Characterization of the effects of attachment material, wastewater, and inoculation strategy on prokaryotic and eukaryotic communities in algal biofilms using 16S rRNA gene amplicon sequencing

Algal biofilms are a promising source of sustainable biofuels and other low-cost algal products due to their high biomass densities and ability to treat wastewater. The mixed communities of algae, bacteria, and other organisms that naturally grow in biofilms may further benefit biofilm growth and nutrient removal, but there are few studies which examine them. To investigate how specific growth parameters (reactor inoculation strategy, biofilm attachment matrix, and wastewater pretreatment) influence algal biofilm communities, algal biofilm community profiles were estimated using next-generation sequencing (NGS) of small subunit rRNA gene amplicons. Cell suspensions and biofilms from biofilm bioreactor systems (operated for 10 days; described in Cheung et al. 2022, Journal of Applied Phycology 34) were harvested and extracted for DNA. Relative abundances of prokaryotes and eukaryotes were estimated by next-generation sequencing, using modified 926f and 1392r primers targeting the V6-V8 hypervariable region of 16S rRNA genes. This primer set was selected because it can capture chloroplast 16S rRNA genes and nuclear genes (18S/ITS) from algal species, in addition to bacterial and archaeal 16S rRNA gene fragments, making it potentially suitable to analyze algal and prokaryotic populations simultaneously. A better understanding of mixed algal-prokaryotic communities could aid in the further optimization of wastewater treatment, and to develop strategies for controlling biofilm growth for commercialization of algal biofilm biofuels and bioproducts.