Analysis On Microbiome Structure And Function In Stomach Of Cervidae

In recent years,the research on the microbiome structure of the digestive system has gradually deepened.The coevolutionary effect of the host and its related microbial flora has been recognized by more and more people.In the long-time coevolution with the cervidae,the microbiome in cervidae continuously affects its feeding behavior,and promotes the cervidae to the digestible system that can decompose high fibers.The physiological structure to make the benefit environment required for complex microorganisms.The delicate dynamic balance formed between this symbiotic microbiome and their host to maintain continuous state of evolution.However,it is unknown whether the interactions and functions of microbiome structures in the stomach can reflect the phylogenetic relationships among deer species.Our study,High-throughput Sequencing technology was used to research microbiome in ruminants' stomach.In our research,five different species cervidae(Sika deer,red deer,reindeer,white-lipped deer and roe deer)were chosen to research the microbiome to explore three aspects in their four stomach:1)Analyze the composition of the three major microbial groups of bacteria,archaea,and fungi in the stomach of 5 species of deer,and determine the composition of the microbial flora in each stomach;2)By comparing the differences in the microbial flora in the stomachs of five deer species,we tried to explore the relationship between the high altitude tolerance or cold tolerance of deer species and the microbial flora in the stomach;3)Through the prediction of metabolic pathway functions,explore the functional differences in the three major microbial groups involved in the catabolic process in different deer stomachs.4)Compare the differences in the microbiome in the stomachs of different deer species.Try to find the differences between the difference in community composition and the phylogenetic relationship between deer species.The results are as follows:1.BacteriaTwenty-three phylum were found in five deer species(Sika deer,reindeer,white-lipped deer,red deer,roe deer;belonging to 19,19,21,20,and 15 phylum),Firmicutes and Bacteroidetes are the dominant bacteria.In addition to the dominant phylum,among the four stomachs of the rumen,reticulum,omasum,and abomasum,the Actinobacteria of sika deer abdomen accounts for up to 23.47%,which is much higher than that of other stomachs.Proteobacteria(1.56%,3.84%)are higher than rumen and reticulum(0.29%,0.4%);The proportions of Proteobacteria,Actinobacteria,and Tenericutes of the reindeer abomasum were 10.17%,6.51%,and 1.4%,which were higher than those of other stomachs.Then only 0.73%,lower than other stomachs;The proportion of actinomycota in the white-lipped deer's abomasum is as high as 11.95%,while that of other stomachs is less than 1%.The proportion of TM7 in each stomach increases to the highest abdomen(1.83%),and the proportion of SR1 in rumen and reticulum is SR1.1.78%and 1.76%,respectively,and much higher than omasum and abomasum;The percentage of Proteobacteria in red deer's abomasum is the highest(12.26%),the second highest in the omasum stomach(3.86%),and the proportion of Actinomycota(12.05%)is also higher than in other stomachs;ln addition to the advantages of roe deer and phylum Proteobacteria(accounting for 1.21%)in the abomasum,the proportion of other bacteria in the stomach gates are low.The F/B ratios in stomach microbiome,reindeer,white-lipped deer,and roe deer are much higher than the sika deer and red deer.This means that sika deer and red deer are less capable of acquiring fat than reindeer,white-lipped deer,and roe deer,while their ability to obtain polysaccharides in plants is higher than reindeer,white-lipped deer,and roe deer.172 genus were found in sika deer,reindeer,white-lipped deer,red deer,and roe deer.There are 13 genus with an average abundance above 1%,and 58 genus with an abundance above 0.1%.According to the abundance,the genus is Prevotella(10.10%),Succiniclasticum(3.40%),Ruminococcus(3.00%),Coprococcus(1.50%),Butyrivibrio(1.20%).Sika deer,reindeer,white-lipped deer,red deer,and roe reed peculiar bacteria in the stomach are 5,6,3,4,and 10.Sika deer have higher bacterial flora diversity and abundance than other deer in the same stomach.The diversity of bacterial flora in the same stomach of different deer is higher in the rumen and reticulum of sika deer than in the other;the diversity is highest in the omasum of white-lipped deer and sika deer;the most diverse in the abomasum of roe deer.The composition of the bacterial flora of the fore stomach of the five species of cervidae is similar,and the abundance and number of bacteria are higher than abomasum,but some special bacteria reach the highest in the abomasum.This may be because the fore stomach structure is more suitable for flora growth.Different gastric functions are different,and different oxygen content and physiological effects may also cause this phenomenon.In this study,no reported bacteria group in the reindeer stomach that can break down lichen poisonous usnic acid in the Arctic region has been reported in the reindeer stomach.This indicates that there is a difference between the microbial flora in the reindeer stomach that decomposes lichenic acid and the reindeer in the Arctic.2.ArchaeaTwo phylum were found in sika deer,reindeer,white-lipped deer,red deer,and roe deer.Our research found that the proportion of Euryarchaeota is as high as 99%,which is the absolute dominant species in deer stomach.Except for the dominant phylum,among the four stomachs of rumen,reticulum,omasum,and abomasum,Actinobacteria(0.01%)of sika deer petal abdomen and red deer abomasum are higher than roe deer.Bacteroidetes(0.1%)in omasum stomach and Firmicutes(both above 0.01%)were higher in than other deer.Proteobacteria(0.01%)in rumen in reindeer and roe reed,and Crenarchaeota(0.2%)in sika deer's abomasum were higher than other deer.Proteobacteria were not found in the stomachs of sika deer and red deer,and Actinomycetes were not found in reindeer and white-lipped deer.49 genus were found in Sika deer,reindeer,white-lipped deer,red deer,and pheasant.There are four genus had an average abundance higher than 1%.Methanobrevibacter has the highest abundance and is an absolute dominant genus.The average abundance in all samples over 93%.The average abundance of the four stomachs in red deer was the highest(98.51%),97.39%in the reindeer stomach,96.94%in the white-lipped deer stomach,96.8%in the sika deer stomach,and the lowest in roe deer is only 79.30%.More than 1%of the genus:Methanoplanus found in the stomachs of reindeer,white-lipped deer,and red deer,the abundance of red deer's abdomen accounted for more than 1%,which was 1.41%;Methanimicrococcus(1.44%)in red deer stomach was not found in the roe deer stomach;Methanosphaera in roe deer has the highest proportion in rumen and reticulum(32.74%,24.85%),and the difference is significant.The other genus less than 0.1%,Halorubrum is a unique genus of sika deer rumen;Staphylococcus and SAGMEG-1 are unique genus of roe deer abomasum and reticulum.Methanoculleus,Natronomonas,and Thermogymnomonas are only found in the roe deer and sika deer.Nitrosopumilus was found only in the sika deer's abomasum and reindeer reticulum.CandidatusNitrososphaera,Methanobacterium,vadinCA11 were not found in the stomach of white-lipped deer.Methanolinea is not found only in the roe deer stomach;Olsenella is not found only in the stomachs of white-lipped deer and reindeer.Candidatus Azobacteroides has the highest abundance in sika deer abomasum,higher than other stomachs;Methanosaeta is distributed in the stomachs of five deer,but abundance in red deer abomasum is higher than other stomachs;Methanosarcina is evenly distributed in the four stomachs of sika deer and roe deer are higher than that of other deer.The abundance and diversity of the archaeal flora in the stomach,the white-lipped deer is higher than the sika deer,reindeer,and red deer,and significantly higher than the roe deer.It was found in the heatmap that the archaeal flora in stomach of roe deer was farthest from that of other typical deer,followed by reindeer,and the sika deer and white-lipped deer were most similar.The stomach abundance and diversity of the white-lipped deer in the four stomachs were higher than others.The sika deer is slightly more diverse than the white-lipped deer only in the abomasum.The abundance and diversity of the four stomach archaea flora of roe deer were significantly lower than other deer species.This means that the methane production is the smallest and the white-lipped deer the highest.The analysis of the common OTU found that most of the methanogenic archaea accounted for the highest abundance in the reticulum,confirming the point that the deer's methane is mainly produced in the reticulum.The lowest methanogenic flora inroedeer may be related to the smallest stomach volume.3.FungiFifteen phylum were found in sika deer,reindeer,white-lipped deer,red deer,and roe deer,belong to 10,10,9,11,and 11 phylum.Ascomycota is the absolute dominant phylum.Sika deer omasum,reindeer omasum and abomasum,white-lipped deer reticulum,red deer reticulum,and roe deer rumen,roe deer have the highest abundance,accounting for 74.77%,50.01%,50.01%,93.78%,50.75%,84.2%.There are 14 higher than 1%,Basidiomycota has the highest abundance in sika deer omasum,reindeer reticulum,white-lipped deer abomasum,red deer omasum,and roe deer abomasum(5.97%,6.39%,2.96%,10.54%,4.14%).The abundance of Mucoromycota(1.29%)in the reindeer reticulum accounted for more than 1%.Other phylum less than 1%,Aphelidiomycota is only found in the reindeer rumen.Basidiobolomycota and Kickxellomycota are unique in the red deer rumen and abomasum;Blastocladiomycota is only found in the sika deer abomasum and roe deer rumen;Entorrhizomycota and Zoogoagomycota are peculiar phylums of roe deer rumen and reticulum;Chytridiomycota average abundance of in the stomachs of sika deer and reindeer is higher than that of other deer;Glomeromycota is evenly distributed in the four stomachs of sika deer higher than that of other deer;Glomeromycota,Mortierellomycota,Olpidiomycota,and Rozellomycota were distributed in the four stomachs of the five species of deer.632 genus were found in five species of deer,which 23 were more than 1%,sika deer,reindeer,white-lipped deer,red deer,and roe deer(6,6,2,8 and 6 genus).There are extremely significant differences in the distribution of fungi in the deer's stomach.In the sika deer's stomach,Candida(6.09%)accounted for the highest proportion in rumen;Issatchenkia(18.04%)and Pichia(16.41%)were absolute dominant fungi have the highest proportion;Humicola(1.25%)and Hannaella(1.57%)in omasum,,higher than other stomachs;Diutina(3.02%)in abomasum,higher than other deer.The abundance of Evernia,Penicillium,Talaromyces,andThelebolus in the reindeer's stomach is greater than 1%.Xenopolyscytalum is a reindeer-specific genus,and its abundance in rumen and reticulum accounts for more than 1%(1.44%and 1.39%).Cutaneotrichosporon(1.98%)of reindeer reticulum,higher than other stomachs.Preussia(15.2%)in omasum is higher than other stomachs.There are two fungi in stomach of the white-lipped deer that are higher than 1%,Issatchenkia(27.12%)and Diutina(1.23%)in omasum are higher than other stomachs.Among the dominant genus of red deer stomach abundance more than 1%,Candida,Eleutheromyces,Ophiognomonia(6.16%,3.51%,1.74%)in rumen,higher than other stomachs;Septoria(20.83%)inreticulum,higher than other stomachs;Cutaneotrichosporon(1.34%)and Hannaella(1.58%)in omasum are higher than other stomachs;Papiliotrema(2.8%)and Trichosporon(1.84%)in abomasum are higher than other stomachs.Among the dominant genus of roe deer over 1%in the stomach,Mycophaerella(6.68%)in rumen is higher than other stomachs;Boeremia,Ramularia,and Preussia in reticulum accounted for 1.43%,8.2%,and 2.01%,higher than other stomachs;Ophiognomonia(5.43%)in the omasum is higher than other stomachs;Septoria is the absolute dominant species in the stomach,accounting for 43.65%in the abomasum,which is higher than other stomachs.Among the other genus with an abundance greater than 0.1%,Cetraria,Cladonia,Cryptodiscus,Evernia,Hormonema,Kalmusia,Periconiella,Podosphaera,Scytalidium and Xenochalara are endemic in the reindeer stomach;Cyberlindnera was only find in sika deer stomach;Cyllamyces are endemic in white-lipped deer;Mycodiella,Peltaster is endemic in roe deer;Acremonium,Alternaria,Archaeorhizomyces,Didymella,Epicoccum,Lecanicillium,Mortierella,Penicillium,Plectosphaerella,Saitozyma,Simplicilium,and Tausonia are commonly found in the four stomachs of the five deer species.Fungi research shows that Ascomycota and Basidiomycota are the dominant flora in stomach of cervidae,but the composition of the fungi flora in the stomach is significantly different Our research suggests that the structural composition of the fungi flora is most significantly affected by feeding preferences and habitats.4.Metabolic pathwayIn the KEGG metabolic pathway of five species of Cervidae,bacterial flora has the highest abundance of amino acid and carbohydrate metabolism pathways;Archaeal flora is most abundant in carbohydrate metabolism pathways,followed by amino acid metabolism pathways;fungi fatty acids and lipids have higher biosynthetic pathways.Some dominant bacterial flora in the deer's stomach produce volatile fatty acids through metabolic pathways to regulate growth of epigastric epithelium.Prevotella,Succiniclasticum,Ruminococcus,Butyrivibrio,Clostridium may affect the development of epigastric epithelium maximum.5.CoevolutionA comparative study of the bacteria,archaea and fungi in the stomach shows that although the fungi have significant interspecies differences in the stomachs of the five deer families,their structural composition is most significantly affected by their feeding preferences and habitat.Roe deer having significant differences in microbial community structure and function of genes in terms of metabolic pathways of bacteria and archaea.The reindeer difference is slightly larger than the other;Sika deer's stomach bacteria and abundance are higher than others;White-lipped deer are more abundant in archaeal flora than others.In general,the sika deer,red deer,and white-lipped deer have higher similarities in functional genes of bacterial and archaeal flora structure and metabolic pathways.The degree of difference between the bacterial and archaea community structure and the five species of cervidae showed a good correspondence.This phenomenon confirms to a certain extent that the community structure of bacteria and archaea is closely related to the coevolution of the cervidae.