Abstract: TiO₂ nanoparticles (TiO₂-NPs) have emerged frequently in wastewater treatment systems. To explore the effects of TiO₂-NPs on the microbial communities in the constructed wetland (CW) system filled by volcanic rock, 16S rDNA highthroughput sequencing technology was used to analyze the differences in microbial communities among treatments of different TiO₂-NPs concentrations (0, 60, 110, 160 mg·L^-1) after short-term exposure (18 d). The results showed that the relative abundances of Gammaproteobacteria and Bacteroidia in CW were notaffected (P>0.05), while the dominant species such as Alphaproteobacteria and Nitrospira were more sensitive to TiO₂-NPs, with their relative abundances being significantly reduced (P<0.05). Three concentrations of TiO₂-NPs had distinct ecotoxicities to the microbes in CW. The inhibition of 60 mg·L^-1 TiO₂-NPs on Nitrospira was the greatest after 18 days exposure. The relative abundance of Nitrospira increased slightly at the TiO₂-NPs concentrations of 110 and 160 mg·L^-1. The lowest  microbial community diversity was presented at the treatment of 110 mg·L^-1TiO₂-NPs, indicating the highest inhibition of microbes at this level. Further, such level led to the highest difference in microbial community diversity compared with the control treatment, followed by the TiO₂-NPs concentration of 60 mg·L^-1. In the 110 mg·L^-1 TiO₂-NPs treatment,  five microbial classes showed significant variations (P<0.05), including Deltaproteobacteria and Nitrospira; and 13 families showed significant changes (P<0.05), such as Flavobacteriaceae, Nitrospiraceae and unidentified_Alphaproteobacteria. Our results suggested that TiO₂-NPs might seriously reduce the nitrification efficiency of CW system. The three concentrations of TiO₂-NPs all affected the composition and diversity of bacterial community in CW, but with different effects on various taxa, which would affect the degradation and transformation of pollutants in CW. Our results provide a reference for understanding the potential harm of nanomaterials to microorganisms and the application of CW technology.

Key words: TiO₂-NPs, ecotoxicology, high-throughput sequencing, microbial diversity, community structure.