Single-cell measurement of microbial growth rate with Raman microspectroscopy.

Area of Science:

• Microbial Ecology
• Environmental Geochemistry
• Biotechnology

Background:

• Understanding microbial growth rates is crucial for environmental geochemistry and ecology.
• Measuring single-cell microbial activity in complex environments is challenging.
• Stable isotope probing (SIP) assesses microbial growth by tracking isotopic labels.

Purpose of the Study:

• To evaluate Raman microspectroscopy as a stable isotope probing (SIP) technique for measuring microbial growth.
• To develop and validate a method for quantifying deuterium (2H) incorporation at the single-cell level.
• To demonstrate the capability of Raman-SIP for sensitive microbial growth rate measurements across diverse timescales.

Main Methods:

• Correlative measurements of microbial cells using Raman microspectroscopy and nanoscale secondary ion mass spectrometry (nanoSIMS).
• Growth experiments with varying concentrations of deuterated water to calibrate 2H measurements.
• Application of Raman-derived isotopic calibrations to a numerical model of microbial growth.

Main Results:

• Raman microspectroscopy provides accurate deuterium (2H) measurements for microbial biomass.
• NanoSIMS measurements of 2H are prone to dilution artifacts from hydrogen exchange during sample washing.
• Raman-derived calibrations enable sensitive quantification of microbial growth rates, from hours to years.

Conclusions:

• Raman microspectroscopy is a viable and sensitive SIP technique for single-cell microbial growth analysis.
• This method offers a rapid, non-destructive approach for studying microbial populations in complex matrices.
• Raman-SIP advances the application of single-cell analysis in environmental and ecological research.