Mineralogy of microbially induced calcium carbonate precipitates formed using single cell drop-based microfluidics

Neerja M Zambare,Connie B Chang,Connie B Chang,Nada Y Naser,Nada Y Naser,Nada Y Naser,Robin Gerlach

doi:10.1038/s41598-020-73870-y

Abstract

Microbe-mineral interactions are ubiquitous and can facilitate major biogeochemical reactions that drive dynamic Earth processes such as rock formation. One example is microbially induced calcium carbonate precipitation (MICP) in which microbial activity leads to the formation of calcium carbonate precipitates. A majority of MICP studies have been conducted at the mesoscale but fundamental questions persist regarding the mechanisms of cell encapsulation and mineral polymorphism. Here, we are the first to investigate and characterize precipitates on the microscale formed by MICP starting from single ureolytic E. coli MJK2 cells in 25 µm diameter drops. Mineral precipitation was observed over time and cells surrounded by calcium carbonate precipitates were observed under hydrated conditions. Using Raman microspectroscopy, amorphous calcium carbonate (ACC) was observed first in the drops, followed by vaterite formation. ACC and vaterite remained stable for up to 4 days, possibly due to the presence of organics. The vaterite precipitates exhibited a dense interior structure with a grainy exterior when examined using electron microscopy. Autofluorescence of these precipitates was observed possibly indicating the development of a calcite phase. The developed approach provides an avenue for future investigations surrounding fundamental processes such as precipitate nucleation on bacteria, microbe-mineral interactions, and polymorph transitions.

Highlights

Microbe-mineral interactions are ubiquitous and can facilitate major biogeochemical reactions that drive dynamic Earth processes such as rock formation
We observed microbially induced calcium carbonate precipitation (MICP) starting from single cells of E. coli MJK2 in 25 μm diameter drops at various concentrations of calcium ions (Fig. 2)
The cells performed ureolysis, which led to an increase in alkalinity and subsequent precipitation of CaCO3 according to the reaction scheme outlined in Eqs. (1–5)

Summary

Introduction

Microbe-mineral interactions are ubiquitous and can facilitate major biogeochemical reactions that drive dynamic Earth processes such as rock formation. One example of MMI is microbially induced calcium carbonate precipitation (MICP), a biomineralization process that occurs both in nature and in engineered systems as a result of bacterial a ctivity[9]. Visualization of MMI is typically performed on samples extracted from bench-scale reactor systems, which may not be accurate representations of the original state of the microbes or minerals Such ‘ex situ’ visualizations do not allow for non-destructive, time-resolved observation of microbes and minerals starting from single cells or small groups of cells. In situ visualization of MICP at the microscale in real-time is required to investigate the potential for cell encapsulation as well as other MMIs. Non-invasive visualization of MMIs during biological calcium precipitation at the single-cell level can be achieved by drop-based microfluidics. The ability to study biomineralization in a non-invasive manner can provide valuable insights into the mechanisms underlying large scale precipitation processes and contribute to designing novel bio-based materials

Methods

Results

Conclusion