Micro-compression analysis of biopolymer-producing bacteria using Cupriavidus necator as the model bacterium

With the development of highly sensitive experimental techniques, the mechanical properties of bacterial cells have become an important research topic. However, existing models used to fit experimental data from micro-compression tests often lack accuracy. The aim of this study was to address this limitation by developing a new curve-fitting mathematical model for evaluating the mechanical properties of rod-shaped bacterial cells. The proposed model is based on a thin-shell approach and is specifically designed for the interpretation of single-cell micro-compression experiments. To verify the applicability of the model, single-cell micro-compression tests were performed using a flat-punch nanoindenter tip larger than the bacterial cells. Atomic force microscopy (AFM) was used to obtain detailed morphological information, including precise cell dimensions required for curve fitting. As a model organism, the polyhydroxyalkanoate-producing bacterium Cupriavidus necator H16 was selected due to its ability to accumulate intracellular polyhydroxybutyrate (PHB) granules. For comparison, a mutant strain, C. necator PHB^-4, which lacks PHB production, was also analyzed. The results showed that C. necator H16 cells, with an average PHB content of 72% of dry cell weight, exhibited a Young's modulus approximately 16× higher than that of the PHB^-4 mutant, indicating a substantial contribution of intracellular PHB granules to cell stiffness. AFM analysis further revealed that PHB-producing cells were, on average, larger in volume than the non-producing mutant. The combination of AFM and micro-compression testing enabled comprehensive characterization of bacterial cell mechanics and demonstrated a clear correlation between PHB content and mechanical behaviour.