纯种柯乐猪与巴×柯杂交猪肠道菌群结构的研究

Study on Intestinal Microflora Structure of Pure-Bred Kele Pigs and Berkshire×Kele Hybrid Pigs

本试验对纯种柯乐猪与巴×柯杂交猪肠道菌群结构特性及差异性进行了研究,旨在从猪肠道微生物的角度揭示纯种柯乐猪耐粗饲特性及巴×柯杂交猪对青绿饲料的适应能力,为后期微生物添加剂的设计、柯乐猪杂交利用及标准化养殖提供依据。纯种柯乐猪和巴×柯杂交猪在同等条件下养殖,达屠宰体重后(100 kg左右),随机各选3头屠宰,取十二指肠、空肠、回肠、结肠内容物,进行高通量测序和生物学信息分析。结果显示:纯种柯乐猪与巴×柯杂交猪各肠段Alpha多样性指数差异不显著(P>0.05);Beta多样性分析显示,2个品种猪大部分样品交叉聚集到一起;肠道菌群结构分析显示,纯种柯乐猪和巴×柯杂交猪小肠阶段以厚壁菌门(Firmicutes)、变形菌门(Proteobacteria)和放线菌门(Actinobacteria)为优势菌门,到了结肠阶段则是以拟杆菌门(Bacteroidetes)和厚壁菌门为优势菌门;纯种柯乐猪与巴×柯杂交猪结肠中有大量的纤维分解菌属,巴×柯杂交猪结肠中月形单胞菌属(Selenomonas)和脱硫弧菌属(Desulfovibrio)的相对丰度显著或极显著高于纯种柯乐猪(P<0.05或P<0.01),而消化球菌属(Peptococcus)、聚乙酸菌属(Acetitomaculum)、Leeia的相对丰度则显著低于纯种柯乐猪(P<0.05);LEfSe分析后发现纯种柯乐猪结肠中富集着大量的产短链脂肪酸菌,其中纤维降解菌有聚乙酸菌属、瘤胃球菌科的不可培养瘤胃细菌4C0d_17(uncultured_rumen_bacterium_4C0d_17)、Leeia、拟杆菌纲、拟杆菌目,与纤维降解菌起协同作用的菌群有考拉杆菌属(Phascolarctobacterium)、考拉杆菌属的uncultured_Veillonellaceae_bacterium;巴×柯杂交猪结肠中富集的纤维降解菌有月形单胞菌属、柔嫩梭菌(Clostridium leptum)。上述结果表明,在整个肠道菌群结构上,纯种柯乐猪和巴×柯杂交猪的优势菌在类别上非常接近,体现了环境相同效应,纯种柯乐猪和巴×柯杂交猪的肠道菌群结构相对稳定且相似度高;在属水平上,纯种柯乐猪和巴×柯杂交猪的结肠中纤维降解菌表现出差异,纯种柯乐猪结肠内纤维降解菌的比例较高,说明引入外源血对肠道菌群结构有一定的影响,巴×柯杂交猪对粗纤维的消化能力较弱于柯乐猪。

This experiment studied the structural characteristics and differences of intestinal microflora of pure-bred Kele pigs and Berkshire×Kele hybrid pigs,aiming to reveal the rough feed tolerance characteristics of Kele pigs and the ability to adapt to green feed of Berkshire×Kele hybrid pigs from the perspective of pig gut microbes,and to provide a basis for the design of microbial additives in the later stage,the cross-utilization of Kele pigs and standardized breeding.Pure-bred Kele pigs and Berkshire×Kele hybrid pigs were raised under the same conditions.After reaching the slaughter weight(about 100 kg),three pigs were randomly selected for slaughter,and the contents of the duodenum,jejunum,ileum and colon were taken to perform high-throughput sequencing and biological information analysis.The results showed that there were no significant differences in Alpha diversity indexes in each intestinal segment between Berkshire×Kele hybrid pigs and pure-bred Kele pigs(P>0.05).In Beta diversity analysis,most samples in the two breeds of pigs crossed and gathered together.Intestinal microflora structure analysis showed that,Firmicutes,Proteobacteria,Actinobacteria were the dominant bacterial phyla in the stage of small intestine of Berkshire×Kele hybrid pigs and pure-bred Kele pigs,and Bacteroidetes and Firmicutes were the dominant bacterial phyla in the stage of colon.There were a lot of cellulose decomposition microbes in the colon of Berkshire×Kele hybrid pigs and pure-bred Kele pigs.The relative abundances of Selenomonas and Desulfovibrio in the colon of Berkshire×Kele hybrid pigs were significantly higher than those of pure-bred Kele pigs(P<0.05 or P<0.01),and the relative abundances of Peptococcus,Acetitomaculum and Leeia were significantly lower than those of pure-bred Kele pigs(P<0.05).LEfSe analysis found that a lot of short chain fatty acid producing bacteria enriched in the pure-bred Kele pigs’colon.Among them,Acetitomaculum,uncultured_rumen_bacterium_4C0d_17,Leeia,Bacteroidia and Bacteroidales belonged to fiber-degrading bacteria,and some bacteria had synergistic effects with the fiber-degrading bacteria,they were Phascolarctobacterium and uncultured_Veillonellaceae_bacterium.The fiber-degrading bacteria such as Selenomonas and Clostridium leptum enriched in the Berkshire×Kele hybrid pigs’colon.Above results indicate that,in the structure of entire intestinal microflora,the type of dominant bacteria in pure-bred Kele pigs and Berkshire×Kele hybrid pigs is almost similar,which reflects the same environmental effects,and the structure of the intestinal microflora of pure-bred Kele pigs and Berkshire×Kele hybrid pigs is relatively stable and the similarity is high;at genus level,the fiber-degrading bacteria show some differences between pure-bred Kele pigs and Berkshire×Kele hybrid pigs,and fiber-degrading bacteria accounts for a higher proportion in cecal microflora of pure-bred Kele pigs,which indicates that the introduction of exogenous blood has a certain impact on the intestinal microflora,and Berkshire×Kele hybrid pigs are weaker in digesting crude fiber than pure-bred Kele pigs.