Quantifying Inorganic Nitrogen Assimilation by Synechococcus Using Bulk and Single-Cell Mass Spectrometry: A...

Area of Science:

• Marine microbial ecology
• Biogeochemical cycles
• Stable isotope biogeochemistry

Background:

• Microorganisms are key drivers of oceanic biogeochemical cycles, particularly nitrogen cycling, which limits primary production.
• Bulk methods like EA-IRMS average nitrogen assimilation across microbial communities, obscuring individual cell variability.
• Single-cell techniques, such as SIMS, offer higher resolution for quantifying elemental assimilation.

Purpose of the Study:

• To compare bulk (EA-IRMS) and single-cell (NanoSIMS, ToF-SIMS) methods for quantifying inorganic nitrogen assimilation by the marine cyanobacterium Synechococcus.
• To evaluate the advantages and disadvantages of each technique and highlight their complementary nature.
• To investigate nitrogen assimilation rates and metabolic heterogeneity at the single-cell level.

Main Methods:

• Application of Elemental Analyzer-Isotope Ratio Mass Spectrometry (EA-IRMS) for bulk nitrogen assimilation measurements.
• Utilized NanoSIMS (Nanoscale Secondary Ion Mass Spectrometry) to quantify nitrogen assimilation at the single-cell level.
• Employed ToF-SIMS (Time-of-Flight Secondary Ion Mass Spectrometry) to identify nitrogen incorporation into cellular components.

Main Results:

• EA-IRMS yielded higher average 15N assimilation values for Synechococcus compared to SIMS techniques, potentially due to sample preparation and systematic depletion.
• NanoSIMS revealed significant metabolic heterogeneity in nitrogen assimilation among individual Synechococcus cells.
• ToF-SIMS successfully identified the assimilation of nitrogen into peptides within individual cells.

Conclusions:

• Coupling bulk and single-cell stable isotope-based approaches provides a more comprehensive understanding of microbial nitrogen cycling.
• Single-cell mass spectrometry techniques offer crucial insights into microbial metabolic diversity and function in marine environments.
• This integrated approach enhances our ability to study the metabolic capacity and heterogeneity of microbes in situ.