Bacillus subtilis and Macleaya cordata extract regulate the rumen microbiota associated with enteric methane emission in dairy cows.

BACKGROUND: Ruminant livestock production is a considerable source of enteric methane (CH4) emissions. In a previous study, we found that dietary inclusions of Bacillus subtilis (BS) and Macleaya cordata extract (MCE) increased dry matter intake and milk production, while reduced enteric CH4 emission in dairy cows. The objective of this study was to further elucidate the impact of feeding BS and MCE on rumen methanogenesis in dairy cows using rumen metagenomics techniques. RESULTS: Sixty dairy cows were blocked in 20 groups of 3 cows accordingly to their live weight, milk yield, and days in milk, and within each group, the 3 cows were randomly allocated to 1 of 3 treatments: control diet (CON), control diet plus BS (BS), and control diet plus MCE (MCE). After 75 days of feeding experimental diets, 12 cows were selected from each treatment for collection of rumen samples for the metagenomic sequencing. Results showed that BS decreased ruminal acetate and butyrate, while increased propionate concentrations, resulting in decreased acetate:propionate ratio. The metagenomics analysis revealed that MCE reduced relative abundances of Methanobrevibacter wolinii, Methanobrevibacter sp. AbM4, Candidatus Methanomassiliicoccus intestinalis, Methanobrevibacter cuticularis, Methanomicrobium mobile, Methanobacterium formicicum, and Methanobacterium congolense. Both BS and MCE reduced relative abundances of Methanosphaera sp. WGK6 and Methanosphaera stadtmanae. The co-occurrence network analysis of rumen bacteria and archaea revealed that dietary treatments influenced microbial interaction patterns, with BS and MCE cows having more and stronger associations than CON cows. The random forest and heatmaps analysis demonstrated that the Halopenitus persicus was positively correlated with fat- and protein-corrected milk yield; Clostridium sp. CAG 269, Clostridium sp. 27 14, Haloarcula rubripromontorii, and Methanobrevibacter curvatus were negatively correlated with rumen acetate and butyrate concentrations, and acetate:propionate ratio, whereas Selenomonas rumiantium was positively correlated with those variables. CONCLUSIONS: The present results provided new information for mitigation of enteric methane emissions of dairy cows by feeding BS and MCE to influence rumen microbial activities. This fundamental knowledge is essential for developing enteric CH4 reduction strategies to mitigate climate change and reduce dietary energy waste. Video Abstract.

Candida tropicalis as a novel dietary additive to reduce methane emissions and nitrogen excretion in sheep.

The goal of this study was to investigate Candida tropicalis as a kind of environmentally friendly dietary additive to manipulate ruminal fermentation patterns, reduce methane emissions and nitrogen excretion, and to screen the appropriate dose for sheep. Twenty-four Dorper × thin-tailed Han crossbred ewes (51.12 kg ± 2.23 kg BW) were selected and randomly divided into four groups which were fed Candida tropicalis at dose of 0 (control), 4 × 108 (low dose), 4 × 109 (medium dose), and 4 × 1010 (high dose) colony-forming units (CFU)/d per head, respectively. The experiment lasted 33 days with 21 days for adaptation and 12 days for nutrient digestibility trial and respiratory gases sampling. The results showed that nutrients intake was not affected by Candida tropicalis supplementation (P > 0.05), whereas apparent digestibility of nutrients significantly increased compared with the control group (P < 0.05). Nitrogen and energy utilization increased with Candida tropicalis supplementation (P < 0.05). Compared with the ewes of the control group, rumen fluid pH and NH3-N concentration were not affected (P > 0.05), whereas total volatile fatty acid concentration and molar proportion of propionate were greater (P < 0.05), and molar proportion of acetate and the ratio of acetate to propionate were less (P < 0.05) when the ewes were fed Candida tropicalis. Daily total CH4 production (L/d) and CH4 emissions yield (L/d of CH4 per kg of dry matter intake, metabolic weight, or digestibility dry matter intake) were decreased at the low dose group (P < 0.05). The abundance of total bacteria, methanogen, and protozoa in rumen fluid was significantly higher at medium dose and high dose of Candida tropicalis supplementation (P < 0.05) compared with low dose and the control group. In summary, Candida tropicalis supplementation has a potential to reduce CH4 emissions and nitrogen excretion, and the optimal dose should be 4 × 108 CFU/d per head.

Advances in the Application of Phytogenic Extracts as Antioxidants and Their Potential Mechanisms in Ruminants.

Under current breeding conditions, multiple stressors are important challenges facing animal husbandry in achieving animal wellbeing. For many years, the use of antibiotics has been a social concern in the livestock industry. With the implementation of the non-antibiotics policy, there is an urgent need to find relevant technologies and products to replace antibiotics and to solve the problem of disease prevention during animal growth. Phytogenic extracts have the unique advantages of being natural and extensive sources, having a low residue, and being pollution-free and renewable. They can relieve the various stresses, including oxidative stress, on animals and even control their inflammation by regulating the signaling pathways of proinflammatory cytokines, improving animal immunity, and improving the structure of microorganisms in the gastrointestinal tract, thereby becoming the priority choice for improving animal health. In this study, we reviewed the types of antioxidants commonly used in the livestock industry and their applicable effects on ruminants, as well as the recent research progress on their potential mechanisms of action. This review may provide a reference for further research and for the application of other phytogenic extracts and the elucidation of their precise mechanisms of action.

Effects of Anemoside B4 on Diarrhea Incidence, Serum Indices, and Fecal Microbial of Suckling Calves.

The study was conducted to evaluate the effects of Anemoside B4 on diarrhea incidence, serum indices, and fecal microbial of suckling calves. Sixty newborn Chinese Holstein calves with similar body weight (43.7 ± 3.9 kg) were randomly divided into four groups with 15 calves each, fed the diet which was supplied 0 (CON), 15 (A1), 30 (A2), and 45 (A3) mg/day of Anemoside B4, respectively. The trial period is 56 days. The blood and fecal samples were collected at 28 and 56 days of age. Results show that during the whole trial period, the diarrhea incidence in Group A1, A2, and A3 was significantly lower than that in Group CON (p < 0.05). Compared with the Group CON, Anemoside B4 supplementation significantly decreased the contents of serum D-lactic acid and diamine oxidase at 28-day-old (p < 0.05). At 56-day-old, the content of serum D-lactic acid in Group A3 tended to be higher (0.05 < p < 0.01), and the content of serum diamine oxidase in Group A3 increased significantly, in comparison with Group CON (p < 0.05). Group A3 increased the level of Chao1 and Simpson indices at 28-day-old (0.05 < p < 0.01), and Chao1, Observed_species, Shannon, and Simpson indices at 56-day-old (p < 0.05), in comparison to Group CON. Compared with Group CON, 45 mg / day Anemoside B4 supplementation significantly increased the contents of Bacteroidota (at the phylum level), Prevotella (at the genus level) at 28-day-old (p < 0.05), and the content of Sutterella (at the genus level) at 56-day-old (p < 0.05), promoted the processes of energy metabolism, glycan biosynthesis and metabolism, metabolism of cofactors and vitamins (p < 0.05). A positive correlation was observed between Prevotella and metabolism of cofactors and vitamins, energy metabolism, and glycan biosynthesis and metabolism. A positive correlation was observed between Sutterella and energy metabolism. In conclusion, Anemoside B4 could effectively alleviate calf diarrhea, protect the integrity of intestinal mucosa, and change the structure of intestinal microbiota, indicating the potential value of Anemoside B4 in regulating intestinal microbiota and the prevention of intestinal diseases.

The crucial role of lysine in the hepatic metabolism of growing Holstein dairy heifers as revealed by LC-MS-based untargeted metabolomics.

The objective of this experiment was to evaluate the effect of supplementing rumen-protected Lys based on a Lys-deficient diet on liver metabolism in growing Holstein heifers. The experiment was conducted for 3 months with 36 Holstein heifers (initial body weight: 200 ± 9.0 kg; 7-month-old). Heifers were randomly assigned to 2 diets based on corn, soybean meal, alfalfa hay, and wheat bran: control, Lys-deficient diet (LD; 0.66% Lys in diet), and Lys-adequate diet (LA; 1.00% Lys in diet). The results showed no difference in growth performance between the 2 groups (P > 0.05). However, there was a clear trend of increasing feed conversion rate with Lys supplementation (0.05 < P < 0.01). The serum urea nitrogen concentration was significantly decreased, and the aspartate aminotransferase-to-alanine aminotransferase ratio was significantly decreased by Lys supplementation (P < 0.05). Moreover, growing heifers fed a Lys-adequate diet had lower levels of urine nitrogen excretion and higher levels of the biological value of nitrogen (P < 0.05). Metabolomic analysis revealed that 5 types of phosphatidylcholine and 3 types of ceramide were significantly increased and enriched in sphingolipid metabolism and glycerophospholipid metabolism (P < 0.05). His, Leu, and Asp levels were significantly decreased in the liver following Lys supplementation (P < 0.05). In conclusion, Lys supplementation may promote the synthesis of body tissue proteins, as evidenced by significantly decreased amino acids in the liver and urine N excretion, it also improves hepatic lipid metabolism by providing lipoprotein precursors.

Effects of dietary rumen-protected Lys levels on rumen fermentation and bacterial community composition in Holstein heifers.

This study aimed to evaluate the effects of partial reducing rumen-protected Lys (RPLys) on rumen fermentation and microbial composition in heifers. Three ruminal fistulated Holstein Friesian bulls were used to determine the effective degradability of RPLys using an in situ method at incubation times of 0, 2, 6, 12, 16, 24, 36, and 48 h. Thereafter, 36 Holstein heifers at 90 days of age were assigned to one of two dietary treatments: a theoretically balanced amino acid diet (PC group; 1.21% Lys, 0.4% Met) or a 30% Lys-reduced diet (PCLys group, 0.85% Lys, 0.4% Met). Rumen fluid samples from five heifers in each group were extracted using esophageal tubing on day 90 to determine pH, microprotein, ammonia, volatile fatty acids, and microbial communities. Results showed that the effective ruminal degradability was 25.76%. Furthermore, differences in rumen fermentation parameters and alpha diversity of the microbiota between the two groups were not significant, but beta diversity was significant. Based upon relative abundance analysis, short-chain fatty acid-producing bacteria, including Sharpea, Syntrophococcus, [Ruminococcus]_gauvreauii_group, Acetitomaculum, and [Eubacterium]_nadotum_group belonging to Firmicutes, were significantly decreased in the PCLys group. Spearman's analysis revealed a positive correlation between the butyrate molar proportion and the relative abundance of butyrate-producing bacteria such as [Eubacterium]_nadotum_group, Coprococcus_1, Ruminococcaceae_UCG_013, Pseudoramibacter, and Lachnospiraceae_UCG_010. Phylogenetic Investigation of Communities by Reconstruction of Unobserved States analysis further validated that RPLys deduction influenced energy metabolism. Together, our findings highlight the role of RPLys or Lys in butyrate-producing bacteria. However, the number of bacteria affected by Lys was very limited and insufficient to alter rumen fermentation. Key Points • Reducing 30% Lys via rumen-protected Lys did not affect rumen fermentation parameters and alpha diversity of microbiota of Holstein heifers. It meant that the ruminal fermentation pattern was not changed. • Reducing 30% Lys via rumen-protected lysine significantly decreased relative abundance of short-chain fatty acid-producing bacteria belonging to Firmicutes. • Functions of microorganisms were changed by reducing 30% Lys via rumen-protected Lys, especially amino acid metabolism. It may affect the amino acid composition of microprotein.

Dynamic profiles of fermentation characteristics and bacterial community composition of Broussonetia papyrifera ensiled with perennial ryegrass.

Broussonetia papyrifera (B. papyrifera) has been proposed to improve silage fermentation due to its high content of protein and abundant active plant extracts. Thus, dynamic profiles of fermentation quality and bacterial community of B. papyrifera mixing with perennial ryegrass in different ratios: 100:0, 90:10, 80:20, 70:30, 60:40, and 50:50 were examined during 60-d fermentation. Results showed that adding perennial ryegrass increased soluble carbohydrate content and lactic acid production in silage and decreased pH and population of epiphytic microorganisms. Adding ryegrass exerted a remarkable effect on the silage bacterial community with a dramatic decrease in the abundance of Enterobacter. Spearman's rank correlation showed that silage lactic acid concentration was positively correlated with Lactobacillus and Stenotrophomonas abundance, while ammonia nitrogen concentration was positively correlated with the abundance of Enterobacter. In conclusion, B. papyrifera ensiled with perennial ryegrass could improve B. papyrifera silage quality and provide high-quality forage resources for sustainable ruminant livestock production.

Effect of Broussonetia papyrifera L. silage on blood biochemical parameters, growth performance, meat amino acids and fatty acids compositions in beef cattle.

OBJECTIVE: The study was conducted to investigate the effects of Broussonetia papyrifera L.(B. papyrifera) silage on growth performance, serum biochemical parameters, meat quality, and meat amino acids and fatty acids compositions in beef cattle. METHODS: Sixty-four male Angus beef cattle were assigned to 4 groups with 4 pens in each group and 4 beef cattle in each pen, and fed with the total mixed ration supplemented with 0%, 5%, 10%, or 15% B. papyrifera silage for 100 days (control group, 5% group, 10% group and 15% group) separately. RESULTS: Beef cattle had significantly higher final body weight (BW) in 15% group, higher average daily gain (ADG) and dry matter intake (DMI) in 5% group, 10% group and 15% group, and higher feed conversion ratio (FCR) in 10% group and 15% group. Significantly higher blood superoxide dismutase (SOD) concentration was noted in 15% group, higher blood total antioxidant capacity (TAC) in 10% group and 15% group, lower 8-hydroxydeoxyguanosine (8-OHdG) and malondialdehyde (MDA) in 15% group. Meat had lower pH in 15% group, higher Commission International DeI'Eclairage (CIE) L* in 5% group, 10% group, and 15% group, and lower drip loss in 15% group. Greater concentration of meat polyunsaturated fatty acids (PUFA) was observed in 10% group and 15% group, and docosahexaenoic acid (DHA) in 15% group. CONCLUSION: Diet with 15% B. papyrifera silage could improve performance and increase final BW, ADG, DMI, and FCR, enhance the antioxidant functions by decreasing blood 8-OHdG and MDA and increasing blood SOD and TAC, improve the meat quality by lowing pH and drip loss and increasing CIE L*, increase the meat PUFA and DHA concentration. Polyphenols and flavonoids might be the main components responsible for the antioxidant activity and anti-biohydrogenation in the B. papyrifera silage. And B. papyrifera silage could be used as a new feedstuff in beef cattle nutrition.

The Facilitating Effect of Tartary Buckwheat Flavonoids and Lactobacillus plantarum on the Growth Performance, Nutrient Digestibility, Antioxidant Capacity, and Fecal Microbiota of Weaned Piglets.

Natural plant extracts and probiotics has been proved as the most preferred and effective alternatives for antibiotics in animal feeding. The current study aimed to investigate the facilitating effect of tartary buckwheat flavonoids and Lactobacillus plantarum on the growth performance, nutrient digestibility, antioxidant capacity, and microbiota of weaned piglets. Fifty 35-day-old weaned piglets (7.85 ± 0.67 kg) were randomly divided into five treatments with 10 piglets per treatment. Piglets in the negative control (NC) group were fed a basal diet, and others were fed basal diets supplemented with 40 mg/kg of colistin sulfate (positive control, PC), 109 CFU/kg Lactobacillus plantarum (LP), 40 mg/kg of tartary buckwheat flavonoids (BF), and a combination of 109 CFU/kg Lactobacillus plantarum and 40 mg/kg of tartary buckwheat flavonoids (LB). Supplementation of BF increased the average daily gain of piglets in the BF group (p < 0.05). The nutrient digestibility of piglets in the NC group was lower than that in other groups, while the digestibility of gross energy, dry matter, organic matter, and phosphorus of piglets in the LB group was higher than the other four groups (p < 0.05). Compared with the NC and pC group, supplementation of Lp increased the activity of superoxide dismutase (SOD), glutathione peroxidase (GSH-px), and catalase (CAT), while the BF increased the content of IgA and IgM (p < 0.05). Supplementation of colistin sulfate decreased the alpha diversity index, including chao and observed species, while the addition of Lp or combination of Lp and BF increased the abundance of Selenomonas or Mitsuokella in fecal samples, respectively. The results indicated that supplementation of Lactobacillus plantarum can improve the antioxidant capacity, while tartary buckwheat flavones can increase the growth performance and immune ability of weaned piglets. Moreover, in combination, they promote nutrient digestibility.

The Limiting Sequence and Appropriate Amino Acid Ratio of Lysine, Methionine, and Threonine for Seven- to Nine-Month-Old Holstein Heifers Fed Corn-Soybean M-Based Diet.

An "Amino acid (AA) partial deletion method" was used in this experiment to study the limiting sequences and appropriate ratio of lysine (Lys), methionine (Met), and threonine (Thr) in the diets of 7- to 9-month-old Holstein heifers. The experiment was conducted for three months with 72 Holstein heifers (age = 22 ± 0.5 weeks old; BW = 200 ± 9.0 kg; mean ± standard deviation). Following an initial two weeks adaptation period, heifers were allocated to one of four treatments: a theoretically balanced amino acid diet (positive control [PC]; 1.00% Lys, 0.33% Met, and 0.72% Thr), a 30% Lys deleted diet (partially deleted Lys [PD-Lys]; 0.66% Lys, 0.33% Met, and 0.72% Thr), a 30% Met deleted diet (partially deleted Met, [PD-Met]; 1.00% Lys, 0.22% Met, and 0.72% Thr), and a 30% Thr deleted diet (partially deleted Thr [PD-Thr]; 1.00% Lys, 0.33% Met, and 0.45% Thr). Experimental animals were fed a corn-soybean meal-based concentrate and alfalfa hay. In addition, the animals were provided with supplemental Lys, Met, and Thr (ruminal bypass). The results found no differences in the growth performance and nitrogen retention between PD-Thr treatment and PC treatment (p > 0.05). The average daily gain (p = 0.0013) and feed conversion efficiency (p = 0.0057) of eight- to ninr-month-old heifers were lower in both PD-Lys and PD-Met treatment than those in PC treatment. According to growth performance, Lys was the first limiting AA, followed by Met and Thr. Moreover, nine-month-old Holstein heifers in PD-Lys treatment and PD-Met treatment had higher levels of serum urea nitrogen (p = 0.0021), urea nitrogen (p = 0.0011) and total excreted N (p = 0.0324) than those in PC treatment, which showed that nitrogen retention significantly decreased (p = 0.0048) as dietary Lys and Met levels decreased. The limiting sequence based on nitrogen retention was the same as that based on growth performance. The appropriate ratio of Lys, Met, and Thr in the diet based on nitrogen retention was 100:32:57. In summary, the limiting sequence and appropriate amino acid ratio of Lys, Met, and Thr for seven- to nine-month-old Holstein heifers fed a corn-soybean meal-based diet were Lys > Met > Thr and 100:32:57, respectively.

Influence of dietary supplementation with Bacillus licheniformis and Saccharomyces cerevisiae as alternatives to monensin on growth performance, antioxidant, immunity, ruminal fermentation and microbial diversity of fattening lambs.

Alternatives to antibiotics for improving productivity and maintaining the health of livestock health are urgently needed. The scope of this research was conducted to investigate the effects of two alternatives (Bacillus licheniformis and Saccharomyces cerevisiae) to monensin on growth performance, antioxidant capacity, immunity, ruminal fermentation and microbial diversity of fattening lambs. One hundred and sixty Dorper × Thin-tailed Han sheep (32 ± 3.45 kg BW) were randomly assigned into 5 treatments of n = 32 lambs/group. Lambs in the control group were fed a basal diet (NC) while the other four treatments were fed basal diets supplemented with monensin (PC), Bacillus licheniformis (BL), Saccharomyces cerevisiae (SC), and the combination of Bacillus licheniformis and Saccharomyces cerevisiae with protease (BS), respectively. The experiment lasted for 66 d. Feed intake was recorded every 2 d and lambs were weighed every 20 d. Ten lambs from each group were slaughtered at the end of the trial, and samples of serum and rumen fluid were collected. The results indicated that the dietary regimen did not affect the dry matter intake (DMI). The average daily gain (ADG) of BS treatment was significantly higher than NC group (P < 0.05). Compared with the NC treatment, the other four supplementation treatments increased the concentration of growth hormone (GH), insulin-like growth factor I (IGF-I) and insulin (INS) (P < 0.05). The malondialdehyde (MDA) and total antioxidant capacity (TAOC) showed no significant difference among the 5 treatments while the activity of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) of BS group was significantly increased (P < 0.05). The supplementation regimen decreased the concentration of ammonia Nitrogen (NH3-N) and increased the content of microbial crude proteins (MCP) (P < 0.05). The supplementation of antibiotics and probiotics reduced the concentrations of acetate and increased the concentrations of propionate (P < 0.05). The supplementation treatments increased the relative abundance of Lentisphaerae, Fibrobacteres and Tenericutes at the phylum level, whereas at the genus level, they increased the relative abundance of Fibrobacter (P < 0.05). Overall, this study confirmed the facilitating effect of B. licheniformis, S. cerevisiae and their compounds on growth performance, improve the antioxidant capacity and immune function, and beneficially manipulate ruminal fermentation and microbial diversity of fatting lambs.

Polysaccharide structure and immunological relationships of RG-I pectin from the bee pollen of Nelumbo nucifera.

From the bee pollen of Nelumbo nucifera, we used hot water extraction and chromatographic fractionation on DEAE-cellulose and Sepharose CL-6B to purify polysaccharides (WNPP) to homogeneity. WNPP-2-RG (3.8 × 105 Da) consisted primarily of Rha (11.5%), GalA (12.0%), Gal (41.2%) and Ara (29.7%). Structure analysis showed that this polysaccharide is a RG-I type pectin containing AG-I and AG-II side chains comprised of 1,5-L-Ara (25.6%), t-D-Gal (12.0%), and 1,6-D-Gal (18.3%) as the primary linkage types. Immunological activity assays demonstrated that WNPP-2-RG increased lymphocyte proliferation and macrophage phagocytosis, and induced production of NO. Compared to WNPP-2-RG, its hydrolysates (RG-8H-P) had no effect on lymphocyte proliferation, although they did promote macrophage phagocytosis and decreased NO production. Our results suggest that Gal side chain residues may play a crucial role in macrophage phagocytosis, whereas Ara is unimportant for NO production and neither Gal nor Ara impact lymphocyte proliferation.

Effects of feeding a Moringa oleifera rachis and twig preparation to dairy cows on their milk production and fatty acid composition, and plasma antioxidants.

BACKGROUND: We determined how supplementing the diet of lactating, multiparous Holstein dairy cows with a preparation of Moringa oleifera rachises and twigs affected their milk production and quality and the levels of plasma antioxidants. RESULTS: We found that milk yield increased in cows receiving the 6% (w/w) moringa supplement compared with that of the control. Addition of the moringa supplement increased the concentration of milk fat and decreased the somatic cell count in the milk. However, protein, glucose and total solid and urea nitrogen concentrations in the milk were the same for all treatments. The concentration of glutathione peroxidase increased for cows fed the moringa supplement compared with the control. The percentages of total unsaturated fatty acids, mono-unsaturated fatty acids, and polyunsaturated fatty acids including n-3 polyunsaturated fatty acid increased in the milk of cows fed the moringa supplement compared with those of the controls. CONCLUSION: Addition of the moringa supplement into the diet of lactating multiparous cows improved milk production and health status and modified milk fatty acid profile positively. The results suggested that moringa supplement could be used as a diet supplement for producing high quality and healthier milk. © 2017 Society of Chemical Industry.

Effects of dietary supplementation with two alternatives to antibiotics on intestinal microbiota of preweaned calves challenged with Escherichia coli K99.

The aim of this study was to investigate the effects of dietary supplementation with two alternatives to antibiotics (Candida tropicalis and mulberry leaf flavonoids) on intestinal microbiota of preweaned calves challenged with Escherichia coli K99. Sixty Holstein calves were randomly assigned to 5 treatments: fed a basal diet (N-CON); fed a basal diet and challenged with E.coli K99 (P-CON); fed a basal diet supplemented with C.tropicalis (CT), mulberry leaf flavonoids (MLF), and the combination of the two additives (CM), respectively, and challenged with E.coli K99. The MLF and CM groups had significantly higher average daily grain and feed efficiency, and significantly lower fecal scores compared with the P-CON group after E. coli K99 challenge. The supplementation groups increased the relative abundance, at the phylum level, of Bacteroidetes and Proteobacteria, whereas at the genus level, they increased the relative abundance of Prevotella, Lactobacillus, and Enterococcus. Quantitative PCR revealed that the CT, MLF, and CM groups had significantly lower copy numbers of E.coli K99 compared with the P-CON group. The CT, MLF, and CM treatments reduce days of diarrhea, improve intestinal health, and beneficially manipulate the intestinal microbiota in preweaned calves.

Dietary supplementation with mulberry leaf flavonoids inhibits methanogenesis in sheep.

The effects of flavonoids on methanogenesis and microbial flora in Dorper × thin-tailed Han crossbred ewes were evaluated in two experiments. To investigate the effects of flavonoids on nutrient digestibility and nitrogen balance, 18 ewes (60.0 ± 1.73 kg body weight (BW)) were allotted to two dietary treatments in experiment one, a control diet and the control diet supplemented with flavonoids (2 g/head/day). In experiment two, the effects of supplementary flavonoids on ruminal fermentation and microbial flora were investigated using quantitative polymerase chain reaction with six ewes (67.2 ± 0.79 kg BW) with ruminal cannula assigned to the identical dietary treatments used in experiment one. Supplementary flavonoids improved the apparent digestibility of nitrogen (N, P < 0.001) and neutral detergent fibre (NDF, P = 0.024) and decreased daily CH4 output (P < 0.001). The ruminal pH (P = 0.638) and ammonia (P = 0.690) were not affected by supplementary flavonoids, whereas the total volatile fatty acid (VFA) content increased (P = 0.037). Supplementary flavonoids decreased ruminal populations of protozoans (P = 0.002) and methanogens (P < 0.001) and increased the populations of Fibrobacter succinogenes (P = 0.016). In conclusion, flavonoids improved the digestibility of organic matter and reduced CH4 output by inhibiting the populations of microbes involved in methanogenesis.

Oral administration of Lactobacillus plantarum and Bacillus subtilis on rumen fermentation and the bacterial community in calves.

The objective of this study was to assess the effect of dietary probiotics on rumen fermentation and the bacterial community in dairy calves. Twelve Holstein calves were randomly allocated to three treatments: a basal diet, the basal diet supplemented with Lactobacillus plantarum GF103 (LB) or basal diet supplemented with a mixture of Lactobacillus plantarum GF103 and Bacillus subtilis B27 (LBS). A milk replacer was fed to calves from 8 days of age. A starter and alfalfa hay was offered ad libitum from 21 and 28 days of age, respectively, and the orts were weighted daily. The ruminal fluid was sampled at 56 and 83 days of age to determine the rumen fermentation characteristics. The bacterial community was analyzed by denaturing gradient gel electrophoresis (DGGE) and the number of certain bacteria was quantified by real-time polymerase chain reaction. The ratio of total dry matter intake to average body wieght was higher in the control (P < 0.05). The DGGE fingerprint of the 16S ribosomal RNA gene was affected by the blended probiotics at 83 days of age. The number of Ruminococcus albus was lower in the LB and LBS treatment (P < 0.05). Oral administration of the probiotics affected the rumen bacterial community and the numbers of cellulolytic bacteria decreased.

Antitumor activity of bee pollen polysaccharides from Rosa rugosa.

In the present study, bee pollen polysaccharides from Rosa rugosa (WRPP) were extracted and fractionated. WRPP were purified to neutral (WRPP-N) and acidic polysaccharides (WRPP-1, WRPP-2) with DEAE-Cellulose. WRPP-N were mainly composed of glucose, mannose, arabinose and galactose, indicating the existence of glucan, arabinogalactan (AG) and mannoglucan. WRPP-1 mainly consisted of rhamnose (3.0%), galacturonic acid (12.4%), galactose (24.7%) and arabinose (53.9%), and contained a large proportion of AGs. WRPP-2 consisted of rhamnose (7.8%), galacturonic acid (23.0%), galactose (15%) and arabinose (48.7%), while WRPP-2 contained more galacturonic acid compared to WRPP-1. WRPP-1 and WRPP-2 were composed by type I rhamnogalacturonan (RG-I), homogalacturonan (HG) and AG fragments, while WRPP-2 contained more HG and RG-I. All the fractions had significant anti-proliferative activity in HT-29 and HCT116 cells; the neutral and acidic fractions were shown to have significant synergistic effects which accounted for the antitumor activity of bee pollen polysaccharides from Rosa rugosa in vitro.

The limiting sequence and proper ratio of lysine, methionine and threonine for calves fed milk replacers containing soy protein.

The limiting sequence and relative ratio of lysine (Lys), methionine (Met), and threonine (Thr) for calves about 2 mo of age fed milk replacers (MR) containing soy protein are not clearly defined. The objective of the study was to investigate the effect of supplementing MR containing 22% CP, half from soy protein concentrate (SPC, 40.56% CP, flour) and half from whey proteins, with Lys, Met, and Thr to estimate amino acid (AA) sequence and their relative ratio for calves about 2 mo of age. A method of partial deduction of AA was adopted. Twenty-four newborn calves (half males and half females, 40.7±0.9 kg of BW) were fed 1 of 4 MR diets for 56 d (n = 6/diet). The diets were supplemented with all (positive control) or with 2 of the 3 AAs: Lys, Met and Thr, (i.e., PC (22% CP, 2.34% Lys, 0.72% Met and 1.80% Thr), PC-Lys (22% CP, 1.64% Lys, 0.72% Met and 1.80% Thr), PC-Met (22% CP, 2.34% Lys, 0.50% Met and 1.80% Thr), and PC-Thr (22% CP, 2.34% Lys, 0.72% Met and 1.26% Thr)). Calves were fed thrice daily; starter (20% CP, 1.03% Lys, 0.30% Met and 0.69% Thr), hay (3.23% CP, 0.29% Lys, 0.12% Met and 0.23% Thr) and water were offered free choice. Starter and hay were only offered beginning on d 36 (after 5 wk) and d 43 (after 6 wk), respectively. BW, body size and blood samples measures were taken every two weeks. Three-day total collection of feed refusals, feces, and urine were recorded starting at d 33 and d 54 of age, respectively. From the results, the limiting sequence and relative ratio between the 3 AAs in calves with different diet structures were calculated. The limiting sequence of the 3 AAs were ranked as Lys, Met and Thr; the proper ratio was 100:29:70 for MR-only diet and 100:30:60 for diets consisted of MR, starter and hay. Nitrogen digestion and utilization and nutrient digestibility were negatively affected by AA deletion treatments. From the evidence of this experiment, it did not appear that the AA limiting sequence was selectively altered by differences in diet structures such as would be encountered in practice. The relative ratio between the 3 AAs varied with the offer of starter and hay to calves, and the average ratio was 100:29.5:65 for calves during 2 to 10 wk of age.

Diversity, abundance and characterization of ruminal cysteine phytases suggest their important role in phytate degradation.

A novel class of cysteine phytase showing ability to degrade phytate has recently been isolated from rumen bacteria. To expand our knowledge of this enzyme class, a total of 101 distinct cysteine phytase gene fragments were identified from the ruminal genomic DNA of Bore goats and Holstein cows, and most of them shared low identities (< 50%) with known sequences. By phylogenetic analysis, these sequences were separated into three clusters that showed substantial diversity. The two most abundant cysteine phytase genes of goat rumens were cloned and their protein products were characterized. Four findings were revealed based on our results. (i) Compared with soil and water environment, where ß-propeller phytase is the most important phytate-degrading enzyme, cysteine phytase is the major phytate-degrading enzyme in the anaerobic ruminal environment. (ii) Cysteine phytase fragments in the rumen contents of goat and cow have the same diversity profile, although most of the sequences and their abundance differ in the two species. (iii) Each species has their respective high-abundance genes, which may play major roles for phytate degradation. (iv) Compared with previously reported cysteine phytases that have pH optimum at 4.5, the pH optima of the two most abundant secreted goat cysteine phytases are 6.5 and 6.0, which are within the pH range found in the rumens. This study provides valuable information about the diversity, abundance and enzymatic properties of the ruminal cysteine phytases and emphasizes the important role(s) of these cysteine phytases probably in the terrestrial cycle of phosphorus.