Use of high-resolution pressure nephelometry to measure gas vesicle collapse as a means of determining growth and turgor changes in planktonic cyanobacteria

Previous work has demonstrated that the physical properties of intracellular bacterial gas vesicles (GVs) can be analyzed in vivo using pressure nephelometry. In analyzing the buoyant state of GV-containing cyanobacteria, hydrostatic pressure within a sample cell is increased in a stepwise manner, where the concomitant collapse of GVs due to pressure and the resultant decrease in suspended cells are detected by changes in nephelometric scattering. As the relative pressure at which GVs collapse is a function of turgor pressure and cellular osmotic gradients, pressure nephelometry is a powerful tool for assaying changes in metabolism that affect turgor, such as photosynthetic and osmoregulatory processes. We have developed an updated and automated pressure nephelometer that utilizes visible-infrared (Vis-IR) spectra to accurately quantify GV critical collapse pressure, critical collapse pressure distribution, and cell turgor pressure. Here, using the updated pressure nephelometer and axenic cultures of Microcystis aeruginosa PCC7806, we demonstrate that GV critical collapse pressure is stable during mid-exponential growth phase, introduce pressure-sensitive turbidity as a robust metric for the abundance of gas-vacuolate cyanobacteria, and demonstrate that pressure-sensitive turbidity is a more accurate proxy for abundance and growth than photopigment fluorescence. As cyanobacterium-dominated harmful algal bloom (cyano-HAB) formation is dependent on the constituent cells possessing gas vesicles, characterization of environmental cyanobacteria populations via pressure nephelometry is identified as an underutilized monitoring method. Applications of this instrument focus on physiological and ecological studies of cyanobacteria, for example, cyanoHAB dynamics and the drivers associated with cyanotoxin production in aquatic ecosystems. The increased prevalence of bloom-forming cyanobacteria and associated risk of exposure to cyanobacterial toxins through drinking water utilities and recreational waterways are growing public health concerns. Cost-effective, earlydetection methodologies specific to cyanobacteria are crucial for mitigating these risks, with a gas vesicle-specific signal offering a number of benefits over photopigment fluorescence, including improved detection limits and discrimination against non-gas-vacuolate phototrophs. Here, we present a multiplexed instrument capable of quantifying the relative abundance of cyanobacteria based on the signal generated from the presence of intracellular gas vesicles specific to bloom-forming cyanobacteria. Additionally, as cell turgor can be measured in vivo via pressure nephelometry, the measurement furnishes information about the internal osmotic pressure of gas-vacuolate cyanobacteria, which relates to the metabolic state of the cell. Together these advances may improve routine waterway monitoring and the mitigation of human health threats due to cyanobacterial blooms.