Quantification of Methanogenic Microbes Associated with Tropical Bromeliads: Influences on Function and Abundance

Plants within the family Bromeliaceae possess unique foliage arranged in compact rosettes capable of retaining water. This unusual freshwater environment allows for the growth of anaerobic methanogenic archaea, which are crucial producers of atmospheric methane, a potent greenhouse gas. This study sought to quantify the methane production capacity of the microbial community by measuring, via quantitative PCR, the methyl coenzyme M reductase (mcrA) gene, which encodes for an enzyme that catalyzes the final step in the production of methane. Quantitative PCR was performed on bromeliad tank DNA extracts, using previously designed primers for both mcrA and 16S rRNA (the latter of which was used to measure archaeal abundance). With the assumption that environment influences gene expression, mcrA copy numbers (which ranged from 1.6E6 to 7.1E7 copies per ng of extracted DNA) were compared to environmental variables, including plant height, tank pH, and photosynthetically active radiation (PAR) levels. A significant correlation was observed between mcrA gene copy number and plant height (P = 0.0009) suggesting that the methanogenic potential of the microbial communities increases with host plant height. This was confirmed by analysis of a series of clonal bromeliads, ranging from 2 to 7 cm tank depth, which only assayed positive for mcrA if larger than 4 cm tank depth. Analysis of 16S rRNA QPCR data revealed that archaeal microbial abundance ranged from 2.0E3 to 1.1E6 cells per ml of tank fluid, and generally exhibited a similar pattern of increasing cells with plant height. Finally, analysis of 4 artificial tanks paired with natural bromeliad plants, in the forest for 6-12 months, showed no methanogenic capacity except in two cases, in which the artificial tanks contained a large amount of organic detritus and had low pH values, suggesting that these particular factors may influence the suitability of the bromeliad habitat for methanogenic microbes. Future experimentation concerning actual gene expression will be conducted by measuring RNA levels (cDNA transcripts) of mcrA within the microbial community