Quantification of outer membrane vesicles: a potential tool to compare response in Pseudomonas putida KT2440 to stress caused by alkanols.

The bacterial release of outer membrane vesicles (OMVs) is an important physiological mechanism of Gram-negative bacteria playing numerous key roles. One function of the release of OMVs is related to an increase in surface hydrophobicity. This phenomenon initiates biofilm formation, making bacteria more tolerant to environmental stressors. Recently, it was qualitatively shown for Pseudomonas putida that vesicle formation plays a crucial role in multiple stress responses. Yet, no quantification of OMVs for certain stress scenarios has been conducted. In this study, it is shown that the quantification of OMVs can serve as a simple and feasible tool, which allows a comparison of vesicle yields for different experimental setups, cell densities, and environmental stressors. Moreover, the obtained results provide insight to the underlying mechanism of vesicle formation as it was observed that n-alkanols, with a chain length of C7 and longer, caused a distinct and steep increase in vesiculation (12-19-fold), compared to shorter chain n-alkanols (2-4-fold increase).

Immediate response mechanisms of Gram-negative solvent-tolerant bacteria to cope with environmental stress: cis-trans isomerization of unsaturated fatty acids and outer membrane vesicle secretion.

Bacteria have evolved an array of adaptive mechanisms enabling them to survive and grow in the presence of different environmental stresses. These mechanisms include either modifications of the membrane or changes in the overall energy status, cell morphology, and cell surface properties. Long-term adaptations are dependent on transcriptional regulation, the induction of anabolic pathways, and cell growth. However, to survive sudden environmental changes, bacterial short-term responses are essential to keep the cells alive after the occurrence of an environmental stress factor such as heat shock or the presence of toxic organic solvents. Thus far, two main short-term responses are known. On the one hand, a fast isomerization of cis into trans unsaturated fatty leads to a quick rigidification of the cell membrane, a mechanism known in some genera of Gram-negative bacteria. On the other hand, a fast, effective, and ubiquitously present countermeasure is the release of outer membrane vesicles (OMVs) from the cell surface leading to a rapid increase in cell surface hydrophobicity and finally to the formation of cell aggregates and biofilms. These immediate response mechanisms just allow the bacteria to stay physiologically active and to employ long-term responses to assure viability upon changing environmental conditions. Here, we provide insight into the two aforementioned rapid adaptive mechanisms affecting ultimately the cell envelope of Gram-negative bacteria.