An ancient deletion of a small fragment of a gene alters the gut microbiota

The chimeric lineage in this study is experimental in pigs. Courtesy of Chen Congying

The composition of the intestinal flora varies from individual to individual and is closely related to health. Does an individual’s genetic variation affect the composition of his or her own gut flora? And how is it affected?

Due to the influence of multiple factors, many studies, especially in humans, are difficult to repeat. Scientists have published a number of controversial articles in Nature and its sub-journals. This is a major research hotspot and difficulty in the world.

To this end, Academician Huang Lusheng of the State Key Laboratory of Pig Genetic Improvement and Breeding Technology of Jiangxi Agricultural University independently designed and organized the team to construct a chimeric family for the genetic analysis of important economic traits of pigs for 12 years, and conducted a series of studies under strict and controllable conditions to solve the influence of host genetics on the composition of intestinal flora.

The results were recently published online in the journal Nature. This is the first time in the world that the host genome affects the composition of the intestinal flora in agricultural animals has been found to have causal gene mutations, which is of great reference significance for the cultivation of new breeds of pigs, as well as the study of the pathogenesis of human metabolic disorders, colorectal cancer and other related diseases from the interaction of host genes and intestinal flora. This is also the third research paper published in Nature in the field of international pig genome and genetic breeding in the past 20 years (the previous one was in 2012).

12 years is only to build an experimental population

The gut microbiome is an important part of the biological organism. Humans and mammalian individuals such as pigs inhabit a large number of microorganisms in the intestines, and their components have an important impact on the health of the host and the production of livestock meat, eggs and milk.

Gao Fu, an academician of the Chinese Academy of Sciences, introduced that there is an international academic controversy over whether host genetics affects and how it affects its own intestinal microbial composition. Because it is very difficult to identify causal genetic mutations that affect the intestinal flora inherited by the host.

Huang Lusheng, corresponding author of the paper, said that the digestive system, genetic composition and organ size of pigs are similar to humans, especially compared with mice, and their intestinal flora composition is more similar to humans. Therefore, it is of great reference significance to study the pathogenesis of human diseases and the regulation technology of microbiota by using pigs as model animals, starting from the interaction of intestinal flora and host genes.

However, the pig genome has 300 million base pairs, which is very large, and in order to locate the relevant genes from it, it is necessary to first build a diverse experimental population. This diversity is not only manifested in the genetic diversity and phenotypic diversity of pigs, but also in the diversity of their intestinal flora composition.

In order to establish this experimental group, Academician Huang Lusheng uniquely designed the world’s only domestic pig chimeric family that uses the reincarnation hybridization of four major Western pig breeds and four major Chinese (Asian) pig breeds, and conducted the project in the sixth and seventh generation groups of this chimeric family.

Chen Congying, co-corresponding author of the paper and professor of Jiangxi Agricultural University, told China Science News that hybridization is to allow the genomes of pigs with different genetic backgrounds to fully “hybridize” together and chimerize with each other, so that there are more genetic and phenotypic variations. By the time of hybridization to the sixth and seventh generations, the genome chimerism of the 8 archaeopteryx varieties was already complete and relatively stable, which was conducive to the development of subsequent experiments.

Pigs are generally one generation a year, and it is very difficult to obtain positive and negative hybrid offspring between different pig breeds designed at the beginning, “so it took us twelve years just to establish the experimental population.” Chen Congying said.

Alternative approaches are found to compensate for the limitations of traditional methods

However, since the composition of the intestinal microbiome is affected by multiple factors such as diet and health, to study the relationship between host genetic variation and the composition of the intestinal flora, it is necessary to conduct experiments under controlled conditions.

“Our experiments strictly control feed, feeding, health and environmental conditions.” Chen Congying said that the reason why previous studies are controversial is because the composition of human intestinal flora is often affected by a variety of external factors, and many research conclusions cannot be repeated. The use of pigs to study the intestinal flora can strictly control the influence of other external environmental factors.

Previous studies have also found that the microbiota structure of different parts of the intestine is not different, which makes the study of only stool samples limited.

The team determined stool samples from 1500 experimental individuals in two generations (F6 and F7) at 25 and 120 days after birth, and the intestinal flora composition of each individual’s three intestinal sites (ileum, cecum, feces) after slaughter on day 240.

“This method is more accurate and comprehensive, and can compensate for the limitations of stool samples.” Yang Hui, the first author of the paper and a young associate professor at the College of Biological Sciences and Engineering of Jiangxi Agricultural University, said that the heterogeneity of the composition of the microflora at different stages of development and intestinal parts of the same individual was found, which proved that the composition and abundance of the intestinal flora have a wide range of specificity and diversity of growth stages and intestinal sites.

Further studies have found heritability of intestinal flora composition and abundance.

The team identified 1050 and 955 bacterial classifications as heritable in two generations, with the highest heritability reaching 0.598. “Hereditism is also known as hereditary rate. The higher the heritability of a trait, the greater the expression pattern of the trait is affected by genetic factors and the less affected by environmental factors. Chen Congying explained.

Of these, 450 heritable bacterial classifications were repeatedly validated in both generations, and some of the heritable bacterial classifications were consistent with the findings in the human gut.

“This proves that the composition of the intestinal flora is heritable.” Chen Congying explained that although scientists have not yet fully understood the initial source and colonization of the intestinal flora, it is certain that not all bacteria can colonize in the intestines of individuals. “This is influenced by a variety of factors, but a lot has to do with different individuals having different genotypes, because genotypes affect the type, abundance, and number of colonizable bacteria in their own gut.”

The mutation 3.5 million years ago sends a strong signal

At this point, the solution to the puzzle has just begun. What bacteria do an individual’s genetic mutations affect and how do they affect the composition of their own gut flora?

The team first resequenced 1430 chimeric individuals from two generations to identify more than 30 million host genome variants. Genome-wide association analysis of the 8490 detected bacterial classifications was then performed, and a total of 1527 host genomic variant sites that significantly affected the abundance or presence or absence of 846 bacterial classifications were detected.

Among more than 30 million host gene mutation sites, they found that the association between ABO blood group genes and microgroups was very strong, so they decided to conduct an in-depth study of the ABO blood group gene system.

Results Identified a 2.3 kb deletion variant in the ABO blood group system of domestic pigs. “This gene fragment deletion variant directly leads to differences in the abundance of erysipelas filamentobacteriaceae-related bacteria in the intestines of domestic pigs.” Chen Congying said.

The team further studied and analyzed the molecular mechanism by which the deletion variant of 2.3 kb affects the abundance of erysipelas filaments associated with the family in the intestines of domestic pigs. They found that this gene deletion variant (individuals with blood type O) caused the N-acetylgalactamine transferase of the ABO protein to lose activity. Therefore, N-acetyl galactosamine cannot be added to the highly glycosylated mucin in the intestinal mucus, resulting in a decrease in the concentration of N-acetyl galactosamine in the intestines of blood type O individuals, thereby affecting the growth of bacteria with N-acetylgalactamine as the carbon source. The most important of these bacteria is Erysipelas.

Wu Jinyan, co-first author of the paper and Jiangxi Agricultural University, introduced that based on the ancestral eight domestic pig breeds of chimeric family groups, Asian wild boar, European wild boar, Sumatran wild boar, Philippine warthog, Indonesian Javan warthog and African common warthog, Traceability of the 2.3 kb deletion of the ABO gene found that the deletion mutation was present in the above animals, and the breakpoint sequence of the mutation was consistent in all samples, thus confirming that the 2.3 kb deletion of the ABO gene in the pig genus belonged to an ancient mutation that occurred 3.5 million years ago.

Gao Fu said that this is the first causal mutation of the host genome that affects the composition of the intestinal flora in agricultural animals (pigs, cattle, sheep, chickens, etc.) identified internationally.

Li Defa, an academician of the Chinese Academy of Engineering, pointed out that this is also the first research result published in nature in the field of animal husbandry in China. This achievement is of great reference significance for the development of new technologies for host genetics to regulate feed conversion efficiency and promote growth by influencing the intestinal flora, and to cultivate new varieties of grain-saving and fast-growing pigs.

The study is not only of great significance in terms of results, but also of great reference significance for further studying the interaction between host genes and intestinal flora, thereby promoting pig health, improving growth rate, feed utilization and reproduction. At the same time, since erysipelas filamentous bacteria are associated with metabolic disorders, cholesterol metabolism and colon/rectal cancer in humans, the results of this study also provide a reference for the pathogenesis of the above related diseases in humans in the future. (Source: China Science Daily Li Chen)

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