Microbial community colonization and assembly in a wastewater treatment sand biofilter

Kylie Langlois, School of Marine and Atmospheric Sciences, Stony Brook University, El Segundo, CA and Jackie L. Collier, School of Marine and Atmospheric Sciences, Stony Brook University, Stony Brook, NY

Background/Question/Methods
Sand biofilters are a common feature of nitrogen-removing on-site wastewater treatment systems (OWTS). Sand provides an oxic, low-carbon environment for microbial autotrophic nitrification (conversion of ammonia to nitrite, then to nitrate). Analysis of an established sand biofilter showed that its microbial community is strongly structured with depth by deterministic processes like environmental selection and competition. To discover how neutral processes might affect assembly of communities with depth, triplicate sand columns (18” depth, 5.75” ID clear polycarbonate) were dosed with septic tank effluent (STE) for 60 days. We hypothesized that the STE microbial community that colonized the sand columns would be structured by environmental selection, or alternatively by neutral processes. STE contained 32.5 mg ammonium (NH4+) L-1 and 0.17 mg nitrite+nitrate (NOx) L-1 while column effluent contained 0.2 mg N L-1 or less ammonium and ~18 mg N L-1 NOx on average for all columns. At the conclusion of the experiment, all columns were destructively sampled at 0, 3, 6, 9, and 12 inches and the microbial community characterized by 16S rRNA high throughput amplicon sequencing and open-source bioinformatics (QIIME2, R).
Results/Conclusions
At each depth, whole community structure was not significantly different across columns except one column at 0 and 3 inches (MRPP, multi-response permutation procedure). Beta diversity (weighted UniFrac metric) was clearly structured by depth, regardless of column. Therefore, whole community diversity was primarily structured by deterministic processes, but the top depths may be relatively more influenced by neutral processes. All functional groups except potential ammonia oxidizing microbes (AOM) differed only by depth while AOM differed by both depth and column (ANOVA), an indication that AOM may be influenced by both deterministic and neutral processes at the functional group level. At the genus level, most nitrifying genera (Nitrosomonas, Candidatus Nitrosotenuis, Ca. Nitrosotalea, Ca. Nitrotoga) displayed depth-dependent patterns of relative abundance. So did two clusters of potential denitrifying genera, a clear influence of deterministic processes. At the ASV level, most nitrifying and denitrifying ASV relative abundances differed significantly by depth rather than by column (ANOVA). While the effects of neutral processes were clearly detected, deterministic processes were the main factors that structured sand biofilter community assembly.