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Jean-marie Laplace - One of the best experts on this subject based on the ideXlab platform.
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purification characterization and subunits identification of the diol dehydratase of Lactobacillus collinoides
FEBS Journal, 2002Co-Authors: Nicolas Sauvageot, Axel Hartke, Yanick Auffray, Vianney Pichereau, Loic Louarme, Jean-marie LaplaceAbstract:The three genes pduCDE encoding the diol dehydratase of Lactobacillus collinoides, have been cloned for overexpression in the pQE30 vector. Although the three subunits of the protein were highly induced, no activity was detected in cell extracts. The enzyme was therefore purified to near homogeneity by ammonium sulfate precipitation and gel filtration chromatography. In fractions showing diol dehydratase activity, three main bands were present after SDS/PAGE with molecular masses of 63, 28 and 22 kDa, respectively. They were identified by mass spectrometry to correspond to the large, medium and small subunits of the dehydratase encoded by the pduC, pduD and pduE genes, respectively. The molecular mass of the native complex was estimated to 207 kDa in accordance with the calculated molecular masses deduced from the pduC, D, E genes (61, 24.7 and 19,1 kDa, respectively) and a alpha2beta2gamma2 composition. The Km for the three main substrates were 1.6 mm for 1,2-propanediol, 5.5 mm for 1,2-ethanediol and 8.3 mm for glycerol. The enzyme required the adenosylcobalamin coenzyme for catalytic activity and the Km for the cofactor was 8 micro m. Inactivation of the enzyme was observed by both glycerol and cyanocobalamin. The optimal reaction conditions of the enzyme were pH 8.75 and 37 degrees C. Activity was inhibited by sodium and calcium ions and to a lesser extent by magnesium. A fourth band at 59 kDa copurified with the diol dehydratase and was identified as the propionaldehyde dehydrogenase enzyme, another protein involved in the 1,2-propanediol metabolism pathway.
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Characterisation of the diol dehydratase pdu operon of Lactobacillus collinoides
FEMS Microbiology Letters, 2002Co-Authors: Nicolas Sauvageot, Cécile Muller, Axel Hartke, Yanick Auffray, Jean-marie LaplaceAbstract:The three genes (pduCDE) encoding the diol dehydratase of Lactobacillus collinoides were sequenced. They exhibited strong identities with the ddrABC and pduCDE genes of Klebsiella oxytoca and Salmonella enterica, respectively. These genes are part of a putative operon with at least four other genes. An eighth open reading frame was identified as homologous to the pocR gene (encoding the operon regulatory protein). Although the enzyme was detected in exponential growth phase, PduCDE activity was increased at the end of exponential phase in presence of 1,2-propanediol.
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glycerol metabolism in Lactobacillus collinoides production of 3 hydroxypropionaldehyde a precursor of acrolein
International Journal of Food Microbiology, 2000Co-Authors: Nicolas Sauvageot, Jean-marie Laplace, Kamila Gouffi, Yanick AuffrayAbstract:Lactobacillus collinoides is a lactic acid bacterium commonly found in fermenting apple juice. Although this bacterium is not particularly involved in malolactic conversion, the presence of L. collinoides in cider may have serious consequences on the product. L. collinoides is indeed considered to be responsible for the transformation of glycerol to 3-hydroxypropionaldehyde (3-HPA), a precursor of acrolein that spoils the product quality by generating bitter tastes. The purpose of our work was to evaluate the influence of environmental and culture conditions on the conversion of glycerol to 3-HPA in L. collinoides, and to obtain a DNA probe of the gene coding for glycerol dehydratase, the enzyme responsible for this conversion.
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characterization of Lactobacillus collinoides response to heat acid and ethanol treatments
Applied Microbiology and Biotechnology, 1999Co-Authors: Jean-marie Laplace, Nicolas Sauvageot, Axel Hartke, Yanick AuffrayAbstract:Tolerance to stress and cross-protection in Lactobacillus collinoides were examined after exposure to ethanol, acid or heat shock. Ethanol and heat-adapted cells demonstrate induced homologous␣tolerance and cross-resistance to acid stress. No cross-protection of acid-adapted cells against ethanol and heat stresses was observed. Heat was the only pretreatment leading to cross-protection against the two other stresses. Whole-cell protein extract analysis revealed that each treatment induced a battery of stress proteins; the synthesis of some of these polypeptides being induced by more than one condition. The greatest overlap was observed between ethanol and heat treatments. Ten proteins were found to be common to these stresses.
Nicolas Sauvageot - One of the best experts on this subject based on the ideXlab platform.
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purification characterization and subunits identification of the diol dehydratase of Lactobacillus collinoides
FEBS Journal, 2002Co-Authors: Nicolas Sauvageot, Axel Hartke, Yanick Auffray, Vianney Pichereau, Loic Louarme, Jean-marie LaplaceAbstract:The three genes pduCDE encoding the diol dehydratase of Lactobacillus collinoides, have been cloned for overexpression in the pQE30 vector. Although the three subunits of the protein were highly induced, no activity was detected in cell extracts. The enzyme was therefore purified to near homogeneity by ammonium sulfate precipitation and gel filtration chromatography. In fractions showing diol dehydratase activity, three main bands were present after SDS/PAGE with molecular masses of 63, 28 and 22 kDa, respectively. They were identified by mass spectrometry to correspond to the large, medium and small subunits of the dehydratase encoded by the pduC, pduD and pduE genes, respectively. The molecular mass of the native complex was estimated to 207 kDa in accordance with the calculated molecular masses deduced from the pduC, D, E genes (61, 24.7 and 19,1 kDa, respectively) and a alpha2beta2gamma2 composition. The Km for the three main substrates were 1.6 mm for 1,2-propanediol, 5.5 mm for 1,2-ethanediol and 8.3 mm for glycerol. The enzyme required the adenosylcobalamin coenzyme for catalytic activity and the Km for the cofactor was 8 micro m. Inactivation of the enzyme was observed by both glycerol and cyanocobalamin. The optimal reaction conditions of the enzyme were pH 8.75 and 37 degrees C. Activity was inhibited by sodium and calcium ions and to a lesser extent by magnesium. A fourth band at 59 kDa copurified with the diol dehydratase and was identified as the propionaldehyde dehydrogenase enzyme, another protein involved in the 1,2-propanediol metabolism pathway.
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Characterisation of the diol dehydratase pdu operon of Lactobacillus collinoides
FEMS Microbiology Letters, 2002Co-Authors: Nicolas Sauvageot, Cécile Muller, Axel Hartke, Yanick Auffray, Jean-marie LaplaceAbstract:The three genes (pduCDE) encoding the diol dehydratase of Lactobacillus collinoides were sequenced. They exhibited strong identities with the ddrABC and pduCDE genes of Klebsiella oxytoca and Salmonella enterica, respectively. These genes are part of a putative operon with at least four other genes. An eighth open reading frame was identified as homologous to the pocR gene (encoding the operon regulatory protein). Although the enzyme was detected in exponential growth phase, PduCDE activity was increased at the end of exponential phase in presence of 1,2-propanediol.
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etude du metabolisme du glycerol chez Lactobacillus collinoides aspects physiologiques genetiques et biochimiques
2002Co-Authors: Nicolas SauvageotAbstract:Lactobacillus collinoides LMG 18850 est une bacterie lactique heterofermentaire stricte dont la presence est regulierement retrouvee dans le cidre. Cette bacterie semble etre impliquee dans une alteration organo-leptique du cidre denommee piqure acroleique resultant de la degradation du glycerol en 3-hydroxypropionaldehyde (3-HPA) par l'enzyme diol deshydratase dependante du coenzyme B12. Lors d'une premiere etude, nous avons mis en evidence les capacites de degradation du glycerol par L. collinoides en presence de glucose. Cette degradation conduit a l'apparition d'un pic de 3-HPA durant les premiere heures de phase stationnaire. La production de cet aldehyde correspond a l'epuisement des composes pouvant etre oxydes. Un rapport molaire glycerol/glucose d'environ 3,3 a pu etre determine pour une production optimale. La presence du 3-HPA en forte concentration se revele etre letale pour la cellule. Par une approche genetique, nous avons sequence 9463 pb et identifie 10 cadres ouverts de lecture. Trois de ces genes (pduCDE) correspondent a la grande, moyenne et petite sous-unites de la diol deshydratase et presentent de fortes homologies avec les genes des autres diol deshydratases deja caracterisees. Ces genes font partie d'un operon nomme pdu ou les proteines PduABG et H correspondent respectivement a deux proteines impliquees dans la formation des carboxysomes (A et B), a la grande et petite sous-unites de la proteine de reactivation (G et H). . . [etc. ]
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glycerol metabolism in Lactobacillus collinoides production of 3 hydroxypropionaldehyde a precursor of acrolein
International Journal of Food Microbiology, 2000Co-Authors: Nicolas Sauvageot, Jean-marie Laplace, Kamila Gouffi, Yanick AuffrayAbstract:Lactobacillus collinoides is a lactic acid bacterium commonly found in fermenting apple juice. Although this bacterium is not particularly involved in malolactic conversion, the presence of L. collinoides in cider may have serious consequences on the product. L. collinoides is indeed considered to be responsible for the transformation of glycerol to 3-hydroxypropionaldehyde (3-HPA), a precursor of acrolein that spoils the product quality by generating bitter tastes. The purpose of our work was to evaluate the influence of environmental and culture conditions on the conversion of glycerol to 3-HPA in L. collinoides, and to obtain a DNA probe of the gene coding for glycerol dehydratase, the enzyme responsible for this conversion.
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characterization of Lactobacillus collinoides response to heat acid and ethanol treatments
Applied Microbiology and Biotechnology, 1999Co-Authors: Jean-marie Laplace, Nicolas Sauvageot, Axel Hartke, Yanick AuffrayAbstract:Tolerance to stress and cross-protection in Lactobacillus collinoides were examined after exposure to ethanol, acid or heat shock. Ethanol and heat-adapted cells demonstrate induced homologous␣tolerance and cross-resistance to acid stress. No cross-protection of acid-adapted cells against ethanol and heat stresses was observed. Heat was the only pretreatment leading to cross-protection against the two other stresses. Whole-cell protein extract analysis revealed that each treatment induced a battery of stress proteins; the synthesis of some of these polypeptides being induced by more than one condition. The greatest overlap was observed between ethanol and heat treatments. Ten proteins were found to be common to these stresses.
Aline Lonvaudfunel - One of the best experts on this subject based on the ideXlab platform.
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organisation of the genes encoding glycerol dehydratase of Lactobacillus collinoides Lactobacillus hilgardii and Lactobacillus diolivorans
Sciences Des Aliments, 2002Co-Authors: Andrea Gorga, Olivier Claisse, Aline LonvaudfunelAbstract:Les souches Lactobacillus collinoides IŒB 9527, Lactobacillus hilgardii IŒB 0003, et la souche IŒB 0004 appartenant a l'espece recemment decrite Lactobacillus diolivorans qui degradent le glycerol en anaerobiose ont ete etudiees. Deux amorces nucleotidiques degenerees, deduites des alignements de sequence de genes codant la glycerol deshydratase de Citrobacter freundii, Klebsiella pneumoniae, Klebsiella oxytoca, Salmonella typhimurium et Clostridium pasteurianum ont ete utilisees pour des amplifications par PCR. Les trois amplifiats de 279 pb de L. collinoides, L. hilgardii et L. diolivorans montrent de fortes homologies avec les parties de genes de glycerol deshydratases deja sequences. Les trois amplifiats ont ete utilises pour dessiner des amorces specifiques pour des reactions de PCR-inverse de facon a etudier les genes entiers des trois especes. Les fragments amplifies ont ete clones et sequences et plusieurs cadres ouverts de lecture ont ete identifies. Pour les trois especes, l'organisation des genes codant les enzymes de la degradation du glycerol suggere l'arrangement en operons. Des proteines deduites de certains ORF sont tres semblables aux proteines de la famille des carboxysomes. Les autres sont similaires aux trois sous-unites glycerol ou propanediol deshydratase dependantes d'adenosylcobalamine de C. freundii, K. pneumoniae, K. oxytoca et S. typhimurium.
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primers and a specific dna probe for detecting lactic acid bacteria producing 3 hydroxypropionaldehyde from glycerol in spoiled ciders
Journal of Food Protection, 2001Co-Authors: Olivier Claisse, Aline LonvaudfunelAbstract:Of the 40 strains isolated from several spoiled ciders where glycerol was degraded, 36 were identified as Lactobacillus collinoides, three were Lactobacillus hilgardii, and one was Lactobacillus mali. However, only 30 L. collinoides and two L. hilgardii could degrade glycerol. The glycerol dehydratase activity was shown. The main product of the transformation was 1,3 propanediol. Two DNA primers GD1 and GD2 were chosen in the region encoding one of the subunits of glycerol dehydratase of Citrobacter freundii, Klebsiella pneumoniae, Klebsiella oxytoca, Salmonella Typhimurium, and Clostridium pasteurianum. A 279-bp amplicon in polymerase chain reaction amplification was obtained with the genomic L. collinoides IOEB 9527 DNA as template. The amino acid sequence deduced from the amplicon DNA sequence showed a very high similarity and identity with the gene of gram-negative and C. pasteurianum species. After labeling, the amplicon was used as DNA probe in dot-blot hybridization with the genomic DNA of all the ...
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assimilation of glycerol by a strain of Lactobacillus collinoides isolated from cider
Food Microbiology, 2000Co-Authors: Olivier Claisse, Aline LonvaudfunelAbstract:A collection of glycerol-degrading lactic acid bacteria from spoiled cider was isolated. The most widespread species was Lactobacillus collinoides, a heterofermentative species which mainly produced the D -lactic acid isomer from glucose. Conditions of glycerol assimilation by such bacteria were investigated for strain Lb. collinoides IOEB 9527. It could not be cultivated with glycerol alone as carbon source. Sugars were necessary for growth, and fructose was preferred to glucose. Glycerol was degraded, mainly to 1,3-propanediol. Small amounts of acrolein, produced from 3-hydroxypropion-aldehyde (3-HPA) during distillation of samples, were detected. When Lb. collinoides was cultivated with glucose or fructose alone, it produced the usual compounds of the heterofermentative pathway, notably D -lactic acid and small amounts of L -lactic acid, acetic acid and ethanol. When glycerol was present, the amounts of lactic acid were surprisingly much lower and acetic acid + ethanol much higher than expected. Oxidation of D -lactic acid, and to a lesser extent of L -lactic acid, was coupled to degradation of glycerol to 1,3-propanediol. This co-metabolism leading to acetic acid provided energy. Consequently, in fermented beverages spoiled by such strains, glycerol is degraded and 3-HPA plus acetic acid are produced. Both components are undesirable for health and sensorial quality.
Yanick Auffray - One of the best experts on this subject based on the ideXlab platform.
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purification characterization and subunits identification of the diol dehydratase of Lactobacillus collinoides
FEBS Journal, 2002Co-Authors: Nicolas Sauvageot, Axel Hartke, Yanick Auffray, Vianney Pichereau, Loic Louarme, Jean-marie LaplaceAbstract:The three genes pduCDE encoding the diol dehydratase of Lactobacillus collinoides, have been cloned for overexpression in the pQE30 vector. Although the three subunits of the protein were highly induced, no activity was detected in cell extracts. The enzyme was therefore purified to near homogeneity by ammonium sulfate precipitation and gel filtration chromatography. In fractions showing diol dehydratase activity, three main bands were present after SDS/PAGE with molecular masses of 63, 28 and 22 kDa, respectively. They were identified by mass spectrometry to correspond to the large, medium and small subunits of the dehydratase encoded by the pduC, pduD and pduE genes, respectively. The molecular mass of the native complex was estimated to 207 kDa in accordance with the calculated molecular masses deduced from the pduC, D, E genes (61, 24.7 and 19,1 kDa, respectively) and a alpha2beta2gamma2 composition. The Km for the three main substrates were 1.6 mm for 1,2-propanediol, 5.5 mm for 1,2-ethanediol and 8.3 mm for glycerol. The enzyme required the adenosylcobalamin coenzyme for catalytic activity and the Km for the cofactor was 8 micro m. Inactivation of the enzyme was observed by both glycerol and cyanocobalamin. The optimal reaction conditions of the enzyme were pH 8.75 and 37 degrees C. Activity was inhibited by sodium and calcium ions and to a lesser extent by magnesium. A fourth band at 59 kDa copurified with the diol dehydratase and was identified as the propionaldehyde dehydrogenase enzyme, another protein involved in the 1,2-propanediol metabolism pathway.
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Characterisation of the diol dehydratase pdu operon of Lactobacillus collinoides
FEMS Microbiology Letters, 2002Co-Authors: Nicolas Sauvageot, Cécile Muller, Axel Hartke, Yanick Auffray, Jean-marie LaplaceAbstract:The three genes (pduCDE) encoding the diol dehydratase of Lactobacillus collinoides were sequenced. They exhibited strong identities with the ddrABC and pduCDE genes of Klebsiella oxytoca and Salmonella enterica, respectively. These genes are part of a putative operon with at least four other genes. An eighth open reading frame was identified as homologous to the pocR gene (encoding the operon regulatory protein). Although the enzyme was detected in exponential growth phase, PduCDE activity was increased at the end of exponential phase in presence of 1,2-propanediol.
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glycerol metabolism in Lactobacillus collinoides production of 3 hydroxypropionaldehyde a precursor of acrolein
International Journal of Food Microbiology, 2000Co-Authors: Nicolas Sauvageot, Jean-marie Laplace, Kamila Gouffi, Yanick AuffrayAbstract:Lactobacillus collinoides is a lactic acid bacterium commonly found in fermenting apple juice. Although this bacterium is not particularly involved in malolactic conversion, the presence of L. collinoides in cider may have serious consequences on the product. L. collinoides is indeed considered to be responsible for the transformation of glycerol to 3-hydroxypropionaldehyde (3-HPA), a precursor of acrolein that spoils the product quality by generating bitter tastes. The purpose of our work was to evaluate the influence of environmental and culture conditions on the conversion of glycerol to 3-HPA in L. collinoides, and to obtain a DNA probe of the gene coding for glycerol dehydratase, the enzyme responsible for this conversion.
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characterization of Lactobacillus collinoides response to heat acid and ethanol treatments
Applied Microbiology and Biotechnology, 1999Co-Authors: Jean-marie Laplace, Nicolas Sauvageot, Axel Hartke, Yanick AuffrayAbstract:Tolerance to stress and cross-protection in Lactobacillus collinoides were examined after exposure to ethanol, acid or heat shock. Ethanol and heat-adapted cells demonstrate induced homologous␣tolerance and cross-resistance to acid stress. No cross-protection of acid-adapted cells against ethanol and heat stresses was observed. Heat was the only pretreatment leading to cross-protection against the two other stresses. Whole-cell protein extract analysis revealed that each treatment induced a battery of stress proteins; the synthesis of some of these polypeptides being induced by more than one condition. The greatest overlap was observed between ethanol and heat treatments. Ten proteins were found to be common to these stresses.
Olivier Claisse - One of the best experts on this subject based on the ideXlab platform.
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organisation of the genes encoding glycerol dehydratase of Lactobacillus collinoides Lactobacillus hilgardii and Lactobacillus diolivorans
Sciences Des Aliments, 2002Co-Authors: Andrea Gorga, Olivier Claisse, Aline LonvaudfunelAbstract:Les souches Lactobacillus collinoides IŒB 9527, Lactobacillus hilgardii IŒB 0003, et la souche IŒB 0004 appartenant a l'espece recemment decrite Lactobacillus diolivorans qui degradent le glycerol en anaerobiose ont ete etudiees. Deux amorces nucleotidiques degenerees, deduites des alignements de sequence de genes codant la glycerol deshydratase de Citrobacter freundii, Klebsiella pneumoniae, Klebsiella oxytoca, Salmonella typhimurium et Clostridium pasteurianum ont ete utilisees pour des amplifications par PCR. Les trois amplifiats de 279 pb de L. collinoides, L. hilgardii et L. diolivorans montrent de fortes homologies avec les parties de genes de glycerol deshydratases deja sequences. Les trois amplifiats ont ete utilises pour dessiner des amorces specifiques pour des reactions de PCR-inverse de facon a etudier les genes entiers des trois especes. Les fragments amplifies ont ete clones et sequences et plusieurs cadres ouverts de lecture ont ete identifies. Pour les trois especes, l'organisation des genes codant les enzymes de la degradation du glycerol suggere l'arrangement en operons. Des proteines deduites de certains ORF sont tres semblables aux proteines de la famille des carboxysomes. Les autres sont similaires aux trois sous-unites glycerol ou propanediol deshydratase dependantes d'adenosylcobalamine de C. freundii, K. pneumoniae, K. oxytoca et S. typhimurium.
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primers and a specific dna probe for detecting lactic acid bacteria producing 3 hydroxypropionaldehyde from glycerol in spoiled ciders
Journal of Food Protection, 2001Co-Authors: Olivier Claisse, Aline LonvaudfunelAbstract:Of the 40 strains isolated from several spoiled ciders where glycerol was degraded, 36 were identified as Lactobacillus collinoides, three were Lactobacillus hilgardii, and one was Lactobacillus mali. However, only 30 L. collinoides and two L. hilgardii could degrade glycerol. The glycerol dehydratase activity was shown. The main product of the transformation was 1,3 propanediol. Two DNA primers GD1 and GD2 were chosen in the region encoding one of the subunits of glycerol dehydratase of Citrobacter freundii, Klebsiella pneumoniae, Klebsiella oxytoca, Salmonella Typhimurium, and Clostridium pasteurianum. A 279-bp amplicon in polymerase chain reaction amplification was obtained with the genomic L. collinoides IOEB 9527 DNA as template. The amino acid sequence deduced from the amplicon DNA sequence showed a very high similarity and identity with the gene of gram-negative and C. pasteurianum species. After labeling, the amplicon was used as DNA probe in dot-blot hybridization with the genomic DNA of all the ...
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assimilation of glycerol by a strain of Lactobacillus collinoides isolated from cider
Food Microbiology, 2000Co-Authors: Olivier Claisse, Aline LonvaudfunelAbstract:A collection of glycerol-degrading lactic acid bacteria from spoiled cider was isolated. The most widespread species was Lactobacillus collinoides, a heterofermentative species which mainly produced the D -lactic acid isomer from glucose. Conditions of glycerol assimilation by such bacteria were investigated for strain Lb. collinoides IOEB 9527. It could not be cultivated with glycerol alone as carbon source. Sugars were necessary for growth, and fructose was preferred to glucose. Glycerol was degraded, mainly to 1,3-propanediol. Small amounts of acrolein, produced from 3-hydroxypropion-aldehyde (3-HPA) during distillation of samples, were detected. When Lb. collinoides was cultivated with glucose or fructose alone, it produced the usual compounds of the heterofermentative pathway, notably D -lactic acid and small amounts of L -lactic acid, acetic acid and ethanol. When glycerol was present, the amounts of lactic acid were surprisingly much lower and acetic acid + ethanol much higher than expected. Oxidation of D -lactic acid, and to a lesser extent of L -lactic acid, was coupled to degradation of glycerol to 1,3-propanediol. This co-metabolism leading to acetic acid provided energy. Consequently, in fermented beverages spoiled by such strains, glycerol is degraded and 3-HPA plus acetic acid are produced. Both components are undesirable for health and sensorial quality.
