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United States Department of Agriculture

Agricultural Research Service

Research Project: ANAEROBIC MICROBIOLOGICAL PROCESSES IN ANIMAL MANURE MANAGEMENT Title: Detection of methanogenic archaea in stored swine manure by direct mcrA PCR and sequence analysis

Authors
item Spence, Cheryl
item Whitehead, Terence
item Cotta, Michael

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: June 27, 2008
Publication Date: N/A

Technical Abstract: Storage of swine manure is associated with the production of a variety of odors and emissions which are products of anaerobic metabolism of the indigenous bacteria in the manure. These emissions can pose problems to the health and production efficiency of the animals, as well as the health and comfort of human workers. Consolidated storage of swine manure also leads to production of large quantities of methane, a greenhouse gas that contributes to global warming. Emissions from animal waste account for about 30% of the total U.S. methane emissions from agriculture. In the U.S., methane emissions from lagoons and manure storage pits are estimated to be over 40 Tg/year. Methane is a product of the anaerobic metabolism of methanogenic archaea. Little is known about the population of methanogens in stored swine manure and surprisingly, few methanogens have been isolated from this environment. This study was initiated to identify the methanogenic archaea population of stored swine manure using a culture-independent technique targeting a methanogen-specific functional gene, methyl coenzyme-M reductase subunit A (mcrA). Methyl coenzyme-M reductase catalyzes the final step of methane production and is ubiquitous in methanogens. Total DNA was isolated from stored swine manure from local swine facilities and used as a template for direct PCR. Amplification with a degenerate primer set flanking a variable region of the mcrA gene generated a PCR product between 464 and 491 bp. These fragments were used to generate a mcrA clone library, and 50 clones were selected at random for sequencing and comparative phylogenetic analysis. A predominant number of clones were from the Methanobacteriales and Methanococcales orders with similarities to Methanosphaera and Methanobrevibacter species. Many clones showed little similarity to any identified methanogens suggesting that these sequences represent novel, as yet unidentified methanogenic archaea. This study is the first mcrA-targeted analysis of the methanogenic archaea population of stored swine manure. The presence of previously unidentified methanogens suggests a new source of microbial diversity and new and novel archaea. Results from this study will be used to further develop molecular methods to monitor different populations of methanogens in this environment.

Last Modified: 10/23/2014
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