Lactobacillus sakei is a food-borne bacterium naturally found in meat and fish products. A study was performed to examine the intraspecies diversity among 73 isolates sourced from laboratory collections in several different countries. Pulsed-field gel electrophoresis analysis demonstrated a 25% variation in genome size between isolates, ranging from 1,815 kb to 2,310 kb. The relatedness between isolates was then determined using a PCR-based method that detects the possession of 60 chromosomal genes belonging to the flexible gene pool. Ten different strain clusters were identified that had noticeable differences in their average genome size reflecting the natural population structure. The results show that many different genotypes may be isolated from similar types of meat products, suggesting a complex ecological habitat in which intraspecies diversity may be required for successful adaptation. Finally, proteomic analysis revealed a slight difference between the migration patterns of highly abundant GapA isoforms of the two prevailing L. sakei subspecies (sakei and carnosus). This analysis was used to affiliate the genotypic clusters with the corresponding subspecies. These findings reveal for the first time the extent of intraspecies genomic diversity in L. sakei. Consequently, identification of molecular subtypes may in the future prove valuable for a better understanding of microbial ecosystems in food products.In foods, the need for microorganisms to adapt to different technological and ripening processes may result in the evolution of strain differences. Unfortunately, intraspecies genetic variations among food-borne bacteria are a largely unexplored area, so we have little understanding of the interactive mechanisms taking place within complex microbial communities existing in food ecosystems. This is particularly relevant in the case of Lactobacillus sakei, a meatborne lactic acid bacterium potentially useful as a meat biopreservative (6, 39). L. sakei has been isolated from a range of meat and fish products, where it is the predominant Lactobacillus species (8). Ecologically, meat can be viewed as a diverse and changing environment that influences the growth potential of a variety of bacterial species during storage (27). An implication of survival in such an environment is that meat-borne bacteria may diverge genetically as they evolve mechanisms to acclimatize and compete in local microenvironments. Indeed, L. sakei strains are known to display a range of key phenotypic differences that have resulted in difficulties in their classification (23,35), and DNA-DNA reassociation analyses have revealed very low levels of relatedness (as low as 72%) between otherwise well-characterized L. sakei strains, indicating that the species exhibits important elements of genetic heterogeneity (7). However, it is not yet known if a strong relationship exists between the niche competition properties of L. sakei in meat products and the genetic diversity between strains.Currently, L. sakei is divided into two subspecies b...