In vitro investigation of the effects of Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 exopolysaccharides on tight junction damage caused by influenza virus infection.

It is a problem that influenza virus infection increases susceptibility to secondary bacterial infection in lungs leading to lethal pneumonia. We previously reported that exopolysaccharides (EPS) derived from Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 (OLL1073R-1) could prevent against influenza virus infection followed by secondary bacterial infection in vitro. Therefore, the present study assessed whether EPS derived OLL1073R-1 protects the alveolar epithelial barrier disfunction caused by influenza virus infection. After A549 cells treated with EPS or without EPS were infected influenza virus A/Puerto Rico/8/34 (IFV) for 12 h, the levels of tight junction genes expression and inflammatory genes expression were measured by reverse transcription polymerase chain reaction. As results, EPS treatment could protect against low-titer IFV infection, but not high-titer IFV infection, followed by suppression of the increased expression of inflammatory cytokine gene levels and recovery of the decrease in the expression level of ZO-1 gene that was caused by low-titer IFV infection, leading to an improvement trend in the barrier function. Our findings showed that EPS derived from OLL1073R-1 could inhibit low-titer IFV infection leading to maintenance of the epithelial barrier function through the suppression of inflammatory cytokine genes expression.

Fermentation characteristics and postacidification of yogurt by Streptococcus thermophilus CICC 6038 and Lactobacillus delbrueckii ssp. bulgaricus CICC 6047 at optimal inoculum ratio.

This study aimed to investigate the symbiosis between Streptococcus thermophilus CICC 6038 and Lactobacillus delbrueckii ssp. bulgaricus CICC 6047. In addition, the effect of their different inoculum ratios was determined, and comparison experiments of fermentation characteristics and storage stability of milk fermented by their monocultures and cocultures at optimal inoculum ratio were performed. We found the time to obtain pH 4.6 and ΔpH during storage varied among 6 inoculum ratios (1:1, 2:1, 10:1, 19:1, 50:1, 100:1). By the statistical model to evaluate the optimal ratio, the ratio of 19:1 was selected, which exhibited high acidification rate and low postacidification with pH values remaining between 4.2 and 4.4 after a 50-d storage. Among the 3 groups included in our analyses (i.e., the monocultures of S. thermophilus CICC 6038 [St] and Lb. bulgaricus CICC 6047 [Lb] and their cocultures [St+Lb] at 19:1), the coculture group showed higher acidification activity, improved rheological properties, richer typical volatile compounds, more desirable sensor quality after the fermentation process than the other 2 groups. However, the continuous accumulation of acetic acid during storage showed that acetic acid was more highly correlated with postacidification than d-lactic acid for the Lb group and St+Lb group. Our study emphasized the importance of selecting an appropriate bacterial consortium at the optimal inoculum ratio to achieve favorable fermentation performance and enhanced postacidification stability during storage.

Role of fumarate reductase on the fermentation properties of Lactobacillus delbrueckii ssp. bulgaricus.

Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus are symbiotic starters widely used in yogurt fermentation. They exchange metabolites to meet their nutritional demands during fermentation, promoting mutual growth. Although S. thermophilus produces fumaric acid, and the addition of fumaric acid has been shown to promote the growth of L. bulgaricus monoculture, whether fumaric acid produced by S. thermophilus is used by L. bulgaricus during coculture remains unclear. Furthermore, the importance of fumaric acid metabolism in the growth of L. bulgaricus is yet to be elucidated. Therefore, in this study, we investigated the importance of fumaric acid metabolism in L. bulgaricus monocultures and coculture with S. thermophilus. We deleted the fumarate reductase gene (frd), which is responsible for the metabolism of fumaric acid to succinic acid, in L. bulgaricus strains 2038 and NCIMB 701373. Both Δfrd strains exhibited longer fermentation times than their parent strains, and fumaric acid was metabolized to malic acid rather than succinic acid. Coculture of Δfrd strains with S. thermophilus 1131 also resulted in a longer fermentation time, and the accumulation of malic acid was observed. These results indicated that fumaric acid produced by S. thermophilus is used by L. bulgaricus as a symbiotic substance during yogurt fermentation and that the metabolism of fumaric acid to succinic acid by fumarate reductase is a key factor determining the fermentation ability of L. bulgaricus.

The interaction between Lactobacillus delbrueckii ssp. bulgaricus M58 and Streptococcus thermophilus S10 can enhanced texture and flavor profile of fermented milk: Insights from metabolomics analysis.

Lactobacillus delbrueckii ssp. bulgaricus M58 (M58) and Streptococcus thermophilus S10 (S10) are 2 dairy starter strains known for their favorable fermentation characteristics. Therefore, this research aimed to study the effects of 1-d low-temperature ripening on the physicochemical properties and metabolomics of fermented milk. Initially, the performance of single (M58 or S10) and dual (M58+S10) strain fermentation was assessed, revealing that the M58+S10 combination resulted in a shortened fermentation time, a stable gel structure, and desirable viscosity, suggesting positive strain interactions. Subsequently, non-targeted metabolomics analyses using LC-MS and GC-MS were performed to comparatively analyze M58+S10 fermented milk samples collected at the end of fermentation and after 1-d low-temperature ripening. The results showed a significant increase in almost all small peptides and dodecanedioic acid in the samples after one day of ripening, while there was a substantial decrease in indole and amino acid metabolites. Moreover, notable increases were observed in high-quality flavor compounds, such as geraniol, delta-nonalactone, 1-hexanol,2-ethyl-, methyl jasmonate, and undecanal. This study provides valuable insights into the fermentation characteristics of the dual bacterial starter consisting of M58 and S10 strains and highlights the specific contribution of the low-temperature ripening step to the overall quality of fermented milk.

Comparative transcriptomic analysis of the flavor production mechanism in yogurt by traditional starter strains.

The synergistic fermentation of milk by Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus is one of the key factors that determines the quality of yogurt. In this study, the mechanism whereby yogurt flavor compounds are produced by a mixture of S. thermophilus SIT-20.S and L. delbrueckii ssp. bulgaricus SIT-17.B were investigated by examining the flavor production, growth, and gene transcription of these strains. The results showed that yogurt produced by a 10:1 mixture of the aforementioned strains had the highest abundance of acetoin, whereas yogurt produced by a 1:1 mixture had the highest abundance of diacetyl and acetaldehyde. In addition, the growth of S. thermophilus SIT-20.S was enhanced in the 10:1 mixture. Transcriptomic analysis revealed differentially expressed genes in the flavor-compound-related pathways of S. thermophilus SIT-20.S and L. delbrueckii ssp. bulgaricus SIT-17.B in yogurts produced by 10:1 and 1:1 mixtures compared with those produced by either strain alone. Mixed fermentations regulated the expression of genes related to glycolysis, resulting in an increase of pyruvate, which is an important precursor for diacetyl and acetoin synthesis. The gene encoding the acetoin reductase (SIT-20S_orf01454) was decreased in S. thermophilus SIT-20.S, which ensured the accumulation of acetoin. In addition, the gene encoding the acetaldehyde dehydrogenase (SIT-20S_orf00949) was upregulated in S. thermophilus SIT-20.S, and the expression of alcohol dehydrogenase (SIT-20S_orf01479; SIT-17B_orf00943) was downregulated in both strains, maintaining the abundance of acetaldehyde. In addition, the gene encoding the NADH oxidase (SIT-17B_orf00860) in L. delbrueckii ssp. bulgaricus SIT-17.B were upregulated, which promoted the accumulation of diacetyl and acetoin. Overall, we characterized the mechanism by which S. thermophilus and L. delbrueckii ssp. bulgaricus synergistically generated yogurt flavor compounds during their production of yogurt and highlighted the importance of appropriate proportions of fermentation starters for improving the flavor of yogurts.

Copper inhibits postacidification of yogurt and affects its flavor: A study based on the Cop operon.

Yogurt and its related products are popular worldwide. During transportation and storage, Lactobacillus delbrueckii ssp. bulgaricus in yogurt continues to metabolize to form lactic acid, the postacidification phenomenon of yogurt. Postacidification of yogurt is a widespread phenomenon in the dairy industry. Many scholars have done research on controlling the postacidification process, but few report on the molecular mechanisms involved. In this study, we used a molecular-assisted approach to screen food additives that can inhibit postacidification and analyzed its effects on yogurt quality as well as its regulatory mechanism from multi-omics perspectives in combination. The copper ion was found to upregulate the expression of the LDB_RS05285 gene, and the copper transporter-related genes were regulated by copper. Based on the metabolic-level analysis, copper was found to promote lactose hydrolysis, accumulate a large amount of glucose and galactose, inhibit the conversion of glucose to lactic acid, and reduce the production of lactic acid. The significantly greater abundance of l-isoleucine and l-phenylalanine increased the abundance of 3-methylbutyraldehyde (∼1.2 times) and benzaldehyde (∼7.9 times) to different degrees, which contributed to the formation of the overall flavor of yogurt. Copper not only stabilizes the acidity of yogurt, but also it improves the flavor of yogurt. Through this established method involving quantitative and correlation analyses at the transcriptional and metabolic levels, this study provides guidance for the research and development of food additives that inhibit postacidification of yogurt and provide a reference for studying the changes of metabolites during storage of yogurt.

The effect of glutathione biosynthesis of Streptococcus thermophilus ST-1 on cocultured Lactobacillus delbrueckii ssp. bulgaricus ATCC11842.

Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus are the main species used for yogurt preparation. Glutathione (GSH) can be synthesized by S. thermophilus and plays a crucial role in combating environmental stress. However, the effect of GSH biosynthesis by S. thermophilus on cocultured L. delbrueckii ssp. bulgaricus is still unknown. In this study, a mutant S. thermophilus ΔgshF was constructed by deleting the GSH synthase. The wild strain S. thermophilus ST-1 and ΔgshF mutants were cocultured with L. delbrueckii ssp. bulgaricus ATCC11842 by using Transwell chambers (Guangzhou Shuopu Biotechnology Co., Ltd.), respectively. It was proven that the GSH synthesized by S. thermophilus ST-1 could be absorbed and used by L. delbrueckii ssp. bulgaricus ATCC11842, and promote growth ability and stress tolerance of L. delbrueckii ssp. bulgaricus ATCC11842. The biomass of L. delbrueckii ssp. bulgaricus ATCC11842 cocultured with S. thermophilus ST-1 or ΔgshF (adding exogenous GSH) increased by 1.8 and 1.4 times compared with the biomass of L. delbrueckii ssp. bulgaricus ATCC11842 cocultured with S. thermophilus ΔgshF. Meanwhile, after H2O2 and low-temperature treatments, the bacterial viability of L. delbrueckii ssp. bulgaricus cocultured with S. thermophilus ΔgshF, with or without GSH, was decreased by 41 and 15% compared with that of L. delbrueckii ssp. bulgaricus cocultured with S. thermophilus ST-1. Furthermore, transcriptome analysis showed that the expression levels of genes involved in purine nucleotide and pyrimidine nucleotide metabolism in L. delbrueckii ssp. bulgaricus ATCC11842 were at least 3 times increased when cocultured with S. thermophilus (fold change > 3.0). Moreover, compared with the mutant strain ΔgshF, the wild-type strain ST-1 could shorten the fermented curd time by 5.3 hours during yogurt preparation. These results indicated that the GSH synthesized by S. thermophilus during cocultivation effectively enhanced the activity of L. delbrueckii ssp. bulgaricus and significantly improved the quality of fermented milk.

Exopolysaccharides from Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 prevent influenza virus infection and attenuate secondary bacterial infection risk.

The present study assessed the inhibitory action of exopolysaccharides (EPS) produced by Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 against influenza virus infection followed by secondary bacterial infection. We found that the presence of 200 or 400 µg ml-1 of EPS significantly protected against influenza virus infection in a dose-dependent manner when A549 cells were treated with EPS before infection but not after it. The expression of carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM-1), an adhesion molecule for bacteria adherence, on A549 cells was significantly enhanced during influenza virus infection compared with viral-non-infected A549 cells. However, this upregulated CEACAM-1 expression was significantly decreased by EPS treatment before viral infection in association with the reduction in the virus titre in A549 cells. In a bacterial adhesion assay using Staphylococcus aureus, the bacterial adherence to viral-infected A549 cells was significantly greater than that to viral-non-infected A549 cells, and the increased bacterial adherence induced by influenza virus infection tended to be decreased by EPS treatment before the infection. Our findings show that EPS treatment before viral infection can inhibit influenza virus infection and alleviate secondary bacterial infection through decreased CEACAM-1 expression.

Analysis of the complete genome sequence of Lactobacillus delbrueckii ssp. bulgaricus with post-acidification capacity and its influence on yogurt in storage.

In recent years, yogurt has been one of the most popular fermented dairy products and is sold worldwide. In this study, pH and titrated acid changes of 4 strains of Lactobacillus delbrueckii ssp. bulgaricus fermented milk during storage were detected. The difference between L. bulgaricus KLDS1.1011 and KLDS1.0207 was significant, with the latter exhibiting reduced acidity levels. Therefore, we determined the complete genome sequence of the 2 strains. Then the expression of specific genes and common genes related to glucose metabolism and proteolysis of L. bulgaricus KLDS1.1011 and KLDS1.0207 were detected by quantitative real-time reverse-transcription PCR. Analysis indicated that the key enzymes in glycometabolism and proteolysis of L. bulgaricus KLDS1.1011 were significantly different than those of L. bulgaricus KLDS1.0207. The contents of lactose and glucose decreased during storage of L. bulgaricus fermented milk, as determined by HPLC, and the contents of lactic acid and galactose increased, with L. bulgaricus KLDS1.1011 increasing less. With skim milk as a raw material, L. bulgaricus KLDS1.1011, KLDS1.0207, and Streptococcus thermophilus S1 were used as fermentation strains to yield yogurt at 42°C, and sensory evaluation was compared with yogurt fermented by commercial starter cultures. Yogurt from L. bulgaricus KLDS1.1011 was the highest-rated. Therefore, the study may provide guidelines for the development of yogurt starters.

Consumption of OLL1073R-1 yogurt improves psychological quality of life in women healthcare workers: secondary analysis of a randomized controlled trial.

BACKGROUND: We conducted a randomized controlled trial to investigate the effects of consumption of yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 in women healthcare workers. In a previous study we used these data to investigate hypothesized preventive effects against flu, however any effects on improving mental quality of life were not analyzed at that time. In the present study, we focus on that aspect. METHODS: The participants (961 women; mainly nurses, aged 20-71 years) were randomly allocated to either the yogurt group (n = 479) or the control group (n = 482). Participants in the yogurt group drank 112 mL of OLL1073R-1 yogurt for 16 weeks, while those in the control group did not consume any yogurt. All participants were prohibited from consuming other yogurt or fermented dairy products during the study period. The participants answered the Pittsburgh Sleep Quality Index (PSQI), Short Form-8 Health Survey (SF-8), and Gastrointestinal Symptom Rating Scale (GSRS) questionnaires at baseline and after 16 weeks. RESULTS: The PSQI score showed significant improvement after the intake of yogurt (p < 0.01). SF-8 results showed significant intervention effects in the General Health and Vitality scores (p = 0.02 and p = 0.01, respectively). In other subscales of SF-8, we did not observe significant effects of the yogurt. In the GSRS, daily intake of yogurt exerted a preventive effect on constipation (p = 0.03). CONCLUSIONS: Consumption of yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 enhances subjective psychological quality of life by improving quality of sleep and gastrointestinal condition among women healthcare workers.

Effect of fumaric acid on the growth of Lactobacillus delbrueckii ssp. bulgaricus during yogurt fermentation.

Yogurt is traditionally fermented by a symbiotic starter culture of Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus. These bacteria exchange metabolites with each other to meet their nutritional demands during protocooperation, resulting in a shorter fermentation time. In this study, we investigated whether fumaric acid functions as a symbiotic agent to promote the growth of Lb. bulgaricus by evaluating 8 strains of Lb. bulgaricus and 7 strains of Strep. thermophilus. All the tested Lb. bulgaricus strains metabolized the added fumaric acid into succinic acid during monoculture in milk, and 6 strains (75%) showed shorter fermentation time compared with the control. The addition of malic acid showed similar trends as that of fumaric acid, indicating that the reverse tricarboxylic acid cycle was functioning in Lb. bulgaricus. All 7 Strep. thermophilus strains tested produced fumaric acid during monoculture in milk. Further, in Lb. bulgaricus 2038, the gene expression of fumarate reductase that converts fumaric acid to succinic acid, was higher in the coculture with Strep. thermophilus 1131 than in the monoculture. These findings indicate that fumaric acid produced by Strep. thermophilus can function as a symbiotic substance during yogurt fermentation to stimulate the growth of Lb. bulgaricus.

Effect of lactose hydrolysis on the milk-fermenting properties of Lactobacillus delbrueckii ssp. bulgaricus 2038 and Streptococcus thermophilus 1131.

Yogurt is a well-known nutritious and probiotic food and is traditionally fermented from milk using the symbiotic starter culture of Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus. However, yogurt consumption may cause health problems in lactose-intolerant individuals, and the demand for lactose-free yogurt has been increasing. The standard method to prepare lactose-free yogurt is to hydrolyze milk by lactase; however, this process has been reported to influence the fermentation properties of starter strains. This study aimed to investigate the fermentation properties of an industrial starter culture of L. bulgaricus 2038 and S. thermophilus 1131 in lactose-hydrolyzed milk and to examine the metabolic changes induced by glucose utilization. We found that the cell number of L. bulgaricus 2038, exopolysaccharide concentration, and viscosity in the coculture of L. bulgaricus 2038 and S. thermophilus 1131 was significantly increased in lactose-hydrolyzed milk compared with that in unhydrolyzed milk. Although the cell number of S. thermophilus 1131 showed no difference, production of formic acid and reduction of dissolved oxygen were enhanced in lactose-hydrolyzed milk. Further, in lactose-hydrolyzed milk, S. thermophilus 1131 was found to have increased the expression of NADH oxidase, which is responsible for oxygen reduction. These results indicated that glucose utilization promoted S. thermophilus 1131 to rapidly reduce the dissolved oxygen amount and produce a high concentration of formic acid, presumably resulting in the increased cell number of L. bulgaricus 2038 in the coculture. Our study provides basic information on the metabolic changes in starter strains in lactose-hydrolyzed milk, and demonstrates that lactose-free yogurt with increased cell number of L. bulgaricus can be prepared without delay in fermentation and decrease in the cell number of S. thermophilus.

Exploring the industrial potential of Lactobacillus delbrueckii ssp. bulgaricus by population genomics and genome-wide association study analysis.

Lactobacillus delbrueckii ssp. bulgaricus is one of the most commonly used starter cultures for yogurt production. However, how its genetic background affects acid production capacity is unclear. This study investigated the industrial potential of L. delbrueckii ssp. bulgaricus using population genomics and GWAS analysis. To meet our goal, population genetics and functional genomics analyses were performed on 188 newly sequenced L. delbrueckii ssp. bulgaricus strains isolated from naturally fermented dairy products together with 19 genome sequences retrieved from the NCBI database. Four distinct clusters were identified, and they were correlated with the geographical sites where the samples were collected. The GWAS analysis about acidification fermentation results with sucrose-fortified reconstituted skim milk revealed a significant association between l-lactate dehydrogenase (lldD; Ldb2036) and the bacterial acid production rate. Our study has broadened the understanding of the population structure and genetic diversity of L. delbrueckii ssp. bulgaricus by identifying potential targets for further research, development, and use of lactic acid bacteria in the dairy industry.

Phenotypic, fermentation characterization, and resistance mechanism analysis of bacteriophage-resistant mutants of Lactobacillus delbrueckii ssp. bulgaricus isolated from traditional Chinese dairy products.

Bacteriophage infection is a large factor in dairy industrial production failure on the basis of pure inoculation fermentation, and developing good commercial starter cultures from wild dairy products and improving the environmental vigor of starter cultures by enhancing their phage resistance are still the most effective solutions. Here we used a spontaneous isolation method to obtain bacteriophage-resistant mutants of Lactobacillus delbrueckii ssp. bulgaricus strains that are used in traditional Chinese fermented dairy products. We analyzed their phenotypes, fermentation characteristics, and resistance mechanisms. The results showed that bacteriophage-insensitive mutants (BIM) BIM8 and BIM12 had high bacteriophage resistance while exhibiting fermentation and coagulation attributes that were as satisfying as those of their respective parent strains KLDS1.1016 and KLDS1.1028. According to the attachment receptor detection, mutants BIM8 and BIM12 exhibited reduced absorption to bacteriophage phiLdb compared with their respective bacteriophage-sensitive parent strains because of changes to the polysaccharides or teichoic acids connected to their peptidoglycan layer. Additionally, genes, including HSDR, HSDM, and HSDS, encoding 3 subunits of a type I restriction-modification system were identified in their respective parent strains. We also discovered that HSDR and HSDM were highly conserved but that HSDS was variable because it is responsible for the DNA specificity of the complex. The late lysis that occurred only in strain KLDS1.1016 and not in strain KLDS1.1028 suggests that the former and its mutant BIM8 also may have an activatable restriction-modification mechanism. We conclude that the L. bulgaricus BIM8 and BIM12 mutants have great potential in the dairy industry as starter cultures, and their phage-resistance mechanism was effective mainly due to the adsorption interference and restriction-modification system.

Antibacterial activity of selenium-enriched lactic acid bacteria against common food-borne pathogens in vitro.

Selenium (Se) is an essential trace element for human health and animal nutrition. The aim of this study was to evaluate the inhibitory activities of Se-enriched lactic acid bacteria (LAB), Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus, against pathogenic Salmonella typhimurium, Escherichia coli, Staphylococcus aureus, and Listeria monocytogenes in vitro. The results indicated that the accumulation amount of Se by Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus reached 12.05 ± 0.43 µg/mL and 11.56 ± 0.25 µg/mL, respectively, accompanied by the relative maximum living cells when sodium selenite was 80 µg/mL. Oxford cup double plate assay showed that bacterial culture solution and cell-free culture supernatant (CFCS) from Se-enriched LAB exerted stronger antibacterial activity than those from the non-Se strains. The growth of pathogenic bacterial culture with CFCS at any growth stages was worse than that without CFCS; moreover, the inhibiting effect of CFCS of Se-enriched LAB was more significant than that of non-Se strains. Results from a scanning electron microscope equipped with energy dispersion X-ray spectrometry showed that elemental Se nanoparticles, which characteristically energy peak around 1.42 keV, were deposited on the cell surfaces of Lactobacillus delbrueckii ssp. bulgaricus. In addition, CFCS of Se-enriched LAB induced more serious cell structure damage of pathogenic bacteria than did non-Se LAB.

Screening for proteolytically active lactic acid bacteria and bioactivity of peptide hydrolysates obtained with selected strains.

In a screening for proteolytically active lactic acid bacteria, three strains, Lactobacillus delbrueckii ssp. lactis 92202, Lactobacillus helveticus 92201, and Lactobacillus delbrueckii ssp. bulgaricus 92059, showed the highest activities following growth in milk. All three strains degraded α- and ß-casein, but did not hydrolyse κ-casein. HPLC analysis of skim milk fermentation revealed increasing amounts of peptides after 5 and 10 h with Lb. d. ssp. bulgaricus 92059. Hydrolysates obtained with Lb. d. ssp. lactis 92202 and Lb. d. ssp. bulgaricus 92059 revealed the highest angiotensin-converting enzyme-inhibitory effect. The effect was dose dependent. Almost no effect (<10%) was seen for Lb. helveticus 92201. For Lb. d. ssp. bulgaricus 92059, maximal inhibition of approx. 65% was reached after 25 h of fermentation. In an in vitro assay measuring potential immunomodulation, hydrolysates of the three strains yielded anti-inflammatory activities in the presence of TNF-α. However, the effects were more pronounced at lower hydrolysate concentrations. In the absence of TNF-α, slight pro-inflammatory effects were observed. The hydrolysate of Lb. d. ssp. bulgaricus 92059, when purified by means of solid-phase extraction, exhibited pro-inflammatory activity. Sour whey containing Lb. d. ssp. bulgaricus 92059 cells showed pro-inflammatory activity while cell-free sour whey was clearly anti-inflammatory. In the purified hydrolysate, 20 different α- and ß-casein (CN)-derived peptides could be identified by LC-MS. Most peptides originated from the central and C-terminal regions of ß-casein. Peptide length was between 9 (ß-CN(f 59-67)) and 22 amino acids (ß-CN(f 117-138)).

Differential enumeration of subpopulations in concentrated frozen and lyophilized cultures of Lactobacillus delbrueckii ssp. bulgaricus.

Differential enumeration of subpopulations in concentrated frozen and lyophilized cultures of Lactobacillus delbrueckii ssp. bulgaricus ND02 derived from 2 propagation procedures was determined. The subpopulations consisted of 3 categories (physiological states): viable cells capable of forming colonies on agar plates (VC+), viable cells incapable of forming colonies on agar plates (VC-), widely referred to as viable but nonculturable (VBNC) cells, and nonviable or dead cells (NVC). Counts of VC+ were recorded using a conventional plate count procedure. A fluorescent vital staining procedure that discriminates between viable (VC+ and VC-) and NVC cells was used to determine the number of viable and nonviable cells. Both propagation procedures had 2 variables: in procedure (P)1, the propagation medium was rich in yeast extract (4.0%) and the pH was maintained at 5.7; in P2, the medium was devoid of yeast extract and the pH was maintained at 5.1. The results showed that post-propagation operations-concentration of cells by centrifugation and subsequent freezing or lyophilization of cell concentrate-induced different degrees of transience from VC+ to VC- states in cells derived from P1 and P2. Compared with cells derived from P2, cells from P1 were more labile to stress associated with centrifugation, freezing, and lyophilization, as revealed by differential counting.

Effects of yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 on the IgA flow rate of saliva in elderly persons residing in a nursing home: A before-after non-randomised intervention study.

OBJECTIVE: The aim of this study was to investigate the alterations in the salivary IgA levels of elderly persons administered yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) OLL1073R-1, which has been reported to reduce the risk of colds. BACKGROUND: Salivary immunoglobulin (Ig)A plays an important role in the defence of the oral cavity mucous membrane against foreign antigens and pathogens. Accordingly, low levels of salivary IgA are associated with an increased risk of upper respiratory tract infection. Furthermore, salivary IgA secretion has been reported to decrease with age. Recently, several studies have reported that certain strains of Lactobacillus and their products can modulate the immune response, but there are currently few studies on the effects of on the IgA level in human saliva. MATERIALS AND METHODS: This was a before-after non-randomised intervention study. Thirty-seven elderly persons (mean age, 82.7 years) residing in a single nursing home ingested 112 g of the yogurt every morning for 12 weeks. The participants' saliva was collected before and after 4, 8 and 12 weeks of yogurt intake. RESULTS: Our results showed that yogurt intake affected the concentration of IgA in the saliva (P < .0001). Additionally, yogurt intake and the body weight of the participants affected the IgA flow rate of saliva (P = .0003 and .03, respectively). CONCLUSION: Continuous intake of yogurt fermented with L. bulgaricus OLL1073R-1 may help improve the mucosal immune function in elderly people with weakened immune systems.

Relationships between functional genes in Lactobacillus delbrueckii ssp. bulgaricus isolates and phenotypic characteristics associated with fermentation time and flavor production in yogurt elucidated using multilocus sequence typing.

Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) is well known for its worldwide application in yogurt production. Flavor production and acid producing are considered as the most important characteristics for starter culture screening. To our knowledge this is the first study applying functional gene sequence multilocus sequence typing technology to predict the fermentation and flavor-producing characteristics of yogurt-producing bacteria. In the present study, phenotypic characteristics of 35 L. bulgaricus strains were quantified during the fermentation of milk to yogurt and during its subsequent storage; these included fermentation time, acidification rate, pH, titratable acidity, and flavor characteristics (acetaldehyde concentration). Furthermore, multilocus sequence typing analysis of 7 functional genes associated with fermentation time, acid production, and flavor formation was done to elucidate the phylogeny and genetic evolution of the same L. bulgaricus isolates. The results showed that strains significantly differed in fermentation time, acidification rate, and acetaldehyde production. Combining functional gene sequence analysis with phenotypic characteristics demonstrated that groups of strains established using genotype data were consistent with groups identified based on their phenotypic traits. This study has established an efficient and rapid molecular genotyping method to identify strains with good fermentation traits; this has the potential to replace time-consuming conventional methods based on direct measurement of phenotypic traits.

Growth and acid production of Lactobacillus delbrueckii ssp. bulgaricus ATCC 11842 in the fermentation of algal carcass.

Algal carcass is a low-value byproduct of algae after its conversion to biodiesel. Dried algal carcass is rich in protein, carbohydrate, and multiple amino acids, and it is typically well suited for growth and acid production of lactic acid bacteria. In this study, Lactobacillus delbrueckii ssp. bulgaricus ATCC 11842 was used to ferment different algal carcass media (ACM), including 2% ACM, 2% ACM with 1.9% glucose (ACM-G), and 2% ACM with 1.9% glucose and 2g/L amino acid mixture (ACM-GA). Concentrations of organic acids (lactic acid and acetic acid), acetyl-CoA, and ATP were analyzed by HPLC, and activities of lactate dehydrogenase (LDH), acetokinase (ACK), pyruvate kinase (PK), and phosphofructokinase (PFK) were determined by using a chemical approach. The growth of L. bulgaricus cells in ACM-GA was close to that in the control medium (de Man, Rogosa, and Sharpe). Lactic acid and acetic acid contents were greatly reduced when L. bulgaricus cells were grown in ACM compared with the control medium. Acetyl-CoA content varied with organic acid content and was increased in cells grown in different ACM compared with the control medium. The ATP content of L. bulgaricus cells in ACM was reduced compared with that of cells grown in the control medium. Activities of PFK and ACK of L. bulgaricus cells grown in ACM were higher and those of PK and LDH were lower compared with the control. Thus, ACM rich in nutrients may serve as an excellent substrate for growth by lactic acid bacteria, and addition of appropriate amounts of glucose and amino acids can improve growth and acid production.