Mixotrophic cyanobacteria are critical active diazotrophs in polychlorinated biphenyl-contaminated paddy soil

Biological nitrogen fixation by diazotrophs is a crucial biogeochemical process in global terrestrial ecosystems, especially in nitrogen-limited, organic-contaminated soils. The metabolic activities of diazotrophs and their ability to supply fixed nitrogen may facilitate the transformation of organic pollutants. However, the active diazotrophic communities in organic-contaminated soils and their potential metabolic functions have received little attention. In the current study, the relationship between biological nitrogen fixation and polychlorinated biphenyl (PCB) metabolism was analyzed in situ in paddy soil contaminated with a representative tetrachlorobiphenyl (PCB52). ¹⁵N-DNA stable isotope probing was combined with high-throughput sequencing to identify active diazotrophs, which were distributed in 14 phyla, predominantly Cyanobacteria (23.40%). Subsequent metagenome binning and functional gene mining revealed that some mixotrophic cyanobacteria (e.g. FACHB-36 and Cylindrospermum) contain essential genes for nitrogen fixation, PCB metabolism, and photosynthesis. The bifunctionality of Cylindrospermum sp. in nitrogen fixation and PCB metabolism was further confirmed by metabolite analyses of Cylindrospermum sp. from a culture collection as a representative species, which showed that Cylindrospermum sp. metabolized PCB and produced 2-chlorobiphenyl and 2,5-dihydroxybenzonic acid. Collectively, these findings indicate that active diazotrophs, particularly mixotrophic cyanobacteria, have important ecological remediation functions and are a promising nature-based in situ remediation solution for organic-contaminated environments.