Effects Of High And Low Proportion Cottonseed Shell Diet On Rumination,Digestive Metabolism And Microbial Flora Of Dolang Sheep

This experiment was conducted to investigate the effects of feeding high and low proportion cottonseed shell diets on rumen microbial community structure,ruminating behavior,rumen fermentation parameters,cellulase and nutrient apparent digestibility of Dolang sheep.The experiment aim was to provide a scientific basis for the efficient utilization of ruminant roughage and the improvement of the utilization rate of cottonseed hull feed.The experiment was designed as a feeding experiment,the pre-feeding period was 30 days and the normal feeding period was 30days.Finally,by analyzing the data of ruminant behavior,nutrient apparent digestibility,ADG,rumen fermentation parameters and rumen microbial community structure of Dolang sheep fed high and low proportion cottonseed shell diets,the adaptive mechanism of Dolang sheep to high proportion cottonseed hull diets was revealed.The specific test results are divided into the following4 parts:Part 1:Effects of feeding high and low proportion cottonseed shell diets on rumination behavior of Dolang sheepThis experiment studied the effects of two kinds of diets with high and low ratio of cottonseed shell on the rumination behavior of Dolang sheep,and explored the adaptability of Dolang sheep to the diets with high ratio of cottonseed shell,so as to provide scientific data for improving the utilization rate of bulk cottonseed shell diets.In the experiment,twenty Dolang sheep with healthy body and permanent rumen fistula were evenly divided into two groups according to the principle of similar body weight（28±2.2kg）.The high proportion of Cottonseed Shell Group（H）and the low proportion of Cottonseed Shell Group（L）,with 10 sheep in each group.The proportion of cottonseed shell was 58%（H）and 20%（L）respectively.The pre-feeding period was 30 days.The ruminant behavior of Dolang sheep was recorded by manual observation in day 13 to day 15 of the normal feeding period,and the rumination behavior data of the experimental sheep were recorded by a timer.Results showed:Feeding high proportion cottonseed hull diets can significantly increase the rumination duration of Dolang sheep（P<0.05）,and the number of ruminations increased but the difference was not significant（P>0.05）.The feeding of high proportion cottonseed hull diets did not affect the circadian rhythm and rumination of Dolang sheep.The rumination behavior of Dolang sheep in Group H and L was mainly concentrated at night,The rumination at night time in both groups H and L was higher than that in the daytime rumination,and the proportion of rumination at night in the total rumination time was 63.97%（H）and 60.24%（L）and the proportion of daytime rumination was 36.03%（H）and 39.76%（L）.The rumination duration per hour in Group H and Group L showed similar changes.After feeding,the rumination duration per hour in Group H and Group L decreased firstly and then increased.After morning feeding（9:00）,the rumination duration per hour decreased firstly and then increased,and decreased to the lowest after5 h of morning feeding,and reached the peak after 8 h.After feeding in the afternoon（21:00）1 h after the lowest,after 9 h reached the peak of rumination,and in most of the time period of the day Group H rumination time per hour were higher than Group L.Part 2:Effects of feeding high and low proportion cottonseed shell diets on rumen fermentation parameters and cellulase activity of Dolang sheepThe experiment design and feeding management are the same as part 1.The rumen fluid was collected at 0,2,4,6 and 8 h after morning feeding in day 9 to day 11 of the normal feeding period to measure the p H value,NH₃-N and the concentration and volatile fatty acid（VFA）.The rumen contents were collected before morning feeding for cellulase activity determination in day 20 of the normal feeding period.The results showed:After morning feeding,the rumen p H values of Group H and Group L decreased firstly and then increased,decreased to the lowest value after 2 h of morning feeding,and reached the highest value after 8 h of morning feeding.The p H values of Group H were higher than those of Group L in different periods,and the difference is significant at0～2 h after morning feeding（P<0.05）.After morning feeding,the rumen NH₃-N concentrations of Group H and Group L decreased firstly and then increased.After morning feeding,the rumen NH₃-N concentrations gradually decreased.The rumen NH₃-N concentrations of Group H and Group L decreased to the lowest at 4 h and 6 h after morning feeding,respectively,and then slowly recovered.The NH₃-N concentrations of Group L were higher than those of Group H in different time periods.The NH₃-N concentrations were extremely significant differences at 0,2,4 and 8 h after morning feeding（P<0.01）,was significant differences at 6 h after morning feeding（P<0.05）.The concentrations of Acetate,Propionate,Pentanoate and total VFA in rumen showed a downward trend after morning feeding.The concentration of Acetate in Group H was significantly lower than that in Group L at 0 and 2 h after morning feeding（P<0.05）.The Propionate concentration in Group H was lower than that in Group L at different time periods,and the difference was significant at 2 h after morning feeding（P<0.05）.The Butyric concentrations in Group H were higher than those in Group L at 0 h,4 h,6 h and 8 h after morning feeding,and lower than those in Group H at 2 h,but there were no significant differences（P>0.05）.The concentration of Pentanoate in Group H was lower than that in Group L at different times,and the concentration of valeric acid in Group H was significantly lower than that in Group L at 0,4 and 6 h after morning feeding（P<0.05）.The total VFA concentrations in Group H were lower than those in Group L at different time periods,and the total VFA concentrations in Group H were significantly lower than those in Group L at 0～2 h after morning feeding（P<0.05）.Feeding a high proportion of cottonseed husks significantly increased the activity of Carboxymethyl Cellulase in the rumen of dolang sheep（P<0.05）.Part 3:Effects of feeding high and low proportion cottonseed shell diets on apparent digestibility of Dolang sheepThe experiment design and feeding management are the same as part 1.The apparent digestibility of nutrients was measured by the total feces collection method on the 1～8 d of the normal feeding period.The experimental animals were weighed on an empty stomach（before morning feeding）before and at the end of the normal feeding period to determine the ADG.The results showed that the apparent digestibility and average daily gain of Dolang sheep were affected by feeding a high proportion of cottonseed hull diets.Dry matter digestibility in Group H was significantly lower than that in Group L（P<0.05）.The digestibility of CP,NDF,OM and ADG was lower than that in Group L,but there was no significant difference（P>0.05）.Part 4:Effects of feeding high and low proportion cottonseed shell diets on rumen microflora structure of Dolang sheepThe experiment design and feeding management are the same as part 1.The rumen fluid was collected before morning feeding in day 25 to day 27 of the normal feeding period.The composition and structure of rumen bacteria,fungi and protozoa were determined by three-generation full-length extenders and 16Sr RNA high-throughput sequencing.The results showed that there was a significant difference in the rumen microflora structure between the Group H and the Group L after feeding with high and low proportions of cottonseed hulls（P<0.05）.The relative abundance of Bacteroides in Group H was significantly higher than that in Group L at phylum level（P<0.05）.The relative abundance of Euryarchaeota was significantly lower than that in Group L（P<0.05）.The relative abundance of Bacteroides in Group H was significantly higher than that in group L（P<0.05）,while the relative abundance of Eubacterium,Lachnoclostridium,Selenomonas and Methanobrevibacter in Group H was significantly lower than that in Group L（P<0.05）.The relative abundance of total methanogens in Group H was significantly lower than that in Group L（P<0.05）.The relative abundances of YE315Methanobrevibacter sp.YE315,Methanobrevibacter millerae,Methanobrevibacter gottschalkii,and Methanobrevibacter oralis were all significantly lower than those in Group L（P<0.05）.There was no significant difference in the relative abundance of anaerobic fungi fed to a high proportion of cottonseed hulls（P>0.05）,but at the species level,the relative abundance of Neocallimastix californiae was significantly lower in Group H than in Group L（P<0.05）The two groups of rumen protozoa are mainly composed of two families:Ophryoscolecidae and Isotrichidae.Six genera:Entodinium,Eudiplodinium,Epidinium,Isotricha,Polyplastron,Ophryoscolex and ten species.There was no significant difference in the relative abundance of total ciliates and total rumen protozoa between Group H and Group L（P>0.05）,and there was no significant difference at the species level（P>0.05）.In the carbohydrate enzyme family,a total of 359 carbohydrate enzyme families were detected in the rumen contents of Group H and Group L,including 9 enzyme families with significant differences between the two treatment groups（GH2,GH9,GH51,GH97,CBM6,CE6,CE10,dockerin and AA6）.Among them,the relative abundance of seven enzyme families（GH2,GH9,GH51,GH97,CBM6,CE6,CE10）increased significantly in Group H（P<0.05）,and decreased significantly in two groups（dockerin and AA6）（P<0.05）.The increased enzyme family functions in Group H mainly included the activities of polysaccharides and glycoside hydrolases,and the decrease was mainly the key scaffold proteins for the formation of benzoquinone reductase and fibrosomes.ConclusionAbove all,feeding high proportion cottonseed hull diets increased rumen p H,decreased rumen NH₃-N concentration,decreased propionic acid and valeric acid concentration,and increased the activity of Carboxymethyl Cellulase.Although DM and ADF apparent digestibility were reduced,there were no significant differences in CP,OM apparent digestibility and ADG.The adaptability of Dolang sheep to high proportion cottonseed hull diets was to prolong the rumination time and Improve NH₃-N utilization,fully digest the high proportion cottonseed hull diets with higher NDF content,and enrich higher abundance rumen bacteria and polysaccharide and glycoside hydrolysis bacteria to participate in microbial degradation.