Effect of Abscisic Acid on Growth, Fatty Acid Profile, and Pigment Composition of the Chlorophyte Chlorella (Chromochloris) zofingiensis and Its Co-Culture Microbiome

Abstract

Microalga Chlorella (Chromochloris) zofingiensis has been gaining increasing attention of investigators as a potential competitor to Haematococcus pluvialis for astaxanthin and other xanthophylls production. Phytohormones, including abscisic acid (ABA), at concentrations relevant to that in hydroponic wastewater, have proven themselves as strong inductors of microalgae biomass productivity and biosynthesis of valuable molecules. The main goal of this research was to evaluate the influence of phytohormone ABA on the physiology of C. zofingiensis in a non-aseptic batch experiment. Exogenous ABA stimulated C. zofingiensis cell division, biomass production, as well as chlorophyll, carotenoid, and lipid biosynthesis. The relationship between exogenous ABA concentration and the magnitude of the observed effects was non-linear, with the exception of cell growth and biomass production. Fatty acid accumulation and composition depended on the concentration of ABA tested. Exogenous ABA induced spectacular changes in the major components of the culture microbiome of C. zofingiensis. Thus, the abundance of the representatives of the genus Rhodococcus increased drastically with an increase in ABA concentration, whereas the abundance of the representatives of Reyranella and Bradyrhizobium genera declined. The possibilities of exogenous ABA applications for the enhancing of the biomass, carotenoid, and fatty acid productivity of the C. zofingiensis cultures are discussed.

Keywords: bacterial community; carotenoids; chlorophyll; fatty acid profile; growth parameters; microalgae; phytohormone.

Figure 1

Cell density of Chlorella zofingiensis cultured in a range of ABA concentrations of 1 µM to 50 µM during 16 days of experiment. Data are shown as the mean ± SD, n = 12. Control cultures did not contain ABA.

Figure 2

Representative changes in optical absorbance spectra of the pigment extracts (see Methods) from the cells of Chlorella zofingiensis incubated for (a) 3, (b) 7, or (c) 16 days in the presence of different ABA concentrations. The spectra normalized to the red Chl absorption maximum (left scale) are shown together with the difference spectra (right scale) obtained by subtracting the control (0 µM ABA) spectrum from the spectrum of a corresponding ABA treatment. The ABA concentrations are shown on the graphs.

Figure 3

The effect of abscisic acid on the composition of major fatty acids (>3% of total FAs) in the cell of Chlorella zofingiensis on the 3rd, 7th, and 16th days of experiment (d3, d7, and d16, respectively). Statistically significant differences from the controls (p < 0.05) are indicated, where *—significantly greater than the control and +—significantly lower than the control. Data are shown as the mean ± SD, n = 4. Control cultures did not contain ABA.

Figure 4

The effect of abscisic acid on the composition of minor fatty acids (1–3% of total FAs) in the cell of Chlorella zofingiensis on the 3rd, 7th, and 16th days of experiment (d3, d7, and d16, respectively). Statistically significant differences from the controls (p < 0.05) are indicated, where *—significantly greater than the control and +—significantly lower than the control. Data are shown as the mean ± SD, n = 4. Control cultures did not contain ABA.

Figure 5

The influence of abscisic acid on the distribution of fatty acid classes in the cell of Chlorella zofingiensis during the experiment on the 3rd, 7th, and 16th days of the experiment (d3, d7, and d16, respectively). Statistically significant differences from the controls (p < 0.05) are indicated, where *—significantly greater than the control and +—significantly lower than the control. Data are shown as the mean ± SD, n = 4. Control cultures did not contain ABA.

Figure 6

Metagenomic analysis of the Chlorella zofingiensis cultures incubated for 16 days in the presence of different ABA concentrations: 0—0 μM, 1—1 μM, 5—5 μM, 10—10 μM, 50—50 μM. Interception between microbiomes of cultures is presented as a Venn diagram (a). The box plot represents mean values (n = 4) of microbial community biodiversity (Shannon index, blue) and evenness (Simpson index, orange) in studied cultures (b). Comparison of the cultures’ similarity is estimated by the Morisita index of β-diversity and depicted as a heatmap (c). Cultures’ microbiomes similarity was observed on a genus level on a clustered heatmap (d).