Effect Of Arginine And Inulin On Intestinal Morphology,Caecal Microflora And Growth Performace Of Meat Rabbits

In rabbit production,diarrhea seriously threatens the health of the rabbits.The highest morbidity and mortality of rabbits always occur from weaning to 60 days age.More than 85% of them are related to intestine.Once the rabbits are involved in intestinal problems,there are almost no time for them to receive treatment.So early prevention and increased intestinal immunity are the key to solve the problem.In recent years,people pay more and more attention to the immune modulating agents such as L-arginine and inlulin.This study aims to investigate the effects of L-arginine and inulin on intestinal morphology,cecal microflora and growth performance of rabbits,and to provide a reference for the application of arginine and inulin in rabbit production.Experiment 1: The development of intestinal morphology and microflora structure of rabbits after weaningNearly 80 weaned Hyla rabbits with samilar age and weighing were selected.At 0,3,7,14 and 28 d after weaning,8 rabbits were slaughtered respectively and then the cecum were quickly removed.We aimed to study the development of intestinal morphology,cecal fermentation and microflora structure based on 16 S rRNA of rabbits after weaning.The results were as follows:Intestinal morphology: duodenal villus height at 3 d after weaning decreased by 9.1%(P<0.05)compared with the 0 d.On 14 d after weaning,villus height was restored to a height of 0 d.Villus height was up to a maximum at 28 d.Crypt depth increased from 0 d after weaning and reached a maximum at 7 d.V/C was the lowest in the duodenum at 7 d after weaning,significantly lower than that of 0 d(P<0.05).The villus height of jejunum decreased gradually after weaning during 0-7 d,reaching the lowest at 7 d(P>0.05)and the highest at 28 d(P<0.05).The crypt depth of jejunum was gradually thickened(P>0.05)from 0-7 d,and then decreased at 14 d.V/C reached the lowest at 7 d after weaning,significantly lower than that of weaning at 14 and 28 d(P<0.05).Volatile fatty acids: pH values in 0,3 d had significant difference with 7,14,28 d(P<0.05).There was no significant difference in ammonia nitrogen at 0,3,7 and 14 d after weaning.The yield of acetic acid at 14 and 28 d was significantly higher than that of 0,3 and 7 d(P<0.05),while the content of propionic acid at weaning day was significantly higher than that of other groups(P<0.05).The total volatile fatty acid yield was highest at 28 d after weaning,significantly higher than that in other groups(P<0.05).Analysis of microbial community structure in caecum:OTUs statistics and diversity analysis: in respect of the reads numbers,25 samples were obtained a total of 150,399 sequences and the sequence numbers of each group were not significantly different(P>0.05).The number of OTUs of each rabbit in caecum microorganism was in the range of 15,000-19,000,but no significant differences was found between the groups(P>0.05).Ace,Chao1 and Shannon index increased gradually and reached the highest at 28 d after weaning.The Ace and Chao1 values of 28 d after weaning were significantly higher than that of other days(P<0.05).The Shannon values at 28 d after weaning were higher than that of 0 and 14 d(P<0.05).Analysis of microbial community structure in caecum:At the phylum level,10 was detected over the 0.01%.The sum of the relative abundance of Bacteroidetes and Firmicutes was more than 96%.At 0 d,the relative abundance of Firmicutes was significantly lower than that of 3,7,14,and 28 d(P<0.05).The relative abundance of Bacteroidetes at 3,5,7,and 14 d was 12.1%,8%,13.1% and 4.2%,respectively,which was significantly lower than that of weaning at 0 d(35.2%,P<0.05).The relative abundance of Cyanobacteria was 0.001% and 1.2% at 0 and 28 d,respectively.The relative abundance of Bacteria_Unclassified at 28 d was significantly higher than that of 0,3,7 and 14 d(P<0.05).The abundance of Proteobacteria,Tenericutes and Verrucomicrobia was not affected by age(P>0.05).At the level of family community,34 families were detected with microbial abundance of more than 0.01% microorganisms,except for unknown bacteria.The relative abundance of 8 families was more than 1%.Three families of higher microbial abundance were Ruminococcaceae(30.9%-54.6%)and Lachnospiraceae(26.6%-29.8%)and Clostridiales(5.1%-9.1%).There were 12,14 families,whose abundance was about 0.1%-1% and 0.01%-0.1%,respectively.The relative abundance of Ruminococcaceae at 0,3,7 d was significant lower than that at 14 and 28 d.The relative abundance of Eubacteriaceae increased with the increase of weaning age,but there was significant difference between 28 d and 0 d(P<0.05).Porphyromonadaceae and Bacteroidaceae decreased with the increase of weaning age.At the genus level,97 genus were found with the abundance of more than 0.01%,except for unknown bacteria.There were 18 genus with the relative abundance of 0.1%-1%,and 69 genus with the relative abundance of 0.01%-1%.Based on the analysis of the differences of dominant bacteria in each group,it was found that the relative abundance of Ruminococcaceae increased with the increase of age.There was no significant difference between 0,3 d and 7,14 d in Ruminococcaceae,but the abundance of these groups was significantly lower than that of 28 d(P<0.05).With the increase of age after weaning,Bacteroides decreased gradually,and there was a significant difference between 28 d and 0,3 d.Experiment two: the effects of arginine and inulin on intestinal morphology and microbial community structure of rabbitsA total of 160 weaned hybrid Hyla rabbits(35-day-old)were selected for this experiment.After 10 days of dietary adaptation,rabbits were randomly divided into 4 treatments and each treatment had 5 replicates of 8 rabbits each.The treatments were control,Arg group(control+1% arginine),Inu group(control+4% inulin)and Arg+Inu group(control+1% arginine +4% inulin)with a two-factor design.The nutrient levels were almost the same.The experiment lasted 21 days to investigate the effects of arginine and inulin on the intestinal morphology,caecum fermentation and microbial community structure on the rabbits.The main results were as follows:Intestinal morphology: Dietary arginine and inulin had no effect on the duodenal villus height(P>0.05),but significantly decreased the crypt depth and increased the ratio of villus height to crypt depth(P<0.05).There was no interaction between arginine and inulin(P>0.05).Dietary arginine and inulin did not affect the jejunal villus height,crypt depth(P>0.05),but inulin significantly increased villus height / crypt depth(P<0.001).Cecal fermentation: Dietary arginine and inulin significantly reduced cecal pH value and ammonia nitrogen(P<0.05).Arginine significantly improved the yield of butyric acid(P<0.05),with no influence on the yield of acetic acid,propionic acid and total volatile fatty acids(P>0.05).Inulin could significantly increase the yield of acetic acid,butyric acid and total volatile fatty acids(P<0.05),but had no effect on the yield of propionic acid(P>0.05).Analysis of microbial community structure in caecum:OTUs statistical analysis and diversity: There was a total of 116,080 16 S rRNA sequence.The number of samples OUT in the control group were lower than those in the arginine,inulin and arginine + inulin groups,but there was no significant difference between the groups(P>0.05).Dietary supplementation of arginine and inulin had no significant effect on Ace,diversity index(Chao1)and Shannon index(Shannon)of 16 S rRNA(P>0.05)in cecal contents.Analysis of microbial community structure in caecum:At the level of the phylum,more than 13 phyla was found with the abundance above 0.01%.The proportion of Firmicutes accounted for 69%-80%,followed by Bacteroidetes(16%-26%).The Proteobacteria accounted for about 1%-4%,and no interaction was found between arginine and inulin.Inulin could significantly increase the amount of Actinobacteria(P=0.009),and had no significant effect on other classification(P>0.05).Arginine had no significant effect on the distribution in cecum(P>0.05).At the family level,there were 67 families with the abundance of more than 0.01%.Ruminococcaceae was about 35.8%-43.7% and Lachnospiraceae was 13.8%-22.0%.Arginine and inulin had no significant difference on the families(P>0.05).The abundance of Firmicutes Unclassified(0.23%-0.56%)was on the increase(P=0.01),while the abundance of Defluviitaleaceae was significantly decreased(P=0.005)in Inulin group.45 families was found in Inulin with the relative abundance of 0.01%-0.1%,while inulin group and arginine + inulin group contained a total of 33 families solely.At the genus level,there were 133 genus with the abundance of more than 0.01%.The control group did not have Eubacterium hallii,whereas Eubacterium was found(<0.01%)in arginine,inulin,arginine,and inulin groups.The abundance of Desulfovibrio in arginine group was significantly lower than that in control group(P=0.05),and the interaction between arginine and inulin(P=0.018)was found.There were 98 genus whose relative abundance was in 0.01%-0.1%.There were 45 genus of microorganisms in the inulin group and the arginine + inulin group solely.Experiment three: the effects of arginine and inulin on growth performance,serum biochemical indexes and meat quality of rabbitsA total of 192 weaned hybrid Hyla rabbits(45-day-old)were selected for this experiment.Rabbits were randomly divided into 4 treatments and each treatment had 6 replicates of 8 rabbits each.The treatments were control,Arg group(control+1% arginine),Inu group(control+4% inulin)and Arg+Inu group(control+1% arginine +4% inulin)with a two-factor design.The nutrient levels were almost the same.The experiment lasted 21 days to investigate the effects of arginine and inulin on the growth performance,nutrient metabolism and meat quality on the rabbits.The main results were as follows:Growth performance: Dietary arginine(P=0.017)and inulin(P=0.040)could significantly improve the body weight of rabbits at the end of experiment.Dietary arginine and inulin did not affect the intake of meat rabbit(P>0.05),but could significantly increase the average daily gain(P<0.05).Arginine had no effect on the feed conversion ratio,but inulin could reduce it(P<0.05).Carcass traits:Arginine couldn't improve the evwascerated weight and evwascerated weight with giblet(P>0.05),but Inulin could improve them(P<0.05).There were no significant difference in percentage of evwascerated weight and evwascerated weight with giblet,cardiac index,liver index and kidney index(P>0.05).Nutrient digestibility: Arginine could significantly improve the digestibility of dry matter,crude protein,crude fat and ADF(P<0.05),but had no effects on NDF,calcium and phosphorus(P>0.05).Inulin could significantly increase the digestibility of dry matter and phosphorus,but decrease the digestibility of ADF and crude fat(P<0.05),with no effect on crude protein,NDF and calcium(P>0.05).Serum biochemical indexes: Arginine could significantly increase the level of serum alkaline phosphatase and reduce the content of serum urea nitrogen(P<0.05).However,no significant effect was found on creatinine,calcium and phosphorus(P>0.05).Dietary inulin could decrease serum urea nitrogen(P<0.05)and aspartate aminotransferase(0.05<P<0.1).Both arginine and inulin could significantly decrease serum cholesterol,triglyceride,low density lipoprotein and very low density lipoprotein levels(P<0.001),but had no significant impact on the high density lipoprotein(P>0.05).There wad no interactive effect between arginine and inulin(P>0.05).Immune index: Both dietary arginine and inulin had no significant effect on thymus and spleen index,serum TNF-? and IL-6 of rabbits(P>0.05).Dietary inulin had a trend of increasing serum IL-2(0.05<P<0.10).Muscle quality: In respect to meat color,dietary arginine and inulin had no effect on L*(P>0.05),but significantly increased a*,b* and C(P<0.05).Arginine and inulin had no interaction with L*,a*,b* and C(P>0.05).In terms of meat quality,arginine had a trend to decreas pH value and improve the water holding capacity of the longissimus dorsi muscle(0.05<P<0.10).However,dietary inulin had no effect on pH value,water holding capacity,cooking loss(P>0.05).Dietary arginine and inulin had no interaction on meat quality(P > 0.05).Dietary arginine could significantly increased the content of arginine in longwassimus dorsi muscle(P<0.05)and inulin could increase the content of methionine,tyrosine and proline(P<0.05).No interaction was found between arginine and inulin(P>0.05).The main conclusion were as follows:(1)Intestinal morphology,caecum fermentation state of rabbits restored to the level of weaning day within 14 days after weaning.The diversity of cecal microorganisms was relatively stable at weaning 0-14 days,and the diversity of cecal microorganisms increased significantly at 14-28 days after weaning.(2)Firmicutes and Bacteroidetes were the caecal microbial flora of rabbits accounting for more than 93% of the total sequencing.This pattern was not affected by age,nutrition regulation after weaning.(3)Inulin cound not change the abundance of dominant microflora,but could increase the diversity of cecum microorganism in rabbits.These microorganisms might play a key role in regulating metabolism and intestinal immunity.(4)Arginine and inulin could improve the growth performance of rabbits,improve the meat quality,and had interaction in protein metabolism,lipid metabolism,morphology of intestinal mucosa.