Time‐resolved genetic responses of Lactococcus lactis to a dairy environment

Abstract

Lactococcus lactis is one of main bacterial species found in mixed dairy starter cultures for the production of semi-hard cheese. Despite the appreciation that mixed cultures are essential for the eventual properties of the manufactured cheese the vast majority of studies on L. lactis were carried out in laboratory media with a pure culture. In this study we applied an advanced recombinant in vivo expression technology (R-IVET) assay in combination with a high-throughput cheese-manufacturing protocol for the identification and subsequent validation of promoter sequences specifically induced during the manufacturing and ripening of cheese. The system allowed gene expression measurements in an undisturbed product environment without the use of antibiotics and in combination with a mixed strain starter culture. The utilization of bacterial luciferase as reporter enabled the real-time monitoring of gene expression in cheese for up to 200 h after the cheese-manufacturing process was initiated. The results revealed a number of genes that were clearly induced in cheese such as cysD, bcaP, dppA, hisC, gltA, rpsE, purL, amtB as well as a number of hypothetical genes, pseudogenes and notably genetic elements located on the non-coding strand of annotated open reading frames. Furthermore genes that are likely to be involved in interactions with bacteria used in the mixed strain starter culture were identified.

Dynamic Analysis of the Lactococcus lactis Transcriptome in Cheeses Made from Milk Concentrated by Ultrafiltration Reveals Multiple Strategies of Adaptation to Stresses

“…Repression of the CodY regulon at day 7 was in accordance with the accumulation in UF-cheeses of large amounts of amino acids, including isoleucine, leucine, and valine, which act as corepressors of CodY. Conversely to our results, Bachmann et al observed a global induction of CodYresponsive genes as a result of amino acid starvation in Goudatype cheeses involving the overexpression of various genes of amino acid and peptide transport and metabolism (1). Thus, in the UF-cheese, L. lactis LD61 did not undergo any nitrogen limitation due to the efficient proteolytic system of L. lactis LD61.…”

“…So far, such global approaches in food microbiology and in situ have been poorly documented. Recently, Bachmann et al presented the genetic responses of L. lactis in mixed cultures in mini-Goudatype cheeses (1). By use of a recombinant in vivo expression technology (R-IVET) assay (2), they targeted expressed genes in semihard cheeses and compared them to those in a laboratory medium (M17).…”

Microbial domestication signatures of Lactococcus lactis can be reproduced by experimental evolution

“…Comparative genomics has revealed that the acquisition of genes related to amino acid degradation and transport, as well as the loss of genes involved in the utilization of carbohydrates that do not occur in milk, are prominent dairy-specific adaptations (Siezen et al 2005;Makarova et al 2006;Makarova and Koonin 2007). Moreover, several genes involved in nitrogen metabolism and peptide transport were induced after transition to a dairy environment (Gitton et al 2005;Bachmann et al 2010). These observations, in combination with the stimulation of growth in milk of the plant isolate KF147 by the addition of amino acids (Supplemental Fig.…”

Multifactorial diversity sustains microbial community stability

“…The gad operon was shown to be induced by chloride and the decarboxylase enzyme has an optimum pH of 4.0-5.0 (Sanders et al, 1998), suggesting a role during cheese ripening. The gadR was shown to be induced in cheese by recombinant in vivo expression technology (Bachmann et al, 2010) as well. The superimposition of the metabolic maps suggests that GABA excreted by L. lactis may serve as a substrate in the formation of succinate by L. mesenteroides as TIFN8 genome encodes a predicted GABA permease, a GABA aminotransferase and succinate-semialdehyde dehydrogenase (Figure 3).…”