Lactococcus lactis subsp. lactis biovar diacetylactis strains are used in the dairy industry for generating acetoin and notably diacetyl which imparts a high level of buttery flavor notes. A collection of domesticated and environmental strains was screened for the production of diacetyl or acetoin (D/A), and citrate fermentation. Unexpectedly, both domesticated and environmental strains produced D/A. Domesticated strains belonging to the currently named “biovar diacetylactis” metabolized citrate and produced large amounts of D/A during early growth. They harbored the citP plasmid gene encoding citrate permease and a chromosomal region citM-citI-citCDEFXG involved in citrate metabolism. In these strains, citrate consumption was identified as the major determinant of aroma production. Environmental strains, specifically UCMA5716 and A12, produced as much D/A as the CitP+ strains, though at slightly lower rates. UCMA5716 was found to contain the citM-citI-citCDEFXG cluster but not the citP gene. A12 had neither. In these strains, production rate of D/A was linearly correlated with pyruvate synthesis rate. However, the correlation factor was strain-dependent, suggesting different modes of regulation for pyruvate rerouting towards fermentation end-products and flavors. This work highlights the genetic and metabolic differences between environmental and domesticated strains. The introduction of environmental strains into industrial processes could considerably increase the diversity of starters, enhancing the delivery of new technological properties. ► A collection of L. lactis subsp. lactis strains was screened for diacetyl/acetoin production. ► Whatever their origin, L. lactis subsp. lactis strains produce diacetyl/acetoin. ► Citrate pathway is the major determinant of rapid synthesis of acetoin. ► Production rate of acetoin is linearly correlated with pyruvate synthesis rate. ► This correlation is strain-dependent.