| Pig breeding plays a large proportion in China's aquaculture industry.Piglet diarrhea diseases are a major problem affecting pig production today.Piglet diarrhea is a syndrome of digestive dysfunction in piglets,which can cause secondary infections,resulting in decreased resistance of piglets.The mortality rate is one of the most serious disease groups in piglets,and it is also an important cause of piglet death,which seriously jeopardizes the development of the pig industry.In this study,we analyzed the bacterial mixed infection of diarrhea in piglets of different ages to find out how the structure of intestinal flora changes.The high-throughput sequencing method was used to sequence the 16 sr DNA V4 region of piglet intestinal flora using Illumina Mi Seq 2×250 platform.The sequencing samples were 14 and the amplified and sequenced region was the v4 variable region of bacterial 16 s r RNA.Through data preprocessing and removal of chimeras and non-specific amplified sequences,a total of 903,700 effective sequences were generated in this study,with an average of 56,481 sequences per sample(range: 47222-69896),the average length of the sequence was 252 bp,and the effective data rate for sequencing.It is 98.24%.The sequence of the sequence was randomly sampled,and the curve was obtained by using the number of sequences drawn and the number of OTUs they can represent,that is,the sparse curve.The results showed that the sequencing covered the microbial community was sufficient;the Venn chart shown showed the piglets of different ages.The most unique number of OTUs in the intestine is the number of OTUs in the 0-7 day old diarrhea piglets(A-diarrhea)is 1264,followed by the number of unique OTUs in the 8-14 day old diarrhea piglets(B-diarrhea).The number of OTUs shared by the A-diarrhea and B-diarrhea groups was 400,the number of OTUs in the A-diarrhea and A-normal groups was 300,and the number of OTUs in the B-diarrhea and B-normal groups was 84.This experiment was calculated using mothur software.Alpha diversity indicators,including Shannon Index,Simpson Index,ACE Index,Chao1 Index,Coverage,etc.,Shannon Index average is 2.00(range 0.32-3.22),Simpson Index average It is 0.32(range 0.09-0.87),the average ACE index is 692(range 96-3539),the average value of Chao1 index is 528(range 67-1972),the average coverage rate is 99.77%;clustering based on species abundance samples Graph4 analysis,Hierarchical cluatering analysis based on ? diversity distance matrix,using unweighted pair group method with arithmetic mean(UPGMA)algorithm to construct tree structure,and obtaining tree relationship form for visual analysis The distance between the samples was calculated.The results showed that the distance between A-diarrhea and B-diarrhea group was similar,which proved that the microbial structure was similar in piglet diarrhea;The classification analysis showed that there were 42 species of gates identified at the gate level,of which Firmicutes,Bacteroidetes and Proteobacteria accounted for more than 96% of the total sequence,and a total of 644 species were identified at the genus level,of which seven species accounted for 80 of the total sequence.More than 80%,the seven genera are: Lactobacillus,Escherichia/Shigella,Bacteroides,Parabacteroides,Streptococcus,Enterococcus,Fusobacterium.Piglets 0-7 days old,In the intestinal tract of diarrhea piglets,lactobacillus,Paralactobacillus Leisner,Parabacillus,enterococci,clostridium and prevotella showed a decreasing trend in terms of the bacterial community structure.Respectively by 39.00%,10.39%,9.28%,7.86%,4.01%,1.36% to37.70%,0.01%,8.14%,1.85%,0.33%,1.10%.The three bacteria of escherichia coli/shigella,streptococcus and actinobacillus showed an increasing trend in terms of the flora structure with the piglets of the control group,Increased from 2.41%,1.13% and0.04% to 14.12%,8.17% and 2.02% respectively.The eight dominant bacteria groups in the intestinal tract of piglets,lactobacillus,escherichia coli/shigella,veronicoccus,streptococcus,paraclactobacillus,enterococcus,bifidobacteria and clostridium,showed different degrees of decline compared with those in the intestinal tract of piglets with diarrhea.the intestinal flora structure showed different degrees of decline compared with that of the control group,By 70.14%,8.70%,4.81%,2.21%,1.40%,1.30%,1.14%,1.11%to 53.00%,4.68%,0.08%,0.13%,0.02%,0.30%,0.07%,0.72%,In the intestinal tract of diarrhea piglets,bacteroidetes,parabaceae,streptococcus digestibilis,amino acid coccus,delft bacteria and Paralactobacillus Leisner,These 6 dominant bacteria compared with the control group,there was an upward trend in the intestinal flora structure of piglets,Increased from 0.08% to11.40%and 0.06%to 8.11% and 0.49%to4.63% and0.02%to 4.13%ande 0.01%to 1.22% and 0.45% to1.16% respectively.The above high-throughput sequencing results show that diarrhea piglets and healthypiglets have significant changes in the dominant flora at the genus level.The changes in the structure and quantity of the intestinal flora of these piglets may be an important cause of diarrhea. |
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