A Study On The Gut Microbiome And Cellulolytic Bacterium In Bactrian Camels

The Bactrian camel is well adapted for life in extremely arid conditions,including hot summers and cold winters.Camels forage for their food,which include scarce resources such as eat salt-tolerant vegetation and poisonous plants,as well as dry,prickly,and bitter plants.The forestomach of Bactrian camel includes three chambers:the rumen,reticulum,and the abomasum.Compared with other herbivores,the rumen of camels has a longer retention time.The bacterial community plays important roles in the gastrointestinal tracts(GITs)of animals.However,our understanding of the microbial communities in the GIT of Bactrian camels remains limited.In this study,first,we characterized the bacterial microbiota in different gastrointestinal tract segments and different age of the Bactrian camel.We applied shotgun metagenomics to fecal contents from the Bactrian camel to obtain insights into this microbiome and represents a valuable resource for biomass degrading enzyme discovery.On this basis,we isolated and identified a highly cellulose-decomposing bacterium isolated from the feces of Bactrian camels and the enzymatic characteristics were also analyzed.The analysis confinned the Bactrian camel gut's microbiome as a dense and yet largely untapped source of enzymes with the potential to be used in food processing industry.The main research results are as follows:(1)Characterizing the bacterial microbiota in different gastrointestinal tract segments of the Bactrian camelWe describe the bacterial communities from eight different GIT segments(rumen,reticulum,abomasum,duodenum,ileum,jejunum,caecum,colon)and faeces determined from 11 Bactrian camels using 16S rRNA gene amplicon sequencing.Twenty-seven bacterial phyla were found in the GIT,with Firmicutes,Verrucomicrohia and Bacteroidetes predominating.However,there were significant differences in microbial community composition between segments of the GIT.In particular,a greater proportion of Akkermansia and Unclassified Ruminococcaceae were found in the large intestine and faecal samples,while more Unclassified Clostridiales and Unclassified Bacteroidales were present in the in forestomach and small intestine.Comparative analysis of the microbiota from different GIT segments revealed that the microbial profile in the large intestine was like that in faeces.We also predicted the metagenomic profiles for the different GIT regions.In forestomach,there was enrichment associated with replication and repair and amino acid metabolism,while carbohydrate metabolism was enriched in the large intestine and faeces.These results provide profound insights into the GIT microbiota of Bactrian camels.(2)High-throughput sequencing reveals the core gut microbiome of the Bactrian camel in different ageWe used high-throughput 16S rRNA gene sequencing to examine the temporal variability of the fecal microbiota in Bactrian camels.At two months of age,the fecal microbiota was comprised of Firmicutes,Proteobacteria,and Actinobacteria.At one and three years of age,the fecal microbiota was dominated by Firmicutes,Bacteroidetes,and Verrucomicrobia.At the genus level,Blautia,Fusobacterium,and Bifidobacterium were more abundant at two months of age,as well as Eschericha-Shigella.Ruminococcaceae UCG-005,Christensenellaceae_R-7_group and Akkermansia were the most abundant at one and three years of age.Diversity and stability of the gut microbiota increased with age.There was enrichment for genes associated with immune system diseases at two months of age.This study is the first to investigate the distribution of the gut microbiota in Bactrian camels with different ages and provide a baseline for future camel microbiology research.(3)Metagenomic analysis of the fecal microbial composition and potential function from wild and captive Bactrian camelWe applied shotgun metagenomics to fecal contents from eight captive and three wild Bactrian camels to obtain insights into this microbiome.Nucleotide sequence homology-based CAZymes annotation was performed for fecal metadata against the CAZy database.On average 93.73%were of bacteria,0.4%of fungi,0.89%of archaeal.A scan of the set of ORFs uncovered by the metagenome sequencing revealed 38.3%encoded a glycoside hydrolases,26.3%encoded a gycosyl transferases,13.3%encoded a carbohydrate esterases,12.5%encoded a carbohydrate binding modules,4.5%a encoded auxiliary activities and 2.8%a polysaccharide lyases.these sequences were assigned to 126 distinct families of glycoside hydrolases.The putative GHs were classified according to both their main substrate and their mode of action into endoglucanases,endohemicellulases,debranching enzymes and oligosaccharide degrading enzymes in the wild and captive Bactrian camel.Phylogenetic analysis of CAZyme encoding contigs revealed that a significant proportion of CAZymes were contributed by bacteria belonging to genera Clostridium,Ruminococcus,Bacteroides,Faecalibacterium,Paenibacillus.The results indicated that the Bactrian camels fecal microbiome and the CAZymes are highly complex,compositionally distinct from other ruminants.The unique characteristics of microbiota and the enzymes produced by resident microbes provide opportunities to improve the feed conversion efficiency in ruminants.(4)Screening,identification and enzymatic characteristics analysis of cellulolytic bacterium from Bactrian camelThis experiment aimed to isolate and identify a highly cellulose-decomposing bacterium from the feces of Bactrian camels,and to examine its enzymatic characteristics.A cellulolytic bacterium which could highly decompose cellulose was screened by using carboxymethylcellulose plate method and shake-flask fermentation method.According to the morphological characteristics,physiological and biochemical characteristics and the requences alignment of 16 S rRNA,the strain was identified as Cellulosimicrobium cellulans strain.The characteristics of enzymatic properties were studied.Furthermore,the enzymatic characteristics of the bacterial strain were studied based on the suiTable.reaction carbon source,nitrogen source,pH,temperature,time and inoculum of carboxymethyl cellulose produced by the bacterial strain.The results showed that the crude cellulase of Cellulosimicrobium cellulans strain was most active at pH 6.0 and 50?,and the enzyme has strong thermal stability.The optimum fermentation conditions for enzyme production was:culture temperature 35?,the initial pH 7.0,inoculum volume 10%,fermentation time 72 h.the best nitrogen source of AT9-6 for fermentation was yeast extract and the best cabon for fermentation was maltose.The enzyme produced by this strain has certain alkali resistance and heat resistance which can be applied to the food industry or waste disposal.