The effects of dietary nitrate and of Propionibacterium acidipropionici (PA) on methane and nitrous oxide emissions, methaemoglobinaemia, volatile fatty acid (VFA) concentration and productivity of sheep were studied. It was hypothesised that PA supplementation would increase the rate of nitrite reduction to ammonia in the rumen and therefore reduce risks of methaemoglobinaemia. Fine-wool Merino wethers (n = 28; 31.8 ± 3.7 kg; 11 months of age) were acclimated to four isonitrogenous and isoenergetic diets based on oaten chaff (1.0 kg/day) supplemented with either urea (1.1% of DM; T1 and T2) or a nitrate source (2.0% of DM; T3 and T4) while T2 and T4 were also supplemented with PA (11.5 × 1010 CFU/day). Replacing urea with nitrate lowered methane production (g/day) by 19% and methane yield (g/kg DMI) by 15%, improved clean wool growth by 12% (P < 0.001) and tended to increase skin temperature (P < 0.1). Nitrate increased ruminal acetate to propionate ratio by 27%, increased plasma nitrite and nitrate concentrations and blood methaemoglobin (MetHb) level up to 45% of total haemoglobin. Nitrous oxide emission from sheep confined in respiration chambers was higher (P < 0.001) when nitrate was fed, lowering the net benefit of methane mitigation on global warming potential (CO2 equivalents/kg DMI) by 18%. In contrast, PA had little effect, decreasing total VFA concentration (P < 0.05), increasing rumen pH (P < 0.05) and clean wool growth (P < 0.05) of urea-fed sheep. This study confirmed the beneficial effects of nitrate on net greenhouse gas reduction and wool growth, but showed that methaemoglobinaemia risks may be higher when diets are fed at a restricted level and contain only low levels of readily fermented carbohydrate. PA supplementation was not effective in reducing methaemoglobinaemia, but did increase clean wool growth of urea-fed sheep.

Nitrate (NO3–) supplementation is a promising methane mitigation strategy for ruminants, but can cause nitrite (NO2–) poisoning. Because some nitrite reductases are NADH-dependent, we hypothesised that replacing glucose with glycerol would increase the NADH yield and so enhance nitrite reductase activity and reduce ruminal NO2– accumulation and toxicity risk. We also hypothesised that adapting sheep to dietary NO3– would limit the accumulation of NO2– when NO3– was added to rumen fluid. Changes in NO3– and NO2– catabolism and CH4 production, resulting from supplementation with glycerol to enhance NADH supply, were studied in vitro. In Experiment 1, rumen fluid from sheep adapted to dietary NO3– (2% of DM intake) or urea (1.1% of DM intake) was incubated with NO3– or urea, respectively. Additionally, ground oaten hay was added to incubations alone (control), or with glucose or glycerol. In Experiement 2, sheep were adapted for 9 weeks to dietary NO3– or urea. Nitrate (2% NO3– of substrate DM) was added to incubated digesta from NO3–- or urea-supplemented sheep, while urea (1.1% of substrate DM) was added to digesta from urea-supplemented sheep. In both studies, triplicate incubations were terminated at nine time points up to 24 h. Methane emissions were lower in all NO3– treatments (P < 0.05). Contrary to our hypotheses, both glycerol supplementation (Experiment 1) and prior adaptation to NO3– (Experiment 2) increased NO2– accumulation. In Experiment 1, there was no difference in ruminal NO2– concentration between the unsupplemented control and added glucose treatments. Nitrous oxide accumulated in NO3– treatments only with rumen fluid from sheep adapted to dietary urea (P < 0.05). In summary, NO2– accumulation in vitro was not reduced by adaptation to NO3– or by glucose or glycerol supplementation, disproving the hypotheses regarding the role of NADH availability and of NO2– adaptation in reducing ruminal NO2– accumulation and toxicity risk.

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SummarySupplementing ruminants with nitrate (NO