Fengwei Tian

Quantitative Genetic Background of the Host Influences Gut Microbiomes in Chickens

Host genotype and gender are among the factors that influence the composition of gut microbiota. We studied the population structure of gut microbiota in two lines of chickens maintained under the same husbandry and dietary regimes. The lines, which originated from a common founder population, had undergone 54 generations of selection for high (HW) or low (LW) 56-day body weight, and now differ by more than 10-fold in body weight at selection age. Of 190 microbiome species, 68 were affected by genotype (line), gender, and genotype by gender interactions. Fifteen of the 68 species belong to Lactobacillus. Species affected by genotype, gender, and the genotype by gender interaction, were 29, 48, and 12, respectively. Species affected by gender were 30 and 17 in the HW and LW lines, respectively. Thus, under a common diet and husbandry host quantitative genotype and gender influenced gut microbiota composite.

Production, Purification, and Characterization of a Potential Thermostable Galactosidase for Milk Lactose Hydrolysis from Bacillus stearothermophilus

Beta-galactosidase, commonly named lactase, is one of the most important enzymes used in dairy processing; it catalyzes the hydrolysis of lactose to its constituent monosaccharides glucose and galactose. Here, a thermostable beta-galactosidase gene bgaB from Bacillus stearothermophilus was cloned and expressed in B. subtilis WB600. The recombinant enzyme was purified by a combination of heat treatment, ammonium sulfate fractionation, ion exchange, and gel filtration chromatography techniques. The purified beta-galactosidase appeared as a single protein band in sodium dodecyl sulfate-PAGE gel with a molecular mass of approximately 70 kDa. Its isoelectric point, determined by polyacryl-amide gel isoelectric focusing, was close to 5.1. The optimum temperature and pH for this beta-galactosidase activity were 70 degrees C and pH 7.0, respectively. Kinetics of thermal inactivation and half-life times for this thermostable enzyme at 65 and 70 degrees C were 50 and 9 h, respectively, and the K(m) and V(max) values were 2.96 mM and 6.62 micromol/min per mg. Metal cations and EDTA could not activate this thermostable enzyme, and some divalent metal ions, namely, Fe(2+), Zn(2+), Cu(2+), Pb(2+), and Sn(2+), inhibited its activity. Thiol reagents had no effect on the enzyme activity, and sulfhydryl group blocking reagents inactivated the enzyme. This enzyme possessed a high level of transgalactosylation activity in hydrolysis of lactose in milk. The results suggest that this recombinant thermostable enzyme may be suitable for both the hydrolysis of lactose and the production of galactooligosaccharides in milk processing.

Protective Effects of Lactobacillus plantarum CCFM8610 against Acute Cadmium Toxicity in Mice

ABSTRACT This study evaluated the protective effects of Lactobacillus plantarum CCFM8610, a selected probiotic with good cadmium binding capacity, against acute cadmium toxicity in mice. Ninety mice were divided into prevention and therapy groups. In the prevention groups, CCFM8610 was administered at 109 CFU once daily for 7 days, followed by a single oral dose of cadmium chloride at 1.8 mg cadmium for each mouse. In the therapy groups, the same dose of CCFM8610 was administered for 2 days after an identical single dose of cadmium exposure. Mice that received neither cadmium nor culture or that received cadmium alone served as negative and positive controls, respectively. The effects of both living and dead CCFM8610 on cadmium ion concentrations in feces, liver, and kidney were determined. Moreover, the alterations in reduced glutathione (GSH), malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and histopathology in the liver and kidney were investigated. The results showed that compared to the mice that received cadmium only, CCFM8610 treatment can effectively decrease intestinal cadmium absorption, reduce tissue cadmium accumulation, alleviate renal and hepatic oxidative stress, and ameliorate hepatic histopathological changes. Living CCFM8610 administered after cadmium exposure offered the most significant protection. Our results suggested that CCFM8610 is more effective against acute cadmium toxicity than a simple antioxidant treatment due to its special physiological functions and that it can be considered a new dietary therapeutic strategy against acute cadmium toxicity.

Immobilization of recombinant thermostable β-galactosidase from Bacillus stearothermophilus for lactose hydrolysis in milk

A recombinant thermostable beta-galactosidase from Bacillus stearothermophilus was immobilized onto chitosan using Tris(hydroxymethyl)phosphine (THP) and glutaraldehyde, and a packed bed reactor was utilized to hydrolyze lactose in milk. The thermostability and enzyme activity of THP-immobilized beta-galactosidase during storage was superior to that of free and glutaraldehyde-immobilized enzymes. The THP-immobilized beta-galactosidase showed greater relative activity in the presence of Ca(2+) than the free enzyme and was stable during the storage at 4 degrees C for 6 wk, whereas the free enzyme lost 31% of the initial activity under the same storage conditions. More than 80% of lactose hydrolysis in milk was achieved after 2 h of operation in the reactor. Therefore, THP-immobilized recombinant thermostable beta-galactosidase from Bacillus stearothermophilus has the potential for application in the production of lactose-hydrolyzed milk.

Protective effects of Lactobacillus plantarum CCFM8610 against chronic cadmium toxicity in mice indicate routes of protection besides intestinal sequestration.

ABSTRACT Our previous study confirmed the ability of Lactobacillus plantarum CCFM8610 to protect against acute cadmium (Cd) toxicity in mice. This study was designed to evaluate the protective effects of CCFM8610 against chronic Cd toxicity in mice and to gain insights into the protection mode of this strain. Experimental mice were divided into two groups and exposed to Cd for 8 weeks via drinking water or intraperitoneal injection. Both groups were further divided into four subgroups, control, Cd only, CCFM8610 only, and Cd plus CCFM8610. Levels of Cd were measured in the feces, liver, and kidneys, and alterations of several biomarkers of Cd toxicity were noted. The results showed that when Cd was introduced orally, cotreatment with Cd and CCFM8610 effectively decreased intestinal Cd absorption, reduced Cd accumulation in tissue, alleviated tissue oxidative stress, reversed hepatic and renal damage, and ameliorated the corresponding histopathological changes. When Cd was introduced intraperitoneally, administration of CCFM8610 did not have an impact on tissue Cd accumulation or reverse the activities of antioxidant enzymes. However, CCFM8610 still offered protection against oxidative stress and reversed the alterations of Cd toxicity biomarkers and tissue histopathology. These results suggest that CCFM8610 is effective against chronic cadmium toxicity in mice. Besides intestinal Cd sequestration, CCFM8610 treatment offers direct protection against Cd-induced oxidative stress. We also provide evidence that the latter is unlikely to be mediated via protection against Cd-induced alteration of antioxidant enzyme activities.

Functional significance of bioactive peptides derived from milk proteins.

Bioactive peptides can be defined as protein fragments with potential biological activities. Milk proteins are precursors of many different biologically active peptides. Bioactive peptides from milk proteins are considered potential modulators of various regulatory processes in the body. They mediate physiological functions in cardiovascular, nervous, gastro intestinal and immune systems. The functional significance of bioactivities depends on peptide fragment. Bioactive peptides encrypted in major milk proteins are latent within the sequence of the parent protein molecule. They can be liberated by (i) gastro intestinal digestion of milk, (ii) fermentation of milk with proteolytic starter cultures, and (iii) hydrolysis by proteolytic enzymes. In relation to their mode of action, bioactive peptides may reach target sites at the luminal side of intestinal tract, or after absorption, in peripheral organs. The production, functionalities, and mode of action of bioactive milk peptides as well as latest peptide products and ingredients are reviewed.

Effects of cryoprotectants on viability of Lactobacillus reuteri CICC6226

Freeze-drying is commonly used to preserve probiotics, but it could cause cell damage and loss of viability. The cryoprotectants play an important role in the conservation of viability during freeze-drying. In this study, we investigated the survival rates of Lactobacillus reuteri CICC6226 in the presence of cryoprotectants such as sucrose, trehalose, and reconstituted skim milk (RSM). In addition, we determined the activities of hexokinase (HK), pyruvate kinase (PK), lactate dehydrogenase (LDH), and ATPases immediately following the freeze-drying. The results showed that the differences in HK and PK activities with and without the cryoprotectants during freeze-drying were not significant, but cell viability and activities of LDH and ATPase were significantly different (P<0.01) prior to and after freeze-drying. Meanwhile, the results showed that the maintenance of the membrane integrity and fluidity was improved in the presence of the 10% trehalose or 10% RSM than other treatments during freeze-drying. These results have provided direct biochemical and metabolic evidence of injured cell during freeze-drying. Freeze-drying damaged membrane structure and function of cell and inactivated enzymes (LDH and ATPases). The results imply that LDH and ATPases are key markers and could be used to evaluate the effect of cryoprotectants on viability and metabolic activities of L. reuteri CICC6226 during freeze-drying.

Synthesis of conjugated linoleic acid by the linoleate isomerase complex in food-derived lactobacilli.

To assess strains of lactobacilli for their capacity to produce functional fatty acid‐conjugated linoleic acid. To assess the linoleate isomerase for CLA production in the most efficient CLA producer.

Modulation of peanut-induced allergic immune responses by oral lactic acid bacteria-based vaccines in mice.

Peanut allergy (PNA) has becoming a non-negligible health concern worldwide. Thus far, allergen-specific immunotherapy aimed at inducing mucosal tolerance has widely been regarded as a major management strategy for PNA. The safety profiles and the intrinsic probiotic properties of lactic acid bacteria (LAB) render them attractive delivery vehicles for mucosal vaccines. In the present study, we exploited genetically modified Lactococcus lactis to produce peanut allergen Ara h 2 via different protein-targeting systems and their immunomodulatory potency for allergic immune responses in mice were investigated. By comparison with the strain expressing the cytoplasmic form of Ara h 2 (LL1), the strains expressing the secreted and anchored forms of Ara h 2 (LL2 and LL3) were more potent in redirecting a Th2-polarized to a non-allergic Th1 immune responses. Induction of SIgA and regulatory T cells were also observed at the local levels by orally administration of recombinant L. lactis. Our results indicate that allergen-producing L. lactis strains modulated allergic immune responses and may be developed as promising mucosal vaccines for managing allergic diseases.

Extracellular secretion in Bacillus subtilis of a cytoplasmic thermostable β-galactosidase from Geobacillus stearothermophilus

beta-Galactosidase catalyzes the hydrolysis of beta-galactosides into monosaccharides and is widely used in dairy processing. This study reports the extracellular secretion of a cytoplasmic thermostable beta-galactosidase from Geobacillus stearothermophilus IAM11001 in Bacillus subtilis. This enzyme has potential applications in the dairy industry. It was not secreted in B. subtilis by mediation of 3 general secretory signal peptides, but was secreted extracellularly when it was fused to a twin-arginine signal peptide of B. subtilis phosphodiesterase. Defined and rich culture media were used for recombinant enzyme production, and the extracellular target enzymatic activity reached about 44% of the total enzymatic activity synthesized at 18 h of cultivation in Luria-Bertani medium. As a control of secretion, when the signal peptide coding sequence was absent from the N terminus of the target gene bgaB, the extracellular target enzymatic activity obtained under the same condition of cultivation accounted for less than 7% of the total enzymatic activity synthesized. Results also showed that coexpression of the B. subtilis proteins TatAd and TatCd was indispensable for the secretion of the target enzyme.