Sulfide-oxidizing autotrophic denitrification (SOAD) is a bioprocess performed by sulfur-oxidizing bacteria (SOB) to drive chemolithoautotrophic denitrification by utilizing sulfide as an electron and energy source. Compared to conventional heterotrophic denitrification, SOAD offers a potential option for biological nitrogen removal with less amount of additional organics and greater cost-effectiveness in treating mainstream wastewater with insufficient levels of organics. The moving-bed biofilm reactor (MBBR), with its unique design for effective sludge retention, could be beneficial for the SOAD process in mainstream denitrification. However, knowledge about the SOAD process using the MBBR system remains limited. This thesis aims to provide an investigation on sulfide-oxidizing auto...[ Read more ]

Sulfide-oxidizing autotrophic denitrification (SOAD) is a bioprocess performed by sulfur-oxidizing bacteria (SOB) to drive chemolithoautotrophic denitrification by utilizing sulfide as an electron and energy source. Compared to conventional heterotrophic denitrification, SOAD offers a potential option for biological nitrogen removal with less amount of additional organics and greater cost-effectiveness in treating mainstream wastewater with insufficient levels of organics. The moving-bed biofilm reactor (MBBR), with its unique design for effective sludge retention, could be beneficial for the SOAD process in mainstream denitrification. However, knowledge about the SOAD process using the MBBR system remains limited. This thesis aims to provide an investigation on sulfide-oxidizing autotrophic denitrification via a moving-bed biofilm reactor with the major focuses on (a) the exploration of strategy for biofilm cultivation, (b) the applicability of a moving-bed biofilm reactor for mainstream nitrogen removal driven by sulfide, (c) the impacts of substrates (sulfide, nitrate and nitrite) on biofilm performance and associated biokinetics and biopathways and (d) the dynamics on biofilm microbial community compositions and bioactivities in the long-term continuous operation.

First, two cultivation approaches for rapid SOAD process start-up and moving-bed biofilm formation were investigated. The results indicated that both approaches could achieve stable nitrate removal (≥ 90%) after 20 days. However, the approach with a heterotrophic start-up may be more advantageous than one with an autotrophic start-up for the cultivation of slow-growing SOAD biomass in terms of the duration of the start-up period, sulfide removal performance, the growth and activity of biomass, the amount of biomass attached to carriers and the enrichment of functional bacteria.

Second, the potential for developing a sulfide-oxidizing moving-bed biofilm for mainstream denitrification driven by reduced sulfur compounds (i.e. sulfide and thiosulfate) was investigated. Both the attached biomass concentration and nitrogen-removal rate of this moving-bed biofilm increased in proportion to the increasing nitrogen loading rate, which was caused by decreasing the hydraulic retention time (HRT) from 12 h to 4 h. Under the steady state operation (HRT = 4 h), the sulfide-oxidizing moving-bed biofilm was observed to have an uneven and porous surface on which elemental sulfur (S⁰) accumulated, and the SOB biomass was highly diverse.

Third, the impacts of substrates including sulfide, nitrate and nitrite on the biofilm performance and the biofilm kinetics were evaluated through 12 batch experiments. The results were interpreted by adopting a two-step sulfide (S^2-) to sulfate (SO₄^2-) oxidation model (S^2-→S⁰ and S⁰→SO₄^2-) with all specific rates having a linear regression coefficient of R² > 0.9. The inhibitory kinetics analysis revealed that 1) the maximum treatment capacity (about 480 mg S/(m²·h) and 80 mg N/(m²·h)) was observed at low sulfide level (40 mg S/L), while higher sulfide level (60-150 mg S/L) showed increasing inhibition of the oxidation of both sulfide and sulfur, and of denitrification. 2) The denitrification activity decreased by up to 43% when free nitrous acid reached a maximum of 8.6 μg N/L, whereas the oxidation of sulfide and sulfur did not have any significant effect.

Finally, and crucially, the long-term treatment performance of this moving-bed biofilm and dynamics of corresponding microbial community were reported. This laboratory-scale moving-bed biofilm performing sulfide–oxidizing autotrophic denitrification was tested in a 700 d continuous experiment to treat synthetic saline domestic sewage, with an increase of the surface loading rate from 8 to 50 mg N/(m²·h) by gradually shortening the HRT from 12 h to 2 h. The specific reaction rates of the reactor were eventually increased up to 0.37 kg N/(m³·d) and 0.73 kg S/(m³·d) for nitrate reduction and sulfide oxidation, respectively, with no significant elemental sulfur accumulation visible. The results of the functional microbial community showed that: two sulfur-oxidizing bacteria (SOB) clades, Sox-independent SOB (SOB_I) and Sox-dependent SOB (SOB_II), were found to be responsible for indirect two-step sulfur oxidation (S^2-→S⁰→SO₄^2-) and direct one-step sulfur oxidation (S^2-→SO₄^2-), respectively. It is known that SOB_II biomass-specific electron transfer capacity can be approximately 2.5 times greater than that of SOB_I (38 mmol e^- /(gSOB_II·d) versus 15 mmol e- /(gSOB_I·d)), possibly resulting in the selection of SOB_II over SOB_I under stress conditions (such as a shorter HRT). Further studies on the methods and mechanism of selecting of SOB_II over SOB_I in biofilm reactors are recommended.

In a nutshell, the findings of the thesis shed light on the design and operation of MBBR-based SOAD process, which could be a promising alternative to conventional heterotrophic denitrification process for mainstream nitrogen removal.