Utilisation of maltose and glucose by lactobacilli isolated from sourdough

Abstract The utilisation of glucose and maltose was investigated with Lactobacillus strains isolated from sourdough starters. These preparations have been in continuous use for a long period to produce sourdough from rye, wheat and sorghum. The major metabolic products formed by resting cells from glucose or maltose were lactate, ethanol and acetate. Upon fermentation of maltose, resting cells of Lactobacillus sanfrancisco, L. reuteri, L. fermentum and Lactobacillus ep. released up to 13.8 mM glucose after 8 h. The ratio of released glucose per mol of utilised maltose was up to 1:1. Glucose formation was high when starved cells of L. sanfrancisco and Lactobacillus sp. were used. This is consistent with maltose utilisation via maltose phosphorylase which phosphorylates maltose without the expenditure of ATP and thus allows the cell to waste glucose in the presence of abundant maltose. The glucose formed may be utilised by the lactobacilli or other microorganisms, e.g. yeasts. However, the release of glucose into the medium by sourdough lactobacilli prevents competitors from utilising the abundant maltose by glucose repression. In strains of L. sanfrancisco , maltose utilisation was very effective and not subject to glucose repression. Therefore, they overgrow other microorganisms sharing this habitat. Wild isolates of L. sanfrancisco were initially unable to grow on glucose. Upon growth on maltose such strains required adaptation times of up to 150 h to grow on glucose. After subsequent transfer of glucose-grown cells to fresh medium the strains resumed growth both on glucose or maltose. They readily lost their ability to grow on glucose upon exposure to maltose. L. sanfrancisco exhibited biphasic growth characteristics on media containing glucose, maltose or both carbon sources. Evidence is provided that biphasic growth and metabolite formation are dependent on the redox potential.     

Characterization of lactobacilli isolated from caper berry fermentations

AIMS: To determine the metabolic and functional properties of lactobacilli isolated from caper fermentation. METHODS AND RESULTS: A collection of 58 lactobacilli from fermentation of caper berries (including species of Lactobacillus plantarum, Lactobacillus paraplantarum, Lactobacillus pentosus, Lactobacillus brevis and Lactobacillus fermentum) was studied. Strains were classified in different clusters according to sugar fermentation patterns. Most strains of L. plantarum (the predominant species in the fermentation) clustered in a single group. Analysis of enzymatic activities revealed a high incidence of leucine aminopeptidase, acid phosphatase, beta-galactosidase and beta-glucosidase among the different strains of lactobacilli. A high number of strains were able to degrade raffinose and stachyose. Phytase activity and bile salt hydrolase activity were only detected in certain strains of L. plantarum. CONCLUSIONS: Lactobacilli from caper fermentation are metabolically diverse, and some strains display functional properties of interest. SIGNIFICANCE AND IMPACT OF THE STUDY: Strains of lactobacilli with selected functional properties could be good candidates for future development of commercial starters for industrial caper fermentation.     

An in vitro study of the probiotic potential of a bile-salt-hydrolyzing Lactobacillus fermentum strain,and determination of its cholesterol-lowering properties

This study evaluated the use of a bile-salt-hydrolyzing Lactobacillus fermentum strain as a probiotic with potential hypocholesterolemic properties. The effect of L. fermentum on representative microbial populations and overall metabolic activity of the human intestinal microbiota was investigated using a three-stage continuous culture system. Also, the use of galactooligosaccharides as a prebiotic to enhance growth and/or activity of the Lactobacillus strain was evaluated. Administration of L. fermentum resulted in a decrease in the overall bifidobacterial population (ca. 1 log unit). In the in vitro system, no significant changes were observed in the total bacterial, Lactobacillus, Bacteroides, and clostridial populations through L. fermentum supplementation. Acetate production decreased by 9 to 27%, while the propionate and butyrate concentrations increased considerably (50 to 90% and 52 to 157%, respectively). A general, although lesser, increase in the production of lactate was observed with the administration of the L. fermentum strain. Supplementation of the prebiotic to the culture medium did not cause statistically significant changes in either the numbers or the activity of the microbiota, although an increase in the butyrate production was seen (29 to 39%). Results from this in vitro study suggest that L. fermentum KC5b is a candidate probiotic which may affect cholesterol metabolism. The short-chain fatty acid concentrations, specifically the molar proportion of propionate and/or bile salt deconjugation, are probably the major mechanism involved in the purported cholesterol-lowering properties of this strain.     

Reduction of alpha-galactooligosaccharides in soyamilk by Lactobacillus fermentum CRL 722: in vitro and in vivo evaluation of fermented soyamilk

AIMS: Consumption of soya-derived products has been hampered by the presence of alpha-galactooligosaccharides (alpha-GOS) because mammals lack pancreatic alpha-galactosidase (alpha-Gal) which is necessary for their hydrolysis. These sugars thus reach the large intestine causing gastrointestinal disorders in sensitive individuals. The use of lactic acid bacteria (LAB) expressing alpha-Gal is a promising solution for the degradation of alpha-GOS in soyamilk. METHODS AND RESULTS: The capacity of the LAB Lactobacillus fermentum CRL 722 to properly degrade alpha-GOS was studied in vitro using controlled fermentation conditions and in vivo using a rat model. Lactobacillus fermentum CRL 722 was able to grow on commercial soyamilk and completely eliminated stachyose and raffinose during fermentation because of its high alpha-Gal activity. Rats fed soyamilk fermented by this LAB had smaller caecums compared with rats fed unfermented soyamilk. CONCLUSIONS: Soyamilk fermentation by Lact. fermentum CRL 722 results in the reduction of alpha-GOS concentrations in soyamilk, thus eliminating possible undesirable physiological effects normally associated with its consumption. SIGNIFICANCE AND IMPACT OF THE STUDY: Fermentation with Lact. fermentum CRL 722 could prevent gastrointestinal disorders in sensitive individuals normally associated with the consumption of soya-based products. This LAB could thus be used in the elaboration of novel fermented vegetable products which better suit the digestive capacities of consumers.     

Arabinose fermentation by Lactobacillus plantarum in sourdough with added pentosans and alphaalpha-L-arabinofuranosidase: a tool to increase the production of acetic acid

Sixty-five strains of obligately and facultatively heterofermentative sourdough lactic acid bacteria were screened for their capacity to grow optimally in the presence of arabinose, ribose and xylose as carbon sources. Lactobacillus alimentarius 15F, Lact. brevis 10A, Lact. fermentum 1F and Lact. plantarum 20B showed higher growth rate, cell yield, acidification rate and production of acetic acid when some pentoses instead of maltose were added to the SDB medium. Lactobacillus plantarum 20B used arabinose also in a synthetic medium where complex growth factors such as yeast extract were omitted. Other Lact. plantarum strains did not show the same property. Pentosan extract was treated with alpha-L-arabinofuranosidase from Aspergillus niger or endo-xylanase from Bacillus subtilis to produce hydrolysates containing mainly arabinose and xylose, respectively. In particular, the hydrolysate containing arabinose substantiated the growth and the production of lactic acid and, especially, of acetic acid by Lact. plantarum 20B. Sourdough fermentation by Lact. plantarum 20B with addition of pentosan extract and alpha-L-arabinofuranosidase increased the acidification rate, titratable acidity and acetic acid content compared with traditional sourdough. A facultatively heterofermentative strain, Lact. plantarum 20B, also produced a sourdough with an optimal fermentation quotient.     

Optimization of nutritional requirements for gentamicin production by Micromonospora echinospora

Effect of various fermentation media, carbon sources, nitrogen sources, phosphate concentration and culture requirements includes inoculum levels and age were determined on gentamicin production and biomass dry weight production for Micromonospora echinospora, a gentamicin producing strain. Of the substrates tested, starch as a sole carbon source promoted maximal gentamicin production, while maltose promoted maximal growth. Yeast extract as a sole nitrogen source promoted maximal growth, while soyabean meal for gentamicin production. Increasing phosphate concentration enhanced gentamicin production and observed optimum production at 1.2 g/1 (6% v/v) of phosphate having 72 h old inoculum in the medium. Highest gentamicin production was obtained after cultivation with shaking for 120 h in a medium containing starch 0.75% (w/v), soyabean meal 0.5%, K2HPO4 0.12%, CaCO3 0.4%, FeSO4 0.003% and CoCl2 0.0001%. The gentamicin production was 1.2-fold in this medium as compared to basal medium.     

Influence of Carbohydrates on Growth and Sporulation of Clostridium perfringens Type A

Growth and sporulation of Clostridium perfringens type A in Duncan and Strong (DS) sporulation medium was investigated. A biphasic growth response was found to be dependent on starch concentration. Maximal levels of heat-resistant spores were formed at a starch concentration of 0.40%. Addition of glucose, maltose, or maltotriose to a sporulating culture resulted in an immediate turbidity increase, indicating that biphasic growth in DS medium may be due to such starch degradation products. Amylose and, to a lesser extent, amylopectin resulted in biphasic growth when each replaced starch in the sporulation medium. A levels of heat-resistant spores approximately equal to the control was produced with amylopectin but not amylose as the added carbohydrate. Addition of glucose or maltose to a DS medium without starch at stage II or III of sporulation did not alter the level of heat-resistant spores as compared with the level obtained in DS medium with starch. Omission of starch or glucose or maltose resulted in an approximately 100-fold decrease in the number of heat-resistant spores, although the percentage of sporulation (90%) was unaffected. The role of starch and amylopectin in the formation of heat-resistant spores probably involves the amyloytic production of utilizable short-chain glucose polymers that provide an energy source for the completion of sporulation.     

Growth studies of potentially probiotic lactic acid bacteria in cereal-based substrates

AIMS: The overall growth kinetics of four potentially probiotic strains (Lactobacillus fermentum, Lact. reuteri, Lact. acidophilus and Lact. plantarum) cultured in malt, barley and wheat media were investigated. The objectives were to identify the main factors influencing the growth and metabolic activity of each strain in association with the cereal substrate. METHODS AND RESULTS: All fermentations were performed without pH control. A logistic-type equation, which included a growth inhibition term, was used to describe the experimental data. In the malt medium, all strains attained high maximum cell populations (8.10-10.11 log10 cfu ml(-1), depending on the strain), probably due to the availability of maltose, sucrose, glucose, fructose (approx. 15 g l(-1) total fermentable sugars) and free amino nitrogen (approx. 80 mg l(-1)). The consumption of sugars during the exponential phase (10-12 h) resulted in the accumulation of lactic acid (1.06-1.99 g l(-1)) and acetic acid (0.29-0.59 g l(-1)), which progressively decreased the pH of the medium. Each strain demonstrated a specific preference for one or more sugars. Since small amounts of sugars were consumed by the end of the exponential phase (17-43%), the decisive growth-limiting factor was probably the pH, which at that time ranged between 3.40 and 3.77 for all of the strains. Analysis of the metabolic products confirmed the heterofermentative or homofermentative nature of the strains used, except in the case of Lact. acidophilus which demonstrated a shift towards the heterofermentative pathway. All strains produced acetic acid during the exponential phase, which could be attributed to the presence of oxygen. Lactobacillus plantarum, Lact. reuteri and Lact. fermentum continued to consume the remaining sugars and accumulate metabolic products in the medium, probably due to energy requirements for cell viability, while Lact. acidophilus entered directly into the decline phase. In the barley and wheat media all strains, especially Lact. acidophilus and Lact. reuteri, attained lower maximum cell populations (7.20-9.43 log10 cfu ml(-1)) than in the malt medium. This could be attributed to the low sugar content (3-4 g l(-1) total fermentable sugar for each medium) and the low free amino nitrogen concentration (15.3-26.6 mg l(-1)). In all fermentations, the microbial growth ceased at pH values (3.73-4.88, depending on the strain) lower than those observed for malt fermentations, which suggests that substrate deficiency in sugars and free amino nitrogen contributed to growth limitation. CONCLUSIONS: The malt medium supported the growth of all strains more than barley and wheat media due to its chemical composition, while Lact. plantarum and Lact. fermentum appeared to be less fastidious and more resistant to acidic conditions than Lact. acidophilus and Lact. reuteri. SIGNIFICANCE AND IMPACT OF THE STUDY: Cereals are suitable substrates for the growth of potentially probiotic lactic acid bacteria.     

A strategy to prevent the occurrence of Lactobacillus strains using lactate-tolerant yeast Candida glabrata in bioethanol production

Contamination of Lactobacillus sp. in the fermentation broth of bioethanol production decreases ethanol production efficiency. Although the addition of lactate to the broth can effectively inhibit the growth of Lactobacillus sp., it also greatly reduces the fermentation ability of Saccharomyces cerevisiae. To overcome this conflict, lactate-tolerant yeast strains were screened. Candida glabrata strain NFRI 3164 was found to exhibit both higher levels of lactate tolerance and fermentation ability. Co-cultivation of C. glabrata was performed with Lactobacillus brevis and Lb. fermentum, which were reported as major contaminating bacteria during bioethanol production, in culture medium containing 2% lactate. Under these culture conditions, the growth of Lactobacillus strains was greatly inhibited, but the ethanol production of C. glabrata was not significantly affected. Our data show the possibility of designing an effective fuel ethanol production process that eliminates contamination by Lactobacillus strains through the combined use of lactate addition and C. glabrata.     

Evaluation of nitric oxide production by lactobacilli

Six strains of Lactobacillus fermentum and Lactobacillus plantarum were investigated for nitric oxide (NO) production. First, the potential presence of NO synthase was examined. None of the strains of L. fermentum and L. plantarum examined produced NO from L-arginine under aerobic conditions. Interestingly, all L. fermentum strains expressed strong L-arginine deiminase activity. All L. fermentum strains produced NO in MRS broth, but the NO was found to be chemically derived from nitrite, which was produced by L. fermentum from nitrate present in the medium. Indeed all L. fermentum strains express nitrate reductase under anaerobic conditions. Moreover, one strain, L. fermentum LF1, had nitrate reductase activity under aerobic conditions. It was also found that L. fermentum strains JCM1173 and LF1 possessed ammonifying nitrite reductase. The latter strain also had denitrifying nitrite reductase activity at neutral pH under both anaerobic and aerobic conditions. The LF1 strain is thus capable of biochemically converting nitrate to NO. NO and nitrite produced from nitrate by lactobacilli may constitute a potential antimicrobial mechanism. studied in a rat acute liver injury model (Adawi et al. 1997). The results indicate that Lactobacillus plantarum DSM 9842 may possess NOS (Adawi et al. 1997). However, NO production from L-arginine has not been investigated in pure cultures of L. plantarum. According to the results of a 15N enrichment experiment, traces of (NO2-+NO3-)-N (total oxidised nitrogen: TON), which seemed to be formed by the resting cells of Lactobacillus fermentum IFO3956, appeared to be derived from L-arginine (Morita et al. 1997). Therefore, it was suggested that L. fermentum may possess a NOS. However, NO produced from L-arginine was not directly measured and a NOS inhibitor test was not performed by Morita et al. (1997). It is known that L-arginine deiminase (ADI) in bacteria may convert L-arginine to NH4+ (Cunin et al. 1986), which may be further oxidised to TON via nitrification by bacteria. Therefore, 15N enrichment experiments could not definitely conclude that L. fermentum possess NOS to convert L-arginine directly to NO. In this study, six Lactobacillus strains belonging to L. plantarum and L. fermentum were measured for NO production in MRS broth. The metabolism of nitrate and L-arginine by the Lactobacillus cell suspensions was also studied. The possibility that NO and nitrite production by lactobacilli may be a potential probiotic trait is also discussed.     

Optimization of submerged culture requirements for the production of mycelial growth and exopolysaccharide by Cordyceps jiangxiensis JXPJ 0109

AIMS: The objective of the present study was to investigate the optimal culture requirements for mycelial growth and exopolysaccharide production by Cordyceps jiangxiensis JXPJ 0109 in submerged culture. METHODS AND RESULTS: The effects of medium ingredients (i.e. carbon and nitrogen sources, and growth factor) and other culture requirements (i.e. initial pH, temperature, etc.) on the production of mycelia and exopolysaccharide were observed using a one-factor-at-a-time method. More suitable culture requirements for mycelial growth and exopolysaccharide production were proved to be maltose, glycerol, tryptone, soya bean steep powder, yeast extract, medium capacity 200 ml in a 500-ml flask, agitation rate 180 rev min(-1), seed age 4-8 days, inoculum size 2.5-7.5% (v/v), etc. The optimal temperatures and initial pHs for mycelial growth and exopolysaccharide production were at 26 degrees C and pH 5 and at 28 degrees C and pH 7, respectively, and corresponding optimal culture age were observed to be 8 and 10 days respectively. According to the primary results of the one-factor-at-a-time experiments, the optimal medium for the mycelial growth and exopolysaccharide production were obtained using an orthogonal layout method to optimize further. Herein the effects of medium ingredients on the mycelial growth of C. jiangxiensis JXPJ 0109 were in the order of yeast extract > tryptone > maltose > CaCl2 > glycerol > MgSO4 > KH2PO4 and the optimal concentration of each composition was 15 g maltose (food-grade), 10 g glycerol, 10 g tryptone, 10 g yeast extract, 1 g KH2PO4, 0.2 g MgSO4, and 0.5 g CaCl2 in 1 l of distilled water, while the order of effects of those components on exopolysaccharide production was yeast extract > maltose > tryptone > glycerol > KH2PO4 > CaCl2 > MgSO4, corresponding to the optimal concentration of medium was as follows: 20 g maltose (food-grade), 8 g glycerol, 5 g tryptone, 10 g yeast extract, 1 g KH2PO4, and 0.5 g CaCl2 in 1 l of distilled water. CONCLUSIONS: Under the optimal culture requirements, the maximum exopolysaccharide production reached 3.5 g l(-1) after 10 days of fermentation, while the maximum production of mycelial growth achieved 14.5 g l(-1) after 8 days of fermentation. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on the submerged culture requirements for mycelial growth and exopolysaccharide in C. jiangxiensis, and this two-step optimization strategy in this study can be widely applied to other microbial fermentation processes.     

Improvement of maltotriose fermentation by Saccharomyces cerevisiae

AIMS: To enhance the fermentation of maltotriose by industrial Saccharomyces cerevisiae strains. METHODS AND RESULTS: The capability to ferment maltotriose by an industrial yeast strain that uses this sugar aerobically was tested in shake flasks containing rich medium. While the presence of maltose in the medium did not improve maltotriose fermentation, enhanced and constitutive expression of the AGT1 permease not only increased the uptake of maltotriose, but allowed efficient maltotriose fermentation by this strain. Supplementation of the growth medium with 20 mmol magnesium l(-1) also increased maltotriose fermentation. CONCLUSIONS: Over expression of the AGT1 permease and magnesium supplementation improved maltotriose fermentation by an industrial yeast strain that respired but did not ferment this sugar. SIGNIFICANCE AND IMPACT OF THE STUDY: This work contributes to the elucidation of the roles of the AGT1 permease and nutrients in the fermentation of all sugars present in starch hydrolysates, a highly desirable trait for several industrial yeasts.     

Influence of temperature and backslopping time on the microbiota of a type I propagated laboratory wheat sourdough fermentation

Sourdough fermentation is a cereal fermentation that is characterized by the formation of stable yeast/lactic acid bacteria (LAB) associations. It is a unique process among food fermentations in that the LAB that mostly dominate these fermentations are heterofermentative. In the present study, four wheat sourdough fermentations were carried out under different conditions of temperature and backslopping time to determine their effect on the composition of the microbiota of the final sourdoughs. A substantial effect of temperature was observed. A fermentation with 10 backsloppings (once every 24 h) at 23°C resulted in a microbiota composed of Leuconostoc citreum as the dominant species, whereas fermentations at 30 and 37°C with backslopping every 24 h resulted in ecosystems dominated by Lactobacillus fermentum. Longer backslopping times (every 48 h at 30°C) resulted in a combination of Lactobacillus fermentum and Lactobacillus plantarum. Residual maltose remained present in all fermentations, except those with longer backslopping times, and ornithine was found in almost all fermentations, indicating enhanced sourdough-typical LAB activity. The sourdough-typical species Lactobacillus sanfranciscensis was not found. Finally, a nonflour origin for this species was hypothesized.     

Diversity and technological properties of predominant lactic acid bacteria from fermented cassava used for the preparation of Gari, a traditional African food

Traditional fermentation of cassava is dominated by a lactic acid bacteria (LAB) population. Fermentation is important for improving product flavour and aroma as well as safety, especially by reduction of its toxic cyanogenic glucosides. The production of Gari from cassava in Benin typically occurs on a household or small industrial scale, and consequently suffers from inconsistent product quality and may not always be safe for consumption. Therefore, the diversity of LAB from a typical cassava fermentation for the preparation of Gari, and their technologically relevant characteristics were investigated with a view towards selection of appropriate starter cultures. A total of 139 predominant strains isolated from fermenting cassava were identified using phenotypic tests and genotypic methods such as rep-PCR and RAPD-PCR. DNA-DNA hybridisation and sequencing of the 16S rRNA genes were done for selected strains. Lactobacillus plantarum was the most abundantly isolated species (54.6% of isolates), followed by Leuconostoc fallax (22.3%) and Lactobacillus fermentum (18.0%). Lactobacillus brevis, Leuconostoc pseudomesenteroides and Weissella paramesenteroides were sporadically isolated. The L. plantarum strains were shown to be better acid producers and capable of faster acid production than the L. fallax or L. fermentum strains. The incidence of beta-glucosidase (linamarase) activity was also highest among strains of this species. Production of antagonistic substances such as H2O2 and bacteriocins, however, was more common among L. fallax and L. fermentum strains. Strains of all three species were capable of utilising the indigestible sugars raffinose and stachyose. Therefore, a starter culture containing a mixture of strains from all three species was recommended.     

Microbiological characterization and probiotic potential of koko and koko sour water, African spontaneously fermented millet porridge and drink

AIMS: To identify and examine the diversity of predominant lactic acid bacteria (LAB) in koko and koko sour water (KSW) from different Ghanaian production sites with regard to pattern of fermentation (API 50 CHL), genotype, antimicrobial activity, and resistance to low pH and bile salts. METHODS AND RESULTS: In total 215 LAB were isolated from koko and KSW. The isolates were identified using intergenic transcribed spacers (ITS)-PCR restriction fragment length polymorphism (RFLP), API 50 CHL, restriction enzyme analysis with pulsed-field gel electrophoresis (REA-PFGE) and sequencing of the 16S rRNA gene. The dominating micro-organisms in koko was found to be Weisella confusa and Lactobacillus fermentum, followed by Lact. salivarius and Pediococcus spp. Chemometric data analysis were used to link the LAB species to the different production stages and production sites. At intra-species level the isolates were found to have a great diversity. The isolates were investigated for antimicrobial activity using agar diffusion assays, and acid and bile tolerance. Most isolates showed low levels of antimicrobial activity towards the indicator strain Listeria innocua, but not towards the bacteriocin-sensitive Lact. sakei. Growth of all LAB isolates was unaffected by the presence of 0.3% (v/v) oxgall bile. The isolates were able to survive, but were not able to grow in growth medium adjusted to pH 2.5. CONCLUSIONS: The dominating LAB of koko and KSW were W. confusa and Lact. fermentum showing a pronounced taxonomic biodiversity at sub-species level between stages within the production as well as between production sites. Other species observed in KSW were Lact. salivarius, Ped. pentosaceus, Ped. acidilactici and Lact. paraplantarum. They occurred in levels of 108 CFU ml-1 in fresh KSW and showed uniform antimicrobial activity, and acid and bile tolerance. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study gives a detailed picture of the taxonomy and diversity of LAB in an African-fermented millet product that may have potential as a probiotic product for the local population. The chemometric tools Principal Component Analysis and anova Partial Least Squares Regression were proven to be useful in the analysis of microbial groupings and associations with specific sites and stages in the production of koko and KSW.     

The biodiversity of predominant lactic acid bacteria in dolo and pito wort for the production of sorghum beer

AIM: To quantify and identify the predominant lactic acid bacteria (LAB) in dolo and pito wort processing, and to examine their biodiversity at strain level. MATERIALS AND RESULTS: The processing of dolo and pito wort was studied at four production sites in Burkina Faso and Ghana. The succession of dominant micro-organisms, pH and titratable acidity were determined from sorghum malt through mashing and acidification to final wort. In the sorghum malt and during mashing, the LAB counts were 5.7-7.5 log CFU g(-1). Similar levels of yeasts and gram-negative, catalase-positive bacteria were observed. These levels decreased to 3.7-4.5 log CFU g(-1) and相似文献   

Antagonistic activity of probiotic lactobacilli and bifidobacteria against entero- and uropathogens

AIM: To develop in vitro assays for comparing the antagonistic properties and anti-oxidative activity of probiotic Lactobacillus and Bifidobacterium strains against various entero- and urinary pathogens. METHODS AND RESULTS: The antagonistic activity of five probiotic lactobacilli (Lactobacillus rhamnosus GG, Lactobacillus fermentum ME-3, Lactobacillus acidophilus La5, Lactobacillus plantarum 299v and Lactobacillus paracasei 8700:2) and two bifidobacteria (Bifidobacterium lactis Bb12, Bifidobacterium longum 46) against six target pathogens was estimated using different assays (solid and liquid media, anaerobic and microaerobic cultivation) and ranked (low, intermediate and high). Bacterial fermentation products were determined by gas chromatography, and the total anti-oxidative activity of probiotics was measured using linolenic acid test. Pyelonephritic Escherichia coli was highly suppressed by GG and both bifidobacteria strains. Lactobacilli strains 8700:2, 299v and ME-3 were the most effective against Salmonella enterica ssp. enterica in microaerobic while ME-3 and both bifidobacteria expressed high activity against Shigella sonnei in anaerobic milieu. Lact. paracasei, Lact. rhamnosus and Lact. plantarum strains showed intermediate antagonistic activity against Helicobacter pylori under microaerobic conditions on solid media. The highest anti-oxidative activity was characteristic for Lact. fermentum ME-3 (P < 0.05). No efficient antagonist against Clostridium difficile was found. The positive correlations between the pH, lactic acid production and anti-microbial activity for all tested probiotics were assessed. CONCLUSIONS: Developed experimental assays enable to compare the anti-microbial and -oxidative activity of Lactobacillus and/or Bifidobacterium probiotics, which have been claimed to possess the ability of suppressing the growth of various enteric and urinary pathogens. SIGNIFICANCE AND IMPACT OF THE STUDY: Screening Lactobacillus and Bifidobacterium sp. strains according to their activity in various environmental conditions could precede the clinical efficacy studies for adjunct treatment with probiotics in cure of different gastrointestinal and urinary tract infections.     

Statistical optimization of a high maltose-forming,hyperthermostable and Ca2+-independent alpha-amylase production by an extreme thermophile Geobacillus thermoleovorans using response surface methodology

AIM: Statistical optimization for maximum production of a hyperthermostable, Ca2+-independent and high maltose-forming alpha-amylase by Geobacillus thermoleovorans. METHODS AND RESULTS: G. thermoleovorans was cultivated in 250 ml flasks containing 50 ml of chemically defined glucose-arginine medium (g l(-1): glucose 20; arginine 1.2; riboflavin 150 microg ml(-1); MgSO4. 7H2O 0.2; NaCl 1.0; pH 7.0). The medium was inoculated with 5 h-old bacterial inoculum (1.8x10(8) CFU ml(-1)), and incubated in an incubator shaker at 70 degrees C for 12 h at 200 rev min(-1). The fermentation variables optimized by 'one variable at a time' approach were further optimized by response surface methodology (RSM). The statistical model was obtained using central composite design (CCD) with three variables: glucose, riboflavin and inoculum density. An over all 24 and 70% increase in enzyme production was attained in shake flasks and fermenter because of optimization by RSM, respectively. A good coverage of interactions could also be explained by RSM. The end products of the action of alpha-amylase on starch were maltose (62%), maltotriose (31%) and malto-oligosaccharides (7%). CONCLUSIONS: RSM allowed optimization of medium components and cultural parameters for attaining high yields of alpha-amylase, and further, a good coverage of interactions could be explained. The yield of maltose was higher than maltotriose and malto-oligosaccharides in the starch hydrolysate. SIGNIFICANCE AND IMPACT OF THE STUDY: By applying RSM, critical fermentation variables were optimized rapidly. The starch hydrolysate contained a high proportion of maltose, and therefore, the enzyme can find application in starch saccharification process for the manufacture of high maltose syrups. The use of this enzyme in starch saccharification eliminates the addition of Ca2+.     

Evaluation of numerical analyses of RAPD and API 50 CH patterns to differentiate Lactobacillus plantarum, Lact. fermentum, Lact. rhamnosus, Lact. sake, Lact. parabuchneri, Lact. gallinarum, Lact. casei, Weissella minor and related taxa isolated from kocho and tef

AIM: The study was carried out to assess the agreement of API 50 CH fermentation data of food lactobacilli with their RAPD profiles to determine whether the system could be used alone as a reliable taxonomic tool for this genus. METHODS AND RESULTS: API 50 CH, RAPD and DNA:DNA reassociation data for 42 lactobacilli from tef and kocho were compared with 30 type strains. Discrepancies were observed between the three methods in assigning strains of Lactobacillus plantarum, Lact. fermentum, Weissella minor and Lact. gallinarum, and Lact. fermentum, Lact. amylophilus, Lact. casei subsp. pseudoplantarum and Lact. rhamnosus. DNA reassociation data agreed well with RAPD results. CONCLUSIONS: API 50 CH profiles should be complemented with molecular genetic results for effective identification in Lactobacillus. SIGNIFICANCE AND IMPACT OF THE STUDY: The study suggested less dependability of metabolic data alone as an identification tool.     

Two Lactobacillus strains, isolated from breast milk, differently modulate the immune response

AIMS: The ability of two different Lactobacillus strains (Lactobacillus salivarius CECT5713 and Lactobacillus fermentum CECT5716), isolated from human breast milk, to modulate the immune response was examined. METHODS AND RESULTS: In rodent bone-marrow-derived macrophages (BMDM), the presence of Lact. fermentum CECT5716 induced pro-inflammatory cytokines, in contrast to the activation of IL-10 induced by Lact. salivarius CECT5713. Although both strains reduced the lipopolysaccharide (LPS)-induced inflammatory response in BMDM, the effect of Lact. salivarius CECT5713 was more efficient, probably because of the production of higher amounts of IL-10 cytokine. In vivo assays in mice showed similar results; the consumption of Lact. fermentum CECT5716 enhanced the production of Th1 cytokines by spleen cells and increased the IgA concentration in faeces. However, the consumption of Lact. salivarius CECT5713 induced IL-10 production by spleen cells. CONCLUSION: Therefore, in general, the effect of Lact. fermentum CECT5716 is immunostimulatory in contrast to the anti-inflammatory effect of Lact. salivarius CECT5713. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this study show that two Lactobacillus strains isolated from breast milk can exert different and even opposing effects on immune response demonstrating the specificity of each strain.