PHOSPHATE-RELATED ARTIFACTS IN CARBONATE MINERALIZATION EXPERIMENTS

Sedimentary deposits of calcium carbonate are ubiquitous and may preserve a record of early life on Earth. Precipitation of carbonates and other minerals can be mediated by microorganisms, a process known as organomineralization. One of the many approaches for the study of organomineralization is growth of bacteria in culture. Although culture media commonly are based on natural conditions, the levels of nutrients, especially phosphate, generally are much higher than in the natural systems they are meant to simulate. This study demonstrates that levels of phosphate ranging from 5.8 to 0.1 mM in common laboratory media (exceeding natural concentrations by 3 to 6 orders of magnitude), can affect carbonate mineral morphology (e.g., spherulites, dumbells) and size (i.e., fewer, larger crystals in the presence of phosphate) and affect the formation of other mineral products not related to study objectives, such as apatite and struvite. In the microbial mats forming the modern stromatolites at Highborne Cay in the Bahamas, layers of calcium carbonate are precipitating in association with high rates of sulfate reduction. Sulfate-reducing bacteria (SRB) have also been implicated in the precipitation of calcium carbonate in other natural systems, and in the precipitation of dolomite. Batch-culture studies using SRB from the precipitating layers of the Highborne Cay stromatolites produced both carbonate (aragonite) and, unexpectedly, phosphate (apatite) minerals. Although this occurrence is not a widely reported result in similar culture studies, thermodynamic modeling of this medium and a range of compositions used in organomineralization investigations revealed oversaturation of apatites and of common carbonate minerals in all cases. Other analyses compared the effect of phosphate on other factors that can impact culture, including bacterially derived extracellular polymeric substances (EPS), sulfate, and low-molecular-weight organic compounds (LMWOC). Results from the EPS study showed precipitated struvite (a phosphate mineral), indicating phosphate association with EPS. Different phosphate levels altered calcium carbonate morphologies precipitated in the presence of sulfate and LMWOC. For both the sulfate study and the LMWOC study, phosphate was the dominant factor controlling mineral products. These results illustrate that phosphate in culture media has a pronounced impact on carbonate mineral products and can potentially obscure or alter more subtle effects on mineral precipitation. These effects might lead to misinterpretation of culture studies meant to simulate natural systems.