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Food supplementation with milk fermented by Lactobacillus casei DN-114 001 protects suckling rats from rotavirus-associated diarrhea.

Group A rotavirus is the leading cause of diarrhea among children aged 3-36 mo worldwide. Introducing fermented milk products into the infant diet has been proposed for the prevention or treatment of rotavirus diarrhea. The preventive effect of milk fermented by the Lactobacillus casei strain DN-114 001 was studied in a model of germfree suckling rats supplemented daily from d 2 of life and infected with SA11 rotavirus at d 5 (RF group). One group was supplemented with nonfermented milk (RM) and two uninfected groups (CM and CF) received either nonfermented or fermented milk. Frequency and severity of diarrhea were observed. Rats were killed at various times from 0 to 120 h postinfection (p.i.). Bacteria were measured in the intestine, and rotavirus antigens were detected by ELISA in fecal samples and in different parts of the intestine. Histologic observations were made, including vacuolation, morphology of intestinal villi and number of mucin cells. RM rats had diarrhea for 6 d; compared with the CM group, they had alterations of the intestinal mucosa characterized by cellular vacuolation 48 and 72 h p.i. and a lower number of sulfated mucin cells 72 and 96 h p.i. (P: < 0.05). Early supplementation with fermented milk significantly decreased the clinical signs of diarrhea from 24 to 144 h p.i. (P: < 0.05) and prevented rotavirus infection in all sections of the intestine. Histologic lesions of the small intestine were greatly reduced (P: < 0.05) and the number of mucin cells remained unchanged. The data are discussed with respect to the possibility of reducing rotavirus diarrhea in young children by consumption of fermented milk.

Animals↗

In vitro fermentation properties of selected fructooligosaccharide-containing vegetables and in vivo colonic microbial populations are affected by the diets of healthy human infants.

Salsify (Tragopogon porrifolius) and gobo (Arctium lappa) are root vegetables that contain high concentrations of naturally occurring fructooligosaccharides (FOS), nondigestible oligosaccharides that have prebiotic effects on the intestinal microflora. The objective of this study was to compare colonic microbial populations and the fermentation characteristics of gobo and salsify in breast-fed vs. formula-fed infants. Fecal inoculum from breast-fed and formula-fed infants consuming either no solid foods, cereal, or fruit and vegetable purees were fermented in vitro with sweet potato puree plus gobo, salsify puree or a control vegetable, carrot. Breast-fed and formula-fed infants had similar fecal bacteria concentrations, with the exception of Clostridium perfringens (P < 0.10). Introduction of solid foods into the diet of infants was associated with increased fecal concentrations of bifidobacteria (P < 0.10) and decreased concentrations of total aerobes (P < 0.01), C. perfringens (P < 0.001) and Escherichia coli (P < 0.10). Inoculum from feces of breast-fed infants resulted in greater acetate production, whereas inoculum from feces of formula-fed infants resulted in greater propionate and butyrate production (P < 0.05). Fermentation of FOS-containing vegetables by infant fecal inoculum did not differ significantly from fermentation of carrots as assessed by total short-chain fatty acid (SCFA) production. The addition of solids to the diet of infants was associated with increased production of acetate and total SCFA (P < 0.05). It appears that both the composition and fermentative activity of the colonic microflora of human infants is affected by breast-feeding and solid food consumption, but not by short-term exposure to low concentrations of FOS-containing substrates.

Bacteria, Aerobic↗

Feeding lactose-intolerant children with a powdered fermented milk.

Fresh yogurt has been proposed as a milk substitute for lactase-deficient patients. We investigated the possibility that a dried, low-fat milk fermented by yogurt microorganisms would be effective. Processing of a fermented milk by spray-drying led to a powder with persistence of lactase activity (11.7 units/g) without storage at 4 degrees C and stable at room temperature (20-23 degrees C). The tolerance and absorption of 10.5 g of lactose in a volume of 150 ml was studied in 25 Gabonese lactase-deficient children aged 5-14 years in the form of a standard humanized milk formula followed 1 week later by the powdered fermented milk formula. Results of the breath hydrogen test showed that in 24 of 25 cases, lactose absorption was normal with a maximal rise of hydrogen over baseline of under 13 ppm after ingestion of the powdered fermented milk. The rise was above 20 ppm in the 25 children with the same load of lactose in the form of the standard formula and one-third had symptoms of lactose intolerance. This powdered fermented milk preparation should be considered as a valid approach in programs of nutritional support targeted to countries with a high prevalence of lactase deficiency.

Adolescent↗

Bactericidal activity of a fermented hot-water extract from Stevia rebaudiana Bertoni towards enterohemorrhagic Escherichia coli O157:H7 and other food-borne pathogenic bacteria.

A fermented aqueous extract from Stevia rebaudiana Bertoni showed strong bactericidal activity towards a wide range of food-borne pathogenic bacteria including enterohemorrhagic Escherichia coli O157:H7. The colony-forming ability of the food-borne pathogenic bacteria tested so far was reduced to < 10(-7) when exposed to > or = 40% (v/v) solutions of the fermented extract at 37 C for 2 hr. Secretion of verocytotoxin 1 and 2 by enterohemorrhagic E. coli was also diminished by fermented extract at a concentration of > or = 10% (v/v). In contrast, the fermented extract did not significantly kill Bifidobacteria or Lactobacilli. The active principle(s) of the fermented Stevia extract were bactericidal under acidic conditions.

Bacterial Toxins↗

Formation of vitamins by pure cultures of tempe moulds and bacteria during the tempe solid substrate fermentation.

The formation of water soluble vitamins (vitamin B12, vitamin B6, riboflavin, thiamine, nicotinic acid and nicotinamide) during the tempe solid substrate fermentation was investigated. The role of several strains of Rhizopus oligosporus, R. arrhizus, and R. stolonifer and the role of several bacteria in the vitamin formation process were checked. All fungal and bacterial strains were isolated from Indonesian tempe and soaking water samples. The Rhizopus strains formed riboflavin, nicotinic acid, nicotinamide and vitamin B6. The final concentrations of these substances depended on the different strains involved and on the fermentation time. Isolates of R. oligosporus were generally the best vitamin formers. The moulds did not produce physiologically active vitamin B12. The thiamine content decreased during fermentation. The addition of bacteria, which had been selected in a screening for vitamin B12 production, resulted in an increase of physiologically active vitamin B12. Citrobacter freundii and Klebsiella pneumoniae showed the best formation capabilities. Furthermore, the bacteria produced riboflavin and vitamin B6 in addition to the moulds. The influence of Rhizopus on the vitamin B12 formation of Cit. freundii was also investigated. The vitamin content of tempe that was fermented with the mould and the bacterium was three times as high as a control fermentation with Cit. freundii only.

Bacteria↗

Locating nisin-producing Lactococcus lactis in a fermented meat system.

Antibody-linked probes were used to locate nisin in a fermented meat system. Free nisin or nisin bound to susceptible cells or food components was not detected. Colonies of nisin-producing Lactococcus lactis were stained at all times during growth. The position of nisin-producing L. lactis colonies was noted and compared with the location of spoilage organisms or the distribution of areas with a fermented meat appearance. No relationship between the distribution of starter culture and the location of spoilage organisms or areas of fermentation was observed. In addition to the presence of L. lactis, a rapidly fermentable sugar was also required to obtain a fermented appearance and to reduce the levels of spoilage organisms.

Animals↗

Predominance of Saccharomyces uvarum during spontaneous alcoholic fermentation, for three consecutive years, in an Alsatian winery.

AIMS: The purpose of this study was to determine the origin of the yeasts involved in the spontaneous alcoholic fermentation of an Alsatian wine. METHODS AND RESULTS: During three successive years, must was collected at different stages of the winemaking process and fermented in the laboratory or in the cellar. Saccharomyces yeasts were sampled at the beginning and at the end of the fermentations. Saccharomyces cerevisiae clones were genetically characterized by inter-delta PCR. Non-S. cerevisiae clones were identified as Saccharomyces uvarum by PCR-RFLP on MET2 gene and characterized at the strain level by karyotyping. The composition of the Saccharomyces population in the vineyard, after crushing and in the vat was analyzed. This led to three main results. First, the vineyard Saccharomyces population was rather homogeneous. Second, new non-resident strains had appeared in the must during the winemaking process. Finally, the yeast population in the vat only consisted in S. uvarum strains. CONCLUSION: This 3-year study has enabled us to show the involvement of indigenous S. uvarum in the alcoholic fermentation. SIGNIFICANCE AND IMPACT OF THE STUDY: This study gives a first insight into the polymorphism of S. uvarum strains involved in a spontaneous alcoholic fermentation.

Chromosomes, Fungal↗

Fermentation conditions affecting the bacterial growth and exopolysaccharide production by Streptococcus thermophilus ST 111 in milk-based medium.

AIMS: To study the effect of different fermentation conditions and to model the effect of temperature and pH on different biokinetic parameters of bacterial growth and exopolysaccharides (EPS) production of Streptococcus thermophilus ST 111 in milk-based medium. METHODS AND RESULTS: The influence of temperature and pH was studied through fermentation and modelling. Fermentations under non-pH controlled conditions with S. thermophilus ST 111 indicated that the EPS production was low in milk medium, even if additional nitrogen sources were supplemented. Under pH-controlled conditions, addition of whey protein hydrolysate to the milk medium resulted in a fivefold increase of the EPS production. This medium did not contain polysaccharides interfering with EPS isolation. Primary and secondary modelling of different fermentations revealed an optimum temperature and pH of 40 degrees C and constant pH 6.2, respectively, for growth in milk medium supplemented with whey protein hydrolysate. Maximum EPS production was observed in the range of 32-42 degrees C and constant pH 5.5-6.6. Whereas growth and maximum EPS production were clearly influenced by temperature and pH, the specific EPS production was only affected by stress conditions (T = 49 degrees C). CONCLUSIONS: Addition of whey protein hydrolysate to milk medium resulted in an increased growth and EPS production of S. thermophilus ST 111 under pH-controlled conditions. A modelling approach allowed studying the influence of temperature and pH on the kinetics of both growth and EPS production. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of an appropriate milk-based medium and a combined model of temperature and pH can be of practical importance for the production of yoghurt or other fermented milks as well as for process optimization of the large-scale production of starter strains to be used for their EPS production.

Animals↗

The role of the succinate pathway in sorbitol fermentation by oral Actinomyces viscosus and Actinomyces naeslundii.

The sorbitol fermentation by Actinomyces viscosus and Actinomyces naeslundii was studied with washed sorbitol-grown cells. The fermentation was followed by titration of acids produced at pH 7.0 under anaerobic conditions. Metabolic end-products and intracellular levels of NAD, NADH and glycolytic intermediates during the fermentation were also analyzed. Cell extracts were examined for certain enzyme activities. Bicarbonate was required for acid production from sorbitol and from a mixture of glucose and sorbitol. Malate and fumarate could also support the acid production of A. viscosus. The main end-products were succinate and lactate but not ethanol. Cell extracts showed no activities of alcohol and aldehyde dehydrogenases, but they had activities of malate dehydrogenase and fumarate reductase. In the absence of bicarbonate, malate or fumarate, the intracellular NADH/NAD ratio increased and the levels of 3- and 2-phosphoglycerate and phosphoenolpyruvate decreased. The results indicate that oral sorbitol-fermenting actinomyces lack the ethanol pathway that can contribute to NADH oxidation. To maintain intracellular redox balance during anaerobic sorbitol fermentation, these bacteria can oxidize surplus NADH through a succinate pathway.

Actinomyces↗

Screening of lactic acid bacteria from fermented vegetables by carbohydrate profiling and PCR-ELISA.

AIMS: The aim of this study was to identify potential souring agents, isolated from fermented plant material, by API 50 CHL assay and a molecular method based on polymerase chain reaction and colorimetric hybridization (PCR-ELISA). METHODS AND RESULTS: Forty-two strains of lactic acid bacteria derived from plant material were screened by taking advantage of API 50 CHL and PCR-ELISA. Oligonucleotide probes used for hybridization in PCR-ELISA were specific for lactobacilli, the Leuconostoc family, Lactobacillus pentosus/plantarum and Lactobacillus brevis. The hybrides were detected by a colour-developing reaction. Bacteria isolated from fermented cucumbers were identified as Lact. plantarum-related (Lact. plantarum and Lact. pentosus) and Leuconostoc species. Most of the strains isolated from sauerkraut were identified as Lact. pentosus/plantarum. CONCLUSIONS: Complementary results were obtained in the identification of bacterial strains, isolated from fermented cucumbers and sauerkraut, by API 50 CHL and PCR-ELISA. SIGNIFICANCE AND IMPACT OF THE STUDY: PCR-ELISA proved to be suitable for the screening of large numbers of bacterial isolates from fermented vegetables. This will be useful for the identification of strains suitable for the design of starter cultures for the fermentation of plant material.

Carbohydrates↗

Effect of the addition of inert cellulose substrates to grape must on Saccharomyces cerevisiae diversity and the evolution of alcoholic fermentation.

AIMS: To study the addition of cellulose-based adjuvant as a resource to offset the negative effects produced by grape juice clarification during alcoholic fermentations. METHODS AND RESULTS: The effect of the addition of two kinds of inert cellulose substrates in white wine vinification was investigated in two different musts. In one of these musts, stuck fermentations were detected. One of the types of cellulose examined had a fining effect, which caused a decrease in the number of viable yeasts in the medium and altered the distribution and frequency of the clones, which performed the fermentation. The other cellulose substrate made the medium cloudier but did not alter the distribution of yeasts in comparison with the control. CONCLUSIONS: The behaviour of the inert cellulose substrates on vinification depends on its physical characteristics and its capacity for making the must cloudy. SIGNIFICANCE AND IMPACT OF THE STUDY: The addition of inert cellulose substrates in white wine vinification improves the fermentation process and the quality of wines obtained. This effect is more noticeable in difficult fermentations. One variety of cellulose showed an inhibitory effect on Torulaspora delbrueckii yeasts.

Acids↗

Microbial population dynamics of kimchi, a fermented cabbage product.

Lactic acid bacteria are known to perform significant roles in the fermentation of kimchi, a fermented cabbage product. However, the microbial population dynamics inherent to kimchi fermentation remain to be clearly elucidated. In this study, we have characterized the microbial dynamics via the identification of a total of 970 bacterial isolates, representing 15 species of the genera Lactobacillus, Leuconostoc, and Weissella, all of which were primarily identified by PCR-based restriction enzyme analysis. These population dynamics appear to be influenced markedly by fermentation temperature. Distinct biphasic microbial growth was observed with preliminary 2-day incubation at 15 degrees C, conducted before main fermentation at -1 degrees C. Leuconostoc citreum, as well as Leuconostoc gasicomitatum, predominated during the first growth phase, whereas Weissella koreensis predominated during the second phase. By way of contrast, with preliminary 4-day incubation at 10 degrees C, only W. koreensis grew rapidly from the beginning of the process. Therefore, our findings suggest that a short incubation at 15 degrees C enhances the growth of the less psychrophilic Leuconostoc species, including Lc. citreum, thus delaying the growth of the predominant W. koreensis, which is a more adaptive species at -1 degrees C.

Brassica↗

Effects of yeast proteolytic activity on Oenococcus oeni and malolactic fermentation.

Alcoholic fermentation of synthetic must was performed using either Saccharomyces cerevisiae or a mutant Deltapep4, which is deleted for the proteinase A gene. Fermentation with the mutant Deltapep4 resulted in 61% lower levels of free amino acids, and in 62% lower peptide concentrations at the end of alcoholic fermentation than in the control. Qualitative differences in amino acid composition were observed. Changes observed in amino acids in peptides were mainly quantitative. After alcoholic fermentation, each medium was inoculated with Oenococcus oeni. Malolactic fermentation in the medium with the Deltapep4 strain took 10 days longer than the control. This difference may have been due to a difference in the nitrogen composition of the two media. Free amino acids and amino acids in peptides were poorly consumed by O. oeni. Thus, the qualitative aspects of nitrogen composition, which depend in part on yeast metabolism, may be a determinant for the optimal growth of O. oeni in wine.

Amino Acids↗

The effect of a hindgut fermentation on urea metabolism in sheep nourished by intragastric infusion.

Four female sheep nourished wholly by infusions of volatile fatty acids, buffer and minerals into the rumen and casein into the abomasum were given, in addition, infusions of fermentable carbohydrates into the terminal ileum. The ileal infusions consisted of (1) water alone, (2) 25 g starch + 50 g cellulose, and (3) 50 g starch + 50 g cellulose. Measurement of nutrient digestibility and nitrogen retention were made over 5 days and the kinetics of urea metabolism were measured over 24 h by means of a single injection of [14C]urea. Endogenous urinary nitrogen excretion was measured over a subsequent 5 days when casein was omitted from the infusion mixtures. Increases in hindgut fermentation resulted in a significant increase in faecal nitrogen excretion (P less than 0.01) and a corresponding reduction in urinary urea nitrogen excretion (P less than 0.05). The ileal infusions did not significantly affect urea irreversible loss rate or urea pool size and were also without effect on plasma urea or rumen ammonia concentrations. Urea degradation in the gastrointestinal tract increased by about 2 g/day in progressing from lowest to highest level of hindgut infusion but differences between treatments were not statistically significant. Endogenous urinary nitrogen excretion was not affected by hindgut fermentation and averaged 206 mg N/(kg0.75 day) over the three treatment groups. Faecal nitrogen excretion however increased progressively with increase in ileal infusions (P less than 0.05) and was similar to that seen when nitrogen input was adequate. It is concluded that changes in hindgut fermentation can alter the partition of nitrogen excretion between faeces and urine but the quantities involved are small relative to the total exchange of urea across the digestive tract. The rumen appears to be an important site of urea degradation even when microbial fermentation is absent.

Animal Nutritional Physiological Phenomena↗

Fermentation of high concentrations of maltose by Saccharomyces cerevisiae is limited by the COMPASS methylation complex.

In Saccharomyces cerevisiae, genes encoding maltose permeases and maltases are located in the telomeric regions of different chromosomes. The COMPASS methylation complex, which methylates lysine 4 on histone H3, controls the silencing of telomeric regions. Yeast strains deleted for SWD1, SWD3, SDC1, SET1, BRE2, or SPP1, encoding components of the COMPASS complex, fermented a medium containing 22% maltose with noticeably higher attenuation than did the wild type, resulting in production of up to 29% more ethanol. The least effective strain was spp1. Absence of COMPASS components had no effect on the fermentation of media with 20% glucose, 20% sucrose, or 16% maltose. Deletion of SWD3 resulted in larger amounts of MAL12 transcript, encoding maltase, at the late stages of fermentation of 22% maltose. A similar effect on maltase activity and maltose uptake capability was seen. The lysine 4 residue of histone H3 was trimethylated in wild-type cells at the late stages, while only small amounts of the dimethylated form were detected. Trimethylation and dimethylation of this residue were not detected in strains deleted for SWD1, SWD3, SET1, BRE2, or SDC1. Trimethylated lysine 4 was apparent only at the early stages (48 and 96 h) of fermentation in an spp1 strain. This work indicates that the COMPASS complex represses the expression of maltose utilization genes during the late stages of fermentation of a high concentration of maltose.

Culture Media↗

Fermentation of L-aspartate by a saccharolytic strain of Bacteroides melaninogenicus.

Resting cells of Bacteroides melaninogenicus fermented L-[14C]aspartate as a single substrate. The 14C-labeled products included succinate, acetate, CO2, oxaloacetate, formate, malate, glycine, alanine, and fumarate in the relative percentages 68, 15, 9.9, 2.7, 1.8, 1.0, 0.7, 0.5, and 0.06, respectively, based on the total counts per minute of the L-[14C]aspartate fermented. Ammonia was produced in high amounts, indicating that 96% of the L-aspartate fermented was deaminated. These data suggest that L-aspartate is mainly being reduced through a number of intermediate reactions involving enzymes of the tricarboxylic acid cycle to succinate. L-[14C]asparagine was also fermented by resting cells of B. melaninogenicus to form L-aspartate, which was subsequently, but less actively, fermented.

Amino Acids↗

Fermentation of mucin and plant polysaccharides by strains of Bacteroides from the human colon.

Ten Bacteroides species found in the human colon were surveyed for their ability to ferment mucins and plant polysaccharides ("dietary fiber"). A number of strains fermented mucopolysaccharides (heparin, hyaluronate, and chondroitin sulfate) and ovomucoid. Only 3 of the 188 strains tested fermented beef submaxillary mucin, and none fermented porcine gastric mucin. Many of the Bacteroides strains tested were also able to ferment a variety of plant polysaccharides, including amylose, dextran, pectin, gum tragacanth, gum guar, larch arabinogalactan, alginate, and laminarin. Some plant polysaccharides such as gum arabic, gum karaya, gum ghatti and fucoidan, were not utilized by any of the strains tested. The ability to utilize mucins and plant polysaccharides varied considerably among the Bacteroides species tested.

Bacteroides↗

In vitro ruminal fermentation of organic acids common in forage.

Mixed rumen bacteria from cows fed either timothy hay or a 60% concentrate were incubated with 7.5 mM citrate, trans-aconitate, malate, malonate, quinate, and shikimate. Citrate, trans-aconitate, and malate were fermented at faster rates than malonate, quinate, and shikimate. Acetate was the primary fermentation product for all six acids. Quinate and shikimate fermentations gave rist to butyrate, whereas malate and malonate produced significant amounts of propionic acid. High-pressure liquid chromatography of fermentation products from trans-aconitate incubations revealed a compound that was subsequently identified as tricarballylate. As much as 40% of the trans-aconitate acid was converted to tricarballylate, and tricarballylate was fermented slowly. The slow rate of tricarballylate metabolism by mixed rumen bacteria and its potential as a magnesium chelator suggest that tricarballylate formation could be an important factor in the hypomagnesemia that leads to grass tetany.

Animals↗