Effect of exogenous glucoamylase on ruminal in situ and in vitro dry matter and starch degradability of cereal grains in beef cattle.

The objective of this study was to evaluate the effect of the proportion of pea (0%, 10%, 20%, and 30%), as a partial replacement for soybean meal (SBM) in the fattening concentrate, on ruminal fermentation in lambs. Gas and methane (CH4 ) production, in vitro dry matter degradability (IVDMD), ammonia (NH3 -N), and volatile fatty acid (VFA) production after 24 h of incubation were evaluated. The concentrates were also incubated in the rumen of the wethers for 2, 4, 6, 8, 12, and 24 h to evaluate the effects of pea inclusion on in situ dry matter degradability (DMD), organic matter degradability (OMD), nitrogen degradability (ND), NH3 -N, and VFA production. In the in vitro assay, the inclusion of pea only affected gas production (mL day-1 degraded dry matter), CH4 production (mL day-1 degraded dry matter), and IVDMD (P < 0.05), and tended to affect NH3 -N content (P < 0.10) without affecting VFA production. In the in situ assay, the inclusion of pea increased DMD, OMD, and ND linearly (P < 0.001), whereas pea inclusion decreased NH3 -N content linearly (P < 0.05). Neither total VFA production nor the proportion of acetic acid were affected by pea inclusion (P > 0.05), but the propionic proportion increased with the proportion of pea included. The best level of pea inclusion in the concentrate could not be established based on the results of this study. However, the results showed that the inclusion of pea provides a good alternative protein source. © 2020 Society of Chemical Industry.

Effects of nitrate sources on in vitro methane production and ruminal fermentation parameters in diets differing in starch degradability

Cereal grains are the main source of energy in finishing feedlot diets. Sorghum is a more water-efficient alternative to corn, but sorghum starch is less ruminally digestible. Processing methods can increase grain digestibility, improving animal efficiency. The objective was to determine the effects of grain type and processing methods on in-situ dry matter (DM) degradability (DMD). Five ruminally cannulated Angus crossbred steers (body weight [BW] = 730 ± 119 kg) were pen-fed a starter ration twice daily at 2% of BW during the experiment. Fresh steam-flaked corn (SFC) and steam-flaked sorghum (SFS) samples were collected, air equilibrated for 240 minutes, then densities were measured (410 g/L SFC and 494 g/L SFS). Fresh cracked corn (CC) samples were collected after the roller mill process (4 to 6 pieces per kernel). High-moisture rolled sorghum (HMRS) was harvested at physiological maturity with 28% humidity, processed through a roller mill bagger and stored for 300 d. After opening, samples of HMRS were collected from 3 different areas of the silo. Nylon bags (10 × 20 cm, 50-µm pores) were filled with 20.5g (± 0.5g) of each grain and incubated in duplicate in the ventral sac of the rumen of each steer for 0, 3, 5, 12, 24, 48, 72 and 96h. The 0h bags were incubated for 10 seconds and then retrieved. Bags were immersed in cold water, rinsed 5 times using a washing machine, and dried to evaluate in-situ DMD. The kinetics of rumen degradability including soluble fraction [a], potentially degradable fraction [b], and degradation rate of the b fraction (c; %/hour) were estimated by a first-order asymptotic model using the NLIM procedure of SAS. The effective degradability (isRD) was determined considering a 6%/hour ruminal passage rate. Data was analyzed using the MIXED procedure of SAS, using steer as experimental unit and the random effect of steer (treatment). Differences were set at P ≤ 0.05 using Tukey test. Fraction a was greater for SFC than SFS, that was greater than HMDRS, that was greater than CC (P ≤ 0.01). Fraction b was greater for CC compared to HMDRS (P &amp;lt; 0.001), with no differences between HMDRS and SFS (P = 0.12), and both HMRS and SFS greater than SFC (P ≤ 0.001). Fraction c was greater for SFC compared to CC (P &amp;lt; 0.001), which was greater than SFS (P &amp;lt; 0.001). Fraction c of SFS and HMDRS were similar (P = 0.23). The isRD of SFC was greater than SFS (P &amp;lt; 0.001). No differences in isRD between SFS and CC were detected (P = 0.96), but isRD of SFS and CC were greater than HMDRS (P &amp;lt; 0.001). Steam flaking appears to improve DMD rates relative to other processing methods.

The objective of this assay was to investigate the effect of adding sunflower oil, Nannochloropsis oculata microalgae and their mixture at 0, 1, 2, 3, 4, and 5% to three total mixed rations (TMRs) with different concentrate:forage ratios (40C:60F, 50C:50F, and 60C:40F) on in vitro gas production (GP), methane (CH4) production, and nutrient degradability. Asymptotic GP, GP rate, CH4 concentration/g acid detergent fiber (ADF), dry matter (DM) degradability (DMD), short chain fatty acids (SCFAs), and ruminal bacteria population increased, but neutral detergent fiber (NDF) degradability (NDFD), ADF degradability (ADFD), and protozoa count decreased with increasing concentrate level in the TMR. Methane production/g DM and NDF was higher for 50C:50F TMR. Sunflower oil reduced asymptotic GP, lag time, CH4 production/g ADF, ammonia-N (NH3-N), and SCFA. Compared to the control treatments, additives decreased GP rate, while sunflower oil/N. oculata mixture increased DMD and NDFD. All additives at 5% increased GP rate and lag time and decreased CH4 production/g DM, ADF, and NDF, ruminal NH3-N, and protozoa count. All additives at 2% increased DMD, NDFD and ADFD, SCFA, and bacteria population. Supplementation of TMR, containing different concentrate:forage ratios, with sunflower oil, N. oculata, and sunflower oil/N. oculata mixture at different doses modified in vitro GP, CH4 production, and nutrient degradability.

In Australia, enteric methane production accounts for 68% of national greenhouse gas (GHG) emissions. Methane production (MP) is affected by diet quality. Approximately 50% of Australian beef cattle are grazed in the northern rangelands (Queensland, the Northern Territory, and part of Western Australia). In this region, cattle consume predominantly native C4 grasses, which are susceptible to substantial seasonal changes in nutritional value. The MP from Australian C4 grasses and associated changes in methane and methanogenic archaeal populations within the rumen are not well described. This thesis aimed to quantify methane from C4 tropical pastures and associated rumen microbiota during seasonal changes in forage quality. Four experiments were conducted to achieve these objectives. Experiment 1 (Chapter 3) measured cumulative gas production in vitro by Ankom Gas Production System batch culture method. Bottles containing ground forages and buffered rumen fluid were incubated for up to 48 h and pH, volatile fatty acids (VFAs); total gas production (TGP) methane (CH4), dry matter degradability (DMd) and organic matter degradability (OMd) were measured. Substrates were a variety of tropical C4 grasses (obtained in Experiment 4) ranging in CP from 29 g/ kg DM to 120 g/ kg DM and digestibility from 38 - 60%. A legume was included for comparison. A general linear model determined effect of forage type and time on variables. Regression relationships were determined between nutritive characteristics and gas variables. More gas, methane and total VFA (mmol/L) were produced when forages were more degradable. Positive linear relationships were observed between CP and both TGP and methane. Negative relationships were observed between fibre and methane. Experiment 2, (Chapter 4) determined contribution of inoculum to fermentative characteristics when donor cattle were fed forage diets representative of wet or dry season in northern Australia. The experimental design was the same as for experiment 1. The substrates included the Mitchell grass, and the Lucerne from experiment 1 and two other forages a pasture (PAS) Urochloa mosambicensis (CP 90 g/ kg DM’ DMD 63%) and a low quality hay (LQH) sample of Chloris gayana (CP 31 g/kg DM; DMD 41%). Inoculum was collected from cattle consuming PAS (PASi) and LQH (LQHi) diets. A general linear model was used with forage type, time and inoculum source as fixed effects. Substrates incubated in PASi had greater MP and lower degradability than the same substrates incubated in LQHi. The in vitro experiments confirm that forage quality is the primary factor effecting gas production but under closed batch system conditions, specific effects of inoculum are observed. Two in vivo experiments (Chapter 5 – 8) were conducted to quantify MP and microbial communities associated with seasonal change in forage quality. In Experiment 3, Bos indicus steers were fed a low quality C. gayana hay and then switched to either moderate quality U. mosambicensis pasture (PAS), a high quality C. gayana hay (HQH) or remained on low quality hay (LQH). Individual measurements of DMI and rumen outflow rates were taken and rumen fluid was sampled for analysis of pH, VFAs, NH3-N and microbial populations (using comparison of the 16S rRNA gene). Open circuit respiration chambers measured daily MP. Higher DMI, MP, and rumen outflow rates accompanied higher forage quality in PAS and HQH compared with LQH steers, although all produced approximately 19.8 g methane /kg DMI. Low quality hay produced more methane /kg digestible DMI and none of the diets reached predicted daily emissions compared with the equations in use for the national greenhouse gas inventory. Analysis of the prokaryotic community revealed predominance of the genus Prevotella and an unassigned genus in the family Ruminococcaceae in bacteria, and of Methanobrevibacter and Thermoplasmatales in the archaea. Finally, Experiment 4, a longitudinal design was conducted at Brunette Downs Station (Barkly Tableland, Northern Territory) in which rumen fluid was collected from 10 heifers grazing Mitchell grass in two wet seasons (May 2012, March 2013) and the intervening wet season (August, November 2012). Regardless of sampling month during the year, dominant genera of bacteria and methanogenic archaea were similar, although differentiation in relative abundance between months was observed. Prevotella and Clostridial genera were predominant in the bacteria and Methanobrevibacter and Thermoplasmatales associated lineages in the methanogens. Methylotrophic methanogens appeared to comprise a greater proportion of the population in both in vivo experiments when forage quality was improved. However, one species identifying with Thermoplasmatales was negatively affected by increasing rumen outflow rates. It can be concluded that methane emissions from cattle consuming tropical grasses was lower than predicted. The experimental analysis confirmed Methanobrevibacter to be the dominant methanogen in northern beef cattle, however, Thermoplasmatales affiliated methanogens make up a significant proportion of the methanogenic population in cattle consuming tropical C4 grasses in Australia. Culturing a representative of this clade would greatly improve understanding of methanogenesis in Australian tropically adapted beef cattle.

In vitro experiments were conducted to evaluate the suitability of several mixtures of high tanniniferous non legumes with low tanniniferous legumes on in vitro gas production (IVGP), dry matter degradation, Ammonia-N, methane production and microbial population. Eight treatments were examined in a randomized complete block design using four non-legumes and two legumes (Carallia integerrima×Leucaena leucocephala (LL) (Trt 1), C. integerrima×Gliricidia sepium (GS) (Trt 2), Aporosa lindeliyana×LL (Trt 3), A. lindeliyana×GS (Trt 4), Ceiba perntandra×LL (Trt 5), C. perntandra×GS (Trt 6), Artocarpus heterophyllus×LL (Trt 7), A. heterophyllus×GS (Trt 8). The condensed tannin (CT) content of non legumes ranged from 6.2% (Carallia integerrima) to 4.9% (Ceiba perntandra) while the CT of legumes were 1.58% (Leucaena leucocephala) and 0.78% (Gliricidia sepium). Forage mixtures contained more than 14% of crude protein (CP) while the CT content ranged from 2.8% to 4.0% respectively. Differences (p<0.05) were observed in in vitro gas production (IGVP) within treatments over a 48 h period dominated by C. perntandra×G. sepium (Trt 6). The net gas production (p<0.05) was also high with Trt6 followed by A. heterophyllus×L. leucocephala (Trt 7) and A. heterophyllus×G. sepium (Trt 8). Highest (p>0.05) NH3-N (ml/200 mg DM) production was observed with the A. heterophyllus×G. sepium (Trt 8) mixture which may be attributed with it’s highest CP content. The correlation between IVGP and CT was 0.675 while IVGP and CP was 0.610. In vitro dry matter degradation (IVDMD) was highest in Trt 8 as well. Methane production ranged from 2.57 to 4.79 (ml/200 mg DM) to be synonimous with IVGP. A higher bacteria population (p<0.05) was found in C. perntandra×G. sepium (Trt 6) followed by Artocarpus heterophyllus+G. sepium (Trt 8) and the same trend was observed with the protozoa population as well. The results show that supplementing high tannin non leguminous forages by incremental substitution of legume forage increased gas production parameters, NH3-N, IVDMD and microbial population in the fermentation liquid. Methane production was not significantly affected by the presence of CT or different levels of CP in forage mixtures. Among non legumes, Ceiba perntandra and Artocarpus heterophyllus performed better in mixture with L. leucocephala and G. sepium.

Influence of single-gene mutations, harvest maturity and sample processing on ruminal in situ and post-ruminal in vitro dry matter and starch degradability of corn grain by ruminants

Relationships between chemical composition, dry matter degradation and in vitro gas production of several ruminant feeds

In recent decades supplementation of animal feeds with exogenous fibrolytic enzymes has substantially improved digestibility and animal performance. However, information related to associated methane production is limited and inconsistent. This study evaluated the effect of cellulase and xylanase enzymes on in vitro methane production of Eragrostis curvula hay, maize (Zea mays) stover and a total mixed ration (TMR) at seven levels of the two enzymes. Feed samples were incubated for 2, 12, 24 and 48 h in an in vitro batch culture with buffer and rumen fluid, and fibrolytic enzymes. Gas production was measured using a pressure transducer connected to a data tracker, while methane gas was analysed using a gas chromatograph which was calibrated with standard CH4 and CO2. Increases in the level of enzyme application resulted in increases in gas volume, total volatile fatty acid (VFA) production, dry matter (DM) disappearance and associated increases in methane production. The linear increase in percentage and volume of methane production in tandem with increases in level of enzyme application might be due to increased fermentation, and organic matter degradability that resulted in a shift in VFA production towards acetate. Considering the efficiency of DM and neutral detergent fiber degradation and production of associated VFA with levels of enzymes, the use of 1 mg g−1 DM of enzyme can be a good option for the feeds tested. However, they cannot decrease methane production. It will be very important to consider other hydrogen sinks that can capture directly extra H+ produced by the addition of enzyme so that their supplementation could be very efficient and environmentally sound.

The present study was conducted to evaluate the effect of nitrate supplementation on dry-matter (DM) degradation and ruminal fermentation parameters by using in vitro gas production and in situ technique. In vitro gas production and in situ DM degradation in the presence or absence of nitrate were recorded at all incubation times. At all incubation times, diets incubated with nitrate gave a significantly lower gas production than did the other diets, except at 2-h incubation. Ruminal DM degradation did not differ among the experimental treatments. Furthermore, at most incubation times, total volatile fatty acids in diets containing nitrate were lower than those in the other treatments. Nitrate supplementation considerably increased gas production from the insoluble fraction, whereas it decreased gas production from the quickly soluble fraction, and potential gas production. Moreover, in all incubations, there were significant correlations between gas production and in situ DM-degradation parameters. The control diet had the greatest retained nitrogen content, but the diets containing nitrate had the greatest faecal nitrogen. The results showed that nitrate addition resulted in a lower gas production and volatile fatty acid production in in vitro assay. It was concluded that considering the strong posetive relationship between the two methodologies, the degradability parameters can be predicted from obtained gas production.

Methane is produced by ruminants as the result of microbial digestion, it represents an energy loss to the animal, and it is also a potent greenhouse gas. Mechanistic modelling can lend insight into dietary strategies aimed at reducing methane emissions from cattle, but require proper representation of aspects of underlying rumen fermentation and digestion. Proper prediction of the production of volatile fatty acids (VFA) is central to accurate methane prediction. This study evaluated the newly updated and expanded model of VFA dynamics developed by Bannink et al. (2008), in comparison to a previous model version (Bannink et al., 2006), within a rumen model (Dijkstra et al., 1992; modified by Mills et al., 2001) for its methane prediction ability in beef cattle fed high-grain diets. In an evaluation of the rumen model performed by Kebreab et al. (2008) using the Bannink et al. (2006) VFA stoichiometry, the model performed well on dairy cow data, but poorly on beef cattle data in predicting methane emission. Several modifications were therefore made to the model to adapt it for typical high-grain beef cattle diets and then evaluated for its accuracy to predict methane emissions from feedlot cattle. Passage rate of protozoa was increased proportionally with the grain percent of the diet, and the protozoal proportion of the amylolytic microbial pool was reduced accordingly. This allowed a small cellulolytic microbial pool to remain in the rumen, where it previously went extinct. Also, stoichiometry of rumen VFA production was adjusted for the use of monensin in the observed data. Preliminary results showed that while the representation of some central aspects of rumen fermentation probably improved, the model over-predicted methane production with a root mean square prediction error value of 2.86 MJ/d, with 56% of that error coming from bias and 44% from random sources. Concordance correlation coefficient value was 0.194. Results indicate that other areas of the model require improvement for predicting methane production accurately in high grain feedlot diets, such as hind gut fibre digestibility.

Comparison of two in vitro fermentation gas production methods using both rumen fluid and faecal inoculum from sheep

The effect of Saccharomyces cerevisiae as live cells (LC) or cells extract (CE) on in vitro gas production (GP) kinetics and ruminal fermentation parameters of a total mixed ration (TMR) consisting of commercial concentrate and alfalfa hay [1:1 dry matter (DM)] as a substrate was studied. The TMR was incubated with CE at 1, 2 and 4 mg/g or LC at 0.3, 0.6 and 0.9 mg/g DM for 96 h. Rumen GP was recorded after 6, 12, 19, 24, 48, 72 and 96 h of incubation. Interaction effects were observed (P<0.01) between treatment type and yeast dose for the asymptotic GP and methane (CH4) production. Incubation of yeast CE improved (P<0.01) the asymptotic GP compared to control and LC with greater effects (P<0.01) for the low and the intermediate doses. Yeast CE treatment was more effective (P<0.01) in GP than both of LC and control treatments with greater effect (P<0.01) for the low and the intermediate doses. Treatment type and yeast dose affected (P<0.01) CH4 production, metabolisable energy (ME), and short chain fatty acids (SCFA) without affecting in vitro DM degradability (IVDMD). Higher values (P<0.01) of CH4, ME, SCFA and IVDMD were observed for the yeast CE treatment. It could be concluded that adding yeast S. cerevisiae (CE and LC extract) improved GP and ruminal fermentation parameters, where CE at 0.3 and 0.6 mg/g DM was more effective than the yeast LC.

PurposeThe present experiment aimed to evaluate date palm leaves (DPL) treated without or with fibrolytic enzymes as a feed for ruminants.MethodsThe experiment employed an in vitro wireless gas production system to evaluate the dietary inclusion of DPL as sun-dried, DPL ensiled without or with fibrolytic enzymes for 45 days. The different forms of DPL replaced berseem hay (300 g/kg diet) at 0, 25, 50, 75 and 100% in the diet.ResultsDried DPL linearly decreased the asymptotic total gas production (GP), rate of methane (CH4) and carbon dioxide (CO2) production, and acid detergent fiber degradability, and increased the lag of total GP (P < 0.05). The ensiled DPL also linearly decreased (P < 0.05) the asymptotic total GP, asymptotic CH4, asymptotic CO2 production and the rate of CH4 and CO2 productions, but dry matter degradability and total volatile fatty acid (VFA) concentrations were unaffected. Date palm leaves treated with fibrolytic enzymes linearly decreased the asymptotic total GP, CH4 and CO2 productions, and the rate of CH4 and CO2 production. Ensiling of DPL with fibrolytic enzymes increased (P < 0.05) dry matter and fiber degradability and the concentrations of ruminal ammonia-N and total VFA.ConclusionIt is concluded that DPL treated with fibrolytic enzymes can replace berseem hay up to 100% in the diet to reduce CH4 production from ruminants. Ensiling with fibrolytic enzymes is recommended as a sustainable strategy to reduce environmental pollution and utilization of DPL.Graphical

The objective of this study was to evaluate the sorghum silage with the addition of guandu using in vitro semi automated gas production technique. Total gas production, methane production, the dry matter degradability (DMD) and organic matter (OMD) were estimated. PEG was used to measure the possible effect of tannins of sorghum silage on methane and gas production. The results indicated that the presence of PEG in the sorghum silage in the level of 25% and 50% inclusion of guandu were the highest (P &lt; 0.05) in the gas and methane production, respectively compared with the other levels. For the gas and methane production increment, the results varied, the highest (P &lt; 0.01) increment of gas production and methane production were found with the levels of 25% 50% inclusion of guandu, respectively. DMD and OMD increased with the increasing of pigeon pea levels with sorghum silage, these variables were greater (P&lt; 0.01) wfhen PEG was added compared to without PEG. This evaluation of the silages showed that the presence of tannin in sorghum silage can interfere in the rumen fermentation and methane production in vitro especially identified when the substrates were incubated in the presence of PEG.