Responses in digestion, rumen fermentation and microbial populations to inhibition of methane formation by a halogenated methane analogue

National Institute of Livestock and Grassland Science, Tsukuba, Ibaraki 305-0901, Japan.
The British journal of nutrition (Impact Factor: 3.45). 11/2011; 108(3):482-91. DOI: 10.1017/S0007114511005794
Source: PubMed

ABSTRACT 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 H2 production. The levels of methane production were reduced by 5, 71 and 91 %, and H2 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 H2 gas.

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Available from: Christopher S Mcsweeney, Jul 15, 2014
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    • "It is possible that not all individuals within a herd respond in the same manner to NOP and the possibility that some animals may exhibit adaptation over time cannot be discounted. When evaluating the effects of providing goats with BCM, Mitsumori et al. (2012) "
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    ABSTRACT: The objective was to evaluate whether long-term addition of 3-nitrooxypropanol (NOP) to a beef cattle diet results in a sustained reduction in enteric CH emissions in beef cattle. Eight ruminally cannulated heifers (637 ± 16.2 kg BW) were used in a completely randomized design with 2 treatments: Control (0 g/d of NOP) and NOP (2 g/d of NOP). Treatments were mixed by hand into the total mixed ration (60% forage, DM basis) at feeding time. Feed offered was restricted to 65% of ad libitum DMI (slightly over maintenance energy intake) and provided once per day. The duration of the experiment was 146 d, including an initial 18-d covariate period without NOP use; a 112-d treatment period with NOP addition to the diet, divided into four 28-d time intervals (d 1 to 28, 29 to 56, 57 to 84, and 85 to 112); and a final 16-d recovery period without NOP use. During the covariate period and at the end of each interval and the end of the recovery period, CH was measured for 3 d using whole animal metabolic chambers. The concentration of VFA was measured in rumen fluid samples collected 0, 3, and 6 h after feeding, and the microbial population was evaluated using rumen samples collected 3 h after feeding on d 12 of the covariate period, d 22 of each interval within the treatment period, and d 8 of the recovery period. Average DMI for the experiment was 7.04 ± 0.27 kg. Methane emissions were reduced by 59.2% when NOP was used (9.16 vs. 22.46 g/kg DMI; < 0.01). Total VFA concentrations were not affected ( = 0.12); however, molar proportion of acetate was reduced and that for propionate increased when NOP was added ( < 0.01), which reduced the acetate to propionate ratio (3.0 vs. 4.0; < 0.01). The total copy number of the 16S rRNA gene of total bacteria was not affected ( = 0.50) by NOP, but the copy number of the 16S rRNA gene of methanogens was reduced ( < 0.01) and the copy number of the 18S rRNA gene of protozoa was increased ( = 0.03). The residual effect of NOP for most of the variables studied was not observed or was minimal during the recovery period. These results demonstrated that the addition of NOP to a diet for beef cattle caused a sustained decrease of methanogenesis, with no sign of adaptation, and that these effects were reversed once NOP addition was discontinued.
    Journal of Animal Science 04/2015; 93(4):1780. DOI:10.2527/jas.2014-8726 · 2.11 Impact Factor
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    • "This is in accordance with the hypothesis that rather than the total archaeal biomass it is the species distribution that correlates to changes in methane production in the rumen (Zhou et al., 2010; Abecia et al., 2012). Mitsumori et al. (2012) reported a halflog reduction in the normal methanogens population correlated with >50% reduction in methane, indicating that the relative methanogenic activity of different archaeal species in the rumen plays a greater role in determining methane output than absolute number of methanogens. Therefore, any study of the methanogenesis inhibition should account for the effect on a wide range of ruminal methanogenic Archaea species and not only on those that are believed to be numerically dominant. "
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    ABSTRACT: The aim of this work was to investigate the effect of α-cyclodextrin-propyl propane thiosulfinate complex (CD-PTS) as antimethanogenic compound in ruminants using in vivo and in vitro approaches. The in vivo trial lasted 36 days and was designed to study the effect of CD-PTS on ruminal methane production, fermentation pattern, microbial abundances and nutrient utilization in goats. Twelve adult goats fed with alfalfa hay and concentrate (50:50), were split in two groups: treated with CD-PTS complex at 200 mg of active component/L rumen content per day or without any treatment (control). On d 7, 14, 21 and 28 methane emissions were recorded in chambers and rumen content samples were collected for pH, volatile fatty acid (VFA) and NH3 analyses. On day 14 and 28 rumen samples were collected for quantification of bacterial, protozoal and archaeal numbers by quantitative real-time PCR (qPCR) and to study the archaeal population structure by denaturing gradient gel electrophoresis. In addition, on days 17 and 18 samples of alfalfa hay were placed into nylon bags and incubated in the rumen of each goat for 24 and 48 h to determine the ruminal degradation of dry matter (DM) and neutral detergent fiber (aNDFom). During the last 5 days of the trial, nutrient digestibility, N and energy balances and urinary purine derivative (PD) excretion were determined. Additionally, an in vitro experiment with pure cultures of methanogens was conducted to test propyl propane thiosulfinate (PTS) at a dose of 200 mg of active component/L culture medium and bromochloromethane (BCM) at a concentration of 10 mg/L culture medium against three different archaeal strains: Methanobrevibacter ruminantium, Methanobrevibacter smithii and Methanobrevibacter millerae. In vivo, no significant (P≤0.17) reduction on methane production was observed although a numerical decrease on day 28 was observed with CD-PTS. The dry matter intake (DMI), nutrient digestibility, N and energy balances, purine derivatives, creatinine and estimated microbial N flow were not affected by the treatment (P≤0.18). Likewise, total concentration of the analyzed microbial groups in the rumen, showed no difference (P≤0.33) between treated and non-treated goats. However, on day 28, the structure of the archaeal population in the rumen of goats treated with CD-PTS was different compared with that in control goats. The in vitro culture of methanogens showed a substantial reduction of methane production in all the strains by both BCM and PTS.
    Animal Feed Science and Technology 05/2014; 191. DOI:10.1016/j.anifeedsci.2014.01.008 · 2.00 Impact Factor
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    • "These results could be explained by a mechanism in which Fibrobacter increase in biomass to compensate the decrease in fibrolytic ruminococci and rumen fungi, as shown by Mitsumori et al. (2012) in goats treated with BCM. Mitsumori et al. (2012) also revealed that the rumen ecosystem appeared to adapt to the high H 2 levels by shifting fermentation to propionate and increasing the population of hydrogenconsuming Prevotella spp. The quantification of the 3 main microbial groups in the rumen showed that the reduction of methane emissions was not accompanied by decreased concentration of any microbial group. "
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    ABSTRACT: The aim of this work was to investigate the effect of feeding ethyl-3-nitrooxy propionate (E3NP) and 3-nitrooxypropanol (3NP), 2 recently developed compounds with potential antimethanogenic activity, in vitro and in vivo in nonlactating sheep on ruminal methane production, fermentation pattern, the abundance of major microbial groups, and feed degradability. Three experiments were conducted, 1 in vitro and 2 in vivo. The in vitro batch culture trial (experiment 1) tested 2 doses of E3NP and 3NP (40 and 80 µL/L), which showed a substantial reduction of methane production (up to 95%) without affecting concentration of volatile fatty acids (VFA). The 2 in vivo trials were conducted over 16 d (experiment 2) and 30 d (experiment 3) to study their effects in sheep. In experiment 2, 6 adult nonpregnant sheep, with permanent rumen cannula and fed alfalfa hay and oats (60:40), were treated with E3NP at 2 doses (50 and 500 mg/animal per day). After 7, 14, and 15 d of treatment, methane emissions were recorded in respiration chambers and rumen fluid samples were collected for VFA analysis and quantification of bacterial, protozoal, and archaeal numbers by real-time PCR. Methane production decreased by 29% compared with the control with the higher dose of E3NP on d 14 to 15. A decrease in the acetate:propionate ratio was observed without detrimental effects on dry matter intake. In experiment 3, 9 adult nonpregnant sheep, with permanent rumen cannula and fed with alfalfa hay and oats (60:40), were treated with E3NP or 3NP at one dose (100 mg/animal per day) over 30 d. On d 14 and d 29 to 30, methane emissions were recorded in respiration chambers. Rumen fluid samples were collected on d 29 and 30 for VFA analysis and quantification of bacterial, protozoal, and archaeal numbers by real-time PCR. In addition, on d 22 and 23, samples of oats and alfalfa hay were incubated in the rumen of sheep to determine dry matter ruminal degradation over 24 and 48 h, respectively; no effect was observed (78.6, 78.3, and 78.8% of alfalfa and 74.2, 74.0, and 70.6% of oats in control, E3NP, and 3NP groups, respectively). A reduction in methane production was observed for both additives at d 14 and d 29 to 30. In both treatments, the acetate:propionate ratio was significantly decreased. Likewise, total concentrations of the analyzed microbial groups in the rumen showed no difference among treatments and doses for both experiments. Both tested compounds showed promise as methane inhibitors in the rumen, with no detrimental effects on fermentation or intake, which would need to be confirmed in lactating animals.
    Journal of Dairy Science 04/2014; 97(6). DOI:10.3168/jds.2013-7398 · 2.57 Impact Factor
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