Open Access
Bird, Laurence Robert
Graduate Program:
Doctor of Philosophy
Document Type:
Date of Defense:
September 26, 2016
Committee Members:
  • Katherine H. Freeman , Dissertation Advisor
  • Katherine H. Freeman , Committee Chair
  • Jennifer L. Macalady, Committee Member
  • Christopher H. House, Committee Member
  • Squire J Booker, Outside Member
  • Methanotrophs
  • reverse methanogenesis
  • ANME
  • Coenzyme F430
  • carbon isotopes
  • Reverse methanogenesis
This dissertation explores the molecular and isotopic signatures of methanotrophic Archaea and the molecular signatures of cyanobacteria in low oxygen environments. Archaeal ANerobic MEthaneotrophs (ANME) oxidize methane in anoxic sediment, and prevent methane, a potent greenhouse gas from reaching the atmosphere. This process is hypothesized to take place via the reversal of methanogenesis based on culture and genetic evidence. Coenzyme F430 is a tetrapyrrole used in the last step of methanogenesis, and likely enables the first step in reverse methanogenesis. Therefore, the presence and concentration of F430 in association with AOM serves as a test for the reverse methanogenesis pathway in sediment. In chapter 2, F430 was extracted, quantified, and isotopically analyzed in methanotrophic sediment from Hydrate Ridge and the Santa Monica Basin (west coast U.S.A). The greatest amounts of F430 were recovered where sulfide, sulfate, and methane concentration profiles indicate the greatest AOM activity in the sediment. These sediment horizons also contained the highest ANME-2 aggregate counts. F430 was found to be isotopically distinct from methane and archaeal lipids, but similar to dissolved inorganic carbon (DIC). In the Hydrate Ridge and Santa Monica sediment F430 was ~60‰ enriched in 13C relative to archaeol lipids. In chapter 3, the dual assimilation of methane and DIC is explored with a series of stable isotope labeling experiments using sediment from Hydrate Ridge and the Santa Monica Basin. In experiments using Hydrate Ridge sediments, we observed the 13C label from DIC assimilated into archaeol, while in experiments using Santa Monica Basin sediment the 13C labeled from DIC and methane was assimilated into both F430 and lipids. The amount of DIC assimilated into F430 and lipids ranged from ~50% to 100%, with between 0% to 20% of carbon coming from methane. Due to the amount of labeled methane that is oxidized to DIC we cannot be sure if methane is directly assimilated or first oxidized to DIC. Coenzyme F430 was also only recovered from experiments where methane was added to the headspace, strengthening the link between F430 and methanotrophy. Little Salt Springs is a sinkhole in Florida where a red biofilm in the euxinic water column produces large amounts of bacterialhopanetetrol (BHT), 2-methyl bacterialhopanetetrol (2-MeBHT) and 2-methyl anhydrobacterialhopanetetrol (2-MeAnhydroBHT). The amount of each BHT produced varies seasonally and between years, with the geochemical cause of this variability unknown. In chapter 4, a red cyanobacteria isolated from this biofilm was cultured under a number of different geochemical conditions in an attempt to identify possible causes for variability in bacteriohopanepolyols (BHP) production. No single geochemical control was identified as amounts of BHT and 2-MeAnhydroBHT were similar in all experiments. Future experiments should explore what effects oxygen concentration, fixed nitrogen species, trace metals, microbial community and combinations of different conditions have on BHP production.