Characterization of anthocyanidin-accumulating Lc-alfalfa for ruminants: nutritional profiles, digestibility, availability and molecular structures, and bloat characteristics

Grazing cattle on alfalfa (Medicago sativa L.) would be economically beneficial, but its rapid initial rate of protein degradation results in pasture bloat, low efficiency of protein utilization and excessive N pollution into the environment. Introducing a gene that stimulates the accumulation of mono/polymeric anthocyanidins might reduce the ruminal protein degradation rate and reduce bloat related foam stability. The overall objective of this thesis was to evaluate newly developed anthocyanidin-accumulating Lc-alfalfa progeny for nutritional properties (composition, site of degradation and molecular structure), environmental emissions and bloat characteristics. The objective of the first study was to determine survival and phytochemical and chemical profiles of Lc-alfalfa progeny (BeavLc1, RambLc3 and RangLc4) and their non-transgenic (NT) parental cultivars (Beaver, Rambler and Rangelander). Lc-alfalfa forage accumulated enhanced amounts of anthocyanidin, with an average concentration of 197.4 µg/g DM, while proanthocyanidin (i.e. condensed tannins) were not detected. Both of these metabolites were absent in the NT-parental varieties. Lc-alfalfa progeny had ~3 % less crude protein (CP) and ~3 % more carbohydrates (CHO), which resulted in their 11 g/kg lower N:CHO ratio compared with NT-alfalfa. Total rumen-degradable N:CHO ratio based on chemical analysis was 12.9 g/kg lower in Lc-alfalfa compared with NT-alfalfa. The objective of the second study was to evaluate in vitro degradation, fermentation and microbial-N partitioning of three forage color phenotypes [green, light purple-green (LPG) and purple-green (PG)] within Lc-progeny and their parental green NT-alfalfa varieties. Purple-green-Lc alfalfa accumulated more anthocyanidin than Green-Lc with LPG-Lc intermediate. Gas, methane and ammonia accumulation rates were slower for the two purple-Lc phenotypes compared with NT-alfalfa with Green-Lc intermediate. Effective degradable DM and N were lower in the three Lc-phenotypes compared with NT-alfalfa. Anthocyanidin concentration correlated negatively with gas and methane production rates and effective degradability of DM and N. The objectives of the third study were to evaluate in situ ruminal degradation characteristics and synchronization ratios, and to model protein availability to dairy cattle and net energy for lactation of three Lc-alfalfa progenies, BeavLc1, RambLc3 and RangLc4 and the cultivar AC Grazeland (selected for a low initial rate of ruminal degradation). Anthocyanidin accumulation was on average 163.4 ìg/g DM in the three Lc-progeny while AC Grazeland did not accumulate anthocyanidin. The basic chemical composition of the original samples, soluble and potentially degradable fractions and degradation characteristics of crude protein and carbohydrates were similar in Lc-alfalfa and AC Grazeland. The undegradable in situ crude protein and neutral detergent fiber fraction were, respectively, 1.3 %CP and 4.8 %CHO lower in the three Lc-progeny compared with AC Grazeland. Lc-alfalfa had a 0.34 MJ/kg DM higher net energy for lactation and tended to have a 11.9, 6.9 and 8.4 g/kg DM higher rumen degradable protein, rumen degraded protein balance and intestinal available protein, respectively, compared with AC Grazeland,. The hourly rumen degraded protein balance included an initial and substantial peak (over-supply) of protein relative to energy which was highest in RangLc4 and lowest in RambLc3. The hourly rumen degraded protein balance between 4 and 24 h was similar and more balanced for all four alfalfa populations. The objective of the fourth study was to determine foam formation and stability in vitro from aqueous leaf extracts of three Lc-alfalfa progeny (BeavLc1, RambLc3, RangLc4), parental NT-alfalfa and AC Grazeland (bloat reduced cultivar) harvested in the field at 07:00 or 18:00 h. Anthocyanidin accumulation averaged 247.5 ìg/g DM in the leaves of the three Lc-progeny. There was an interaction between population and harvest time for the foam parameters. Initial foam volume (0 min) and final foam volume (150 min) at 07:00 h were lower for AC Grazeland compared with all other treatments and lower for RangLc4 compared with the other two Lc-progeny at 0 min and NT-alfalfa at 150 min; while from the 18:00 h harvest, initial foam volume was larger for NT-alfalfa and final foam volume was larger for RambLc3 compared with AC Grazeland, BeavLc1 and RangLc4. Foam formation correlated positively (R = 0.30 to 0.44) with leaf DM content, leaf extract protein and ethanol-film content, spectroscopic vibration intensity due to all carbohydrates (CHOVI) and amide I:amide II ratio and negatively (R = -0.33 and -0.34;

Feeding di-ammonium phosphate as a phosphorous source in finishing lambs reduced excretion of phosphorus in feces without detrimental effects on animal performance

Objective Phosphorous (P) sources with greater bioavailability might increase animal production efficiency and decrease environmental pollution. The objective of current study was to determine animal performance, nutrient digestibility, blood metabolites and fecal P concentration in finishing lambs fed a diet with either di-calcium phosphate (DCP) or di-ammonium phosphate (DAP) as a P source. Methods Twelve 4-month-old male lambs (initial body weight 24.87±3.4 kg) were randomly allocated to a diet with either DCP or DAP (~261 g/kg of total diet P) fed ad libitum for 93 days. Diets were iso-nitrogenous and iso-energetic and had same calcium (Ca) and P concentrations. Results The DAP contained 19.7 g/kg of dry matter (DM) Ca, 185.4 g/kg DM P and 14,623 ppm fluorine, while DCP contained 230.3 g/kg DM Ca, 195.2 g/kg DM P and 1,039 ppm fluorine. The diet with DAP contained 60 ppm fluorine while the diet with DCP contained 13 ppm fluorine. Lambs fed the diet with DAP tended to have a greater daily DM intake compared to those fed diet with DCP (p = 0.09). Lambs fed DAP had greater plasma P concentration and alkaline phosphatase activity (p≤0.01) compared with lambs fed DCP. Dry matter and organic matter digestibility of the diets were similar between two treatments at days 60 and 90, while they were greater in lambs fed DCP (p<0.05) at day 30 of the trial. Feeding DAP increased P digestibility (58.7% vs 50.2%; p<0.05) and decreased fecal P concentration in lambs compared with feeding DCP (3.1 vs 3.8 g/kg DM; p<0.05). Conclusion Providing ~261 g/kg of total diet P as DAP in the diet of finishing lambs improved the bioavailability of P in the body and decreased excretion of P in feces without affecting lamb performance

Review: Markers and proxies to monitor ruminal function and feed efficiency in young ruminants

Developing the rumen’s capacity to utilise recalcitrant and low-value feed resources is important for ruminant production systems. Early-life nutrition and management practices have been shown to influence development of the rumen in young animals with long-term consequences on their performance. Therefore, there has been increasing interest to understand ruminal development and function in young ruminants to improve feed efficiency, health, welfare, and performance of both young and adult ruminants. However, due to the small size, rapid morphological changes and low initial microbial populations of the rumen, it is difficult to study ruminal function in young ruminants without major invasive approaches or slaughter studies. In this review, we discuss the usefulness of a range of proxies and markers to monitor ruminal function and nitrogen use efficiency (a major part of feed efficiency) in young ruminants. Breath sulphide and methane emissions showed the greatest potential as simple markers of a developing microbiota in young ruminants. However, there is only limited evidence for robust indicators of feed efficiency at this stage. The use of nitrogen isotopic discrimination based on plasma samples appeared to be the most promising proxy for feed efficiency in young ruminants. More research is needed to explore and refine potential proxies and markers to indicate ruminal function and feed efficiency in young ruminants, particularly for neonatal ruminants

Challenges and opportunities to capture dietary effects in on-farm greenhouse gas emissions models of ruminant systems

This paper reviews existing on-farm GHG accounting models for dairy cattle systems and their ability to capture the effect of dietary strategies in GHG abatement. The focus is on methane (CH4) emissions from enteric and manure (animal excreta) sources and nitrous oxide (N2O) emissions from animal excreta. We identified three generic modelling approaches, based on the degree to which models capture diet-related characteristics: from ‘none’ (Type 1) to ‘some’ by combining key diet parameters with emission factors (EF) (Type 2) to ‘many’ by using process-based modelling (Type 3). Most of the selected on-farm GHG models have adopted a Type 2 approach, but a few hybrid Type 2 / Type 3 approaches have been developed recently that combine empirical modelling (through the use of CH4 and/or N2O emission factors; EF) and process-based modelling (mostly through rumen and whole tract fermentation and digestion). Empirical models comprising key dietary inputs (i.e., dry matter intake and organic matter digestibility) can predict CH4 and N2O emissions with reasonable accuracy. However, the impact of GHG mitigation strategies often needs to be assessed in a more integrated way, and Type 1 and Type 2 models frequently lack the biological foundation to do this. Only Type 3 models represent underlying mechanisms such as ruminal and total-tract digestive processes and excreta composition that can capture dietary effects on GHG emissions in a more biological manner. Overall, the better a model can simulate rumen function, the greater the opportunity to include diet characteristics in addition to commonly used variables, and thus the greater the opportunity to capture dietary mitigation strategies. The value of capturing the effect of additional animal feed characteristics on the prediction of on-farm GHG emissions needs to be carefully balanced against gains in accuracy, the need for additional input and activity data, and the variability encountered on-farm

Measuring enteric methane emissions from individual ruminant animals in their natural environment

Ruminant livestock are an important source of meat, milk, fiber, and labor for humans. The process by which ruminants digest plant material through rumen fermentation into useful product results in the loss of energy in the form of methane gas from consumed organic matter. The animal removes the methane building up in its rumen by repeated eructations of gas through its mouth and nostrils. Ruminant livestock are a notable source of atmospheric methane, with an estimated 17% of global enteric methane emissions from livestock. Historically, enteric methane was seen as an inefficiency in production and wasted dietary energy. This is still the case, but now methane is seen more as a pollutant and potent greenhouse gas. The gold standard method for measuring methane production from individual animals is a respiration chamber, which is used for metabolic studies. This approach to quantifying individual animal emissions has been used in research for over 100 years; however, it is not suitable for monitoring large numbers of animals in their natural environment on commercial farms. In recent years, several more mobile monitoring systems discussed here have been developed for direct measurement of enteric methane emissions from individual animals. Several factors (diet composition, rumen microbial community, and their relationship with morphology and physiology of the host animal) drive enteric methane production in ruminant populations. A reliable method for monitoring individual animal emissions in large populations would allow (1) genetic selection for low emitters, (2) benchmarking of farms, and (3) more accurate national inventory accounting

The Role of Proanthocyanidins Complex in Structure and Nutrition Interaction in Alfalfa Forage

Alfalfa (Medicago sativa L.) is one of the main forages grown in the world. Alfalfa is a winter hardy, drought tolerant, N-fixing legume with a good longevity, high yield, high nutrient levels, high digestibility, unique structural to non-structural components ratio, high dry matter intake, and high animal productivity per hectare. However, its main limitation is its excessively rapid initial rate of protein degradation in the rumen, which results in pasture bloat and inefficient use of protein with consequent excessive excretions of nitrogen into the environment. Proanthocyanidins are secondary plant metabolites that can bind with protein and thereby reduce the rate and extent of ruminal protein degradation. However, these secondary metabolites do not accumulate in alfalfa. This review aims to firstly describe the events involved in the rapid release of protein from alfalfa and its effect on ruminant nutrition, environmental pollution, and pasture bloat; secondly, to describe occurrence, structure, functions and benefits of moderate amounts of proanthocyanidin; and finally, to describe the development of alfalfa which accumulates moderate amounts of proanthocyanidins. The emphasis of this review focuses on the role of proanthocyanidins compounds in structure and nutrition interaction in ruminant livestock systems

The Occurrence, Biosynthesis, and Molecular Structure of Proanthocyanidins and Their Effects on Legume Forage Protein Precipitation, Digestion and Absorption in the Ruminant Digestive Tract

Forages grown in temperate regions, such as alfalfa (Medicago sativa L.) and white clover (Trefolium repens L.), typically have a high nutritional value when fed to ruminants. Their high protein content and degradation rate result, however, in poor utilization of protein from the forage resulting in excessive excretion of nitrogen into the environment by the animal. Proanthocyanindins (also known as condensed tannins) found in some forage legumes such as birdsfoot trefoil (Lotus corniculatus L.), bind to dietary protein and can improve protein utilization in the animal. This review will focus on (1) the occurrence of proanthocyanidins; (2) biosynthesis and structure of proanthocyanidins; (3) effects of proanthocyanidins on protein metabolism; (4) protein precipitating capacity of proanthocyanidins and their effects on true intestinal protein adsorption by ruminants; and (5) effect on animal health, animal performance and environmental emissions

The Role of Proanthocyanidins Complex in Structure and Nutrition Interaction in Alfalfa Forage

Alfalfa (Medicago sativa L.) is one of the main forages grown in the world. Alfalfa is a winter hardy, drought tolerant, N-fixing legume with a good longevity, high yield, high nutrient levels, high digestibility, unique structural to non-structural components ratio, high dry matter intake, and high animal productivity per hectare. However, its main limitation is its excessively rapid initial rate of protein degradation in the rumen, which results in pasture bloat and inefficient use of protein with consequent excessive excretions of nitrogen into the environment. Proanthocyanidins are secondary plant metabolites that can bind with protein and thereby reduce the rate and extent of ruminal protein degradation. However, these secondary metabolites do not accumulate in alfalfa. This review aims to firstly describe the events involved in the rapid release of protein from alfalfa and its effect on ruminant nutrition, environmental pollution, and pasture bloat; secondly, to describe occurrence, structure, functions and benefits of moderate amounts of proanthocyanidin; and finally, to describe the development of alfalfa which accumulates moderate amounts of proanthocyanidins. The emphasis of this review focuses on the role of proanthocyanidins compounds in structure and nutrition interaction in ruminant livestock systems

Protein Structures among Bio-Ethanol Co-Products and Its Relationships with Ruminal and Intestinal Availability of Protein in Dairy Cattle

Abstract: The objectives of this study were to reveal molecular structures of protein among different types of the dried distillers grains with solubles (100 % wheat DDGS (WDDGS); DDGS blend1 (BDDGS1, corn to wheat ratio 30:70%); DDGS blend2 (BDDGS2, corn to wheat ratio 50:50 percent)) and different batches within DDGS type using diffuse reflectance infrared Fourier transform spectroscopy (DRIFT). Compared with BDDGS1 and BDDGS2, wheat DDGS had higher (p &lt; 0.05) peak area intensities of protein amide I and II and amide I to II intensity ratio. Increasing the corn to wheat ratio form 30:70 to 50:50 in the blend DDGS did not affect amide I and II area intensities and their ratio. Amide I to II peak intensity ratio differed (p &lt; 0.05) among the different batches within WDDGS and BDDGS1. Compared with both blend DDGS types, WDDGS had higher α-helix and β-sheet ratio (p &lt; 0.05), while α-helix to β-sheet ratio was similar among the three DDGS types. The α-helix to β-sheet ratio differed significantly among batches within WDDGS. Principal component analysis (PCA) revealed that protein molecular structures in WDDGS differed from those of BDDGS1 and between differentInt. J. Mol. Sci. 2013, 14 16803 batches within BDDGS1 and BDDGS2. The α-helix to β-sheet ratios of protein in al

Feeding lucerne silage to beef cattle at three allowances and four feeding frequencies affects circadian patterns of methane emissions, but not emissions per unit of intake

Abstract. The objective of this study was to determine the circadian variation in methane (CH 4 ) emissions from cattle fed lucerne silage at different feeding levels and feeding frequencies, to assist with interpretation of short &apos;snapshot&apos; CH 4 measurements used for predicting daily emissions. Eight Hereford · Friesian heifers (initially 20 months of age) were used in five consecutive periods (P1-5) of 14 days with CH 4 emissions measured using respiration chambers for two consecutive days at the end of each period. Feed was restricted to intakes of~6, 8, 8, 8 and 11 AE 1.3 (ad libitum) kg lucerne silage dry matter (DM), fed in 2, 2, 3, 4 or ad libitum (refilled twice daily) meals per day in P1-5, respectively. Daily CH 4 production (g/day) was lower in P1 than in P2-4 (P &lt; 0.05), which were lower than in P5 (P &lt; 0.05), but CH 4 yield (24.3 AE 1.23 g/kg DM) was unaffected by treatment. Among the five periods, CH 4 emission rate (g/h) before feeding ranged from 1.8 to 6.5 g/h, time to peak CH 4 production after start of feeding ranged from 19 to 40 min and peak CH 4 production rate ranged from 11.1 to 17.5 g/h. The range in hourly CH 4 emission rates during the day decreased with increasing feed intake level, but was unaffected by feeding frequency. In summary, the circadian pattern of CH 4 emissions was affected by feed allowance and feeding frequency, and variation in CH 4 emission rate was reduced with increasing intake, without affecting average daily yield (g CH 4 /kg DM intake)