ESTABLISHING STROMATOLITE-BUILDING EUKARYOTIC BIOFILM IN A CONTROLLED LABORATORY SETTING

Eukaryotic biofilm dominated by the acidophilic protozoan Euglena mutabilis contributes to the formation of Fe-rich stromatolites in an acid mine drainage (AMD) environment in western Indiana. The biofilm community builds stromatolitic structures by several biogenic processes, including the use of mucilage to bind and trap chemical precipitates. The purpose of the study is to establish the biofilm community in a laboratory setting under controlled conditions to monitor formation of stromatolitic structures. Initially, biofilm attached to limestone and AMD collected from the field site was placed in a shallow container with a pump to simulate flowing water. In this experiment, pH of the AMD rapidly decreased from 3.8 to 1.6, along with rapid oxidation of ferrous iron to create a reddish-orange solution rich in suspended ferric iron minerals, which was detrimental to the biofilm. The suspended Fe minerals are the most likely factor leading to death of the biofilm as opposed to the decrease in pH because E. mutabilis has been observed in the field in AMD as low as pH 1.2. Additionally, we have observed E. mutabilis in the field forming biofilm along the margin of reddish colored AMD pools with a pH <2.0. The suspended Fe particles may have absorbed light in the part of the spectrum needed for photosynthesis, resulting in the death of the biofilm. An artificial AMD solution consisting of FeSO₄, MgSO₄, Al₂(SO₄)₃, (NH₃)₂SO₄, KH₂PO₄, and CaCl₂ also met with limited success because a pale bluish-green precipitate, possibly Fe hydroxide, was produced and lowered pH from 3.7 to 2.3. Artificial AMD was again simulated without the addition of FeSO₄ and with the use of NaHSO₄ and NaOH to buffer pH changes. E. mutabilis biofilm will be added to this artificial AMD solution, and when stabilized, sediment will be distributed over the biolfilm surface to induce the biological processes involved in the production of stromatolites.