DNA stable isotope probing and metagenomics reveal temperature responses of sulfur-driven autotrophic partial denitrification coupled with anammox (SPDA) system.

The sulfur-driven autotrophic partial denitrification coupled with anammox (SPDA) process showed significant advantages in energy conservation and resource recovery in municipal wastewater treatment. However, its application in regions with seasonal temperature fluctuations and high latitudes is challenged by low temperatures. In this study, the feasibility of the SPDA process for treating low-strength municipal wastewater across a wide temperature range (30-10 °C) was systematically investigated. The results demonstrated that thiosulfate-driven autotrophic partial denitrification maintained an efficient nitrate removal rate of 7.82 mg NO-N/gVSS/h and a nitrate to nitrite transformation rate of 62.7 % even at temperatures as low as 10 °C. Molecular ecological network and DNA-SIP revealed that dominant sulfur-oxidizing bacteria (SOB) shifted from norank_f_Hydrogenophilaceae and Thiobacillus at higher temperatures (30-20 °C) to Thiobacillus and Sulfurimonas as temperature decreased, thus ensuring the performance of autotrophic partial denitrification and consistent nitrite supply for anammox. Metagenomic analysis showed that the abundance of functional genes related to sulfur conversion increased almost universally, ensuring a stable electron supply for nitrate reduction through sulfur oxidation at low temperatures. The functional genes responsible for nitrate reduction changed from nar genes at higher temperatures to nap genes at lower temperatures, while a decrease in the abundance of hzs and hdh genes corresponding to reduced anammox performance. This study highlights the stable performance of the sulfur-driven autotrophic denitrification at low temperatures and the reliability of coupling with anammox, extending the applicability of SPDA to a broader geographical range.