EVIDENCE FOR DISSIMILATORY NITRATE REDUCTION TO AMMONIA BY AQUEOUS BIOFILMS IN THE SULFIDIC CAVES OF FRASASSI, ITALY

Open Access
Author:
Labrado, Amanda Leane
Graduate Program:
Geosciences
Degree:
Master of Science
Document Type:
Master Thesis
Date of Defense:
November 14, 2016
Committee Members:
  • Dr. Jennifer Macalady , Thesis Advisor
  • Dr. Katherine Freeman , Committee Member
  • Dr. Christopher House , Committee Member
Keywords:
  • Metagenomics
  • Elemental sulfur
  • Biofilms
Abstract:
Low-diversity microbial communities in Frasassi offer a promising model system for studying biogeochemical cycling in chemosynthetically fueled, low-oxygen, sulfidic environments. Microbial sulfur oxidation occurs in Frasassi as part of a larger sulfur cycle that, above the water table, stimulates cave formation through the production of sulfuric acid. Sulfur-oxidizing microbes commonly use either oxygen or nitrate as electron acceptors. Metagenomic studies of Frasassi biofilms revealed that complete sulfur oxidation pathway and some nitrate reduction genes exist within the microbial community (Hamilton, et al 2015). There are two known dissimilatory nitrate reduction pathways: denitrification and dissimilatory nitrate reduction to ammonia (DNRA). In Frasassi, H2S (100-800 μM) and Sº (20-80% dry mass of biofilms) are available in high concentrations, whereas oxygen (< 2-60 uM) and nitrate (< 0.06 μM) concentrations are much lower. Therefore we expect to find evidence of DNRA as an active nitrate reduction pathway, following the equation: H+ + H2O + NO3- + HS- → NH4+ +SO42- ΔG°= -448 kJ/molrxn Sulfur oxidation would drive the need for nitrate and create large amounts of energy, possibly explaining why nitrate is generally found below detection limit. Using previously binned metagenomic data from Frasassi biofilm sample PC08-64, I searched for homology with functional genes related to sulfur and nitrogen cycling. Genes necessary for DNRA are present in Bin 14, which is almost complete based on the presence of a full complement of amino acid tRNA synthetase genes. Bin 14 also contained a 16S rRNA gene that was closely related to Metallibacterium of the Xanthomonadaceae family. This genus has not been investigated at Frasassi before. Although previous FISH surveys have not detected large unidentified populations, the presence of a complete or nearly complete Metallibacterium genome bin suggests that it is abundant, a hypothesis that could be tested in future work using FISH and by further analyzing EMIRGE reconstructed 16S rRNA data.