Rapid start-up sulfur-driven autotrophic denitrification granular process: Extracellular electron transfer pathways and microbial community evolution

Sulfur-driven autotrophic denitrification (SAD) granular process has significant advantages in treating low-carbon/nitrogen wastewater; however, the slow growth rate of sulfur-oxidizing bacteria (SOB) results in a prolonged start-up duration. In this study, the thiosulfate-driven autotrophic denitrification (TAD) was successfully initiated by inoculating anaerobic granular sludge on Day 7. Additionally, the electron donor was successfully transferred to the cheaper elemental sulfur from Day 32 to Day 54 at the nitrogen loading rate of 176.2 g N m⁻³ d^-1. During long term experiment, the granules maintained compact structures with the α-helix/(β-sheet + random coil) of 29.5–40.1 %. Extracellular electron transfer (EET) pathway shifted from indirect to direct when electron donors were switched thiosulfate to elemental sulfur. Microbial analysis suggested that thiosulfate improved EET involving enzymes activity. Thiobacillus and Sulfurimonas were dominant in TAD, whereas Longilinea was enriched in elemental sulfur-driven autotrophic denitrification. Overall, this strategy achieved in-situ enrichment of SOB in granules, thereby shortening start-up process.