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Arginine deiminase pathway activity and nucleoside catabolism in the fermented-meat starter culture Lactobaciilus sakei

dinsdag, 7 februari, 2012 - 17:00
Campus: Brussels Humanities, Sciences & Engineering campus
Faculteit: Science and Bio-engineering Sciences
D
0.07
Tom Rimaux
doctoraatsverdediging

The industrial production of fermented sausages is mostly performed by the use of starter
cultures. In this way the fermentation process can be controlled and shortened, and final
fermented products with improved quality and safety can be obtained. Lactobacillus sakei is the
most dominant lactic acid bacterium present in most artisan sausage fermentations and is,
therefore, often applied as a starter culture for fermented sausage production.

This study dealt with the identification of several metabolic factors, encoded in the genome
sequence of L. sakei, which could, at least partially, explain the superior dominance of L. sakei
in meat fermentations. Although glucose is the preferential energy source for L. sakei, its
concentration is rapidly depleted during meat fermentation. For this reason, a versatile use of
energy sources other than glucose present in meat can improve the competitiveness and
survival of L. sakei in the meat matrix. Therefore, this study focused on the capacity of L. sakei
to grow on and convert arginine, nucleosides (inosine and adenosine), and glycerol, energy
sources that are constantly released during fermentation and ripening of sausages.

All energy sources, except for glycerol, were consumed by several L. sakei strains
tested in this study. Using a modelling approach, the conversion of arginine and nucleosides
was described in detail for L. sakei CTC 494, a natural and highly competitive isolate from a
Spanish spontaneously fermented sausage. The catabolism of both energy sources operated
maximally under slightly acidic conditions, which are typical for sausage fermentations. The
catabolism of nucleosides resulted in the production of a mixture of organic acids (lactic acid,
acetic acid, and formic acid) and ethanol, while arginine conversion through the arginine
deiminase (ADI) pathway resulted in the production of both citrulline and mainly ornithine.

The final ratio of citrulline to ornithine was modulated by environmental pH, with the highest
production of ornithine under acidic conditions, thereby providing the strain with an energetic
advantage. The genes coding for the ADI pathway in L. sakei CTC 494 were clustered in a
single arc operon (arcABCTDR), showing the exact gene order as in L. sakei 23K. Moreover,
differential expression of these genes was growth phase- and strain-dependent, suggesting a
different role of the ADI pathway in different strains.

This study also unraveled the functional role of a previously uncharacterized gene, which is
located downstream of the other genes of the arc operon in L. sakei CTC 494. This gene was
shown to encode a citrulline/ornithine antiporter that is involved in the uptake of extracellular
citrulline and thus contributes to the further conversion of citrulline into ornithine. Relative
expression of this gene showed a different modulation as a function of environmental pH
compared to the other genes of the arc operon. This differential modulation might result in an
increased flexibility of the operation of the ADI pathway under stress conditions. Finally, this
study identified the presence of an operational agmatine deiminase pathway for only a few
strains of L. sakei, formerly annotated as a second arc operon. These L. sakei strains were thus
able to convert extracellular agmatine into putrescine, both biogenic amines that are undesirable
in meat.

The results obtained in this work contribute to the understanding of the adaptation mechanisms
of L. sakei to the meat matrix, aiding its competitiveness and dominance in meat ecosystems. In
particular, the contribution of the ADI pathway and the catabolism of nucleosides to the general
competitiveness and adaption of L. sakei were studied. The modelling approach allowed a
quantitative description of these metabolic traits, which, when applied consistently, should allow
for an objective comparison of various starter cultures and, hence, a rational choice for their
commercial implementation in fermented sausage production.