Techniques for Quantifying Bacterially Induced Carbonate Mineralization in Escherichia coli

The lack of reliable techniques to accurately quantify bacterial carbonate production and a tractable genetic system limits our ability to thoroughly understand the metabolic basis for bacterial calcification. In this work, we demonstrate bacterially induced CaCO3 precipitation in the model organism, Escherichia coli K-12, and develop new quantitative methods for carbonate mineral production using inductively coupled plasma-optical emission spectroscopy (ICP-OES) and an image analysis approach repurposed for agar cultures. These assays allowed us to assess the impact of various calcium sources and environmental factors on crystalline CaCO3 formation in E. coli. Our data demonstrate that Ca-acetate and Ca-succinate yield the greatest CaCO3 output in E. coli, while a preliminary study demonstrated microclimatic influences on crystal growth and size. This work helps lay the foundation for using E. coli as a model bacterial species to probe the genetic elements that promote calcification in Bacteria, with the potential to identify previously undescribed pathways for precipitation.