Method for synergistically removing multi-element pollutants and antibiotics in water based on anaerobic microorganisms

Abstract

The invention discloses a method for synergistically removing multi-element pollutants and antibiotics in water based on anaerobic microorganisms, which comprises the following two steps: firstly, placing aerobic activated sludge in a sequencing batch reactor for domestication, adding S < 2->, NO2 <-> and low-concentration tetracycline through intermittent water feeding, and carrying out anaerobic fermentation; the tetracycline-resistant anaerobic activated sludge is mainly prepared from desulfurization and denitrification bacteria; tetracycline is added in stages, and collaborative removal of the multi-element pollutants under antibiotic exposure is completed. And the horizontal transfer of the resistance gene is inhibited by adjusting the pH value in a gradient manner. By periodically adjusting the stirring speed, the secretion amount of extracellular polymeric substances (EPS) is increased, rich functional groups of EPS are combined with tetracycline, and adsorption of EPS to tetracycline is promoted. By culturing the tolerance of the activated sludge to tetracycline, the adsorption and degradation capability of the activated sludge to pollutants and tetracycline is improved. The comprehensive treatment of desulfurization, denitrification and antibiotic removal of the domestic wastewater is realized, and the method has relatively high environmental and economic benefits.

Description

Technical Field

[0001] The present invention relates to the technical field of sewage treatment, and in particular to a method for collaboratively removing multiple pollutants and antibiotics in water based on anaerobic microorganisms. Background Art

[0002] With the acceleration of industrial development and urbanization, water pollution is becoming increasingly serious. The multiple pollutants in sewage are complex and can have serious ecological impacts. For example, excessive nitrogen and phosphorus nutrients can easily lead to eutrophication of water bodies, excessive algae growth, and disruption of the ecological balance of water bodies. At the same time, the widespread use of antibiotics in fields such as medicine and animal husbandry has caused them to enter the aquatic environment in large quantities. Tetracycline is widely used due to its antibacterial properties and has become one of the most commonly detected antibiotics in wastewater. Tetracycline residues not only have adverse effects on the growth, development, and reproduction of aquatic organisms, but can also induce the generation and spread of drug-resistant bacteria and drug-resistant genes in the environment, posing a potential threat to human health.

[0003] Currently, traditional wastewater treatment technologies, such as physical precipitation, chemical oxidation, and biological treatment, can remove some pollutants to a certain extent, but their effectiveness in synergistically treating multiple pollutants and antibiotics in water is not ideal. The activated sludge process, a commonly used wastewater treatment technology, degrades pollutants through microbial metabolism while simultaneously removing some pollutants through physical and chemical adsorption. Biological denitrification and desulfurization is a key step in wastewater treatment. The desulfurization and denitrification process, which uses NO₂⁻ as an electron acceptor to oxidize S₂⁻ to SO₂, has the advantages of low sludge production, low energy consumption, and no greenhouse gas emissions. Traditional wastewater treatment processes struggle to effectively degrade antibiotics such as tetracycline, making sewage treatment plants a major source of tetracycline in the environment. While efficiently removing sulfur and nitrogen pollutants, microbial adsorption and degradation of tetracycline by microorganisms has important practical significance for improving the removal rate of antibiotics in wastewater. Furthermore, the hazards of resistance genes cannot be ignored.

[0004] Because antibiotics can inhibit or kill microorganisms in sludge, altering microbial species and community composition, the activated sludge process's ability to remove both pollutants and antibiotics is reduced when treating antibiotic-containing wastewater. Cultivating microbial tolerance to antibiotics can improve their ability to adsorb and degrade them. Therefore, developing an efficient, economical, and environmentally friendly anaerobic microbial-based method for the synergistic removal of multiple pollutants and antibiotics from water is urgent and crucial for improving water quality, protecting the aquatic ecosystem, and ultimately, human health. Summary of the Invention

[0005] In response to the above problems, the present invention studies and designs a method for the synergistic removal of multiple pollutants and antibiotics in water based on anaerobic microorganisms. The technical means adopted by the present invention are as follows:

[0006] A method for collaboratively removing multiple pollutants and antibiotics from water based on anaerobic microorganisms comprises the following steps:

[0007] S1: Sludge directional acclimation stage: Place aerobic activated sludge in a sequencing batch reactor, maintain an anaerobic environment, use intermittent water inlet, control the water inlet stage to 0.5-1h, the stirring stage to 12-13h, the sedimentation stage to 2-3h, and the water outlet stage to 0.5-1h. Set the temperature in the reactor to 20-40℃, use periodic stirring, divided into two parts, fast and slow. The fast part has a stirring speed of 50-70r / min and a time of 2-3h. The slow part has a stirring speed of 15-30r / min and a time of 10-11h. Add acclimation water into the reactor, adjust the pH value from 7 to 7.2, wait until the sludge flocculation state is initially formed, and then adjust the pH value to 7.5. The acclimated water contains S2- and NO2-, with the S2- concentration in the acclimated water being 190-260 mg / L as S and the NO2- concentration being 30-50 mg / L as N. The acclimated water also contains trace elements that can meet the growth needs of microorganisms. When the removal rates of S2- and NO2- continue to rise and stabilize at above 95%, tetracycline at a concentration of 20-60 μg / L is added, and the pH value is slowly raised to 7.5-8. When the activated sludge forms brown sedimentable bacterial flocs and the fluctuation range of the system treatment efficiency is within ±10%, the pH value is then raised to 8-8.5 to inhibit the transfer of resistance genes until the removal rates of S2-, NO2-, and tetracycline are all above 95%.

[0008] S2: Sewage treatment stage: The sewage is treated using a sequencing batch reactor containing the activated sludge acclimated in step S1. Intermittent water inflow is used, with a controlled inflow phase of 0.5-1 hour, a stirring phase of 12-13 hours, a sedimentation phase of 2-3 hours, and a effluent phase of 0.5-1 hour. The temperature in the reactor is set at 25-35°C. Periodic stirring is used, divided into two stages: fast and slow. The fast stage is stirred at a speed of 50-70 r / min for 2-3 hours, and the slow stage is stirred at a speed of 15-30 r / min for 10-11 hours. This periodically increases the EPS content secreted by the activated sludge, thereby enhancing the adsorption and degradation of tetracycline, respectively. The pH is maintained at 8-8.5, and trace elements are added to meet the growth requirements of microorganisms.

[0009] Furthermore, the activated sludge is aerobic activated sludge.

[0010] Furthermore, the acclimation water is artificially prepared water, and the components of the artificially prepared water include tap water, Na2S·9H2O, NaNO2 and tetracycline.

[0011] Furthermore, the activated sludge has a suspended solids concentration of 10-20 g / L and a volatile suspended solids concentration of 5-15 g / L.

[0012] Furthermore, in step S1, the functional bacteria that adsorb and degrade tetracycline in the acclimated activated sludge are auto-aerobic desulfurization and denitrification bacteria, and the dominant bacterial genera thereof include Sulfurovum, Sulfurimonas and Thauera.

[0013] Furthermore, in step S2, the sewage inflow is divided into two stages, each stage lasting 20-35 days. The concentration of tetracycline in the inflow in the first stage is controlled to be 180-220 μg / L, and the concentration of tetracycline in the inflow in the second stage is controlled to be 180-350 μg / L.

[0014] Compared with the prior art, the method for collaboratively removing multiple pollutants and antibiotics in water based on anaerobic microorganisms described in the present invention utilizes the microbial community in the specifically domesticated activated sludge, and by periodically adjusting the stirring rate, strengthens the stress response mechanism of the activated sludge microorganisms, inducing them to secrete more EPS. In the presence of tetracycline, this combined stress can significantly increase the EPS content, especially the content of protein components, and the secreted EPS combines with tetracycline through various mechanisms such as complexation by virtue of its rich functional groups, promoting the adsorption of tetracycline by EPS, achieving the purpose of collaborative treatment of sulfur and nitrogen pollutants and efficient adsorption and degradation of antibiotics (tetracycline). On the basis of efficient removal of pollutants and antibiotics, the pH value is adjusted in stages to help inhibit the transfer of resistance genes. When the pH value is >8, the horizontal transfer of ARGs is inhibited, achieving the goal of active microorganisms collaboratively removing multiple pollutants and antibiotics in water without causing a large amount of transfer of resistance genes, solving the key problem of high-concentration tetracycline inhibiting traditional biological treatment processes, and realizing the comprehensive management of domestic wastewater desulfurization and denitrification and antibiotic removal, with high environmental and economic benefits. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 This is a relationship diagram between the removal rates of S2-, NO2- and tetracycline and the number of treatment days in the directional sludge acclimation stage of step 1 in an embodiment of the present invention.

[0016] Figure 2 This is a relationship diagram between the removal rates of NO2-, S2- and tetracycline and the number of treatment days in the sewage treatment stage of step 2 in an embodiment of the present invention.

[0017] Figure 3 This is a distribution diagram of functional bacteria classified by genus in the system of an embodiment of the present invention.

[0018] Figure 4 This is a diagram showing the relationship between pH stage regulation and changes in the abundance of integron IntI1 and resistance genes according to an embodiment of the present invention.

[0019] Figure 53 is a comparison chart of the changes in extracellular polymer content (protein and polysaccharide) caused by periodic rotation speed regulation in an embodiment of the present invention. DETAILED DESCRIPTION

[0020] This embodiment provides a method for collaboratively removing multiple pollutants and antibiotics in water based on anaerobic microorganisms, comprising the following steps:

[0021] Step 1: Sludge directional domestication stage:

[0022] Aerobic activated sludge was acclimated in two identical sequencing batch reactors. The acclimation method was as follows: maintaining an anaerobic environment, using intermittent water inflow, and conducting the acclimation process in four stages: water inflow, stirring, sedimentation, and effluent. The water inflow stage lasted 0.5-1 hour, the stirring stage 12 hours, the sedimentation stage 2 hours, and the effluent stage 0.5-1 hour. The reactor temperature was set at 30±0.5°C, and periodic stirring was used, divided into two stages: fast and slow. The fast stage was stirred at a speed of 60 r / min for 2 hours, and the slow stage was stirred at a speed of 20 r / min for 10 hours. Wastewater containing S2-, NO2- and tetracycline was introduced into the reactor, with the S2- concentration in the influent being 220 mg / L in terms of S and the NO2- being 45 mg / L in terms of N to acclimate its desulfurization and denitrification function, and 1 mL of trace elements was added per liter of influent to meet the growth of microorganisms. The trace elements added in this embodiment included the following substances in mass concentrations: MnCl4·H2O (0.1 g / L), Na2MoO4·2H2O (0.04 g / L), H3BO3 (0.05 g / L), AlCl3 (0.1 g / L), ZnCl2 (0.2 g / L), MgCl2·4H2O (0.2 g / L), CoCl2·6H2O (0.22 g / L), NiCl2·4H2O (0.08 g / L), CuCl2 (0.07 g / L), and CaCl2 (0.07 g / L). The pH was adjusted from 7 to 7.2. Once the sludge initially formed flocculation, the pH was adjusted to 7.5. When the S2- and NO2- removal rates continued to rise and stabilized at over 95%, a 25 μg / L tetracycline concentration was added to acclimate the bacteria to and degrade tetracycline. The pH of Reactor 1 was slowly raised to 7.5-8. Once brown, settling flocs formed in the activated sludge and the system's treatment efficiency fluctuated within ±10%, the pH was raised to 8.5 to inhibit the transfer of resistance genes. The pH of Reactor 2 was maintained at 7.5. Ultimately, the S2-, NO2-, and tetracycline removal rates all exceeded 95%, completing the acclimatization of the activated sludge dominated by tetracycline-resistant desulfurization and denitrification bacteria and successfully initiating the system.

[0023] Step 2, sewage treatment stage:

[0024] After the system was successfully started in the first step, the tetracycline concentration in the influent was increased to 200μg / L and 300μg / L in two stages. Intermittent water inflow was used, with four stages: inflow, stirring, sedimentation, and effluent. The inflow stage lasted 0.5-1h, the stirring stage lasted 12h, the sedimentation stage lasted 2h, and the effluent stage lasted 0.5-1h. The temperature in the reactor was set at 30±0.5℃. The pH value of the two reactors remained unchanged. Periodic stirring was used, divided into two parts: fast and slow. The fast stage stirring speed was 60r / min for 2h, and the slow stage speed was 20r / min for 10h. The S2- concentration in the influent was 220mg / L as S, and the NO2- as N was 45mg / L. After the reactor 1 system stabilized, samples were taken before, after, and after a cycle of rapid stirring to detect the EPS content and changes in tetracycline concentration. The system detects the concentrations of NO2-, S2- and tetracycline in the effluent once every 24 hours. The removal rates of NO2- and S2- reach more than 90%, and the removal rate of tetracycline reaches more than 80%. The second step of the system is successfully operated.

[0025] The reactor used in this embodiment is a sequencing batch reactor, which is divided into four stages: water inlet, stirring, precipitation, and water outlet. The water inlet stage is 0.5-1h, the stirring stage is 12h, the precipitation stage is 2h, and the water outlet stage is 0.5-1h. The temperature in the reactor is set to 30±0.5℃. In step 1, the concentration of S2-influent is 200mg / L in terms of S, the concentration of NO2-in terms of N is 40mg / L, and the concentration of tetracycline is 25μg / L. In step 2, the concentration of S2-influent is 200mg / L in terms of S, and the concentration of NO2-in terms of N is 40mg / L. Tetracycline is added in two stages, A1 and A2, with concentrations of 200μg / L and 300μg / L, respectively. Each stage is 25-30 days.

[0026] In this embodiment, the activated sludge to be acclimated is taken from the residual aerobic activated sludge in the secondary sedimentation tank of a municipal sewage treatment plant;

[0027] In this embodiment, activated sludge acclimation adopts artificial water distribution, and the artificial water distribution components are tap water, Na2S·9H2O, NaNO2 and tetracycline;

[0028] In this embodiment, the concentration of activated sludge in the reactor is: suspended solid concentration (MLSS) 10-20 g / L, volatile suspended solid concentration (MLVSS) 5-15 g / L, MLVSS / MLSS 0.5-0.75;

[0029] In this embodiment, the main functional bacteria that absorb and degrade tetracycline domesticated by the system are autotrophic desulfurization and denitrification bacteria, and the dominant genera thereof include Sulfurovum, Sulfurimonas and Thauera.

[0030] In this embodiment, in step 1, the sludge directional acclimation stage, the efficient removal of S2- and NO2- is completed and the tolerance and degradation ability of microorganisms to tetracycline are cultivated; in step 2, the sewage treatment stage, the efficient removal of tetracycline is completed on the basis of the efficient removal of S2- and NO2-.

[0031] This example uses staged pH adjustment. During the first sludge directional acclimation phase, the pH is adjusted from 7 to 7.2. Once the sludge flocculation is initially formed, the pH is adjusted to 7.5. After adding 25 μg / L tetracycline, the pH of reactor 1 is slowly raised to a range of 7.5-8. Once the activated sludge forms brown, sedimenting flocs and the system's treatment efficiency fluctuates within ±10%, the pH is then raised from 8 to 8.5. During the sewage treatment phase, the pH is also set at 8.5, achieving the goal of inhibiting the transfer of resistance genes while efficiently removing multiple pollutants and tetracycline. Reactor 2, whose pH is consistently maintained at 7.5, serves as the control group, demonstrating the inhibitory effect of pH on the transfer of resistance genes.

[0032] The reactor in this embodiment adopts periodic stirring, which is divided into two parts, fast and slow. The stirring speed of the fast part is 60r / min, and the time is 2h, and the speed of the slow part is 20r / min, and the time is 10h, so as to periodically increase the EPS content secreted by the activated sludge to strengthen the adsorption and degradation processes respectively.

[0033] In this embodiment, NO2-, S2- and tetracycline in the waste liquid after treatment in reactor 1 were detected:

[0034] Step 1: The start-up phase of the reactor 1 device is 55 days, and the results are shown in Figure 1 , Figure 1 The figure shows the relationship between the removal rates of NO2-, S2- and tetracycline and the number of treatment days. When the removal rates of NO2- and S2- in the device continue to rise and both reach above 95%, it indicates that the acclimation of desulfurization and denitrification bacteria is complete. When 25 μg / L of tetracycline is added, the tetracycline removal rate steadily increases to above 95%, and the acclimation of activated sludge dominated by tetracycline-resistant desulfurization and denitrification bacteria is completed.

[0035] Step 2: The operation of reactor 1 is divided into three stages: A1 and A2. The concentration of tetracycline in the influent is 200 μg / L and 300 μg / L respectively. Each stage lasts for 25-30 days. The results are shown in Figure 2 , Figure 2This is a relationship diagram between the removal rates of S2-, NO2- and tetracycline and the number of treatment days. In the sequencing batch device, the removal rates of S2- and NO2- reached more than 95%, and the removal rate of tetracycline was maintained at more than 90% when the influent concentration was 200μg / L, and stabilized at more than 85% when the influent concentration was 300μg / L. During the operation, the treatment effects of NO2- and S2- were relatively stable, and the reaction system had a strong treatment capacity for tetracycline.

[0036] according to Figure 3 As shown, in order to gain a deeper understanding of the metabolic functions of the system microorganisms, the composition of the microbial flora at the genus level in reactor 1 was analyzed, including the end of the sludge directional acclimation stage and the two stages A1 and A2 of the device operation, which was used to analyze in detail the distribution of functional bacteria in the device on different days.

[0037] The dominant bacterial genera in the system included Sulfurovum, Sulfurimonas, and Thauera, indicating that low-concentration tetracycline acclimation directed the enrichment of functional bacterial communities centered on Sulfurovum, Sulfurimonas, and Thauera.

[0038] Sulfurovum and Sulfurimonas are desulfurizing and denitrifying bacteria within the system. Sulfurovum uses S2- as an electron donor, while Sulfurimonas uses S2- and SO as electron donors, reducing NO2- to N2. They play a key role in maintaining sulfur balance. At a pH of 7.5-8.5, they secrete large amounts of EPS, which facilitates the adsorption of antibiotics and exhibits strong tolerance to them.

[0039] Thauera primarily uses S₂- and NO₂- as substrates for desulfurization and denitrification. It not only decomposes macromolecular organic matter to reduce NO₂- but also oxidizes S₂- to SO₃. It is a dominant bacterial genus in the system, with a strong ability to secrete EPS. Acclimation has conferred it with tetracycline tolerance, resulting in strong stability at 300 μg / L tetracycline, maintaining stable abundance.

[0040] Mesotoga can degrade macromolecular organic compounds such as aromatic compounds and polychlorinated biphenyls. High levels of tetracycline inhibit its growth, and some of its functions are replaced by Candidatus Promineofilum, resulting in a decrease in the abundance of Mesotoga.

[0041] Candidatus Promineofilum is an acid-producing bacterium that also plays a role in secreting EPS to adsorb tetracycline. EPS buffers the high toxicity of tetracycline and protects it from inhibition. Low-concentration tetracycline, as an organic matter, can be decomposed into small molecular organic matter by Candidatus Promineofilum. In the treatment of tetracycline-containing wastewater, it forms a functional network with Sulfurovum and Thauera bacteria to improve treatment efficiency.

[0042] according to Figure 4 As shown in the figure, in order to reflect the inhibitory effect of stage-by-stage pH regulation on resistance genes, qPCR technology was used to detect the integron IntI1 gene and typical tetracycline resistance genes (tetB, tetX and tetM) in the activated sludge in the two reactors at the end of the sludge directional acclimation stage and the end of the sewage treatment stage. The results showed that the abundance of the integron IntI1 gene and typical tetracycline resistance genes in the reactor was greater when the pH value was 7.5 than that when the pH value was 8.5, which indicates that the environment with a pH of 8.5 inhibits the transfer of resistance genes.

[0043] according to Figure 5 As shown in the figure, to demonstrate the effect of periodic stirring on EPS secretion, during the sewage treatment stage, after the system stabilized, samples were taken from three sections: before, after, and after a cycle of rapid stirring. The EPS content and tetracycline concentration were tested. The results showed that under the stimulation of rapid stirring, microbial EPS secretion reached a peak, especially a sharp increase in protein (PN) content, which is precisely the protective substance produced by microorganisms under stress. At the same time, the tetracycline concentration in the aqueous phase dropped sharply, demonstrating that the large amount of EPS in the reactor was rapidly adsorbing tetracycline. During the slow stirring period, the EPS amount slowly decreased, and the tetracycline concentration in the aqueous phase remained low or even continued to slowly decline, indicating that the activated sludge microorganisms were biodegrading tetracycline in the aqueous phase and EPS. This shows that periodic stirring can enhance the system's ability to adsorb and degrade tetracycline.

[0044] The embodiments described above are merely descriptions of preferred implementations of the present invention and are not intended to limit the scope of the present invention. Without departing from the spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should fall within the scope of protection determined by the claims of the present invention.

Claims

1. A method for collaboratively removing multiple pollutants and antibiotics in water based on anaerobic microorganisms, characterized in that: The following steps are involved: S1: Sludge directional acclimation stage: Place aerobic activated sludge in a sequencing batch reactor, maintain an anaerobic environment, use intermittent water inlet, control the water inlet stage to 0.5-1h, the stirring stage to 12-13h, the sedimentation stage to 2-3h, and the water outlet stage to 0.5-1h. Set the temperature in the reactor to 20-40℃, use periodic stirring, divided into two parts: fast and slow. The fast part has a stirring speed of 50-70r / min and a time of 2-3h. The slow part has a stirring speed of 15-30r / min and a time of 10-11h. Add acclimation water into the reactor, adjust the pH value from 7 to 7.2, and wait for the sludge to flocculent. The coagulation state is initially formed, and then the pH value is adjusted to 7.

5. The acclimated water contains S2- and NO2-. The S2- concentration in the acclimated water is 190-260 mg / L in terms of S, and the NO2- concentration is 30-50 mg / L in terms of N. The acclimated water also contains trace elements that can meet the growth of microorganisms. When the removal rate of S2- and NO2- continues to rise and stabilizes at more than 95%, tetracycline with a concentration of 20-60 μg / L is added, and the pH value is slowly raised to 7.5-8. When the activated sludge forms brown sedimentation micelles and the fluctuation range of the system treatment efficiency is within ±10%, the pH value is then raised to 8- 8.5, inhibit the transfer of resistance genes until the removal rates of S2-, NO2- and tetracycline are all above 95%; S2: Sewage treatment stage: The sewage is treated using a sequencing batch reactor containing the activated sludge acclimated in step S1. Intermittent water inflow is used, with a controlled inflow phase of 0.5-1 hour, a stirring phase of 12-13 hours, a sedimentation phase of 2-3 hours, and a effluent phase of 0.5-1 hour. The temperature in the reactor is set at 25-35°C. Periodic stirring is used, divided into two stages: fast and slow. The fast stage is stirred at a speed of 50-70 r / min for 2-3 hours, and the slow stage is stirred at a speed of 15-30 r / min for 10-11 hours. This periodically increases the EPS content secreted by the activated sludge, thereby enhancing the adsorption and degradation of tetracycline, respectively. The pH is maintained at 8-8.5, and trace elements are added to meet the growth requirements of microorganisms.

2. The method for collaboratively removing multiple pollutants and antibiotics in water based on anaerobic microorganisms according to claim 1, characterized in that: The activated sludge is aerobic activated sludge.

3. The method for collaboratively removing multiple pollutants and antibiotics in water based on anaerobic microorganisms according to claim 1, characterized in that: The acclimation water is artificially prepared water, and the components of the artificially prepared water include tap water, Na2S·9H2O, NaNO2 and tetracycline.

4. The method for collaboratively removing multiple pollutants and antibiotics in water based on anaerobic microorganisms according to claim 1, characterized in that: The activated sludge has a suspended solid concentration of 10-20 g / L and a volatile suspended solid concentration of 5-15 g / L.

5. The method for collaboratively removing multiple pollutants and antibiotics in water based on anaerobic microorganisms according to claim 1, characterized in that: In step S1, the functional bacteria that adsorb and degrade tetracycline in the acclimated activated sludge are auto-aerobic desulfurization and denitrification bacteria, and the dominant bacterial genera thereof include Sulfurovum, Sulfurimonas and Thauera.

6. The method for collaboratively removing multiple pollutants and antibiotics in water based on anaerobic microorganisms according to claim 1, characterized in that: In step S2, the sewage inflow is divided into two stages, each stage lasting 20-35 days. The concentration of tetracycline in the inflow in the first stage is controlled to be 180-220 μg / L, and the concentration of tetracycline in the inflow in the second stage is controlled to be 180-350 μg / L.

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