Investigation On The Diversity And Phylogenetic Of Methanogens And The Relationship Between Methanogenesis And Acetogenesis In The Hindgut Of Pigs

Methanogens play an important role in the hindgut of monogastric animals,and they catalyze the transfer of hydrogen and carbondioxide into methane, the maintenance of a low partial pressure of hydrogen by methanogenesis has a profound influence on the fermentation of organic materials.However,to our knowledge, analysis of the 16S rRNA genes of the methanogens from swine feces have not been reported.Therefore,in the present study,the 16S rRNA gene clone libraries were constructed to investigate the diversity of methanogens in the feces of pigs based on the16S rRNA gene method,and PCR-DGGE was used to study the changes of methanogens community in colon of piglets from seven days to 35 days afte birth; finally,the relationship between methanogenesis and acetogenesis in the hindgut of piglets in vitro fermentation was investigated using the batch culture techniques in vitro. This dissertation was described in the following three sections.1 Phylogenetic analysis of methanogens from the pig fecesOne methanogenic 16S rRNA gene library was constructed from feces of pigs to reveal the methanogen population compositions using methanogen-specific primers Met86F/Met1340R.Seventy-three clones were randomly picked up from the library. Sequence examination of these clones revealed 10 different phylotypes,The phylogenetic analysis indicated that archaea found in the libraries were all clustered into the Euryarchaeota.The data from the tree showed that those sequences belonged to three monophyletic groups.Phylotypes LGM2 and LGM7 grouped within the Methanobacteriales. Phylotypes LGM8, LGM6,LGM4 and LGM9 grouped within the Methanosphaera. Other phylotypes grouped together.The results show that the dominant methanogen in the feces of pigs was Methanobrevibacter sp.. The next were uncultured archaeon clone and Methanosphaera stadtmanae.2 PCR based DGGE analysis the methanogens community in the gut of pigletsMolecular methods based on the 16S rRNA gene were used to study methanogenic communities in gut of piglets from 7 to 35 (two weeks after weaning) days.The DGGE profiles showed the bands in DGGE prifle from the samples before weaning were very simple.In 35 days,some new predominant bands appeared.And the sequence analysis showed that these bands belong to the Methanobacteria and novel group of uncultivated Achaean. The results showed that the methanogens community is very stabel among the first 24 days after birth,and more diversity was appeared in 35 days, this indicated that the ages or diet may be cause for the flunction.3 Investigation on the relationship between methanogenesis and acetogenesis in the hindgut of pigs in vitro fermentationThis study was conducted to investigate the relationship between methane and acetate production in the hindgut of pigs.The batch cultures techniques were used to study the changes of the VFA concentration,methane production and gas production after BES (0,10,20 mM) addition.The result showed,as compared with the control group,the acetate and butyrate production increased significantly for 20mM group(P< 0.05);the acetate and butyrate production had a little increase for 10mM group(P>0.05).The results indicated that reductive acetogenesis may become a significant pathway of acetate formation in the absence of methanogenesis.