Anaerobic consumers of monosaccharides in a moderately acidic fen

Abstract

16S rRNA-based stable isotope probing identified active xylose- and glucose-fermenting Bacteria and active Archaea, including methanogens, in anoxic slurries of material obtained from a moderately acidic, CH(4)-emitting fen. Xylose and glucose were converted to fatty acids, CO(2), H(2), and CH(4) under moderately acidic, anoxic conditions, indicating that the fen harbors moderately acid-tolerant xylose- and glucose-using fermenters, as well as moderately acid-tolerant methanogens. Organisms of the families Acidaminococcaceae, Aeromonadaceae, Clostridiaceae, Enterobacteriaceae, and Pseudomonadaceae and the order Actinomycetales, including hitherto unknown organisms, utilized xylose- or glucose-derived carbon, suggesting that highly diverse facultative aerobes and obligate anaerobes contribute to the flow of carbon in the fen under anoxic conditions. Uncultured Euryarchaeota (i.e., Methanosarcinaceae and Methanobacteriaceae) and Crenarchaeota species were identified by 16S rRNA analysis of anoxic slurries, demonstrating that the acidic fen harbors novel methanogens and Crenarchaeota organisms capable of anaerobiosis. Fermentation-derived molecules are conceived to be the primary drivers of methanogenesis when electron acceptors other than CO(2) are absent, and the collective findings of this study indicate that fen soils harbor diverse, acid-tolerant, and novel xylose-utilizing as well as glucose-utilizing facultative aerobes and obligate anaerobes that form trophic links to novel moderately acid-tolerant methanogens.

FIG. 1.

Effects of supplemental xylose (A1) and glucose (B1) on the production of organic acids (A2 and B2) and gases (A3 and B3) by anoxic fen slurries (n = 3). Concentrations were corrected with values obtained from control slurries (i.e., unsupplemented anoxic slurries). Gases are given in mM, i.e., mmol produced per liter of liquid phase. Arrows indicate the addition of monosaccharides. Dotted lines indicate early and late sampling time points for RNA gradients.

FIG. 2.

DGGE profile of fractions 4 to 11 of bacterial RNA gradients obtained from the [¹³C]xylose experiment at day 13. Numbers at the top of the lanes correspond to fractions 4 to 11. Arrows identify bands that occur only in the “heavy” fractions (4 and 5) or are more intensive than in the “light” fractions (6-11).

FIG. 3.

T-RFLP profiles of bacterial cDNA of “heavy” fraction 4 and “light” fraction 10 obtained from the [¹³C]xylose experiment (A) or the [¹³C]glucose experiment (B) at days 2 (A1), 4 (B1), and 13 (A2 and B2).

FIG. 4.

Consensus phylogenetic tree of bacterial 16S rRNA sequences obtained in this study (bold). Accession numbers are provided after sequence identifiers, and the relative abundance levels (percentages) of OTUs in gene libraries are given in parentheses. Codes: X4 and X13, sequences retrieved from “heavy” fractions at days 4 and 13, respectively, of the [¹³C]xylose experiment; G2 and G13, sequences retrieved from “heavy” fractions at days 2 and 13, respectively, of the [¹³C]glucose experiment; B, sequences obtained by direct 16S rRNA analysis of fen soil; n.d., not detected. Scale bar represents 10% sequence dissimilarity. The outgroup is Thermocrinis albus (accession no. AJ278895).

FIG. 5.

Consensus phylogenetic tree of archaeal 16S rRNA sequences (bold) obtained at day 13 of the [¹³C]xylose experiment. Accession numbers are provided after sequence identifiers, and relative abundance levels (percentages) of OTUs in gene library X13 are given in parentheses. Scale bar represents 10% sequence dissimilarity. The outgroup is Thermoplasma acidophilum (accession no. M20822).