Tracing Intestinal Bacteria Of Newborn Piglets And Their Interaction With Viruses

Early life is a critical window period for the colonization of intestinal microbiota in animals.The colonized gut microbiota during this early stage have an important influence on the growth of the animals and the development of the immune system.In this theses,16 S r RNA sequencing and bioinformatics technology were used to analyze the colonization of gut bacterial community in the newborn piglets.Then the relative contribution of different microbial sources to the colonization of intestinal bacteria in piglets was analyzed by systematically collecting microorganism from sows and the pig pens.Then,the colonization of virus in the gut of piglets were studied by viral metagenomics.Finally,the results of colonization of intestinal bacteria and virus communities were integrated to analyze the interaction between intestinal bacteria and viruses in newborn piglets.The major results are as follow:Part one: Colonization of intestinal bacterial community in newborn pigletsTo study the colonization of intestinal bacterial community in newborn piglets,fecal samples from 60 head of newborn piglets from the same breeding house in a farm in nine time points after birth(0,1,3,5,7,14,21,24 and 28 d)were collected and analyzed using 16 S r RNA sequencing analysis to determine the of intestinal bacteria community structure changes.The results showed that the diversity of intestinal bacterial community in newborn piglets decreased initially and then increased gradually.Facultative anaerobes in the intestinal tract are preferentially propagated and then gradually replaced by obligate anaerobes.Intestinal flora structure change with piglet age increase remarkably.Relative abundance of bacteria that can efficiently utilize oligosaccharides from breast milk such as Bacteroides,Streptococcus and Clostridium reduced significantly after weaning.However,the relative abundance of bacteria that can efficiently utilize plant-derived polysaccharide such as Prevotella,Ruminococcus and Parabacteroides increased significantly,and enrichment of plant polysaccharide source metabolism related gene pathways.The maturity of intestinal bacteria increased gradually with the increase of age,and the difference of microflora structure between individuals decreased gradually.All these indicated that the colonization sequence and composition structure evolution of intestinal bacterial community in newborn piglets were selected by intestinal microenvironment and feeding structure.Part two: Tracing the intestinal bacterial community of newborn pigletsIn order to study the main source of intestinal bacteria in piglets,a total of 20 sows and 60 newborn piglets delivered at the same time were selected to analyze the relative contribution of microorganisms from the sow(vagina,feces,areola skin,milk)and housing environment(air,water,slatted floor)using 16 S r RNA sequencing combined with bioinformatics technology.The results showed that the similarity of microbiota between piglets and vagina of sows decreased with age,but the similarity between the gut microbiota of piglet and the intestinal microbiota of sow increased with age.Source Tracker analysis found that the sow vagina is the main source of aerobic and facultative anaerobic bacteria in piglets,and the relative contribution rate to the intestinal microbiota of 0?3 days of piglets is as high as 69.00%?86.94%,then gradually reduced to 0.28%.At the same time,the relative contribution of the sow's intestine and slatted floor gradually increased to 62.12% and 33.75%,respectively at day 28,and became the main source of piglet intestinal bacteria.These results suggest that most of the microorganism from vagina of sows only temporarily colonized in the intestine of piglets,while the microorganism in the intestine of sows has a longer term effect on the colonization of piglet intestinal bacterial communities.Part three: Colonization of viruses in newborn pigletTo study the composition and structure of the intestinal virus community in neonatal piglets,the DNA of virus community in fecal samples of piglets at 5 different time points(3,7,14,21 and 28 d)were analyzed using virus metagenomics techniques.The results showed that phage was predominant in the intestinal DNA virus community of piglets,accounting for 83.93% of the total virus sequence,and it was mainly composed of Siphoviridae(34.47±5.56%),Microviridae(19.68±9.62%)and Myoviridae(16.01±3.01%).The richness and diversity of intestinal phage decreased with the age of piglets,while the richness and diversity of eukaryotic DNA virus increased gradually.The structure of intestinal virus community changed significantly with the age,and different samples had relative uniqueness.The functional composition of intestinal DNA virus community of piglets of different age varied significantly,mainly including signal transduction mechanisms(10.43±3.56%),chromatin structure and dynamics(10.25±2.36%)and defense mechanisms(9.28±4.74%).Part four: interaction between intestinal bacteria and viruses in newborn pigletsIn order to study the interaction between the intestinal bacterial and viral community in newborn piglets,the 16 S r RNA sequencing data and the virus metagenomics sequencing data were analyzed jointly.The results showed that there was a significant negative correlation between intestinal phage and bacterial community diversity in piglets,in which the diversity of intestinal bacterial community increased gradually,while the diversity of phage decreased gradually with age.There is a complex interaction between intestinal phage and bacteria in piglets,and the interaction is mainly positive at the early stage,and gradually negative with the increase of age.In conclusion:(1)The intestinal microbial ecosystem of newborn piglets rapidly succession with the increase of age,the diversity of gut microbiota decreased first and then increased,the maturity and similarity of gut microbiota gradually increased,the facultative anaerobe colonized in priority is gradually replaced by obligate anaerobe under the influence of intestinal microenvironment,and the composition and function of intestinal microbial community tend to be stable.(2)The colonization of gut microbiota of piglets was mainly driven by vertical transfer of sow's microorganisms,and had the characteristics of bacterial source and time specificity.Intestinal microbiota of sow gradually replaced the priority colonization of the vaginal microbiota,and had stronger ecological adaptability and listing contribution in the intestinal tract of piglets.(3)The intestinal phage of newborn piglets driven the diversity of the virome,which was negatively correlated with the change of bacterial diversity.The structure of virome of piglets had obvious individual,and mainly composed of Siphoviridae,Microviridae and Myoviridae.