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Early adaptation to oxygen is key to the industrially important traits of Lactococcus lactis ssp. cremoris during milk fermentation.

Cretenet M, Le Gall G, Wegmann U, Even S, Shearman C, Stentz R, Jeanson S - BMC Genomics (2014)

Bottom Line: In oxygen metabolism, the over-expression of all the genes of the nrd (ribonucleotide reductases) operon or fhu (ferrichrome ABC transports) genes was particularly significant.In carbon metabolism, the presence of oxygen led to an early shift at the gene level in the pyruvate pathway towards the acetate/2,3-butanediol pathway confirmed by the kinetics of metabolite production.An early and transitional adaptation to oxidative stress was revealed for L. lactis subsp. cremoris MG1363 in the presence of initially high levels of oxygen.

View Article: PubMed Central - PubMed

Affiliation: Institute of Food Research, Norwich Research Park, Colney, Norwich NR4 7UA, UK. sophie.jeanson@rennes.inra.fr.

ABSTRACT

Background: Lactococcus lactis is the most used species in the dairy industry. Its ability to adapt to technological stresses, such as oxidative stress encountered during stirring in the first stages of the cheese-making process, is a key factor to measure its technological performance. This study aimed to understand the response to oxidative stress of Lactococcus lactis subsp. cremoris MG1363 at the transcriptional and metabolic levels in relation to acidification kinetics and growth conditions, especially at an early stage of growth. For those purposes, conditions of hyper-oxygenation were initially fixed for the fermentation.

Results: Kinetics of growth and acidification were not affected by the presence of oxygen, indicating a high resistance to oxygen of the L. lactis MG1363 strain. Its resistance was explained by an efficient consumption of oxygen within the first 4 hours of culture, leading to a drop of the redox potential. The efficient consumption of oxygen by the L. lactis MG1363 strain was supported by a coherent and early adaptation to oxygen after 1 hour of culture at both gene expression and metabolic levels. In oxygen metabolism, the over-expression of all the genes of the nrd (ribonucleotide reductases) operon or fhu (ferrichrome ABC transports) genes was particularly significant. In carbon metabolism, the presence of oxygen led to an early shift at the gene level in the pyruvate pathway towards the acetate/2,3-butanediol pathway confirmed by the kinetics of metabolite production. Finally, the MG1363 strain was no longer able to consume oxygen in the stationary growth phase, leading to a drastic loss of culturability as a consequence of cumulative stresses and the absence of gene adaptation at this stage.

Conclusions: Combining metabolic and transcriptomic profiling, together with oxygen consumption kinetics, yielded new insights into the whole genome adaptation of L. lactis to initial oxidative stress. An early and transitional adaptation to oxidative stress was revealed for L. lactis subsp. cremoris MG1363 in the presence of initially high levels of oxygen. This enables the cells to maintain key traits that are of great importance for industry, such as rapid acidification and reduction of the redox potential of the growth media.

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Related in: MedlinePlus

Lactococcus lactisssp.cremorisstrain MG1363 + pLP712 plasmid cultivated in LM17 at 30°C under O2 condition (black lines) and N2 condition (grey lines) for [1.] dissolved oxygen concentration (—) and redox evolution (−−−), for [2.] growth kinetics: enumerations (—) and OD600(−−−) and for [3.] acidification kinetics. Values are means of biological independent triplicates (n = 3) and bars represent the standard deviations.
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Fig1: Lactococcus lactisssp.cremorisstrain MG1363 + pLP712 plasmid cultivated in LM17 at 30°C under O2 condition (black lines) and N2 condition (grey lines) for [1.] dissolved oxygen concentration (—) and redox evolution (−−−), for [2.] growth kinetics: enumerations (—) and OD600(−−−) and for [3.] acidification kinetics. Values are means of biological independent triplicates (n = 3) and bars represent the standard deviations.

Mentions: The kinetics of growth (OD and enumerations), acidification (pH), reduction (Eh7) and dissolved oxygen consumption (dissolved oxygen concentration) were monitored over 24 h for two conditions: initial hyper-oxygenized condition (O2) and no oxygen condition (N2) (Figure 1). Despite completely different gas conditions, no differences could be observed during the exponential growth phase, while substantial differences in culturability could be observed during the stationary growth phase. We defined 4 stages based on the oxygen concentration during growth under condition O2 (Figure 1). In stage A, the medium was initially saturated with oxygen (Figure 1.1) and the bacteria were in the early exponential growth phase (Figure 1.2), with minor changes in the pH (Figure 1.3) and the redox potential (Figure 1.1). In stage B, oxygen had totally been consumed and both culture media (under conditions O2 and N2) were anaerobic; bacteria were between mid- and late-exponential growth phase, the rate of acidification was at a maximum level and the redox potential decreased drastically since the oxygen concentration dropped abruptly to 0%. Until the strain MG1363 reached the stationary growth phase, its kinetics of growth and acidification was not affected by oxygen. Stage C started when the cells entered the early stationary growth phase reaching the maximum population density; acidification kinetics in conditions O2 and N2 were superimposed until a minimum pH was reached. In contrast to condition N2, the oxygen concentration in condition O2 increased again up to 50%, leading in turn to an increased redox potential. This increase of oxygen occurred because the bioreactor head-space was still filled with pure oxygen which dissolved into the LM17 medium and was presumably no longer consumed by the cells at this stage. Finally, during stage D the dissolved oxygen concentration stabilized at 50% of the initial value in condition O2 while there was still no oxygen detectable in condition N2. The L. lactis MG1363 was then in the late stationary growth phase, the minimum pH was reached (4.4 for condition N2 and 4.5 for condition O2) and the redox potential was stable (+100 mV for condition N2 and +490 mV for condition O2). The main difference between the two conditions occurred during this stage: the culturability loss was much higher under condition O2 (−2 Log10 CFU ml−1) than under condition N2 (−0.5 Log10 CFU ml-1).Figure 1


Early adaptation to oxygen is key to the industrially important traits of Lactococcus lactis ssp. cremoris during milk fermentation.

Cretenet M, Le Gall G, Wegmann U, Even S, Shearman C, Stentz R, Jeanson S - BMC Genomics (2014)

Lactococcus lactisssp.cremorisstrain MG1363 + pLP712 plasmid cultivated in LM17 at 30°C under O2 condition (black lines) and N2 condition (grey lines) for [1.] dissolved oxygen concentration (—) and redox evolution (−−−), for [2.] growth kinetics: enumerations (—) and OD600(−−−) and for [3.] acidification kinetics. Values are means of biological independent triplicates (n = 3) and bars represent the standard deviations.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4289295&req=5

Fig1: Lactococcus lactisssp.cremorisstrain MG1363 + pLP712 plasmid cultivated in LM17 at 30°C under O2 condition (black lines) and N2 condition (grey lines) for [1.] dissolved oxygen concentration (—) and redox evolution (−−−), for [2.] growth kinetics: enumerations (—) and OD600(−−−) and for [3.] acidification kinetics. Values are means of biological independent triplicates (n = 3) and bars represent the standard deviations.
Mentions: The kinetics of growth (OD and enumerations), acidification (pH), reduction (Eh7) and dissolved oxygen consumption (dissolved oxygen concentration) were monitored over 24 h for two conditions: initial hyper-oxygenized condition (O2) and no oxygen condition (N2) (Figure 1). Despite completely different gas conditions, no differences could be observed during the exponential growth phase, while substantial differences in culturability could be observed during the stationary growth phase. We defined 4 stages based on the oxygen concentration during growth under condition O2 (Figure 1). In stage A, the medium was initially saturated with oxygen (Figure 1.1) and the bacteria were in the early exponential growth phase (Figure 1.2), with minor changes in the pH (Figure 1.3) and the redox potential (Figure 1.1). In stage B, oxygen had totally been consumed and both culture media (under conditions O2 and N2) were anaerobic; bacteria were between mid- and late-exponential growth phase, the rate of acidification was at a maximum level and the redox potential decreased drastically since the oxygen concentration dropped abruptly to 0%. Until the strain MG1363 reached the stationary growth phase, its kinetics of growth and acidification was not affected by oxygen. Stage C started when the cells entered the early stationary growth phase reaching the maximum population density; acidification kinetics in conditions O2 and N2 were superimposed until a minimum pH was reached. In contrast to condition N2, the oxygen concentration in condition O2 increased again up to 50%, leading in turn to an increased redox potential. This increase of oxygen occurred because the bioreactor head-space was still filled with pure oxygen which dissolved into the LM17 medium and was presumably no longer consumed by the cells at this stage. Finally, during stage D the dissolved oxygen concentration stabilized at 50% of the initial value in condition O2 while there was still no oxygen detectable in condition N2. The L. lactis MG1363 was then in the late stationary growth phase, the minimum pH was reached (4.4 for condition N2 and 4.5 for condition O2) and the redox potential was stable (+100 mV for condition N2 and +490 mV for condition O2). The main difference between the two conditions occurred during this stage: the culturability loss was much higher under condition O2 (−2 Log10 CFU ml−1) than under condition N2 (−0.5 Log10 CFU ml-1).Figure 1

Bottom Line: In oxygen metabolism, the over-expression of all the genes of the nrd (ribonucleotide reductases) operon or fhu (ferrichrome ABC transports) genes was particularly significant.In carbon metabolism, the presence of oxygen led to an early shift at the gene level in the pyruvate pathway towards the acetate/2,3-butanediol pathway confirmed by the kinetics of metabolite production.An early and transitional adaptation to oxidative stress was revealed for L. lactis subsp. cremoris MG1363 in the presence of initially high levels of oxygen.

View Article: PubMed Central - PubMed

Affiliation: Institute of Food Research, Norwich Research Park, Colney, Norwich NR4 7UA, UK. sophie.jeanson@rennes.inra.fr.

ABSTRACT

Background: Lactococcus lactis is the most used species in the dairy industry. Its ability to adapt to technological stresses, such as oxidative stress encountered during stirring in the first stages of the cheese-making process, is a key factor to measure its technological performance. This study aimed to understand the response to oxidative stress of Lactococcus lactis subsp. cremoris MG1363 at the transcriptional and metabolic levels in relation to acidification kinetics and growth conditions, especially at an early stage of growth. For those purposes, conditions of hyper-oxygenation were initially fixed for the fermentation.

Results: Kinetics of growth and acidification were not affected by the presence of oxygen, indicating a high resistance to oxygen of the L. lactis MG1363 strain. Its resistance was explained by an efficient consumption of oxygen within the first 4 hours of culture, leading to a drop of the redox potential. The efficient consumption of oxygen by the L. lactis MG1363 strain was supported by a coherent and early adaptation to oxygen after 1 hour of culture at both gene expression and metabolic levels. In oxygen metabolism, the over-expression of all the genes of the nrd (ribonucleotide reductases) operon or fhu (ferrichrome ABC transports) genes was particularly significant. In carbon metabolism, the presence of oxygen led to an early shift at the gene level in the pyruvate pathway towards the acetate/2,3-butanediol pathway confirmed by the kinetics of metabolite production. Finally, the MG1363 strain was no longer able to consume oxygen in the stationary growth phase, leading to a drastic loss of culturability as a consequence of cumulative stresses and the absence of gene adaptation at this stage.

Conclusions: Combining metabolic and transcriptomic profiling, together with oxygen consumption kinetics, yielded new insights into the whole genome adaptation of L. lactis to initial oxidative stress. An early and transitional adaptation to oxidative stress was revealed for L. lactis subsp. cremoris MG1363 in the presence of initially high levels of oxygen. This enables the cells to maintain key traits that are of great importance for industry, such as rapid acidification and reduction of the redox potential of the growth media.

Show MeSH
Related in: MedlinePlus