| Methane production in the rumen represents a reduction on theefficiency of feed utilization,but also methane eructated by ruminants may causeirreversible change on environment.Thus mitigating the emission of ruminal methaneis becoming a serious problem.This study was carried out to investigate the effectsof roughage combinations on methane production,ruminal fermentation pattern andmicrobial population in lactating cows and dry cows using sulfur hexafluoride(SF6)tracer method and real-time PCR technique,whether to change the ruminal microbialspecies and fermentation types and reduce methane emission by regulating theproportion of roughage in the diet were discussed in this paper.Three healthy Holstein lactating dairy cows and three dry cows fixed withpermanent rumen fistulas with the similar age and parity were chosen to be treatedwith diet A（10kg corn silage and6kg alfalfa hay）,diet B(17kg corn silage and4kgalfalfa hay) and diet C（23kg corn silage and2kg alfalfa hay) respectively，the ratioof forage and concentrate in the three diets were50:50, the energe level and aciddetergent fiber contents of the three diets were similar,crude protein levels wererespectively15.13%,14.20%and13.32%,neutral detergent fiber contents wererespectively49.42%,51.36%and53.28%.The trial last2months,and each trial lasted20days,the first1~14days of each trial was the pre-feeding period,the methane gassamples from nose and mouth were collected on15~19days and rumen fluid werecollected at different time points in the last day of each trial to determine the value ofruminal pH,the concentration of ammonia nitrogen,microbial crude protein andvolatile fatty acid(acetate acid,propionate acid and butyrate acid).The surplus rumenfluid samples from different times were mixed together,DNA was extracted fromsamples after centrifuging and rumen microorganisms analyzed by real-time PCRtechnique.The numbers of rumen methanogens,protozoa,fungus, F.succinogenres, R.flavefaciens and B.fibrisolvens relative to total bacterial16SrDNA weremeasured.The results showed that:1. The different combination of corn silage and alfalfa hay had a significant effect onthe methane production, the methane production of diet B was significant lower18.26%and8.59%than diet A and diet C in lactating dairy cows(P<0.05)respectively; and the methane production of diet B was significant lower21.20%and8.79%than diet A and diet C in dry cows(P<0.05) respectively;methane productionsin lactating cows and dry cows feeding the same diet had no significantdifferences(P>0.05).2. Rumen fermentation indexes of three diets had significant differences (exceptbutyric acid),the value of ruminal pH,the concentration of ammonia nitrogenconcentration,acetic acid and acetic acid/propionate acid in the diet B and C werelower significantly than diet A(P<0.01); the concentration of propionate acid andmicrobial protein were higher (P<0.01)(P<0.05);the concentration of butyric acid inthe three diets had no significant differences(P>0.05); the rumen fermentation indexesin lactating cows and dry cows feeding the same diet had no significantdifferences(P>0.05).Ruminal pH,ammonia nitrogen concentration,acetic acid,propionate acid and the ratio of acetate to propionate acid of the three diets atdifferent time points had no significant differences(P>0.05);the microbial proteinconcentration of lactating cows feeding diet B and C at different time points had asignificant difference(P<0.01); the microbial protein concentration of dry cows in dietC at different time points had a significant difference acetic acid and propionate acidof dry cows in diet A had a significant difference (P<0.05); butyric acid had asignificant difference (P<0.01); the value of ruminal pH,the concentration ofammonia nitrogen and microbial crude protein at6:00and18:00were higher thanother time points.3.The numbers of rumen methanogens,protozoa, fungus, F.succinogenes,R.flavefaciens and B.fibrisolvens were decreased gradually with the proportion ofcorn silage increased, there was no significant difference in the three diets(exceptfungus)(P>0.05);the number of fungus in diet C was significantly decreased than dietB and C (P<0.01);the numbers of F.succinogenes, R.flavefaciens and B.fibrisolvensrelative to total bacterial16SrDNA had a significant differences (P<0.01);the numberof microbial community was most in diet A and lest in diet C. There was no significant effect of microbial populations on different Physiological Stage（lactatingperiod and dry period） of dairy cows.In summary,it is the most conducive to rumen fermentation and the methaneemission is lest in the addition quantity of17kg corn silage and4kg alfalfa hay oflactating dairy cows and dry cows in the three diets. |
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