Ellis Joshua T, Tramp Cody, Sims Ronald C, Miller Charles D
Department of Biological Engineering, Utah State University, 4105 Old Main Hill, Logan, UT 84322, USA.
ISRN Microbiol. 2012 Jun 21;2012:753892. doi: 10.5402/2012/753892. Print 2012.
The microbial diversity and metabolic potential of a methanogenic consortium residing in a 3785-liter anaerobic digester, fed with wastewater algae, was analyzed using 454 pyrosequencing technology. DNA was extracted from anaerobic sludge material and used in metagenomic analysis through PCR amplification of the methyl-coenzyme M reductase α subunit (mcrA) gene using primer sets ML, MCR, and ME. The majority of annotated mcrA sequences were assigned taxonomically to the genera Methanosaeta in the order Methanosarcinales. Methanogens from the genus Methanosaeta are obligate acetotrophs, suggesting this genus plays a dominant role in methane production from the analyzed fermentation sample. Numerous analyzed sequences within the algae fed anaerobic digester were unclassified and could not be assigned taxonomically. Relative amplicon frequencies were determined for each primer set to determine the utility of each in pyrosequencing. Primer sets ML and MCR performed better quantitatively (representing the large majority of analyzed sequences) than primer set ME. However, each of these primer sets was shown to provide a quantitatively unique community structure, and thus they are of equal importance in mcrA metagenomic analysis.
利用454焦磷酸测序技术，分析了一个3785升厌氧消化器中以废水藻类为食的产甲烷菌联合体的微生物多样性和代谢潜力。从厌氧污泥材料中提取DNA，并通过使用引物组ML、MCR和ME对甲基辅酶M还原酶α亚基（mcrA）基因进行PCR扩增，将其用于宏基因组分析。大多数注释的mcrA序列在分类学上被归类到甲烷八叠球菌目中的甲烷鬃毛菌属。甲烷鬃毛菌属的产甲烷菌是专性乙酸营养型，这表明该属在分析的发酵样品的甲烷产生中起主导作用。在以藻类为食的厌氧消化器中，许多分析序列未分类，无法在分类学上进行归类。确定每个引物组的相对扩增子频率，以确定每个引物组在焦磷酸测序中的效用。引物组ML和MCR在定量方面表现更好（代表了绝大多数分析序列），优于引物组ME。然而，这些引物组中的每一个都显示出提供了一个定量上独特的群落结构，因此它们在mcrA宏基因组分析中具有同等重要性。
