Differential effects of Lactobacillus acidophilus and Lactobacillus plantarum strains on cytokine induction in human peripheral blood mononuclear cells.

Lactic acid bacterial strains have received interest for their immunomodulating activities and potential use in probiotic products. A wide variety of strain-dependent properties have been reported, but comparative studies at the species level are scarce. The objective of this study was to assess the immunomodulatory effect of Lactobacillus species on the cytokine profiles and proliferative response of human peripheral blood mononuclear cells (hPBMC), and in particular, on the comparison between the species Lactobacillus acidophilus and Lactobacillus plantarum. hPBMC from healthy donors were stimulated in the presence or absence of the lactic acid bacteria, and cytokine production, surface marker staining, proliferation and cell death were determined after 1 and 4 days of culture. All Lactobacillus strains tested were capable of inducing the production of interleukin (IL)-1beta, IL-10, interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha). The bacterial strains did not differentially influence the amount of proliferating, viable, apoptotic and necrotic cells. Generally, L. plantarum showed a significantly higher induction capacity of IFN-gamma, IL-12 and TNF-alpha compared with L. acidophilus. We conclude that the variation in immunomodulatory effects between species is even larger than the variation between the strains of the same species. In addition, we demonstrate that L. plantarum strains are most potent in skewing the T-cell differentiation toward a putative Th1 response.

Branched chain aldehydes: production and breakdown pathways and relevance for flavour in foods.

Branched aldehydes, such as 2-methyl propanal and 2- and 3-methyl butanal, are important flavour compounds in many food products, both fermented and non-fermented (heat-treated) products. The production and degradation of these aldehydes from amino acids is described and reviewed extensively in literature. This paper reviews aspects influencing the formation of these aldehydes at the level of metabolic conversions, microbial and food composition. Special emphasis was on 3-methyl butanal and its presence in various food products. Knowledge gained about the generation pathways of these flavour compounds is essential for being able to control the formation of desired levels of these aldehydes.

Transcriptional monitoring of steady state and effects of anaerobic phases in chemostat cultures of the filamentous fungus Trichoderma reesei.

BACKGROUND: Chemostat cultures are commonly used in production of cellular material for systems-wide biological studies. We have used the novel TRAC (transcript analysis with aid of affinity capture) method to study expression stability of approximately 30 process relevant marker genes in chemostat cultures of the filamentous fungus Trichoderma reesei and its transformant expressing laccase from Melanocarpus albomyces. Transcriptional responses caused by transient oxygen deprivations and production of foreign protein were also studied in T. reesei by TRAC. RESULTS: In cultures with good steady states, the expression of the marker genes varied less than 20% on average between sequential samples for at least 5 or 6 residence times. However, in a number of T. reesei cultures continuous flow did not result in a good steady state. Perturbations to the steady state were always evident at the transcriptional level, even when they were not measurable as changes in biomass or product concentrations. Both unintentional and intentional perturbations of the steady state demonstrated that a number of genes involved in growth, protein production and secretion are sensitive markers for culture disturbances. Exposure to anaerobic conditions caused strong responses at the level of gene expression, but surprisingly the cultures could regain their previous steady state quickly, even after 3 h O2 depletion. The main effect of producing M. albomyces laccase was down-regulation of the native cellulases compared with the host strain. CONCLUSION: This study demonstrates the usefulness of transcriptional analysis by TRAC in ensuring the quality of chemostat cultures prior to costly and laborious genome-wide analysis. In addition TRAC was shown to be an efficient tool in studying gene expression dynamics in transient conditions.

Flavour formation by lactic acid bacteria and biochemical flavour profiling of cheese products.

Flavour development in dairy fermentations, most notably cheeses, results from a series of (bio)chemical processes in which the starter cultures provide the enzymes. Particularly the enzymatic degradation of proteins (caseins) leads to the formation of key-flavour components, which contribute to the sensory perception of dairy products. More specifically, caseins are degraded into peptides and amino acids and the latter are major precursors for volatile aroma compounds. In particular, the conversion of methionine, the aromatic and the branched-chain amino acids are crucial. A lot of research has focused on the degradation of caseins into peptides and free amino acids, and more recently, enzymes involved in the conversion of amino acids were identified. Most data are generated on Lactococcus lactis, which is the predominant organism in starter cultures used for cheese-making, but also Lactobacillus, Streptococcus, Propionibacterium and species used for surface ripening of cheeses are characterised in their flavour-forming capacity. In this paper, various enzymes and pathways involved in flavour formation will be highlighted and the impact of these findings for the development of industrial starter cultures will be discussed.

Identification, cloning, and characterization of a Lactococcus lactis branched-chain alpha-keto acid decarboxylase involved in flavor formation.

The biochemical pathway for formation of branched-chain aldehydes, which are important flavor compounds derived from proteins in fermented dairy products, consists of a protease, peptidases, a transaminase, and a branched-chain alpha-keto acid decarboxylase (KdcA). The activity of the latter enzyme has been found only in a limited number of Lactococcus lactis strains. By using a random mutagenesis approach, the gene encoding KdcA in L. lactis B1157 was identified. The gene for this enzyme is highly homologous to the gene annotated ipd, which encodes a putative indole pyruvate decarboxylase, in L. lactis IL1403. Strain IL1403 does not produce KdcA, which could be explained by a 270-nucleotide deletion at the 3' terminus of the ipd gene encoding a truncated nonfunctional decarboxylase. The kdcA gene was overexpressed in L. lactis for further characterization of the decarboxylase enzyme. Of all of the potential substrates tested, the highest activity was observed with branched-chain alpha-keto acids. Moreover, the enzyme activity was hardly affected by high salinity, and optimal activity was found at pH 6.3, indicating that the enzyme might be active under cheese ripening conditions.

Chemical conversion of alpha-keto acids in relation to flavor formation in fermented foods.

Formation of flavor compounds from branched-chain alpha-keto acids in fermented foods such as cheese is believed to be mainly an enzymatic process, while the conversion of phenyl pyruvic acid, which is derived from phenylalanine, also proceeds chemically. In this research, the chemical conversion of alpha-keto acids to aldehydes with strong flavor characteristics was studied, with the main focus on the conversion of alpha-ketoisocaproic acid to the aldehyde 2-methylpropanal, and a manganese-catalyzed reaction mechanism is proposed for this conversion. The mechanism involves keto-enol tautomerism, enabling molecular oxygen to react with the beta-carbon atom of the alpha-keto acid, resulting in a peroxide. This peroxide can react in several ways, leading to unstable dioxylactone or noncyclic intermediates. These intermediates will break down into an aldehyde and oxalate or carbon oxides (CO and CO(2)). All the alpha-keto acids tested were converted at pH 5.5 and in the presence of manganese, although their conversion rates were rather diverse. This chemical reaction might provide new ways for controlling cheese flavor formation with the aim of acceleration of the ripening process or diversification of the flavor characteristics.

Concomitant extracellular accumulation of alpha-keto acids and higher alcohols by Zygosaccharomyces rouxii.

Alpha-keto acids are key intermediates in the formation of higher alcohols, important flavor components in soy sauce, and produced by the salt-tolerant yeast Zygosaccharomyces rouxii. Unlike most of the higher alcohols, the alpha-keto acids are usually not extracellularly accumulated by Z. rouxii when it is cultivated with ammonium as the sole nitrogen source. To facilitate extracellular accumulation of the alpha-keto acids from aspartate-derived amino acid metabolism, the amino acids valine, leucine, threonine and methionine were exogenously supplied during batch and A-star cultivations of (routants of) Z. rouxii. It was shown that all alpha-keto acids from the aspartate-derived amino acid metabolism, except alpha-ketobutyrate, could be extracellularly accumulated. In addition, it appeared from the concomitant extracellular accumulation of alpha-keto acids and higher alcohols that in Z. rouxii, valine, leucine and methionine were converted via Ehrlich pathways similar to those in Saccharomyces cerevisiae. Unlike these amino acids, threonine was converted via both the Ehrlich and amino acid biosynthetic pathways in Z. rouxii.