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Quorum sensing is involved in the regulation of multicellular behavior through communication via small molecules. Given the high number and diversity of the gastrointestinal microbiota, it is postulated that members of this community communicate to coordinate a variety of adaptive processes. AI-2 is suggested to he a universal bacterial signaling molecule synthesized by the LuxS enzyme, which forms an integral part of the activated methyl cycle. We have previously reported that the well-documented probiotic strain Lactobacillus rhamnosus GG, a human isolate, produces AI-2-like molecules. In this study, we identified the luxS homologue of L. rhamnosus GG. luxS seems to be located in an operon with a yxjH gene encoding a putative cohalaminindependent methionine synthase. In silico analysis revealed a methionine-specific T box in the leader sequence of the putative yxjH-luxS operon. However, transcriptional analysis showed that luxS is expressed mainly as a monocistronic transcript. Construction of a luxS knockout mutant confirmed that the luxS gene is responsible for AI-2 production in L. rhamnosus GG. However, this mutation also resulted in pleiotropic effects on the growth of this fastidious strain. Cysteine, pantothenate, folio acid, and biotin could partially complement growth, suggesting a central metabolic role for luxS in L. rhamnosus GG. Interestingly, the luxS mutant also showed a defect in monospecies biofilm formation. Experiments with chemically synthesized (S)-4,5-dihydroxy2,3-pentanedione, coculture with the wild type, and nutritional complementation suggested that the main cause of this defect has a metabolic nature. Moreover, our data indicate that suppressor mutations are likely to occur in luxS mutants of L. rhamnosus GG. Therefore, results of luxS-related studies should be carefully interpreted.

The potential advantages of providing heterologous superoxide dismutases (SODs) to some of the intestinal lactobacilli are examined. It was seen that the enzyme provided clear protection against the inhibitory effect of hydrogen peroxide on the growth of Lactobacilli gasseri and L. acidophilus.

Byline: Markus J. Brandt (1), Walter P. Hammes (1), Michael G. Ganzle (2) Keywords: Rye sourdough; Lactobacillus sanfranciscensis; Candida humilis; Modelling Abstract: The effects of temperature, pH, inoculum level, and NaCl on the growth and metabolism of Lactobacillus sanfranciscensis and Candida humilis in rye sourdough were determined. The temperature optima for growth of C. humilis and L. sanfranciscensis were 28 and 32 degC, respectively. Yeast growth was inhibited at 35 degC. The pH did not affect yeast growth in the range 3.5--5.5, whereas growth of L. sanfranciscensis was inhibited at pH 4.0. A NaCl concentration of 4% (flour base) inhibited growth of L. sanfranciscensis but not C. humilis. The effects of the process parameters on the formation of lactate, acetate, ethanol, and CO.sub.2 by the organisms were generally in agreement with their effects on growth. However, decreased formation of acetate by L. sanfranciscensis was observed at 35 degC although lactate and ethanol formation were not affected. In conclusion, the study provides a rationale for the stable persistence of L. sanfranciscensis and C. humilis in traditional sourdoughs and will facilitate the optimisation of sourdough fermentations in traditional and new applications. Author Affiliation: (1) Institut fur Lebensmitteltechnologie, Fachgebiet Allgemeine Lebensmitteltechnologie und -mikrobiologie, Universitat Hohenheim, 70599, Stuttgart, Germany (2) Lehrstuhl fur Technische Mikrobiologie, Technische Universitat Munchen, Weihenstephaner Steig 16, D-85350, Freising, Germany Article History: Registration Date: 15/12/2003 Received Date: 31/10/2003 Online Date: 06/02/2004

A study was conducted to determine whether supplementing milk with whey protein hydrolysates (WPHs) would enhance the growth of several cultures of probiotic lactobacilli and bifidobacteria. The result reveals that WPH as a supplement for milk being cultured for yogurt can increase the growth of probiotic bacteria.

To evaluate the contribution of intracellular peptidases to the growth of the 14-amino-acid (aa) auxotroph Lactobacillus helveticus CNRZ32, single--and multiple-peptidase-deletion mutants were constructed. Two broadspecificity aminopeptidases (PepC and PepN) and X-prolyl dipeptidyl aminopeptidase (PepX) were inactivated through successive cycles of chromosomal gene replacement mutagenesis. The inactivation of all three peptidases in JLS247 ([DELTA]pepC [DELTA]pepN [DELTA]pepX) did not affect the growth rate in amino acid-defined medium. However, the peptidase mutants generally had decreased specific growth rates when acquisition of amino acids required hydrolysis of the proteins in milk, the most significant result being a 73% increase in generation time for JLS247. The growth rate deficiencies in milk were overcome by amino acid supplements with some specificity to each of the peptidase mutants. For example, milk supplementation with Pro resulted in the most significant growth rate increase for [DELTA]pepX strains and a 7-aa supplement (Asn, Cys, Ile, Pro, Set, Thr, and Val) resulted in a JLS247 growth rate indistinguishable from that of the wild type. Our results show that characterization of the activities of the broad-specificity aminopeptidases had little predictive value regarding the amino acid supplements found to enhance the milk growth rates of the peptidase mutant strains. These results represent the first determination of the physiological roles with respect to specific amino acid requirements for peptidase mutants grown in milk.

A model is described that can estimate the growth of Lactobacillus sakei by accounting for growth inhibition caused by nutrient depletion and the accumulation of lactic acid and other toxic end products. The production of bacteriocin is also affected by nutrient availability.

Researchers have developed a medium that can support the growth of Lactobacillus delbrueckii subsp. bulgaricus, other lactobacilli, and Streptococcus thermophilus. The medium is called milieu proche du lait (MPL) and 22 tested strains grew faster on lactose than on glucose, mannose, and fructose.

Results demonstrate that the growth of five Lactobacilli species in a wheat mash is affected by urea hydrogen peroxide (UHP) in the presence of yeast. Data show that Lactobacilli growth can be contained by UHP at 30 millimole per liter of mash to favor vigorous fermentation by the yeast.

MRS broth is used in studying the in vitro cell growth and bacteriocin production of the Lactobacillus sake starter strain CTC 494 during laboratory fermentations. The CTC 494 strain has previously been isolated from dry fermented Spanish sausages. The bacteriocin produced has been named sakacin K and has a bacteriolytic effect on Listeria monocytogenes. The data obtained from the fermentations are used in deriving a predictive model to describe the influence of temperature and pH on microbial behavior.

The growth of Lactococcus lactis and bacteriocin production in salt-enriched defined medium has been studied. The osmoprotective effect of glycine betaine (GB) on bacterial growth and on lacticin 481 production has also been examined. Lacticin 481 is a bacteriocin produced by L. lactis subsp. lactis ADRIA 85LO30 which inhibits food spoilage bacterium Clostridium tyrobutyricum. Results show GB improves the growth of L. lactis under hyperosmotic constraints but contributes to a reduction in bacteriocin production.

An attempt was made to validate a model demonstrating the effects of pH, temperature, sodium chloride concentration as well as accumulated metabolic end product on microbial growth. In particular, a model for the growth of sourdough microflora was developed using the representatives, Candida milleri and two strains of Lactobacillus sanfranciscensis. Predictions made by the model agreed with literature data available on sourdough. This suggests that the most important factors for microbial stability in sourdough fermentation have been accounted for.

The sugar-glycerol cofermentation process in Lactobacillus brevis and L. buchneri was analyzed. The presence of glycerol induced biphasic growth rates, resulting in increased growth yields. This could be attributed to the presence of 1,3-propanediol, an intermediate in glycerol metabolism, which channeled intermediate metabolites towards NADH-producing reactions. This additional NADH allows lactate to be scavenged for ATP generation, producing acetate via pyruvate. The conversion of lactate to pyruvate probably involves NAD-independent lactate dehydrogenases, which are induced under conditions of reduced intracellular NADH levels.

The expression of the genes during the growth in milk and MRS medium is examined by using microarrays. The results have shown that phosphoserine is used as an energy source during the growth of Lactobacillus helveticus CNRZ32.

The optimized electroporation protocol was used to implement the nisin-controlled expression (NICE) system in Lactobacillus rhamnosus GG in order to tag Lactobacillus rhamnosus GG with the green fluorescent protein (GFP). The GFP-tagged strain was used for determining the sensitivity of Lactobacillus rhamnosus GG to human beta-defensins-2 but not to human beta-defensin-1.

Research has been conducted on micro-organisms subjected to abiotic stresses. The authors have investigated the ability of cells to become tolrtant and adapt to the environmental conditions, and they present the results.

Research has been conducted on Streptococcus thermophilus and Lactobacillus bulgaricus. The scientists report that the role of cell-wall proteinases from these bacteria in bacterial growth in milk has been investigated via the study of the negative mutant constructed for the cell-wall proteinase gene.