Six plant sources of hydrolyzable tannins (HT) or HT and condensed tannins (CT; designated as HT1, HT2, HT3, HT + CT1, HT + CT2, and HT + CT3) were evaluated to determine their effects in vitro on CH(4) production and on ruminal archaeal and protozoa populations, and to assess potential differences in biological activities between sources containing HT only or HT and CT. Samples HT1, HT2, and HT3 contained only HT, whereas samples HT + CT1, HT + CT2, and HT + CT3 contained HT and CT. In experiment 1, in vitro incubations with samples containing HT or HT + CT resulted in a decrease in CH(4) production of 0.6 and 5.5%, respectively, compared with that produced by incubations containing the added tannin binder polyethylene glycol-6000. Tannin also suppressed the population of methanogenic archaea in all incubations except those with HT2, with an average decrease of 11.6% in HT incubations (15.8, 7.09, and 12.0 in HT1, HT2, and HT3) and 28.6% in incubations containing HT + CT (35.0, 40.1, and 10.8 in HT + CT1, HT + CT2, and HT + CT3) when compared with incubations containing added polyethylene glycol-6000. The mean decrease in protozoal counts was 12.3% in HT and 36.2% in HT + CT incubations. Tannins increased in vitro pH, reduced total VFA concentrations, increased propionate concentrations, and decreased concentrations of iso-acids. In experiment 2, when a basal diet was incubated with graded levels of HT + CT1, HT + CT2, and HT + CT3, the total gas and CH4 production and archaeal and protozoal populations decreased as the concentration of tannins increased. Our results confirm that tannins suppress methanogenesis by reducing methanogenic populations in the rumen either directly or by reducing the protozoal population, thereby reducing methanogens symbiotically associated with the protozoal population. In addition, tannin sources containing both HT and CT were more potent in suppressing methanogenesis than those containing only HT.

The effects of the anti-methanogenic compound, bromochloromethane (BCM), on rumen microbial fermentation and ecology were examined in vivo. Japanese goats were fed a diet of 50 % Timothy grass and 50 % concentrate and then sequentially adapted to low, mid and high doses of BCM. The goats were placed into the respiration chambers for analysis of rumen microbial function and methane and H 2 production. The levels of methane production were reduced by 5, 71 and 91 %, and H 2 production was estimated at 545, 2941 and 3496 mmol/head per d, in response to low, mid and high doses of BCM, respectively, with no effect on maintenance feed intake and digestibility. Real-time PCR quantification of microbial groups showed a significant decrease relative to controls in abundance of methanogens and rumen fungi, whereas there were increases in Prevotella spp. and Fibrobacter succinogenes, a decrease in Ruminococcus albus and R. flavefaciens was unchanged. The numbers of protozoa were also unaffected. Denaturing gradient gel electrophoresis and quantitative PCR analysis revealed that several Prevotella spp. were the bacteria that increased most in response to BCM treatment. It is concluded that the methane-inhibited rumen adapts to high hydrogen levels by shifting fermentation to propionate via Prevotella spp., but the majority of metabolic hydrogen is expelled as H 2 gas.Key words: Rumen: Methane: Hydrogen: Bromochloromethane: Goats Enteric fermentation in livestock accounts for 19 % of anthropogenic sources of methane, a potent greenhouse gas (1) , for which rumen fermentation is the largest source of methane production. In rumen fermentation, several pathways involving both hydrogen-producing and -consuming steps are involved in the conversion of feedstuffs into various fermentation end products such as SCFA (2,3) . Although metabolic hydrogen in the rumen is incorporated in fermentation end products by bacteria, methanogenic archaea (methanogens) consume the greater majority of metabolic hydrogen to obtain energy for their metabolism and finally release methane, which accounts for 2 -12 % loss of the metabolic energy from feed (1,3,4) . Therefore, management of metabolic hydrogen and methane production in the rumen is an important factor to be considered, when developing strategies to reduce greenhouse gas emissions and improve efficiency of energy utilisation from feed.It is known that many chemical agents such as ionophores (e.g. monensin), unsaturated fatty acids, sulphate, nitrate, fumarate and halogenated methane analogues (e.g. bromochloromethane (BCM)) are able to reduce methane production from ruminants (1,4 -6) . BCM is one of the most effective inhibitors and apparently reduces methane production by interfering with the cobamide-dependent methyl transferase step of methanogenesis (7,8) . BCM complexed in cyclodextrin (CD; BCM-CD) results in the sustained inhibition of methane production when fed to ruminants (9 -11) . Moreover, an in vitro continuous fermentation system simulating rumen fermentation demon...

The effects of cashew nut shell liquid (CNSL) feeding on methane production and rumen fermentation were investigated by repeatedly using 3 Holstein nonlactating cows with rumen fistulas. The cows were fed a concentrate and hay diet (6:4 ratio) for 4 wk (control period) followed by the same diet with a CNSL-containing pellet for the next 3 wk (CNSL period). Two trials were conducted using CNSL pellets blended with only silica (trial 1) or with several other ingredients (trial 2). Each pellet type was fed to cows to allow CNSL intake at 4 g/100 kg of body weight per day. Methane production was measured in a respiration chamber system, and energy balance, nutrient digestibility, and rumen microbial changes were monitored. Methane production per unit of dry matter intake decreased by 38.3 and 19.3% in CNSL feeding trials 1 and 2, respectively. Energy loss as methane emission decreased from 9.7 to 6.1% (trial 1) and from 8.4 to 7.0% (trial 2) with CNSL feeding, whereas the loss to feces (trial 1) and heat production (trial 2) increased. Retained energy did not differ between the control and CNSL periods. Digestibility of dry matter and gross energy decreased with CNSL feeding in trial 1, but did not differ in trial 2. Feeding CNSL caused a decrease in acetate and total short-chain fatty acid levels and an increase in propionate proportion in both trials. Relative copy number of methyl coenzyme-M reductase subunit A gene and its expression decreased with CNSL feeding. The relative abundance of fibrolytic or formate-producing species such as Ruminococcus flavefaciens, Butyrivibrio fibrisolvens, and Treponema bryantii decreased, but species related to propionate production, including Prevotella ruminicolla, Selenomonas ruminantium, Anaerovibrio lipolytica, and Succinivibrio dextrinosolvens, increased. If used in a suitable formulation, CNSL acts as a potent methane-inhibiting and propionate-enhancing agent through the alteration of rumen microbiota without adversely affecting feed digestibility.

On a global scale agriculture and in particular enteric fermentation in ruminants is reported to produce about one fourth (21 to 25%) of the total anthropogenic emissions of methane (CH 4 ). Methane is produced during the anaerobic fermentation of hydrolyzed dietary carbohydrates in the rumen and represents an energy loss to the host besides contributing to emissions of greenhouse gases into the environment. However, there appears to be uncertainty in the CH 4 estimation from livestock due to the limited availability of data to document the variability at the farm level and also due to the significant impact of diet on the enteric CH 4 production. The methane mitigation strategies require robust prediction of emissions from rumen. There are many methods available which would be suitable for measuring CH 4 produced from the various stages of animal production. However, several factors need to be considered in order to select the most appropriate technique like the cost, level of accuracy required and the scale and design of the experiments to be undertaken. Selection of any technique depends on the accuracy as each one has its advantages and disadvantages. Screening of mitigation strategies may be evaluated using individual animal before large-scale trials on groups of animals are carried out. In this review various methods for the estimation of methane production from ruminants as well as for the determination of methane production potential of ruminant feeds are discussed. The advantages and disadvantages of the methods starting from respiration chamber, ventilated hood, facemask, sulphur hexafluoride (SF 6 ) tracer technique, prediction equations and meteorological methods to in vitro methods are detailed.

SUMMARYA series of studies were carried out to measure the methane (CH4) production by Japanese goats fed 19 different diets (D1–D19) varying in nutritive composition in the open circuit respiration chamber (RC) and to compare them with CH4 estimated by the in vitro gas production test (IVGPT). Adult Japanese goats (>2 years old) with a mean body weight of 26±5·4 kg were used in these experiments. Each diet was fed to four randomly selected goats and feeding was carried out at 1·1 maintenance (M) as per National Research Council (NRC) (1981) for goats. Average CH4 emission by goats in the RC ranged from 0·23 to 0·39 (mean value 31 ml/g dry matter intake (DMI)); when it was expressed as a proportion of gross energy or, with methane conversion rate (MCR), it ranged from 5·0 to 8·2, with an average of 6·6. Incorporation of by-products like sweet potato vine silage (SPVS) (P=0·016), dried pumpkin (P=0·052) and brewers' grain in the diet suppressed (P<0·01) methanogenesis in goats, when compared with that of standard farm diet (D1). The CH4 output measured in the RC was very close to that estimated from the gas collected after 24 h and higher after 48 h of in vitro incubation. Although composition of the diets' acid detergent fibre (ADF) had a significant effect on methane emission, methane output estimated by IVGPT was very close to that measured in the RC demonstrating that this system could be used to estimate the CH4 production potential from diets in preparing a database and also in the planning of mitigation strategies in small ruminants to improve their performance as well as to reduce greenhouse gas emissions.