Structural studies of the exopolysaccharide consisting of a nonasaccharide repeating unit isolated from Lactobacillus rhamnosus KL37B

A novel structure of exopolysaccharide from the lactic acid bacteria (LAB) Lactobacillus rhamnosus KL37B, from the human intestinal flora, is described. During the structural investigation of the exopolysaccharide it was found that the repeating unit is a nonasaccharide, which is the largest repeating unit found in LAB exopolysaccharides to date. The polysaccharide material was prepared by TCA extraction of a bacterial cell mass, purified by anion-exchange and gel permeation chromatography and characterized using chemical and enzymatic methods. On the basis of monosaccharide and methylation analysis and also 1D and 2D ¹H and ¹³C NMR spectroscopy the exopolysaccharide was shown to be composed of the following nonasaccharide repeating unit:The physicochemical cell surface study and adhesive properties indicated distinct surface properties of Lactobacillus rhamnosus strain KL37B with high adhesive abilities to Caco-2 cells, hydrophobicity and slime production, in comparison to other Lactobacillus strains used as controls.     

Structural determination of a neutral exopolysaccharide produced by Lactobacillus delbrueckii ssp. bulgaricus LBB.B332

The neutral exopolysaccharide produced by Lactobacillus delbrueckii ssp. bulgaricus LBB.B332 in skimmed milk was found to be composed of d-glucose, d-galactose, and l-rhamnose in a molar ratio of 1:2:2. Linkage analysis and 1D/2D NMR (1H and 13C) studies carried out on the native polysaccharide as well as on an oligosaccharide generated by a periodate oxidation protocol, showed the polysaccharide to consist of linear pentasaccharide repeating units with the following structure: -->3-alpha-D-Glcp-(1-->3)-alpha-D-Galp-(1-->3)-alpha-L-Rhap-(1-->2)-alpha-L-Rhap-(1-->2)-alpha-D-Galp-(1-->.     

Structure of a neutral exopolysaccharide produced by Lactobacillus delbrueckii ssp. bulgaricus LBB.B26

The neutral exopolysaccharide produced by Lactobacillus delbrueckii ssp. bulgaricus LBB.B26 in skimmed milk was found to be composed of d-glucose and d-galactose in a molar ratio of 2:3. Linkage analysis and 1D/2D NMR ((1)H and (13)C) studies performed on the native polysaccharide, and on an oligosaccharide obtained from a partial acid hydrolysate of the native polysaccharide, showed the polysaccharide to consist of branched pentasaccharide repeating units with the following structure. [structure: see text]     

Structural determination of the complex exopolysaccharide from the virulent strain of Cryphonectria parasitica

The structure of a new exopolysaccharide from the virulent strain of Cryphonectria parasitica was elucidated by means of 2D NMR spectroscopy and selective degradations (mild hydrolysis and acetolysis). The polysaccharide is built up of mannose, galactose and rhamnose and has a rather complex non-repetitive structure that can be idealised as follows:     

Structural studies on the exopolysaccharide from Erwinia persicina

The Gram-negative bacterial strain HKI 0380 was isolated from biofilms located on palaeolithic rock paintings in the Cave of Bats in Zuheros, southern Spain. It was identified as the phytopathogenic Erwinia persicina and attracted attention due to the production of considerable quantities of slime. The acidic exopolysaccharide produced by the E. persicina was studied after O-deacylation by sugar and methylation analyses, along with (1)H and (13)C NMR spectroscopy. The following structure of the branched pentasaccharide repeating unit of the O-deacylated exopolysaccharide was established: [carbohydrate structure: see text].     

Draft genome sequences and description of Lactobacillus rhamnosus strains L31, L34, and L35

Lactobacillus rhamnosus is a facultative, lactic acid bacterium in the phylum Firmicutes. Lactobacillus spp. are generally considered beneficial, and specific strains of L. rhamnosus are validated probiotics. We describe the draft genomes of three L. rhamnosus strains (L31, L34, and L35) isolated from the feces of Thai breastfed infants, which exhibit anti-inflammatory properties in vitro. The three genomes range between 2.8 – 2.9 Mb, and contain approximately 2,700 protein coding genes.     

Phosphatidylinositol-specific phospholipase C activity in Lactobacillus rhamnosus with capacity to translocate

Phosphatidylinositol-specific phospholipase C (PI-PLC) activity was investigated in 25 different lactic acid bacteria (LAB) strains belonging to the genera Lactobacillus, Weisella, and Enterococcus. PI-PLC activity was detected in 44% of the strains studied in culture medium without carbon source. From the PI-PLC positive strains, Lactobacillus rhamnosus ATCC 7469 was selected for translocation studies. Healthy mice were orally administered with a daily dose of 2.0 x 10(9) of viable L. rhamnosus suspension. Viable bacteria were detected in liver and spleen of mice fed with LAB for 7 days. Bacterial colonies isolated from liver were biochemically characterized, and further subjected to randomly amplified polymorphic DNA. Amplification patterns of five strains displayed identical profiles to L. rhamnosus. PI-PLC activity was determined in the strains recovered from liver.     

In vitro mutagen binding and antimutagenic activity of human Lactobacillus rhamnosus 231

In vitro mutagen binding ability of human Lactobacillus rhamnosus 231 (Lr 231) was evaluated against acridine orange (AO), N-methyl-N’-nitro-N-nitrosoguanidine (MNNG), 2-amino-3, 8-dimethylimidazo-[4,5-f]-quinoxaline (MeIQx) and 4-nitro-o-phenylenediamine (NPD). Binding of AO by Lr 231 is due to adsorption, thereby leading to removal of mutagen in solution and is instantaneous, pH- and concentration-dependent. Whereas, binding of MNNG and MeIQx by Lr 231 results into biotransformation leading to detoxification with subsequent loss of mutagenicity as determined by spectral analysis, thin layer chromatography and Ames test. Binding of mutagen by Lr 231 was dependent on culture age and optimum binding of AO, MNNG and MeIQx was observed to occur with 24 h old culture. Cells of Lr 231 were subjected to different chemical treatments prior to binding studies. Results indicated cell wall component such as cell wall polysaccharide, peptidoglycan, carbohydrates and proteins plays an important role in adsorption of AO, also involving hydrophilic and ionic interactions. Binding, biotransformation and detoxification of MNNG and MeIQx by Lr 231 was dependent on cell surface characteristics mainly involving carbohydrates, proteins, teichoic acid/lipoteichoic acid, hydrophobic interaction and presence of thiol group. L. rhamnosus 231 bound MNNG instantaneously. More than 96 (p < 0.01) and 70% (p < 0.05) cells remained viable after mutagen binding and various pretreatments respectively. This study shows Lr 231 exhibits ability to bind and detoxify potent mutagens, and this property can be useful in formulating fermented foods for removal of potent mutagens.     

Determination of the structure and molecular weights of the exopolysaccharide produced by Lactobacillus acidophilus 5e2 when grown on different carbon feeds

Lactobacillus acidophilus 5e2 when grown on skimmed milk, skimmed milk supplemented with sodium formate and skimmed milk supplemented with glucose secretes a branched heteropolysaccharide having a weight average molecular weight less than 450 kDa. The exopolysaccharide has a heptasaccharide repeat unit and is composed of D-glucose, D-galactose and N-acetyl-D-glucosamine in the molar ratio 3:3:1. Using chemical techniques and 1D and 2D-NMR spectroscopy the polysaccharide has been shown to possess the following repeat unit structure:     

Diacetyl production and growth of Lactobacillus rhamnosus on multiple substrates

Lactobacillus rhamnosus is a heterolactic acid bacterium, which can be used to produce flavour compounds like diacetyl and acetoin. Various startegies have been applied to improve the growth rate and diacetyl yield. The use of multiple substrates affected growth as well as the yield of diacetyl. Growth on a medium containing glucose demonstrated a diauxic growth profile, with the second phase of growth being on the product, lactic acid. L. rhamnosus also grew on a medium containing citrate. Growth on medium containing glucose+citrate demonstrated simultaneous utilization of carbon sources. L. rhamnosus did not grow in a medium containing acetate and also did not co-metabolize it with glucose. Maximum specific growth rate ( _max) was found to increase in the case of simultaneous utilization of glucose+citrate (0.38 h^–1) as compared to glucose as the sole carbon source (0.28 h^–1). The yields of diacetyl were also found to increase for glucose + pyruvate and glucose + citrate (0.10 and 0.05 g g^–1 of glucose, respectively) as compared to glucose alone (0.01 g g^–1 of glucose). The productivity of diacetyl on medium containing glucose and citrate was double that of a medium containing only citrate, although the yields were comparable.     

Lactic acid fermentation by cells of Lactobacillus rhamnosus immobilized in polyacrylamide gel

Summary The process of lactic acid fermentation of lactose to lactic acid by Lactobacillus rhamnosus ATCC 7469 has been studied. The following processes have been explored: growth kinetics, as well as lactose utilization, production of lactic acid and further degradation of lactic acid. The immobilization experiments were conducted with microbial cells entrapped in polyacrylamide gels. Gels with different ratios of the monomer (acrylamide) and the cross-linking agent (N,N′-methylene-bis-acrylamide) have been tested. These were used in a repeat-batch process. The current processes inside and outside the gel particles were subjects of examination. The evolution of the activity of immobilized cells with repeated use showed that the particles served mainly as a donor of cells for the free culture. In all experiments a very high degree of conversion, 85–90% was observed. After several runs however, the particles were exhausted for microbial cells. A kinetic model of the process of lactic acid production was developed. This model allowed the evaluation of the effect of microbial growth and diffusion limitations inside the gel particles on the process rate and the separate contribution of the free and immobilized cells to the overall fermentation process upon multiple use.     

Structural studies on exopolysaccharides produced by three different propionibacteria strains

The exopolysaccharides produced by three propionibacteria strains, Propionibacterium freudenreichii 109, Propionibacterium freudenreichii 111, and Propionibacterium thoenii 126, grown on whey-based media, were found to be charged heteropolymers, composed of D-glucose, D-mannose, and D-glucuronic acid in molar ratios of 2:2:1. By means of methylation analysis, mass spectrometry, partial acid hydrolysis, and 1D/2D NMR (1H and 13C) studies, it was determined that all three exopolysaccharides contain the same branched, pentasaccharide repeating unit: [Formula: see text].     

鼠李糖乳杆菌遗传转化方法的优化

采用原生质体电转化和PEB电转化方法向鼠李糖乳杆菌LA-04-01中导入外源基因。在培养过程中使用甘氨酸和青霉素辅助溶菌酶处理细胞制备原生质体,得到300个/μg的转化子。使用PEB缓冲液处理细胞,使转化率稳定达到6×105个/μg,建立了针对鼠李糖乳杆菌简便高效的转化方法,为进一步的基因操作提供了便利。     

Physicochemical properties of exopolysaccharide produced by Lactobacillus kefiranofaciens ZW3 isolated from Tibet kefir

An exopolysaccharide (EPS) producing strain, ZW3, was isolated from Tibet kefir grain and was identified as Lactobacillus kefiranofaciens. FT-IR spectroscopy revealed the presence of carboxyl, hydroxyl, and amide groups, which correspond to a typical heteropolymeric polysaccharide. The GC analysis of ZW3 EPS revealed that it was glucogalactan in nature. Exopolymer showed similar flocculation stability like xanthan gum but better than guar gum with a melting point of 93.38 degrees C which is lower than xanthan gum (153.4 degrees C) and guar gum (490.11 degrees C). Compared with other commercially available hydrocolloids like xanthan gum, guar gum and locust gum ZW3 EPS showed much better emulsifying capability.     

Immune enhancement in rainbow trout (Oncorhynchus mykiss) by potential probiotic bacteria (Lactobacillus rhamnosus)

The present study assessed the immune enhancement of fish by a lactic acid bacterium (LAB) Lactobacillus rhamnosus (ATCC 53103). The bacterium was administered orally at five different doses 7.9 x 10(4) (LAB4), 2.1 x 10(6) (LAB6), 2.8 x 10(8) (LAB8), 1.9 x 10(10) (LAB10) and 9.7 x 10(10) (LAB11) CFU/g feed to rainbow trout for two weeks and the feed was changed to un-supplemented diet. From the onset of feeding supplemented diets at 1, 2, 3 and 4 weeks, blood and mucus samples were taken. During the LAB feeding period L. rhamnosus persisted in the fish intestine and in the tank water in high numbers. However, L. rhamnosus disappeared from the intestine, skin mucus and tank water within one week after the change to the non-supplemented feed. In comparison to untreated control fish, respiratory burst activity of blood cells was raised significantly in the LAB4 treated group on week 2. Serum-mediated killing of Escherichia coli was increased significantly in group LAB6 on week 2. Serum immunoglobulin levels were significantly raised only in LAB8 group on week 1 and in LAB4 and LAB8 at the end of the trial. The results show that rainbow trout immune parameters were enhanced by using probiotic bacteria.     

Complete genome sequencing of exopolysaccharide-producing Lactobacillus plantarum K25 provides genetic evidence for the probiotic functionality and cold endurance capacity of the strain

Lactobacillus plantarum (L. plantarum) K25 is a probiotic strain isolated from Tibetan kefir. Previous studies showed that this exopolysaccharide (EPS)-producing strain was antimicrobial active and cold tolerant. These functional traits were evidenced by complete genome sequencing of strain K25 with a circular 3,175,846-bp chromosome and six circular plasmids, encoding 3365 CDSs, 16 rRNA genes and 70 tRNA genes. Genomic analysis of L. plantarum K25 illustrates that this strain contains the previous reported mechanisms of probiotic functionality and cold tolerance, involving plantaricins, lysozyme, bile salt hydrolase, chaperone proteins, osmoprotectant, oxidoreductase, EPSs and terpenes. Interestingly, strain K25 harbors more genes that function in defense mechanisms, and lipid transport and metabolism, in comparison with other L. plantarum strains reported. The present study demonstrates the comprehensive analysis of genes related to probiotic functionalities of an EPS-producing L. plantarum strain based on whole genome sequencing.     

Influence of culture conditions on exopolysaccharide production by Lactobacillus rhamnosus strain C83

The influence of culture conditions on growth and exopolysaccharide (EPS) production by Lactobacillus rhamnosus strain C83 was investigated using fermentor batch cultures. A temperature shift (from 37 to 25 °C) at the beginning of the exponential growth phase (0–5 h) enhanced EPS production. Furthermore, an optimal environmental pH of 6·2–7·2 improved the performance of strain C83 and partially anaerobic culture conditions (pO₂ from 0 to 10%) triggered EPS over-production by the cells. The sugar composition of this polymer was independent of culture conditions, such as carbon source, medium composition, temperature, pH, pO₂ and growth phases. In all cases, galactose and glucose were the principal components.     

Structural studies of the exopolysaccharide produced by Lactobacillus rhamnosus strain GG (ATCC 53103)

The structure of the extracellular polysaccharide (EPS) from Lactobacillus rhamnosus strain GG has been investigated. In combination with component analysis, NMR spectroscopy shows that the polysaccharide is composed of hexasaccharide repeating units. Sequential information was obtained by two-dimensional (1)H,(1)H-NOESY, and (1)H,(13)C-HMBC NMR techniques. The structure of the repeating unit of the EPS from Lactobacillus rhamnosus strain GG was determined as: [carbohydrate structure: see text]