Investigation of Resonant Mass Measurement for Physiological Analysis of Microorganisms

Introduction: Information on microbial physiology has traditionally been obtained using imaging methods such as light, fluorescence and electron microscopy. Other methods, such as flow cytometry, can provide data on dynamic physiological changes occurring in microbial populations with single-cell resolution. However, physiological properties such as mass are typically measured using bulk-phase techniques, which yield values that are averaged across entire populations. Filling this gap, the LifeScale-R instrument (Affinity Biosensors, LLC) enables flow-through mass measurement of individual microbes using resonant cantilever technology.

Purpose: We sought to explore use of resonant mass measurement for physiological characterization of stress-filamented Salmonella spp. and for early detection of germ tube formation in Candida albicans. 

Methods: Five Salmonella serovars and E. coli (non-Salmonella control) were filamented under conditions of salt- or cold-stress and reversion of filaments to smaller, individual cells was monitored over time after removal of these stressors. Mean mass and mass distribution of Salmonella samples were monitored and compared with forward- and side-scatter data collected via flow cytometry. For analysis of Candida albicans, yeast cells were inoculated into rich media containing serum and were measured over time to determine if mass-based measurements could enable early detection of germ tube emergence. Non-C. albicans germ tube-negative control strains were also examined.

Results: Mean mass and mass distribution measurements made using the LifeScale-R clearly highlighted the post-stress physiological differentiation of Salmonella filaments into smaller, individual cells. LifeScale-R data correlated well with parallel light scatter data collected using flow cytometry. For C. albicans, incubation in germ tube-inducing media resulted in increasing cell mass as a function of time, with visible elongation of cells, however, these cells proved too large for successful analysis in the LifeScale-R as it is currently configured.  

Significance: This study demonstrates the utility of resonant mass measurement for single-cell analysis of dynamic changes in microbial physiology.