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Three strains of the freshwater microalgae used for wastewater treatment, Chlorella vulgaris and Chlorella sorokiniana co-immobilized separately in alginate beads with the microalgae-growth-promoting bacterium Azospirillum brasilense Cd, resulted in significant changes in microalgal-population size, cell size, cell cytology, pigment, lipid content, and the variety of fatty acids produced in comparison with microalgae immobilized in alginate without the bacterium. Cells of C. vulgaris UTEX 2714 did not change in size, but the population size within the beads significantly increased. On the other hand, C. vulgaris UTEX 395 cells grew 62% larger, but their numbers did not increase. The population of C. sorokiniana UTEX 1602 increased, but not their cell size. The content of pigments chlorophyll a and b, lutein, and violoaxanthin increased in all microalgal species. The lipid content also significantly increased in all three strains, and the number of different fatty acids in the microalgae increased from four to eight. This study indicates that the microalgae-growth-promoting bacterium induced significant changes in the metabolism of the microalgae.

When the freshwater microalga Chlorella sorokiniana and the plant growth-promoting bacterium Azospirillum brasilense were deployed as free suspensions in unsterile, municipal wastewater for tertiary wastewater treatment, their population was significantly lower compared with their populations in sterile wastewater. At the same time, the numbers of natural microfauna and wastewater bacteria increased. Immobilization of C. sorokiniana and A. brasilense in small (2-4 mm in diameter), polymer Ca-alginate beads significantly enhanced their populations when these beads were suspended in normal wastewater. All microbial populations within and on the surface of the beads were evaluated by quantitative fluorescence in situ hybridization combined with scanning electron microscopy and direct measurements. Submerging immobilizing beads in wastewater created the following sequence of events: (a) a biofilm composed of wastewater bacteria and A. brasilense was created on the surface of the beads, (b) the bead inhibited penetration of outside organisms into the beads, (c) the bead inhibited liberation of the immobilized microorganisms into the wastewater, and (d) permitted an uninterrupted reduction of ammonium and phosphorus from the wastewater. This study demonstrated that wastewater microbial populations are responsible for decreasing populations of biological agents used for wastewater treatment and immobilization in alginate beads provided a protective environment for these agents to carry out uninterrupted tertiary wastewater treatment.

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