PCR-DGGE技术分析蛋鸡MHC基因对肠道细菌种群结构的影响

 【目的】研究蛋鸡MHC（Major Histocompatibility Complex，主要组织相容性复合物）基因对不同周龄和不同部位肠道细菌种群结构的影响。【方法】使用基于16S rDNA的PCR-DGGE技术，结合特异性和共性条带割胶回收DNA进行克隆和测序，对2，4，6和8周龄商品蛋鸡和15I5系蛋鸡嗉囊、十二指肠、空肠、回肠和盲肠内含物细菌群落的结构和多样性进行比较，鉴定8周龄蛋鸡部分特异性和共性群落成员。【结果】商品蛋鸡和15I5系蛋鸡盲肠细菌的多样性最高，其次是回肠和空肠，嗉囊和十二指肠的细菌多样性比较低，且随着蛋鸡周龄增加，各肠段细菌多样性呈上升趋势，MHC基因对细菌多样性无显著影响。DGGE图谱聚类分析表明，蛋鸡MHC基因对4、6周龄肠道细菌的影响最大，对2周龄影响最小;而且MHC基因对消化道前段细菌的影响小于对后段影响。序列分析发现，8周龄商品蛋鸡和15I5系蛋鸡均有大量的三得利乳杆菌（Lactobacillus suntoryeus），是蛋鸡消化道前段的优势细菌;商品蛋鸡空肠、回肠和盲肠中的特异性条带主要是Ochrobactrum spp，索氏梭菌（Clostridium sordellii），超巨巨单胞菌（Megamonas hypermegale）和大量不可培养的细菌;15I5系蛋鸡的特异性条带主要是不可培养的细菌。【结论】蛋鸡MHC基因可影响不同周龄、不同肠段细菌群落的结构。

Influence of MHC Genotype on the Bacterial Community in the Layer Gastrointestinal Tract Analyzed by PCR-DGGE

【Objective】 The purpose of this study was to investigate the influence of MHC(Major Histocompatibility Complex) gene on the predominant bacteria community in the layer gastrointestinal tract. 【Method】 The bacterial community and diversity in the crop, duodenal, jejunal, ileal and ceacal digesta at the age of 2, 4, 6 and 8 weeks of commercial layers and line 15I5 layers was studied by using 16S rDNA-based denaturing gradient gel electrophoresis (DGGE) and identification of some common and special bands in DGGE profile at 8 weeks. 【Result】 The highest level of diversity was found in cecum of commercial layers and line 15I5 layers, followed by ileum and jejunum, while the lowest levels of diversity were found in crop and duodenum. The diversity of bacteria increased with the age at every region of gastrointestinal (GI) tract. MHC gene didn’t influence the diversity of bacteria. The dendrograms revealed that MHC gene heavily influenced the bacteria composition at the age of 4 weeks and 6 weeks, followed by 8 weeks, while the highest level of similarity was found in 2 weeks. At the same time significant differences were observed between two lines in the lower GI tract than that in the upper GI tract. In addition, some amplicons in the DGGE pattern of two layer lines were identified by cloning and sequence analysis. Lactobacillus suntoryeus were predominant in the upper GI tract of two layer lines. The special bacterial in jejunal, ileal and ceacal digesta of commercial layers were Ochrobactrum spp，Clostridium sordellii，Megamonas hypermegale and plenty of uncultured bacterial. Most of the special bacteria in 15I5 layers were uncultured bacteria. 【Conclusion】 The results of this study indicate that MHC gene influence the composition of the dominant bacterial population in every region of GI tract of layers at different ages.