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The effects of two kinds of Escherichia coli strains, wild-type E. coli W3110 or E. coli nir-Ptac, which has enhanced nitrite reduction activity, on in vitro CH4 production and nitrate and nitrite reduction in cultures of mixed ruminal microorganisms was investigated using continuous incubation systems. Escherichia coli nir-Ptac, a derivative of wild-type E. coli W3110, was constructed by replacing self promoter of nir BD operon encoding subunits of nitrite reductase in E. coli W3110 by tac promoter to make the expression of nir BD higher and constitutive. The nitrite reductase activity of E. coli nir-Ptac was approximately twice as high as E. coli W3110. The culture media consisted of 400 mL of strained ruminal fluid taken from two nonlactating Holstein cows receiving a basal diet of orchardgrass hay at maintenance level (55 g of DM/kg of BW0.75 daily), and 400 mL of autoclaved artificial saliva. Treatments were arranged in two separate 3 x 3 factorials consisting of nitrate (NaNO3; 0, 5, or 10 mM) without E. coli or inoculated with E. coli W3110 or E. coli nir-Ptac, or nitrite (NaNO2; 0, 1 or 2 mM) without E. coli or inoculated with E. coli W3110 or E. coli nir-Ptac. The control culture contained no chemical or microbial additives. Escherichia coli cells were inoculated into in vitro mixed ruminal cultures at approximately 2 x 10(8) to 10(9) cells/mL. Methane production by ruminal microorganisms was decreased markedly (P < 0.001) by the addition of nitrate and nitrite, and by the inoculation of cultures with E. coli W3110 or E. coli nir-Ptac (P < 0.01). With mixed nitrite-containing cultures, E. coli nir-Ptac inhibited (P < 0.001) in vitro nitrite accumulation and CH4 production more than E. coli W3110, which may be due to the tac promoter-enhanced nitrite reductase activity of E. coli nir-Ptac accelerating electrons to be consumed for nitrite reduction rather than CH4 biosynthesis. In conclusion, anaerobic cultures of E. coli W3110 or E. coli nir-Ptac may decrease CH4 production in the rumen. The inoculation of E. coli W3110 or, especially, E. coli nir-Ptac to mixed ruminal microorganisms may decrease nitrite toxicity when ruminants consume high-nitrate-containing forages and when nitrite is applied to abate ruminal CH4 production.

The effects of two kinds of Escherichia coli (E. coli) strain, wild-type E. coli W3110 and E. coli nir-Ptac, which has enhanced NO 2 reduction activity, on oral CH 4 emission and NO 3 toxicity in NO 3 -treated sheep were assessed in a respiratory hood system in a 4 £ 6 Youden square design. NO 3 (1·3 g NaNO 3 / kg 0·75 body weight) and/or E. coli strains were delivered into the rumen through a fistula as a single dose 30 min after the morning meal. Escherichia coli cells were inoculated for sheep to provide an initial E. coli cell density of optical density at 660 nm of 2, which corresponded to 2 £ 10 10 cells/ml. The six treatments consisted of saline, E. coli W3110, E. coli nir-Ptac, NO 3 , NO 3 plus E. coli W3110, and NO 3 plus E. coli nir-Ptac. CH 4 emission from sheep was reduced by the inoculation of E. coli W3110 or E. coli nir-Ptac by 6 % and 12 %, respectively. NO 3 markedly inhibited CH 4 emission from sheep. Compared with sheep given NO 3 alone, the inoculation of E. coli W3110 to NO 3 -infused sheep lessened ruminal and plasma toxic NO 2 accumulation and blood methaemoglobin production, while keeping ruminal methanogenesis low. Ruminal and plasma toxic NO 2 accumulation and blood methaemoglobin production in sheep were unaffected by the inoculation of E. coli nir-Ptac. These results suggest that ruminal methanogenesis may be reduced by the inoculation of E. coli W3110 or E. coli nir-Ptac. The inoculation of E. coli W3110 may abate NO 3 toxicity when NO 3 is used to inhibit CH 4 emission from ruminants.Escherichia coli W3110: Escherichia coli nir-Ptac: Methane emission: Nitrate

Four nonlactating, ruminally cannulated Holstein cows were used in a 4 x 4 Latin square design, balanced for residual effects, to evaluate the effects of supplementing dairy cow diets with yeast culture (Trichosporon sericeum; YC), galacto-oligosaccharides (GOS), or the mixture of YC and GOS on ruminal fermentation, microbial N supply, in situ degradation, and energy and nitrogen metabolism. Treatments were arranged in a 2 x 2 factorial as follows: 1) basal diet, 2) basal diet plus 10 g/d YC, 3) basal diet plus 2% GOS, 4) basal diet plus a mixture of 10 g/d YC and 2% GOS. Nitrogen losses in urine were lower, and retained N was higher, for cows supplemented with a mixture of YC and GOS. Ruminal pH was lower in cows supplemented with GOS alone compared with other treatments. Total VFA concentration was higher in cows fed control and GOS-supplemented diets than in those fed YC containing diets. The molar proportion of propionate was higher, and the molar proportion of acetate was lower, in cows fed control diets. Microbial N supply was higher in cows fed control diets. There were no major positive effects of supplements observed in this study. However, supplementation of a mixture of YC and GOS had a tendency for synergistic effects on N metabolism and in situ degradation of a soluble fraction of oat straw DM and CP of concentrates compared with supplementation of YC or GOS alone.

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