Search for microbial signatures within human and microbial calcifications using soft x-ray spectromicroscopy

Area of Science:

• Biomineralization research
• Microbiology
• Nanotechnology

Background:

• The origin of arterial and renal calcification is poorly understood.
• Putative nanobacteria, detected in human calcified tissues, are debated due to difficulties in distinguishing them from mineral deposits.
• Advanced characterization techniques are needed to analyze the organic and mineral content of these structures.

Purpose of the Study:

• To investigate the microbiologic nature of self-replicating calcified nanoparticles from human samples.
• To differentiate between biomineralized microbes and non-biologic mineral nucleation.
• To apply submicrometer scale techniques for characterizing organic and mineral content.

Main Methods:

• Scanning transmission X-ray microscopy (STXM) was used to examine calcifying bacteria and cultured nanoparticles.
• Near-edge X-ray absorption fine structure (NEXAFS) spectroscopy was applied at the C K-edge, N K-edge, and Ca L(2,3)-edge.
• Analysis was performed at a 25 nm spatial scale.

Main Results:

• Calcified reference bacteria (C. crescentus, R. tataouinensis) showed unique NEXAFS spectral signatures distinct from nonbiologic hydroxyapatite.
• NEXAFS analysis of human-derived calcified nanoparticles revealed evidence of organic components, likely proteins, associated with hydroxyapatite.
• A biochemical signature, including proteins, polysaccharides, nucleic acids, and hydroxyapatite, was defined for cultured calcified bacteria.

Conclusions:

• NEXAFS at 25 nm spatial scale can define a biochemical signature for cultured calcified bacteria.
• Preliminary findings suggest human-derived nanoparticles share spectroscopic characteristics with calcified proteins.
• This supports the hypothesis of a microbial origin for certain calcifications.