Evaluation of the genetic polymorphism among Lactococcus lactis subsp. cremoris strains using comparative genomic hybridization and multilocus sequence analysis Amel Taïbi a , Nassra Dabour a,b , Maryse Lamoureux c , Denis Roy a , Gisèle LaPointe a, ⁎ a STELA Dairy Research Centre, Institute of Nutraceuticals and Functional Foods, Pavillon des services, Université Laval, Québec, QC, Canada, G1V 0A6 b Department of Dairy Science and Technology, Faculty of Agriculture, University of Alexandria, Alexandria, Egypt c Agropur, Inc., 4700 Armand-Frappier Street, St. Hubert, QC, Canada, J3Z 1G5 abstract article info Available online xxxx Keywords: Lactococcus lactis subsp. cremoris Starters Cheese Genetic diversity Microarray-based comparative genomic hybridization Multilocus sequence analysis Genetic diversity of Lactococcus lactis subsp. cremoris provides an important reservoir of industrial functions. Knowledge of strain diversity is an important step for the selection of starter cultures, because technological and sensorial attributes are strain-dependent and it may help to distinguish strains with particular technological properties and performances. In the present study, microarray-based comparative genomic hybridization (CGH) and multilocus sequence analysis (MLSA) were used to investigate the genetic variation among eight strains of Lactococcus lactis subsp. cremoris. The CGH analysis allows strain grouping and identification of absent or divergent genes involved in metabolism, amino acid biosynthesis, osmoregulation and proteolysis. The MLSA clustering of strains based on the partial sequence of eight genes shows good correlation with the CGH grouping. Strains HP, ATCC 19257 and Wg2 were clustered together, followed by E8, and finally SK11 was in a separate cluster. The combined information provides genetic markers for distinguishing between strains and their variants. By selecting strains according to their genetic profiles, compatible and complementary mixed starters can be formulated, opening new avenues for industrial applications. © 2010 Elsevier B.V. All rights reserved. 1. Introduction In the dairy industry, lactococci are widely used for the production of a large variety of fermented milks and cheeses as a principle component of the starter culture (Macura and Townsley, 1983; Oberman and Libudzisz, 1998). Their importance resides mainly in lactic acid formation, the production of minor flavor and preservation components and proteolysis (Broom and Limsowtin, 1998; Kieronc- zyk et al., 2003). Lactococcal cultures play a key role for determining the quality of fermented dairy products with respect to shelf-life and sensory quality (Smit et al., 2005). To date, two principal subspecies of lactococci are used intensively in dairy starters, Lactococcus lactis subsp. lactis and Lactococcus lactis subsp. cremoris. For Canadian Cheddar cheese manufacture, L. lactis subsp. cremoris are preferred over L. lactis subsp. lactis because of their improved contribution to flavor production through degradation of bitter peptides (Salama et al., 1991; Sandine, 1988; Vedamuthu et al., 1966). The diversity of lactococcal strains provides an important reservoir for industrial starters and may contribute to flavor differences and specific features detected for the same product. Many L. lactis subsp. lactis and cremoris strains are known to carry plasmids encoding important traits such as lactose catabolism, citrate utilization, proteinase production, bacteriocin production and immunity, bacte- riophage resistance, exopolysaccharide production, as well as heavy metal resistance (Davidson et al., 1996; McKay, 1983). As mobile elements, plasmids can be lost or acquired and contribute to an important polymorphism among lactococcal strains, even within the same culture. Moreover, genomic diversity is the origin of differences in strain performance. Knowledge about strain diversity at the subspecies level is an important step for starter culture selection, because technological and sensorial potential can be strain-depen- dent characters. Several molecular techniques, including DNA–DNA hybridization, small subunit (SSU) rRNA gene sequencing, and PCR fingerprint analysis have been used to differentiate between L. lactis subsp. lactis and L. lactis subsp. cremoris (Erlandson and Batt, 1997; Mangin et al., 1999; Pu et al., 2002; Salama et al., 1991). However, little is known about the genomic diversity at the subspecies level of the L. lactis subsp. cremoris strains. Genomic comparison at the intra-subspecies level was performed between L. lactis subsp. cremoris strains of the NCDO712 family using pulsed-field gel electrophoresis (PFGE) (Campo et al., 2002). Identical profiles were reported for most of the strains tested, suggesting that the differences can be attributed to plasmid content and minor chromosomal mutation or rearrange- ments (Campo et al., 2002). Samarzija et al. (2002) investigated the genetic diversity of new isolates of L. lactis subsp. cremoris strains from International Journal of Food Microbiology xxx (2010) xxx–xxx ⁎ Corresponding author. Tel.: + 1 418 656 2131x3100; fax: + 1 418 656 3353. E-mail address: gisele.lapointe@fsaa.ulaval.ca (G. LaPointe). FOOD-05054; No of Pages 9 0168-1605/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.ijfoodmicro.2010.02.020 Contents lists available at ScienceDirect International Journal of Food Microbiology journal homepage: www.elsevier.com/locate/ijfoodmicro ARTICLE IN PRESS Please cite this article as: Taïbi, A., et al., Evaluation of the genetic polymorphism among Lactococcus lactis subsp. cremoris strains using comparative genomic hybridization and..., International Journal of Food Microbiology (2010), doi:10.1016/j.ijfoodmicro.2010.02.020