生长环境中微生物丰富程度对仔猪免疫系统的调控机制研究

The human hygiene hypothesis believes that low exposure to pathogens early in life can increase the risk for immune-mediated diseases, suggesting that the richness of microbiome in the growing environment can affect the development of the animal's early immune system, while the regulatory mechanism is still unclear. In this study, newborn piglets will be used as experimental animals, and firstly they will be controlled to grow in four different growing environments (sterile captivity, normal captivity, arch soil captivity, arch soil free-range), regularly collect feces, blood and intestines samples, and analyze the effects of different growth environments on immune indicators and intestinal morphology. Using 16s sequencing to systematically analyze the regulation and mechanism of different growth environments on early colonization of gut microbiome; using macrogenomics combined with macrotranscriptome to further verify the relationship between microbial community structure and function in piglets under different growth environments, and identify the relationship between environment and microbial function; finally, the immune index and microbial indicators are integrated through ADONIS analysis and multi-factor correlation analysis. This study will reveal the regulation and mechanism of the richness of environment microbiome in the early colonization of pigs intestinal microbes and early development of the immune system, meanwhile it can screen out the some microbes associated with the development of the early immune system and lay the foundation for improving the disease resistance of pigs by using fecal transplantation in the future and give a new basis for the immunological mechanism of the hygiene hypothesis.

人类“卫生假说”认为生命早期与病原菌的接触越少，生命后期出现免疫介导疾病的风险就越大，暗示生长环境中微生物的丰富程度可影响动物早期免疫系统的发育，但调控机制尚不清楚。本研究以仔猪为实验动物，控制其在不同微生物丰富程度的环境（无菌圈养、正常圈养、拱土圈养、拱土散养）中生长，规定时间采集粪便、血液和肠道样本，分析不同生长环境对免疫指标、肠道形态等的影响；利用16s测序，分析不同生长环境对肠道菌群早期定植的调控作用和机制；利用宏基因组结合宏转录组进一步验证不同生长环境下幼年猪微生物群落结构与功能的关系，明确环境与菌群功能之间的关联性，最后通过ADONIS和多因子关联分析将免疫指标和微生物指标进行整合，揭示环境中微生物的丰富程度和猪肠道微生物早期定植、免疫系统早期发育的调控机制，筛选出与早期免疫系统发育相关的菌群，为今后利用粪便移植提高猪抗病力奠定基础，同时为“卫生假说”的免疫学机制提供新的依据。

卫生假说是指如果机体在生命早期通过“不卫生接触”的感染越少，日后发展出过敏性疾病的机率就会越高。表明生长环境中微生物的丰富程度，对哺乳动物免疫系统发育具有一定的调节作用。围绕生长环境中微生物的丰富程度与动物早期肠道菌群建立、免疫能力提升的关系，项目以梅山仔猪为实验动物，分析环境中微生物丰富程度对仔猪免疫系统的调控机制研究。细胞因子检测显示不同组别组之间细胞免疫存在显著差异。此外，我们对梅山猪的十二指肠组织进行代谢组学和转录组学分析，鉴定出差异表达基因（DEG）、差异累积代谢产物（DAM）和差异信号通路。两组共获得1113个DEG、577个阳性（POS）DAM和372个阴性（NEG）DAM。随后，我们构建了mRNAs和与免疫应答相关的代谢产物（如9-顺式-瑞甲酸、（9Z，11E）-（13S）-13-羟基十八碳-9,11-二烯酸、（10E，12Z）-（9S）-9-羟基十八烷-10,12-二烯酸等）相互作用网络。功能富集评估表明，基因代谢产物主要参与一些免疫途径，包括IgA产生的肠道免疫网络和PPAR信号传导。同时根据筛选出的差异菌株，研制出一种猪用发酵饲料，能够提高育肥猪生长性能、增强机体免疫力、改善血液生化指标。本研究结果加强了我们对梅山猪在不同环境下激活的mRNA和代谢产物的理解，并将为未来探索“卫生假说”的免疫机制提供新的视角和方向。