Fate, inducibility, and behavior of Latilactobacillus curvatus temperate phage TMW 1.591 P1 during sausage fermentation.

AIMS: Temperate phages insert their genome into the host's chromosome. As prophages, they remain latent in the genome until an induction event leads to lytic phage production. When this occurs in a starter culture that has been added to food fermentation, this can impair the fermentation success. This study aimed to analyze prophage inducibility in the Latilactobacillus curvatus TMW 1.591 strain during meat fermentation and investigate whether an induction signal before cryopreservation is maintained during storage and can lead to phage-induced lysis after culture activation. METHODS AND RESULTS: A prophage-free isogenic derivative of the model starter organism, L. curvatus TMW 1.591, was developed as a negative control (L. curvatus TMW 1.2406). Raw meat fermentation was performed with the wild-type (WT) and phage-cured strains. The WT strain produced high numbers of phages (5.2 ± 1.8 × 107 plaque-forming units g-1) in the meat batter. However, the prophage did not significantly affect the meat fermentation process. Induction experiments suggested an acidic environment as a potential trigger for prophage induction. Phage induction by ultraviolet light before strain cryopreservation remains functional for at least 10 weeks of storage. CONCLUSIONS: Intact prophages are active during meat fermentation. However, in this study, this has no measurable consequences for fermentation, suggesting a high resiliency of meat fermentation against phages. Inadequate handling of lysogenic starter strains, even before preservation, can lead to phage introduction into food fermentation and unintended host lysis.

Impact of different sugars and glycosyltransferases on the assertiveness of Latilactobacillus sakei in raw sausage fermentations.

Latilactobacillus sakei comprises a biodiversity of strains, which display different assertiveness upon their application as starter cultures in raw sausage fermentation. While the assertiveness of winning partner strains has been referred to competitive exclusion based on genomic settings enabling occupation of multiple niches of the sausage habitat, single strain assertiveness of L. sakei remained unexplained. In this study we assessed the impact of the expression of a glycosyltransferase enabling the production of a glucan from sucrose to the assertiveness of L. sakei TMW 1.411, which expresses a plasmid-encoded glycosyltransferase (gtf). In a sausage fermentation model wild type L. sakei TMW 1.411 and its plasmid-cured mutant were employed in competition with each other and with other Latilactobacillus sakei strains. To differentiate any effects resulting from general sugar utilization from those of glucan formation, the experiments were carried out with glucose, fructose, and sucrose, respectively. It was shown that the type of sugar affects the individual strains behaviour, and that the wild type was more competitive than the mutant in the presence of any of these sugars. In direct competition between wild type and mutant, a clear competitive advantage could also be demonstrated for the strain possessing the plasmid with the glycosyltransferase. Since this competitive advantage was observed with all sugars, not just sucrose, and Gtf expression has been shown as independent of the employed sugar, it is suggested that possession of the gtf gene-carrying plasmid confers a competitive advantage. It appears that the Gtf contributes to competitive exclusion and the establishment of colonization resistance, to a larger extent by an adhesive functionality of the Gtf on the cellular surface than by the production of glucan. Hence, gtf genes can be used as a possible additional marker for the selection of assertive L. sakei starter strains in sausage fermentation.

Utilization of optimized processing conditions for high yield synthesis of conjugated linoleic acid by L. plantarum AB20-961 and L. plantarum DSM2601 in semi-dry fermented sausage.

The aim of this study was to utilize optimized processing conditions to obtain the highest conjugated linoleic acid (CLA) contents in semi-dry fermented sausages produced with L. plantarum AB20-961 and L. plantarum DSM 2601. Optimized conditions were 5.7 meat pH, 5% hydrolyzed safflower oil addition, 108 CFU/g added starter culture, fermentation time of 73 h for L. plantarum DSM2601 and 79 h for L. plantarum AB20-961, 24 °C fermentation temperature, 65 °C internal cooking temperature and 90% relative humidity. Results indicated that CLA contents in sausages were increased 21% by L. plantarum AB20-961 and 121% by L. plantarum DSM2601 after fermentation compared to initial CLA level determined on manufacturing day (P < .05). After fermentation, an increased CLA content of sausages remained stable during heat processing and storage. Sausages incorporated with L. plantarum strains and hydrolyzed safflower oil had the highest TBARS and PUFA levels, and the lowest pH and moisture content (P < .05). Differences were not found in sensorial and other physicochemical properties among sausage treatment groups. This study demonstrated that high CLA content can be achieved in sausages by utilizing optimum processing conditions described above and starter cultures (L. plantarum AB20-961 and L. plantarum DSM2601) without any adverse effects on quality of the final product.

Acid stress response of Staphylococcus xylosus elicits changes in the proteome and cellular membrane.

AIMS: Coagulase-negative Staphylococcus xylosus strains are used as starter organisms for sausage fermentation. As those strains have to cope with low pH-values during fermentation, the aim of this study was to identify the acid adaptation mechanisms of S. xylosus TMW 2.1523 previously isolated from salami. METHODS AND RESULTS: A comparative proteomic study between two different acid tolerant mutants was performed. Therefore, both S. xylosus mutants were grown pH-static under acid stress (pH 5·1) and reference conditions (pH 7·0). Proteomic data were supported by metabolite and cell membrane lipid analysis. Staphylococcus xylosus acid stress adaptation is mainly characterized by a metabolic change towards neutral metabolites, enhanced urease activity, reduced ATP consumption, an increase in membrane fluidity and changes in the membrane thickness. CONCLUSION: This study corroborates mechanisms as previously described for other Gram-positive bacteria. Additionally, the adjustment of membrane structure and composition in S. xylosus TMW 2.1523 play a prominent role in its acid adaptation. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates for the first time changes in the membrane lipid composition due to acid stress adaptation in staphylococci.

Monitoring of assertive Lactobacillus sakei and Lactobacillus curvatus strains using an industrial ring trial experiment.

AIMS: In a previous study, we used a 5-day fermenting sausage model to characterize assertiveness of Lactobacillus curvatus and Lactobacillus sakei starter strains towards employ autochthonous contaminants. In this work, we probed those findings and their transferability to real sausage fermentation including the drying process in an industrial ring trial experiment. METHODS AND RESULTS: Raw fermented sausages ('salami') were produced with three L. curvatus and four L. sakei strains as starter cultures in cooperation with three manufacturers from Germany. We monitored pH, water activity and microbiota dynamics at strain level over a total fermentation and ripening time of 21 days by MALDI-TOF-MS identification of isolates. The principal behaviour of the strains in real sausage fermentations was the same as that one observed in the 5-day model system delineating single strain assertiveness of a bacteriocin producer from co-dominance of strains. CONCLUSIONS: The water activity decrease, which is concomitant with the sausage ripening process has only limited impact on the assertiveness and survival of the starter strains. SIGNIFICANCE AND IMPACT OF THE STUDY: Results of a 5-day model can provide insight in the assertiveness of a specific starter strain in sausage fermentation.