Detection and classification of the integrative conjugative elements of Lactococcus lactis.

Lactococcus lactis is widely applied by the dairy industry for the fermentation of milk into products such as cheese. Adaptation of L. lactis to the dairy environment often depends on functions encoded by mobile genetic elements (MGEs) such as plasmids. Other L. lactis MGEs that contribute to industrially relevant traits like antimicrobial production and carbohydrate utilization capacities belong to the integrative conjugative elements (ICE). Here we investigate the prevalence of ICEs in L. lactis using an automated search engine that detects colocalized, ICE-associated core-functions (involved in conjugation or mobilization) in lactococcal genomes. This approach enabled the detection of 36 candidate-ICEs in 69 L. lactis genomes. By phylogenetic analysis of conserved protein functions encoded in all lactococcal ICEs, these 36 ICEs could be classified in three main ICE-families that encompass 7 distinguishable ICE-integrases and are characterized by apparent modular-exchangeability and plasticity. Finally, we demonstrate that phylogenetic analysis of the conjugation-associated VirB4 ATPase function differentiates ICE- and plasmid-derived conjugation systems, indicating that conjugal transfer of lactococcal ICEs and plasmids involves genetically distinct machineries. Our genomic analysis and sequence-based classification of lactococcal ICEs creates a comprehensive overview of the conserved functional repertoires encoded by this family of MGEs in L. lactis, which can facilitate the future exploitation of the functional traits they encode by ICE mobilization to appropriate starter culture strains.

High throughput screening of technological and biopreservation traits of a large set of wild lactic acid bacteria from Brazilian artisanal cheeses.

This study aimed to evaluate technological (acidification, proteolysis, lipolysis, resistance to low pH, NaCl, and bile salts) and biopreservation (antimicrobial activity against foodborne pathogens) features of 1002 LAB by high throughput screening (HTS) methods. The LAB was isolated from 11 types of Brazilian artisanal cheeses (BAC) marketed in the main 5 producing regions. Remarkable intra-species variability in acidification rates have been found, which was most pronounced between isolates from Mina's artisanal cheeses, Caipira and Coalho cheeses. Lacticaseibacillus paracasei and Levilactobacillus brevis showed the fastest acidification rate; however, all isolates showed slower acidification rates than a lactococcal control strain (4.3 × lower). When testing inhibitory effects, > 75% of LAB isolates could inhibit the growth of Staphylococcus aureus ATCC 19095 and Listeria monocytogenes ATCC 7644. Two of these isolates, identified as Lactiplantibacillus plantarum and Lentilactobacillus buchneri, the sterile and neutral supernatants alone, were sufficient to inhibit L. monocytogenes growth. Principal component analysis (PCA) allowed the identification of functional groups based on proteolytic and lipolytic activity, osmotic stress resistance, and inhibition of L. monocytogenes. The type of cheese the isolates were recovered from influenced properties such as anti-listerial compounds and lipolytic enzyme production. The use of HTS and multivariate statistics allowed insights into a diverse set of LAB technological and biopreservation properties. These findings allow a profound knowledge of the heterogeneity of a large set of isolates, which can be further used to design starter cultures with varied and combined properties, such as biopreservation and technological features. Besides that, HTS makes it possible to analyze a vast panel of LAB strains, reducing costs and time within laboratory analysis, while avoiding the loss of information once all LAB are tested at the same time (differently from the traditional labor-intensive approach, in which a few numbers of strains is tested per time).