Linking metabolites in eight bioactive forage species to their in vitro methane reduction potential across several cultivars and harvests.

An in vitro Hohenheim gas test was conducted to analyze the fermentation end-products from 17 cultivars of eight polyphenol containing forage species. The polyphenol composition and proanthocyanidin (PA) structural features of all the cultivars were analyzed with UPLC-MS/MS in leaves of vegetative or generative plants. The samples were incubated with and without polyethylene glycol (PEG, a tannin-binding agent) to separate the tannin-effect on methane (CH4, ml/200 mg DM) production from that of forage quality. Sulla and big trefoil, two particularly PA rich species, were found to have the highest CH4 reduction potential of up to 47% when compared to the samples without PEG. However, concomitant reduction in gas production (GP, ml/200 mg DM) of up to 44% was also observed. An increase in both GP and CH4 production under PEG treatments, confirms the role of tannins in CH4 reduction. Moreover, PA structural features and concentration were found to be an important source of variation for CH4 production from PA containing species. Despite having low polyphenol concentrations, chicory and plantain were found to reduce CH4 production without reducing GP. Additionally, interspecies variability was found to be higher than intraspecies variability, and these results were consistent across growth stages, indicating the findings' representativeness.

Diets for Dairy Cows with Different Proportions of Crude Protein Originating from Red Clover Silage versus Soybean Meal: Ruminal Degradation and Intestinal Digestibility of Amino Acids.

The purpose was to assess the effect of exchanging crude protein (CP) of soybean meal (SBM) with red clover silage (RCS) in total mixed rations (TMR) on ruminal degradation and intestinal digestibility (ID) of essential amino acids (EAA). Four TMR and their individual feed components were studied. The TMR were composed of forage and concentrates (75:25), with proportions of RCS in TMR of 0.15, 0.30, 0.45, and 0.60 on a dry matter basis, resulting in diet groups RCS15, RCS30, RCS45, and RCS60, respectively. The ruminal degradation of EAA was determined using the nylon bag technique. For this, samples of TMR and their individual feed components were ruminally incubated for 16 h. The feed residues of TMR obtained after 16 h of incubation were used for the determination of ID of EAA using the mobile-bag technique. Increasing RCS and reducing SBM proportions linearly increased (p < 0.01) the in situ ruminal degradation of individual EAA from 75.5% to 83.5%. The degradation of EAA followed the trend of CP degradation among TMR, except for Cys that was lower (p < 0.05) than that of CP in RCS60 (79.7% vs. 86.3%). The degradation of EAA in individual feed ingredients not always corresponded to the degradation of CP and was feed dependent. Increasing the proportions of RCS in the TMR linearly reduced (p < 0.001) the ID of EAA (except for Ile) from 78.2% to 67.3%. However, the ID of EAA did not always reflect the ID of CP, and in general, the differences between the ID of CP and EAA increased as RCS increased in the TMR. The ID values of most of the EAA were similar (p > 0.05) to ID of CP in RCS15 and RCS30, while they mostly differed (p < 0.05) in RCS45 and RCS60, and being higher for EAA than CP (except for Cys that was lower than CP, p < 0.05). Similar trends were observed for intestinal absorbable AA, resulting in higher values (p < 0.05) of intestinal absorbable for all EAA than of CP in diet RCS60. In conclusion, increasing levels of RCS in TMR reduced the extent of EAA flow into the small intestine, the ID of EAA, and consequently the intestinal absorbable EAA. Therefore, accurate determination of metabolizable AA must be considered for optimal diet formulation when including high proportions of RCS in diets of high-producing dairy cows.

Assessing the Potential of Diverse Forage Mixtures to Reduce Enteric Methane Emissions In Vitro.

Methane emissions from ruminants are a major contributor to agricultural greenhouse gas emissions. Thus, eight different forage species were combined in binary mixtures with Lolium perenne in increasing proportions, in vitro, to determine their methane reduction potential in ruminants. Species were sampled in two consecutive years where possible. The aims were: a) to determine if mixtures with specific forages, particularly those rich in plant specialized metabolites (PSM), can reduce methane emissions compared to ryegrass monocultures, b) to identify whether there is a linear-dose effect relationship in methane emissions from the legume or herb addition, and c) whether these effects are maintained across sampling years. Results showed that all dicot species studied, including the non-tannin-containing species, reduced methane production. The tannin-rich species, Sanguisorba minor and Lotus pedunculatus, showed the greatest methane reduction potential of up to 33%. Due to concomitant reductions in the forage digestibility, Cichorium intybus yielded the lowest methane emissions per digestible forage unit. Contrary to total gas production, methane production was less predictable, with a tendency for the lowest methane production being obtained with a 67.5% share of the legume or herb partner species. Thus, linear increments in the partner species share did not result in linear changes in methane concentration. The methane reduction potential differed across sampling years, but the species ranking in methane concentration was stable.

Influence of quercetin on the global DNA methylation pattern in pigs.

The plant flavonol quercetin causes multiple health-promoting effects in human and animals. In this study, we investigated the possible effects of quercetin on global DNA methylation in growing pigs after 7 weeks of feeding a quercetin-enriched diet. The results indicated that a trend for an improved feed conversion was observed in the quercetin fed group compared to the control group. Furthermore, quercetin influenced global DNA methylation by upregulating DNA methyltransferase 1 (DNMT1) in both mRNA and protein expressions in a tissue specific manner. The observed bioactivity of quercetin against the global methylation marker in pigs indicates that quercetin could be a potential inducer of DNA methylation which might be of economic significance for effective nutritional interventions affecting the health and productivity of animals.

Contribution of different rumen microbial groups to gas, short-chain fatty acid and ammonium production from different diets-an approach in an in vitro fermentation system.

In this study, the relative contribution of different microbial groups to ruminal metabolism was investigated for different diets. The rumen microbial cultures included whole rumen fluid, fungi + protozoa, bacteria + protozoa, protozoa and bacteria + fungi and were established by physical and chemical methods. Gas production, short-chain fatty acid (SCFA) and ammonium production were measured at 24 hr in in vitro incubations using the Hohenheim gas test (HGT) procedure. Seven donor animal diets with different concentrate-to-roughage ratios (C:R: 10:90, 30:70, 50:50, 70:30, 70:30BC (BC = NaHCO3 ), 90:10 and 90:10BC) and five HGT diets (C:R: 10:90, 30:70, 50:50, 70:30 and 90:10) were formulated. Incubations in the HGT were always based on inoculum from sheep diets with the respective C:R ratio. Gas and ammonium production increased (p < 0.001) as a result of a gradual increase in concentrate proportion of the diets. In general, SCFA production followed the same trend. Whole rumen fluid and bacteria + fungi produced approximately 50% higher gas volume than protozoa and fungi + protozoa fractions, whereas gas production with bacteria + protozoa was at an intermediate level. Coculture of protozoa either with bacteria or with fungi produced more ammonium. Populations without bacteria were characterized by a particularly high acetate/propionate ratio. Although an interaction between microbial group and diet was observed for several variables, no clear direction could be established. Manipulating rumen fluid by selectively suppressing specific rumen microbial groups may be a helpful tool in elucidating their role in nutrient degradation and turnover in vitro.

Bioavailability of Quercetin from Onion Extracts after Intraruminal Application in Cows.

The aim of the present study was to investigate the bioavailability of quercetin from onion bulb (OB) and onion skin (OS) extracts in ruminants. Three non-lactating cows equipped with a permanent rumen fistula intraruminally received equimolar amounts of quercetin as either aglycone, rutin, or OB or OS extract, respectively, at a dose of 50 mg of quercetin equivalents/kg of body weight. Blood samples were drawn before and frequently within the 24 h period after application of the respective substance. Quercetin and quercetin metabolites with an intact flavonol structure (kaempferol, isorhamnetin, and tamarixetin) were analyzed in plasma samples by high-performance liquid chromatography with fluorescence detection. All quercetin sources administered resulted in a fast increase of the plasma concentrations of quercetin and total flavonols (sum of quercetin and its metabolites), followed by a rapid decline, whereby significant higher concentrations occurred with OB extract and rutin compared to quercetin aglycone and OS extract, respectively. The results clearly demonstrate a higher systemic availability of quercetin from OB extract and rutin. Taken together, OB extract with a high content of quercetin glucosides is an interesting source for the application of quercetin to ruminants.

Postruminal digestion of starch infused into the abomasum of heifers with or without exogenous amylase administration.

The effect of an exogenous amylase on postruminal digestion of starch infused into the abomasum of cattle was studied. Four rumen-cannulated heifers were fed 5.5 kg DM/d of a diet without starch, and assigned randomly to a crossover design. The experiment consisted of 2 periods lasting 23 d each with 10 d for adaptation to the diet followed by 13 d of abomasal infusion and sample collection. During the first 3 d of each infusion phase, isotonic saline solution was infused (1 liter/h) for measurement of baseline values in feces, followed by daily infusions of 880 g DM corn starch (1 kg/10 liters of water) without or with the addition of 2% of amylase. Titanium dioxide (10 g/d) was ruminally administered for estimation of fecal excretion. Digestion of starch in small intestine was calculated as the difference between the amounts of infused starch, disappeared from hindgut and fecal excretion. The apparent disappearance of starch from the hindgut was estimated based on the increment of microbial nitrogen (N) excretion due to starch infusion (1 g microbial N/100 g fermented starch) compared to baseline values. The concentration of purine bases in feces was used to estimate excretion of microbial N. Microbial N excretion increased with starch infusion (P < 0.05) but was not influenced by amylase (P = 0.81). Starch disappearance from the small intestine was not improved by amylase (P = 0.78) and averaged 85%. Amylase affected neither blood concentration of glucose (P = 0.80) nor of insulin (P = 0.26), but glucagon was lower without (P < 0.0001) than with amylase. The infusion of starch increased fecal excretion of total VFA (acetate, propionate, and butyrate) by 53% (P < 0.05), which indicates increased carbohydrate fermentation in the hindgut and incomplete digestion of starch in the small intestine. However, the excretion of total VFA was not affected by amylase (P = 0.66). Lactate excretion was higher at the second day of starch infusion (P < 0.05) without than with amylase, which suggests lower flow of starch from the small intestine to the hindgut due to a possible effect of amylase addition in animals not adapted to starch digestion. However, lactate excretion returned near to baseline values within 2 d, which was probably due to increase of lactate-utilizing bacteria and the adaptation of the microbial population in the hindgut. Further studies with higher starch levels and addition of amylase are recommended.

Systemic Absorption of Catechins after Intraruminal or Intraduodenal Application of a Green Tea Extract in Cows.

Green tea catechins have various potential health benefits in humans including anti-inflammatory, anti-oxidative and hepato-protective effects. If present in the circulation, they might have similar effects in ruminants, which are exposed to oxidative stress and fatty liver disease such as dairy cows during the periparturient phase. However, the bioavailability of a substance is a prerequisite for any post absorptive effect in vivo. This study aimed to investigate the appearance of catechins from a green tea extract (GTE) in cattle plasma after intraruminal and intraduodenal administration because absorption is of major importance regarding the bioavailability of catechins. The studies were performed in 5 rumen-fistulated non-lactating heifers and 6 duodenally fistulated lactating dairy cows, respectively, equipped with indwelling catheters placed in a jugular vein. The GTE was applied intraruminally (10 and 50 mg/kg BW, heifers) or duodenally (10, 20 and 30 mg/kg BW, dairy cows) in a cross-over design with a 2 d washout period between different dosages. Blood samples were drawn following the GTE administration at various pre-defined time intervals. The concentration of the major GTE catechins (gallocatechin, epigallocatechin, catechin, epicatechin, epigallocatechin-gallate, epicatechin-gallate) in plasma samples were analysed by HPLC with electrochemical detection. Irrespective of the dose, almost none of the catechins originally contained in the GTE were detected in plasma samples after intraruminal application. In contrast, intraduodenal administration of GTE resulted in increased plasma concentrations of epicatechin, epigallocatechin, epigallocatechin gallate in a dose-dependent manner. Thus, we can conclude that intraruminally or orally administered catechins are intensively metabolized by ruminal microorganisms.

Increase in West Nile virus infections imported to Germany in 2012.

Following the first confirmed imported West Nile virus (WNV) infection in 2011, the number of imported WNV infections to Germany increased in 2012. Two cases of West Nile neuroinvasive disease (WNND) and two cases of West Nile fever (WNF) were reported. The WNND cases were imported from Montenegro and Greece, including the first fatal case for Germany. The WNF cases were imported from Tunisia and Egypt. The cases were unambiguously confirmed according to laboratory criteria of European Centre for Disease Prevention and Control (ECDC). In this report we summarize the clinical and laboratory findings in order to sensitize physicians in Germany for this imported viral disease.