Fragmentation and aggregation of cyanobacterial colonies

TL;DR

This study tackles how fluid flow shapes fragmentation and aggregation of cyanobacterial colonies, focusing on Microcystis, to develop a framework linking hydrodynamics to colony-size dynamics. The authors combine a cone-and-plate rheometer with direct microscopic imaging of lab cultures and lake samples to track colony-size distributions under controlled dissipation rates $\dot{\varepsilon}$ and to distinguish division-formed and aggregation-formed colonies. They show that division-formed colonies exhibit strong EPS-mediated resistance and fragment by erosion at high $\dot{\varepsilon}$, whereas aggregation-formed colonies form weaker bonds and are readily fragmented or fail to contribute to large-size growth; field samples demonstrate higher EPS-related resistance than lab cultures. A two-category population-balance model with erosion for $C_2$ and equal-fragments for $C_1$ reproduces observed size distributions and yields a phase diagram predicting regimes where cell division, aggregation, or fragmentation dominates under varying $\phi$ and $\dot{\varepsilon}$, with implications for bloom management and biotechnological applications.

Abstract

Fluid flow has a major effect on the aggregation and fragmentation of bacterial colonies. Yet, a generic framework to understand and predict how hydrodynamics affects colony size remains elusive. This study investigates how fluid flow affects the formation and maintenance of large colonial structures in cyanobacteria, using an experimental technique that precisely controls hydrodynamic conditions. We performed experiments on laboratory cultures and lake samples of the cyanobacterium Microcystis, while their colony size distribution was measured simultaneously by direct microscopic imaging. We demonstrate that EPS-embedded cells formed by cell division exhibit significant mechanical resistance to shear forces. However, at elevated hydrodynamic stress levels (exceeding those typically generated by surface wind mixing) these colonies experience fragmentation through an erosion process. We also show that single cells can aggregate into small colonies due to fluid flow. However, the structural integrity of these flow-induced colonies is weaker than that of colonies formed by cell division. We provide a mathematical analysis to support the experiments and demonstrate that a population model with two categories of colonies describes the measured size distributions. Our results shed light on the specific conditions wherein flow-induced fragmentation and aggregation of cyanobacteria are decisive and indicate that colony formation under natural conditions is mainly driven by cell division, although flow-induced aggregation could play a role in dense bloom events. These findings can be used to improve prediction models and mitigation strategies for toxic cyanobacterial blooms and also offer potential applications in other areas such as algal biotechnology or medical settings where the dynamics of biological aggregates play a significant role.

Figures (14)

• Figure 1: Methodology used to observe effects of fluid flow on Microcystis colonies. (A) Experimental setup consisting of a (cone-and-plate) controlled flow setup combined with inverted microscopy. The conical upper surface was rotated by a rheometer head, while the stationary glass slide below the sample allowed optical access for the microscope. (B) Examples of microscopy images. Colony size distributions were calculated after image processing of the captured frames. (C) Changes in size distributions (and other complementary measurements) over time were used to identify aggregation and fragmentation of cyanobacterial colonies. (D) The majority of the measurements were conducted using a laboratory culture of Microcystis strain V163. Colonies collected from Lake Gaasperplas (Netherlands), dominated by the morphospecies M. aeruginosa were also used.
• Figure 2: Kinetics of fragmentation of Microcystis strain V163 colonies under cone-and-plate shear flow. The laboratory culture was filtered to select mainly large colonies, and the total biovolume fraction was adjusted to $\phi = 10^{-4}$. Suspensions were subjected to an intense dissipation rate ($\dot{\varepsilon} = 5.8~ {m^2/s^3}$) in panels A-E. (A) The initial size distribution of colonies, expressed as biovolume fraction of the relative colony diameter (normalized by single cell diameter). The size distribution had a bimodal shape, with large colonies ($l>5$, in yellow) and small colonies ($l\leq5$, in green, composed mostly of single cells, dimers, and some trimers, as depicted by the inset). (B) Median diameter of small and large colonies as a function of time. Symbols indicate the experimental data, while the lines indicate the predictions from the population model given by Eq. \ref{['eq:dndt']} with an erosion (dashed) or equal fragments (dot-dash) hypothesis for large division-formed colonies. Small aggregates colonies follow the equal fragments hypothesis. Bars and shaded region indicate limits of 25th and 75th percentiles. (C) Most large colonies have been fragmented after 1 hour of shear flow, but the bimodal shape of the colony size distribution remained. (D) Biovolume fraction of small colonies (i.e., biovolume of small colonies over the total biovolume) as a function of time. A shift is observed from large to small colonies, captured well by the erosion model, but not by the equal fragments model. (E) The rate of change in biovolume distribution at $t = 0$h. Negative values indicate loss of colonies by fragmentation, while positive values indicate newly created fragments. The distribution suggests an erosion mechanism, as depicted by the cartoon inside the plot. (F) Fragmentation frequency as a function of the relative diameter of colonies for three values of dissipation rate. The inset shows details for moderate dissipation rates. Error bars indicate the standard deviation and the lines indicate the predictions by Eq. \ref{['eq:breakkernel']}. Best fit parameters: $\alpha_1 = 0.023$, $S_1= 0.034$ , $q_1 = 4.5$; erosion: $S_2= 31$ , $q_2 = 4.1$ ; equal fragments: $S_2= 33$ , $q_2 = 6.3$.
• Figure 3: Kinetics of aggregation for a single-cell suspension of Microcystis strain V163 at a moderate dissipation rate of $\dot{\varepsilon} = 0.019~{m^2/s^3}$ and a total biovolume fraction of $\phi = 10^{-4}$. (A) Initial size distribution of the suspension as a function of the relative diameter, composed mostly of single cells. (B) Median diameter of colonies formed by aggregation of single cells as a function of time. Symbols indicate the experimental data, while the lines indicate the predictions from the population model given by Eq. \ref{['eq:dndt']} with an equal fragment hypothesis for small aggregates colonies. Bars and shaded region indicate limits of 25th and 75th percentiles. (C) After 1 hour of shear flow, the size distribution has shifted towards slightly larger diameters. (D) Time behavior of a suspension of large division-formed colonies under the same moderate dissipation rate and total biovolume fraction. The bimodal size distribution is separated into large (yellow) and small (green) colonies. Dashed lines and shaded regions indicate predictions from the population model given by Eq. \ref{['eq:dndt']}. Best fit parameters: $\alpha_1 = 0.023$, $S_1= 0.034$ , $q_1 = 4.5$, $S_2= 31$ , $q_2 = 4.1$.
• Figure 4: Kinetics of the fragmentation of colonies in field samples of Microcystis spp. at an intense dissipation rate ($\dot{\varepsilon} = 5.8~ {m^2/s^3}$) and total biovolume fraction of $\phi = 10^{-4}$. (A) Comparison of the fragmentation frequency as a function of colony size for the laboratory culture (Microcystis strain V163) and the field samples (Microcystis spp.). Error bars indicate the standard deviation. (B) Brightfield microscopy images of colonies in a Nigrosin-dyed medium (dark region) show evidence of a thick EPS layer (bright region) surrounding a field colony. (C) Initial size distribution of colonies in field samples as a function of the relative diameter. The size distribution had a bimodal shape, with small colonies (green) and large colonies (yellow). (D) The median diameter of the colonies in each subpopulation as a function of time. Bars indicate 25th and 75th percentiles. (E) After 1 hour of shear flow, the small colonies have aggregated slightly, while the large colonies keept their size distribution. (F) The biovolume fraction of small colonies remained nearly constant during the experiment.
• Figure 5: Phase diagram indicating the dominant colony formation mechanism as a function of the dissipation rate $\dot{\varepsilon}$ and the cyanobacterial abundance (expressed by the total biovolume fraction $\phi$). (I) Colonies grow only by cell division at low dissipation rates and total biovolume fractions. (II) As the biovolume fraction increases, aggregation enhances colony growth. (III) For moderate dissipation rates, aggregated colonies are fragmented, and only cell division can increase colony size. (IV) Fragmentation of colonies dominates at intense dissipation rates, irrespective of whether these colonies were formed by aggregation or cell division. Bars on the right side indicate typical values of dissipation rate observed for natural wind-mixing, bubble plumes in artificially mixed lakes and laboratory-scale setups such as cone-and-plate systems and stirred tanks. The dashed arrowed line indicates the transition from an early bloom (left bullet - WHO alert level 1 chorus2021toxic) to a dense surface scum (right bullet - typical scum biovolume fraction wu2020recovery) under typical wind mixing.