeni to complete MLF. Assessment of 67 progeny from a hybrid S. cerevisiae strain (SBxGN), co-inoculated with a single O. oeni strain, SB3, revealed a major QTL linked to MLF completion by O. oeni. This QTL encompassed a well-known translocation, XV-t-XVI, that results in increased SSU1 expression and is functionally linked with numerous phenotypes including lag phase duration and sulphite export and production. A reciprocal hemizygosity assay was performed to elucidate the effect of the gene SSU1 in the SBxGN background. Our results revealed a strong effect of SSU1 haploinsufficiency on O. oeni’s ability to complete malolactic fermentation during co-inoculation and pave the way for the implementation of QTL mapping projects for deciphering the genetic bases of microbial interactions.