Method for culturing anaerobic ammonium oxidation flora tolerant to perfluorinated compounds and application of anaerobic ammonium oxidation flora

Abstract

The invention belongs to the technical field of environmental microorganisms, and particularly relates to a method for culturing a perfluorinated compound tolerant anaerobic ammonium oxidation flora and application of the method, the obtained anaerobic ammonium oxidation flora can tolerate impact of 1 ng / L-100 mg / L of perfluorinated compounds, the concentration is far higher than that of the perfluorinated compounds existing in sewage at present, and the perfluorinated compounds can tolerate the impact of 1 ng / L-100 mg / L of the anaerobic ammonium oxidation flora. And in the presence of a high-concentration perfluorinated compound, the total nitrogen removal rate of 85% or above can still be maintained, so that the problem of perfluorinated compound stress possibly faced by the anaerobic ammonia oxidation process in the application process is completely solved. In addition, the method is simple and easy to implement, the concentration of the perfluorinated compound is increased in a gradient mode, sludge discharging and low-temperature-starvation stimulation means are combined, and the cost is low; the cultured anaerobic ammonium oxidation flora not only can be used for treating sewage, but also can be used for repairing river sediment polluted by perfluorinated compounds.

Description

Technical Field

[0001] This invention belongs to the field of environmental microbiology technology, and specifically relates to a method for culturing anaerobic ammonia-oxidizing bacteria. Background Technology

[0002] In recent years, the harm of emerging pollutants to human health and the ecological environment has attracted great attention from government departments and researchers. Perfluorinated compounds, in particular, have received significant attention due to their significant recalcitrant nature, thermal stability, and biotoxicity. In 2009, the Stockholm Convention listed perfluorooctane sulfonic acid (PFOS) in Annex B for restriction, and subsequently in 2019, PFOS and its salts were listed in Annex A, prohibiting their production, use, import, and export. my country has also included them in the "List of Key Controlled Emerging Pollutants" for both 2022 and 2023. Perfluorinated compounds are widely used in industries such as fire protection, textiles, papermaking, electroplating, and fluoropolymer materials. Although restrictions have already been placed on the production and use of two major types of perfluorinated compounds...

[0003] On the other hand, anaerobic ammonia oxidation (AAO) is a novel nitrogen removal process developed in recent years. Patent 201510802682.2 discloses a method for enriching and cultivating anaerobic ammonia-oxidizing bacteria, using dissolved oxygen as a control measure for phased enrichment cultivation and using nitrification level as a transition point, which helps the growth and accumulation of AAO bacteria. Patent 201711445202.7 discloses a method for acclimating sulfate-tolerant AAO bacteria, achieving sulfate tolerance acclimation of common AAO bacteria by gradually increasing sulfate concentration. However, the acclimation process relies solely on stable nutrient supply conditions and lacks evaluation of the long-term operational stability and application effects of the acclimated bacteria. Anaerobic ammonia oxidation has already been applied in engineering and has broad application prospects due to its advantages of saving carbon sources and reducing energy consumption. In this context, AAO will inevitably face the stress of perfluorinated compounds during its application. Since AAO bacteria are sensitive to environmental conditions, directly treating wastewater containing perfluorinated compounds will lead to decreased nitrogen removal efficiency and system collapse. Currently, no relevant patents have been published. Therefore, enhancing the tolerance of anaerobic ammonia oxidation systems to perfluorinated compounds and maintaining efficient denitrification performance is the focus of current research. Summary of the Invention

[0004] To address the above problems, this invention proposes a method for cultivating anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds and its application.

[0005] The technical solution of this invention is implemented as follows: This invention application protects a method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds, the steps of which are as follows: (1) Enrichment culture: First, anaerobic ammonia oxidizing bacteria are enriched in the reactor to increase the abundance of anaerobic ammonia oxidizing bacteria. Simulated wastewater containing ammonia nitrogen, nitrous oxide and inorganic carbon source is used as influent in the reactor. The reaction time is adjusted according to the total nitrogen removal rate. After circulation, the reactor is stably operated to obtain the enriched anaerobic ammonia oxidizing bacteria. (2) Low concentration acclimatization: Perfluorinated compounds are added in a gradient to the anaerobic ammonia oxidation bacteria culture enriched in step (1) until the concentration of perfluorinated compounds reaches 1-10 mg / L, or the total nitrogen removal rate is always <80% and has a downward trend. Then the low concentration acclimatization is ended and the anaerobic ammonia oxidation sludge after low concentration acclimatization is obtained. (3) Low temperature-starvation stimulation: After the anaerobic ammonia oxidation sludge acclimated in step (2) is allowed to settle and the supernatant is discharged, the precipitate is refrigerated and sealed to obtain low temperature-starvation stimulation anaerobic ammonia oxidation sludge; during this process, no substrate is provided, so that the sludge is in a starvation state, which promotes the consumption of extracellular polymers and endogenous carbon sources, reduces its hindrance to mass transfer, and promotes the release of quorum sensing signal molecules, thereby enhancing tolerance and stress resistance. After that, it is taken out and placed in the original reactor to continue operation, using the low temperature environment to reduce the expression of related metabolic gene copy number of other denitrifying bacteria competing with anaerobic ammonia oxidation bacteria and reduce the activity of related enzymes. (4) High concentration enhancement: The anaerobic ammonia oxidation sludge stimulated by low temperature-starvation in step (3) is placed in the reactor and continues to operate at room temperature with the original substrate to restore its activity until the total nitrogen removal rate recovers to >80% or the change range is <5% for 7 consecutive days, which is considered to be basically stable. Then, high concentrations of perfluorinated compounds are added to the influent in a gradient to verify the sludge's tolerance to perfluorinated compounds. At the same time, high concentration stimulation is used to further enhance its tolerance until the perfluorinated compound concentration reaches 50-100 mg / L and it operates stably for more than 30 days to obtain anaerobic ammonia oxidation bacteria that are tolerant to perfluorinated compounds.

[0006] Preferably, in step (1) above, the molar concentration ratio of nitrous oxide to ammonia nitrogen in the influent is 1-1.05:1, and the mass concentration ratio of inorganic carbon source to ammonia nitrogen is 2-4:1.

[0007] Preferably, in step (1) above, the reaction time is adjusted according to the total nitrogen removal rate. If the total nitrogen removal rate is >10% after running for more than 30 days, the operation continues. If the total nitrogen removal rate is <10% and the change is <5% for more than 7 consecutive days, other conditions are kept unchanged, and the influent nitrous oxide concentration is adjusted to make the influent nitrous oxide / ammonia nitrogen ratio 0.45-0.55:1. The operation continues until the total nitrogen removal rate is >60% and the change is <5% for more than 7 consecutive days. Then the nitrous oxide concentration is restored to the influent nitrous oxide / ammonia nitrogen ratio of 1-1.05:1. The operation continues until the total nitrogen removal rate is >85% and the operation is stable for more than 30 days to achieve a stable operating state and obtain the enriched anaerobic ammonia-oxidizing bacteria.

[0008] Preferably, the above-mentioned gradient addition step is a stage for each gradient. After a stage runs for 20-30 days, if the total nitrogen removal rate is >80%, or the total nitrogen removal rate changes by <5% for more than 7 consecutive days, the concentration of perfluorinated compounds is increased to the next gradient.

[0009] Preferably, in step (2) above, the concentration of the perfluorinated compound added in a gradient is 1 ng / L-10 mg / L, and the concentration is gradually increased from 1 ng / L to 10 mg / L; the daily sludge discharge during the low-concentration acclimatization process is 1%-2% of the effective volume of the reactor.

[0010] Preferably, in step (3) above, the refrigerated and sealed placement is carried out at 2-6℃ for 20-30 days.

[0011] Preferably, the concentration of the perfluorinated compound added in step (4) above is 10-100 mg / L, and the concentration is gradually increased in a gradient. It can be set to 10 mg / L, 50 mg / L, 100 mg / L, or 50 mg / L, 100 mg / L.

[0012] Secondly, this invention applies to protect anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds obtained by the above-described method.

[0013] Thirdly, this invention application protects the use of the above-mentioned anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds in the synergistic removal of total nitrogen and perfluorinated compounds from wastewater.

[0014] Preferably, the total nitrogen removal rate in the above wastewater is >85%, the ammonia nitrogen removal rate is >90%, the nitrous oxide removal rate is >90%, and the removal rate is >90% when the concentration of perfluorinated compounds in the influent is below 1 mg / L.

[0015] Fourthly, this invention applies for protection of the application of the aforementioned anaerobic ammonia-oxidizing bacteria resistant to perfluorinated compounds in the remediation of riverbed sediment. Riverbed sediment refers to deposits in rivers contaminated with perfluorinated compounds.

[0016] The present invention has the following beneficial effects: This invention provides a method for cultivating anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds. The resulting anaerobic ammonia-oxidizing bacteria can withstand perfluorinated compound shocks of 1 ng / L to 100 mg / L, a concentration far exceeding that found in current wastewater. Furthermore, even in the presence of high perfluorinated compound concentrations, it maintains a total nitrogen removal rate of over 85%, thus completely solving the perfluorinated compound stress problem that may be encountered during the application of anaerobic ammonia oxidation processes. In addition, this method is simple and easy to implement, relying solely on a gradient increase in perfluorinated compound concentration combined with sludge discharge and low-temperature starvation stimulation, resulting in low cost.

[0017] The anaerobic ammonia-oxidizing bacteria culture cultivated in this application can be used to treat wastewater, achieving synergistic removal of total nitrogen and perfluorinated compounds (PFOCs). The total nitrogen removal rate is >85%, ammonia nitrogen removal rate is >90%, and nitrous oxide removal rate is >90%. When the influent PFOC concentration is below 1 mg / L, the PFOC removal rate is >90%. It can also be used to remediate riverbed sediment contaminated with PFOCs. After adding tolerant bacteria to sediment contaminated with PFOCs at concentrations of 1 ng / L, 1 μg / L, 1 mg / L, and 10 mg / L, the ammonia nitrogen removal rate increased to 7.5, 10.3, 12.5, and 16.2 times that of the contaminated sediment, respectively; the nitrous oxide removal rate increased to 6.7, 10.0, 12.1, and 16.2 times that of the contaminated sediment, respectively; and the total nitrogen removal rate increased to 7.0, 10.1, 13.3, and 15.4 times that of the contaminated sediment, respectively. Attached Figure Description

[0018] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0019] Figure 1 The changes in denitrification performance during the enrichment of anaerobic ammonia-oxidizing bacteria (using an MBR reactor).

[0020] Figure 2 This study describes the changes in denitrification performance of tolerant bacterial communities under the influence of low concentrations of perfluorobutyric acid (PFBA) during cultivation.

[0021] Figure 3 This study investigates the changes in denitrification performance of tolerant bacterial communities under the influence of high concentrations of perfluorobutyric acid (PFBA).

[0022] Figure 4 The changes in denitrification performance during the enrichment of anaerobic ammonia-oxidizing bacteria (using a UASB reactor).

[0023] Figure 5 This study describes the changes in denitrification performance of tolerant bacterial communities under the influence of low concentrations of perfluorohexanoic acid (PFNA).

[0024] Figure 6 This study describes the changes in denitrification performance of tolerant bacterial communities under the influence of high concentrations of perfluorononanoic acid (PFNA).

[0025] Figure 7 The changes in denitrification performance during the enrichment of anaerobic ammonia-oxidizing bacteria (using another MBR reactor).

[0026] Figure 8This study describes the changes in denitrification performance of tolerant bacterial communities under the influence of low concentrations of perfluorohexanoic acid (PFHxA) during cultivation.

[0027] Figure 9 This study investigated the changes in denitrification performance of tolerant bacterial communities under the influence of high concentrations of perfluorohexanoic acid (PFHxA).

[0028] Figure 10 The changes in denitrification performance during the enrichment of anaerobic ammonia-oxidizing bacteria (using another UASB reactor).

[0029] Figure 11 This study investigates the changes in denitrification performance of tolerant bacterial communities under the influence of low concentrations of perfluoropropionic acid (PFPrA) during cultivation.

[0030] Figure 12 This study investigates the changes in denitrification performance of tolerant bacterial communities under the influence of high concentrations of perfluoropropionic acid (PFPrA) during cultivation.

[0031] Figure 13 The denitrification performance of anaerobic ammonia-oxidizing bacteria without low-concentration acclimation and starvation-stimulation under high-concentration perfluorinated compound shock.

[0032] Figure 14 The denitrification performance of anaerobic ammonia-oxidizing bacteria without starvation-stimulation under high concentrations of perfluorinated compounds was studied.

[0033] Figure 15 The invention utilizes the tolerant bacterial community to repair changes in ammonia nitrogen removal rate in sediment contaminated with perfluorinated compounds.

[0034] Figure 16 The invention utilizes the tolerant bacterial community to repair changes in the nitrite removal rate of sediment contaminated with perfluorinated compounds.

[0035] Figure 17 The invention utilizes the tolerant bacterial community to repair changes in the total nitrogen removal rate of sediment contaminated with perfluorinated compounds. Detailed Implementation

[0036] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0037] Unless otherwise specified, the experimental methods used in the following experimental examples are conventional methods; the materials and reagents used are commercially available unless otherwise specified.

[0038] This invention proposes a method for cultivating anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds, aiming to enhance the tolerance of anaerobic ammonia oxidation systems to perfluorinated compounds, thereby maintaining high denitrification performance and system stability. At the same time, the tolerant bacteria can be used to remediate aquatic systems such as rivers contaminated by perfluorinated compounds and strengthen the nitrogen cycle process.

[0039] Example 1 A method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds mainly includes the following steps: 1. Enrichment Culture: First, anaerobic ammonia-oxidizing bacteria are enriched and cultured in the MBR reactor to increase their abundance. The ammonia nitrogen concentration in the influent is maintained at 95-105 mg / L (average 97 mg / L), and the nitrite concentration is maintained at 96-106 mg / L (average 98.1 mg / L). The influent nitrite / ammonia nitrogen ratio is 1.01, and the inorganic carbon source / ammonia nitrogen ratio is 2. The reaction time is adjusted according to the removal performance, and the cycle is continuously operated. When the operation time is >30 days, if the total nitrogen removal rate is >10%, the operation continues. If the total nitrogen removal rate is <10% and the change is <5% for more than 7 consecutive days, other conditions are kept unchanged, and the influent nitrite concentration is adjusted to make the influent nitrite / ammonia nitrogen ratio 0.45-0.55:1. The operation continues until the total nitrogen removal rate is >60% and the change is <5% for more than one week. Then the nitrite concentration is restored to the influent nitrite / ammonia nitrogen ratio, and the operation continues until the total nitrogen removal rate is >85% and the operation is stable for more than one month to obtain the enriched anaerobic ammonia-oxidizing bacteria.

[0040] Anaerobic ammonia-oxidizing bacteria were cultured in an MBR reactor using the enrichment culture method described above. Figure 1 As shown, by Figure 1 It was observed that after 33 days of operation, the total nitrogen removal rate remained below 10%, with a variation of less than 5% over seven consecutive days (Phase A1). Therefore, starting from day 34, the influent nitrite concentration was reduced to 50-55 mg / L, resulting in an average nitrite / ammonia nitrogen ratio of 0.53. The total nitrogen removal rate increased rapidly, reaching >60% after day 75 and continuing to increase for a week (Phase A2). From day 76 onwards, the nitrite concentration was restored to 100-105 mg / L (average 102 mg / L). After continuous operation until day 77, the total nitrogen removal rate reached 86.6%, and thereafter exceeded 85% for one month (Phase A3).

[0041] 2. Low-concentration acclimation: Perfluorinated compounds (PFBA) are added in a gradient to the stably operating anaerobic ammonia oxidation reactor, starting at 1 ng / L and gradually increasing to 10 mg / L. Each PFBA gradient constitutes a stage. After approximately 30 days of operation for each stage, if the total nitrogen removal rate is >80%, or if the total nitrogen removal rate fluctuates by <5% for more than 7 consecutive days, the influent PFBA concentration is increased to the next gradient until the PFBA concentration reaches 10 mg / L. Alternatively, if the total nitrogen removal rate remains <80% for any stage during this period, the low-concentration acclimation step is terminated, yielding low-concentration acclimated anaerobic ammonia oxidation sludge. During this operation, the daily sludge discharge is 1% of the reactor's effective volume.

[0042] Depend on Figure 2 It was found that when PFBA was added in a gradient within a stable reactor, the average total nitrogen removal rate was 85.9% after 30 days of operation with 1 ng / L PFBA (Phase B1). Then, the PFBA concentration was increased to 1 μg / L, and after another 30 days of operation, the average total nitrogen removal rate was 83.5% (Phase B2). Next, the PFBA concentration was increased to 1 mg / L, and after another 30 days of operation, the average total nitrogen removal rate was 80.6% (Phase B3). Finally, the PFBA concentration was increased to 10 mg / L, and after 30 days of operation, the total nitrogen removal rate was <80%, at which point the low-concentration acclimation step was terminated (Phase B4).

[0043] 3. Low-Temperature Starvation Stimulation: After the anaerobic ammonia oxidation sludge mixture with low concentrations was allowed to settle and the supernatant was discharged, the settled sludge was refrigerated and sealed at 4°C for 21 days without any substrate, thus putting the sludge in a starvation state. This sludge consumed extracellular polymers and endogenous carbon sources, reducing its inhibition of mass transfer and promoting the release of signaling molecules, thereby enhancing its tolerance and stress resistance. Afterward, it was removed and placed in the original reactor for continued operation. The low-temperature environment reduced the expression of relevant metabolic genes and decreased the activity of related enzymes in other denitrifying bacteria that compete with anaerobic ammonia oxidizing bacteria.

[0044] 4. High-Concentration Enhancement: The anaerobic ammonia oxidation sludge, after low-temperature-starvation stimulation, was removed and placed back into the original reactor. It was then operated at room temperature with the original substrate to restore its activity until the total nitrogen removal rate recovered to >80% or the variation over 7 consecutive days was <5%, indicating that its activity was basically stable. Subsequently, high-concentration perfluorinated compounds (PFOCs) of 50-100 mg / L were added to the influent in stages to verify the sludge's tolerance to PFOCs. Simultaneously, high-concentration stimulation was used to further enhance its tolerance. After the total nitrogen removal rate reached >80% or the variation over 7 consecutive days was <5%, the PFOC concentration was increased to proceed to the next stage until the influent PFOC concentration reached 100 mg / L. Stable operation was maintained for one month to obtain a perfluorinated anaerobic ammonia oxidation bacterial community.

[0045] Results of the high-concentration enhancement phase following refrigeration-starvation stimulation are as follows: Figure 3 As shown, by Figure 3 It was found that after 21 days of starvation-refrigeration stimulation, the sludge was removed and placed back into the original MBR reactor to restore its activity. The total nitrogen removal rate increased from below 10% initially to 80% on day 21, at which point its activity was basically stable (stage C1). Subsequently, 50 mg / L of PFBA was added to the influent, and the total nitrogen removal rate remained stable at 78.5% for 30 days (stage C2). Increasing the PFBA concentration to 100 mg / L maintained the total nitrogen removal rate and increased it to over 85%, operating stably for one month (stage C3).

[0046] 5. Synergistic Removal: The anaerobic ammonia-oxidizing bacteria cultured above can achieve synergistic removal of total nitrogen and perfluorinated compounds in wastewater. The total nitrogen removal rate is >85%, the ammonia nitrogen removal rate is >90%, and the nitrous oxide removal rate is >90%. When the perfluorinated compound concentration in the influent is below 1 mg / L, the perfluorinated compound removal rate is >90%.

[0047] Example 2 A method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds mainly includes the following steps: 1. Enrichment Culture: Anaerobic ammonia-oxidizing bacteria were first enriched and cultured in the UASB reactor to increase their abundance. The ammonia nitrogen concentration in the influent was maintained at 93-105 mg / L (average 98.7 mg / L), and the nitrite concentration at 95-109 mg / L (average 102.3 mg / L). The influent nitrite / ammonia nitrogen ratio is 1.05, and the inorganic carbon source / ammonia nitrogen ratio is 4. The reaction time is adjusted according to the removal performance, and the cycle is continuously operated in this manner: if the total nitrogen removal rate is >10% after 30 days, the operation continues; if the total nitrogen removal rate is <10% and the change is <5% for more than 7 consecutive days, other conditions are kept unchanged, and the influent nitrite concentration is adjusted to make the influent nitrite / ammonia nitrogen ratio 0.45-0.55:1; the operation continues until the total nitrogen removal rate is >60% and the change is <5% for more than one week, then the nitrite concentration is restored to the influent nitrite / ammonia nitrogen ratio, and the operation continues until the total nitrogen removal rate is >85% and the operation is stable for more than one month to obtain the enriched anaerobic ammonia-oxidizing bacteria community.

[0048] Anaerobic ammonia-oxidizing bacteria were cultured in a UASB reactor using the above method. Figure 4 After that, by Figure 4 It was observed that after 35 days of operation, the total nitrogen removal rate remained below 10% and fluctuated by less than 5% over seven consecutive days (Phase D1). Therefore, starting from day 36, the influent nitrite concentration was reduced to 49-53 mg / L, resulting in an average nitrite / ammonia nitrogen ratio of 0.52. The total nitrogen removal rate increased rapidly, reaching 63.7% on day 67 and continuing to increase for the following week (Phase D2). From day 74 onwards, the nitrite concentration was restored to 100-105 mg / L (average 102 mg / L). After continuous operation until day 76, the total nitrogen removal rate reached 85.3%, and remained above 85% for the following month (Phase D3).

[0049] 2. Low-concentration acclimatization: Perfluorinated compounds (perfluorononanoic acid, PFNA) are added in a gradient to the stably operating anaerobic ammonia oxidation reactor, starting at 1 ng / L and gradually increasing to 1 mg / L. Each PFNA gradient constitutes a stage. After each stage lasts approximately 22 days, if the total nitrogen removal rate is >80%, or if the total nitrogen removal rate fluctuates by less than 5% for more than 7 consecutive days, the influent PFNA concentration is increased to the next gradient until the PFNA concentration reaches 1 mg / L. Alternatively, if the total nitrogen removal rate remains <80% and shows a continuous downward trend during this period, the low-concentration acclimatization step is terminated. During this operation, the daily sludge discharge is 2% of the reactor's effective volume.

[0050] Depend on Figure 5It can be seen that when PFNA is added in a gradient within a stable reactor, initially at 1 ng / L for 22 days, the average total nitrogen removal rate is 80.6%, with occasional fluctuations below 80% in the last few days and a change of <5% for 7 consecutive days (Phase E1). Then, the PFNA concentration is increased to 1 μg / L, and after another 22 days, the average total nitrogen removal rate is 80.8% (Phase E2). Next, the PFNA concentration is increased to 1 mg / L, and after another 22 days, the average total nitrogen removal rate is 77.0%. During this phase, the total nitrogen removal rate remains <80% and shows a gradual decreasing trend, thus ending the low-concentration acclimatization step (Phase E3).

[0051] 3. Low-Temperature Starvation Stimulation: After the anaerobic ammonia oxidation sludge mixture with low concentrations was allowed to settle and the supernatant was discharged, the settled sludge was refrigerated and sealed at 4°C for 21 days without any substrate, thus putting the sludge in a starvation state. This sludge consumed extracellular polymers and endogenous carbon sources, reducing its inhibition of mass transfer and promoting the release of signaling molecules, thereby enhancing its tolerance and stress resistance. Afterward, it was removed and placed in the original reactor for continued operation. The low-temperature environment reduced the expression of relevant metabolic genes and decreased the activity of related enzymes in other denitrifying bacteria that compete with anaerobic ammonia oxidizing bacteria.

[0052] 4. High-Concentration Enhancement: The anaerobic ammonia oxidation sludge, after low-temperature-starvation stimulation, was removed and placed back into the original reactor. It was then operated at room temperature with the original substrate to restore its activity until the total nitrogen removal rate recovered to >80% or the variation over 7 consecutive days was <5%, indicating that its activity was basically stable. Subsequently, high-concentration perfluorinated compounds (PFOCs) of 50-100 mg / L were added to the influent in stages to verify the sludge's tolerance to PFOCs. Simultaneously, high-concentration stimulation was used to further enhance its tolerance. After the total nitrogen removal rate reached >80% or the variation over 7 consecutive days was <5%, the PFOC concentration was increased to proceed to the next stage until the influent PFOC concentration reached 100 mg / L. Stable operation was maintained for one month to obtain a perfluorinated anaerobic ammonia oxidation bacterial community.

[0053] Results of the high-concentration enhancement phase are as follows Figure 6 As shown, by Figure 6 It was found that after 21 days of starvation-refrigeration stimulation, the sludge was removed and placed back into the original UASB reactor to restore its activity. The total nitrogen removal rate increased to 80% on day 6, then fluctuated slightly, and its activity stabilized after 21 days of operation (stage F1). Subsequently, 50 mg / L of PFNA was added to the influent, and the average total nitrogen removal rate was 85.1% (stage F2). The PFNA concentration was further increased to 100 mg / L, and the total nitrogen removal rate remained stable at an average of 86.3%, and it operated stably for one month (stage F3).

[0054] 5. Synergistic Removal: The anaerobic ammonia-oxidizing bacteria cultured above can achieve synergistic removal of total nitrogen and perfluorinated compounds in wastewater. The total nitrogen removal rate is >85%, the ammonia nitrogen removal rate is >90%, and the nitrous oxide removal rate is >90%. When the perfluorinated compound concentration in the influent is below 1 mg / L, the perfluorinated compound removal rate is >90%.

[0055] Example 3 A method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds mainly includes the following steps: 1. Enrichment Culture: First, anaerobic ammonia-oxidizing bacteria are enriched and cultured in the MBR reactor to increase their abundance. The ammonia nitrogen concentration in the influent is maintained at 85-100 mg / L (average 96 mg / L), and the nitrite concentration is maintained at 92-104 mg / L (average 100 mg / L). The influent nitrite / ammonia nitrogen ratio is 1.04, and the inorganic carbon source / ammonia nitrogen ratio is 3. The reaction time is adjusted according to the removal performance, and the cycle is continuously operated in this manner: if the total nitrogen removal rate is >10% after 30 days, the operation continues; if the total nitrogen removal rate is <10% and the change is <5% for more than 7 consecutive days, other conditions are kept unchanged, and the influent nitrite concentration is adjusted to make the influent nitrite / ammonia nitrogen ratio 0.45-0.55:1; the operation continues until the total nitrogen removal rate is >60% and the change is <5% for more than one week, then the nitrite concentration is restored to the influent nitrite / ammonia nitrogen ratio, and the operation continues until the total nitrogen removal rate is >85% and the operation is stable for more than one month to obtain the enriched anaerobic ammonia-oxidizing bacteria community.

[0056] Anaerobic ammonia-oxidizing bacteria were cultured in an MBR reactor using the above method. Figure 7 ),Depend on Figure 7 It can be seen that after 30 days of operation, the total nitrogen removal rate is >10% (stage G1). Continuing operation until day 76, the total nitrogen removal rate reaches 85.6%, and thereafter the total nitrogen removal rate is >85% for one month (stage G2).

[0057] 2. Low-concentration acclimation: Perfluorinated compounds (perfluorohexanoic acid PFHxA) are added in a gradient to the stably operating anaerobic ammonia oxidation reactor, starting at 1 ng / L and gradually increasing to 10 mg / L. Each perfluorinated compound gradient constitutes a stage. After each stage has been running for approximately 30 days, if the total nitrogen removal rate is >80%, or if the total nitrogen removal rate fluctuates by <5% for more than 7 consecutive days, the influent perfluorinated compound concentration is increased to the next gradient, until the perfluorinated compound concentration reaches 5 mg / L. Alternatively, if the total nitrogen removal rate remains <80% and shows a gradual decreasing trend during this period, the low-concentration acclimation step is terminated. During this operation, the daily sludge discharge is 1% of the reactor's effective volume.

[0058] Depend on Figure 8It can be seen that when PFHxA is added in a gradient within a stable reactor, the average total nitrogen removal rate is 85.9% after 30 days of operation with 1 ng / L added initially (stage H1). Then, the PFHxA concentration is increased to 1 μg / L, and after another 30 days of operation, the average total nitrogen removal rate is 83.5% (stage H2). Next, the PFHxA concentration is increased to 1 mg / L, and after another 30 days of operation, the average total nitrogen removal rate is 80.6% (stage H3). Finally, the PFHxA concentration is increased to 10 mg / L, and after 30 days of operation, the total nitrogen removal rate is <80%, at which point the low-concentration acclimation step is terminated (stage H4).

[0059] 3. Low-Temperature Starvation Stimulation: After the anaerobic ammonia oxidation sludge mixture with low concentrations was allowed to settle and the supernatant was discharged, the settled sludge was refrigerated and sealed at 4°C for 21 days without any substrate, thus putting the sludge in a starvation state. This sludge consumed extracellular polymers and endogenous carbon sources, reducing its inhibition of mass transfer and promoting the release of signaling molecules, thereby enhancing its tolerance and stress resistance. Afterward, it was removed and placed in the original reactor for continued operation. The low-temperature environment reduced the expression of relevant metabolic genes and decreased the activity of related enzymes in other denitrifying bacteria that compete with anaerobic ammonia oxidizing bacteria.

[0060] 4. High-Concentration Enhancement: The anaerobic ammonia oxidation sludge, after low-temperature-starvation stimulation, was removed and placed back into the original reactor. It was then operated at room temperature with the original substrate to restore its activity until the total nitrogen removal rate recovered to >80% or the variation over 7 consecutive days was <5%, indicating that its activity was basically stable. Subsequently, high-concentration perfluorinated compounds (PFOCs) of 50-100 mg / L were added to the influent in stages to verify the sludge's tolerance to PFOCs. Simultaneously, high-concentration stimulation was used to further enhance its tolerance. After the total nitrogen removal rate reached >80% or the variation over 7 consecutive days was <5%, the PFOC concentration was increased to proceed to the next stage until the influent PFOC concentration reached 100 mg / L. Stable operation was maintained for one month to obtain a perfluorinated anaerobic ammonia oxidation bacterial community.

[0061] Results of the high-concentration enhancement phase are as follows Figure 9 As shown, by Figure 9 It can be seen that after 21 days of starvation-refrigeration stimulation, the sludge was removed and placed back into the original MBR reactor to restore its activity. The total nitrogen removal rate increased from less than 10% initially to 80% on day 7, and then remained stable, at which point its activity was basically stable (stage I1). On day 22, 50 mg / L of PFHxA was added to the influent, and the total nitrogen removal rate remained stable at around 86.3% for 30 days (stage I2). Increasing the PFHxA concentration to 100 mg / L maintained the total nitrogen removal rate and increased it to over 85%, and it operated stably for one month (stage I3).

[0062] 5. Synergistic Removal: The anaerobic ammonia-oxidizing bacteria cultured above can achieve synergistic removal of total nitrogen and perfluorinated compounds in wastewater. The total nitrogen removal rate is >85%, the ammonia nitrogen removal rate is >90%, and the nitrous oxide removal rate is >90%. When the perfluorinated compound concentration in the influent is below 1 mg / L, the perfluorinated compound removal rate is >90%.

[0063] Example 4 A method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds mainly includes the following steps: 1. Enrichment Culture: Anaerobic ammonia-oxidizing bacteria were first enriched and cultured in the UASB reactor to increase their abundance. The ammonia nitrogen concentration in the influent was maintained at 90-104 mg / L (average 98.0 mg / L), and the nitrite concentration at 94-106 mg / L (average 100.8 mg / L). The influent nitrite / ammonia nitrogen ratio is 1.03, and the inorganic carbon source / ammonia nitrogen ratio is 2. The reaction time is adjusted according to the removal performance, and the cycle is continuously operated. When the operation time is >30 days, if the total nitrogen removal rate is >10%, the operation continues. If the total nitrogen removal rate is <10% and the change is <5% for more than 7 consecutive days, other conditions are kept unchanged, and the influent nitrite concentration is adjusted to make the influent nitrite / ammonia nitrogen ratio 0.45-0.55:1. The operation continues until the total nitrogen removal rate is >60% and the change is <5% for more than one week. Then the nitrite concentration is restored to the influent nitrite / ammonia nitrogen ratio, and the operation continues until the total nitrogen removal rate is >85% and the operation is stable for more than one month to obtain the enriched anaerobic ammonia-oxidizing bacteria.

[0064] Anaerobic ammonia-oxidizing bacteria were cultured in a UASB reactor using the above method. Figure 10 ),Depend on Figure 10 It was observed that after 33 days of operation, the total nitrogen removal rate remained below 10%, with a variation of less than 5% over seven consecutive days (Phase J1). Therefore, starting from day 34, the influent nitrite concentration was reduced to 52-57 mg / L, resulting in an average nitrite / ammonia nitrogen ratio of 0.55. The total nitrogen removal rate increased slowly, so on day 55, sludge was appropriately discharged to avoid excessively high sludge concentration affecting mass transfer. Afterward, the total nitrogen removal rate increased rapidly, reaching 60.1% after day 73 and continuing to increase for a week (Phase J2). From day 80 onward, the nitrite concentration was restored to 94-106 mg / L (average 102 mg / L), and the total nitrogen removal rate reached 85.9% on day 81. For the following month, the total nitrogen removal rate was >85% (Phase J3).

[0065] 2. Low-concentration acclimation: Perfluorinated compounds (perfluoropropionic acid, PFPrA) are added in a gradient to the stably operating anaerobic ammonia oxidation reactor, starting at 1 ng / L and gradually increasing to 10 mg / L. Each perfluorinated compound gradient constitutes a stage. After each stage runs for approximately 22 days, if the total nitrogen removal rate is >80%, or if the total nitrogen removal rate varies by <5% for more than 7 consecutive days, the influent perfluorinated compound concentration is increased to the next gradient, until the perfluorinated compound concentration reaches 10 mg / L, or if the total nitrogen removal rate remains <80% throughout any stage during this period, then the low-concentration acclimation step is terminated. During this operation, the daily sludge discharge is 1.5% of the reactor's effective volume.

[0066] Depend on Figure 11 It can be seen that when PFPrA is added in a gradient within a stable reactor, the average total nitrogen removal rate is 85.6% after 22 days of operation with an initial concentration of 1 ng / L (stage K1). Then, the PFPrA concentration is increased to 1 μg / L, and after another 22 days of operation, the average total nitrogen removal rate is 81.3% (stage K2). Next, the PFPrA concentration is increased to 1 mg / L, and after another 22 days of operation, the average total nitrogen removal rate is 82.9% (stage K3). Finally, the PFPrA concentration is increased to 10 mg / L, and the total nitrogen removal rate decreases significantly. During this period, the total nitrogen removal rate remains below 80% and gradually decreases, thus ending this stage (stage K4).

[0067] 3. Low-Temperature Starvation Stimulation: After the anaerobic ammonia oxidation sludge mixture with low concentrations was allowed to settle and the supernatant was discharged, the settled sludge was refrigerated and sealed at 4°C for 25 days without any substrate, thus putting the sludge in a starvation state. This sludge consumed extracellular polymers and endogenous carbon sources, reducing its inhibition of mass transfer and promoting the release of signaling molecules, thereby enhancing its tolerance and stress resistance. Afterward, it was removed and placed in the original reactor for continued operation. The low-temperature environment reduced the expression of relevant metabolic genes and decreased the activity of related enzymes in other denitrifying bacteria that compete with anaerobic ammonia oxidizing bacteria.

[0068] 4. High-Concentration Enhancement: The anaerobic ammonia oxidation sludge, after low-temperature-starvation stimulation, was removed and placed back into the original reactor. It was then operated at room temperature with the original substrate to restore its activity until the total nitrogen removal rate recovered to >80% or the variation over 7 consecutive days was <5%, indicating that its activity was basically stable. Subsequently, high-concentration perfluorinated compounds (PFOCs) of 50-100 mg / L were added to the influent in stages to verify the sludge's tolerance to PFOCs. Simultaneously, high-concentration stimulation was used to further enhance its tolerance. After the total nitrogen removal rate reached >80% or the variation over 7 consecutive days was <5%, the PFOC concentration was increased to proceed to the next stage until the influent PFOC concentration reached 100 mg / L. Stable operation was maintained for one month to obtain a perfluorinated anaerobic ammonia oxidation bacterial community.

[0069] Results of the high-concentration enhancement phase following refrigeration-starvation stimulation are as follows: Figure 12 As shown, by Figure 12 It was found that after 25 days of starvation-refrigeration stimulation, the sludge was removed and placed back into the original UASB reactor to restore its activity. The total nitrogen removal rate increased from 0% initially to 81.4% on day 4, after which its activity remained relatively stable (stage L1). Subsequently, 50 mg / L of PFPrA was added to the influent, and the total nitrogen removal rate remained stable at 85.7% for 30 days (stage L2). Increasing the PFPrA concentration to 100 mg / L maintained the total nitrogen removal rate and increased it to over 85%, operating stably for one month (stage L3).

[0070] 5. Synergistic Removal: The anaerobic ammonia-oxidizing bacteria cultured above can achieve synergistic removal of total nitrogen and perfluorinated compounds in wastewater. The total nitrogen removal rate is >85%, the ammonia nitrogen removal rate is >90%, and the nitrous oxide removal rate is >90%. When the perfluorinated compound concentration in the influent is below 1 mg / L, the perfluorinated compound removal rate is >90%.

[0071] Example 5 A method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds mainly includes the following steps: 1. Enrichment Culture: First, enrich and culture anaerobic ammonia-oxidizing bacteria in the reactor to increase their abundance. Maintain a nitrite to ammonia nitrogen molar ratio of 1 in the influent, and provide an inorganic carbon source of 200 mg / L. Adjust the reaction time according to the removal performance, and continue this process continuously. If the total nitrogen removal rate is >10% after 30 days, continue operation. If the total nitrogen removal rate is <10% and the change is <5% for more than 7 consecutive days, keep other conditions unchanged and adjust the influent nitrite concentration to 0.45. Continue operation until the total nitrogen removal rate is >60% and the change is <5% for one week. Then restore the nitrite concentration to 1 and continue operation until the total nitrogen removal rate is >85% and stable operation is maintained for more than one month to obtain the enriched anaerobic ammonia-oxidizing bacteria.

[0072] 2. Low-concentration acclimatization: Perfluorinated compounds are gradually added to the stably operating anaerobic ammonia oxidation reactor, starting from 1 ng / L and increasing to 1 mg / L. Each perfluorinated compound gradient constitutes a stage. After each stage has been running for approximately 20 days, if the total nitrogen removal rate is >80%, or if the total nitrogen removal rate varies by <5% for more than 7 consecutive days, the influent perfluorinated compound concentration is increased to the next gradient, until the perfluorinated compound concentration reaches 10 mg / L, or if the total nitrogen removal rate remains <80% for any stage during this period, then the low-concentration acclimatization step is terminated. During this operation, the daily sludge discharge is 1% of the reactor's effective volume.

[0073] 3. Low-Temperature Starvation Stimulation: After the anaerobic ammonia oxidation sludge-water mixture with low concentration acclimation was allowed to settle and the supernatant was discharged, the settled sludge was refrigerated and sealed at 2°C for 20 days. During this process, no substrate was provided, putting the sludge in a starvation state. This prompted it to consume extracellular polymers and endogenous carbon sources, reducing its inhibition of mass transfer and simultaneously promoting the release of signaling molecules, thereby enhancing its tolerance and stress resistance. Afterward, it was removed and placed in the original reactor for continued operation. The low-temperature environment reduced the expression of relevant metabolic genes of other denitrifying bacteria competing with anaerobic ammonia oxidizing bacteria and decreased the activity of related enzymes.

[0074] 4. High-Concentration Enhancement: The anaerobic ammonia oxidation sludge, after being subjected to low-temperature starvation stimulation, was removed and placed back into the original reactor. It was then operated at room temperature with the original substrate to restore its activity until the total nitrogen removal rate recovered to >80% or the variation over 7 consecutive days was <5%, indicating that its activity was basically stable. Subsequently, high-concentration perfluorinated compounds (PFOCs) of 10-100 mg / L were added to the influent in stages to verify the sludge's tolerance to PFOCs. Simultaneously, high-concentration stimulation was used to further enhance its tolerance. After the total nitrogen removal rate reached >80% or the variation over 7 consecutive days was <5%, the PFOC concentration was increased to proceed to the next stage until the influent PFOC concentration reached 100 mg / L. Stable operation was maintained for one month to obtain a perfluorinated tolerant anaerobic ammonia oxidation bacterial community.

[0075] Example 6 A method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds mainly includes the following steps: 1. Enrichment Culture: First, enrich and culture anaerobic ammonia-oxidizing bacteria in the reactor to increase their abundance. Maintain a nitrite to ammonia nitrogen molar ratio of 1 in the influent, and provide an inorganic carbon source of 200 mg / L. Adjust the reaction time according to the removal performance, and continue this process continuously. If the total nitrogen removal rate is >10% after 30 days, continue operation. If the total nitrogen removal rate is <10% and the change is <5% for more than 7 consecutive days, keep other conditions unchanged and adjust the influent nitrite concentration to 0.50. Continue operation until the total nitrogen removal rate is >60% and the change is <5% for one week. Then restore the nitrite concentration to 1 and continue operation until the total nitrogen removal rate is >85% and stable operation is maintained for more than one month to obtain the enriched anaerobic ammonia-oxidizing bacteria.

[0076] 2. Low-concentration acclimatization: Perfluorinated compounds are gradually added to the stably operating anaerobic ammonia oxidation reactor, starting from 1 ng / L and increasing to 1 mg / L. Each perfluorinated compound gradient constitutes a stage. After each stage has been running for approximately 20 days, if the total nitrogen removal rate is >80%, or if the total nitrogen removal rate varies by <5% for more than 7 consecutive days, the influent perfluorinated compound concentration is increased to the next gradient, until the perfluorinated compound concentration reaches 10 mg / L, or if the total nitrogen removal rate remains <80% for any stage during this period, then the low-concentration acclimatization step is terminated. During this operation, the daily sludge discharge is 1% of the reactor's effective volume.

[0077] 3. Low-Temperature Starvation Stimulation: After the anaerobic ammonia oxidation sludge-water mixture with low concentration acclimation was allowed to settle and the supernatant was discharged, the settled sludge was refrigerated and sealed at 6℃ for 30 days. During this process, no substrate was provided, putting the sludge in a starvation state. This prompted it to consume extracellular polymers and endogenous carbon sources, reducing its inhibition of mass transfer and simultaneously promoting the release of signaling molecules, thereby enhancing its tolerance and stress resistance. Afterward, it was removed and placed in the original reactor for continued operation. The low-temperature environment reduced the expression of relevant metabolic genes of other denitrifying bacteria competing with anaerobic ammonia oxidizing bacteria and decreased the activity of related enzymes.

[0078] 4. High-Concentration Enhancement: The anaerobic ammonia oxidation sludge, after being subjected to low-temperature starvation stimulation, was removed and placed back into the original reactor. It was then operated at room temperature with the original substrate to restore its activity until the total nitrogen removal rate recovered to >80% or the variation over 7 consecutive days was <5%, indicating that its activity was basically stable. Subsequently, high-concentration perfluorinated compounds (PFOCs) of 10-100 mg / L were added to the influent in stages to verify the sludge's tolerance to PFOCs. Simultaneously, high-concentration stimulation was used to further enhance its tolerance. After the total nitrogen removal rate reached >80% or the variation over 7 consecutive days was <5%, the PFOC concentration was increased to proceed to the next stage until the influent PFOC concentration reached 100 mg / L. Stable operation was maintained for one month to obtain a perfluorinated tolerant anaerobic ammonia oxidation bacterial community.

[0079] Comparative Example 1 The difference between this comparative example and Example 2 is that after enrichment culture in step 1 to obtain the enriched anaerobic ammonia-oxidizing bacteria, the low-concentration acclimatization in step 2 and the low-temperature-starvation stimulation in step 3 are not carried out. Instead, a high concentration of PFNA is directly added for high-concentration enhancement in step 4.

[0080] The denitrification performance of anaerobic ammonia-oxidizing bacteria without low-concentration acclimation and low-temperature-starvation stimulation under high-concentration perfluorinated compound shock is as follows: Figure 13 As stated, by Figure 13It can be seen that before the addition of PFNA, the total nitrogen removal rate was stable at around 87.6% (stage M1). After adding 50 mg / L of PFNA, the total nitrogen removal rate decreased from 87% to 62% within 21 days (stage M2). Further addition of 100 mg / L of PFNA resulted in a continuous decrease in the total nitrogen removal rate, eventually reducing it to almost zero (stage M3). This comparative example demonstrates that although the anaerobic ammonia-oxidizing bacteria exhibit good activity after acclimatization, without the two steps of low-concentration acclimatization and low-temperature-starvation stimulation, these bacteria cannot withstand the impact of high concentrations of perfluorinated compounds.

[0081] Comparative Example 2 The difference between this comparative example and Example 2 is that after enrichment culture in step 1 to obtain the enriched anaerobic ammonia-oxidizing bacteria, low-concentration acclimatization was carried out in step 2, but without the low temperature-starvation stimulation in step 3, a high concentration of PFNA was directly added for high-concentration enhancement in step 4.

[0082] The denitrification performance of anaerobic ammonia-oxidizing bacteria without low-temperature-starvation stimulation under high-concentration perfluorinated compound shock is as follows: Figure 14 As stated, by Figure 14 It was observed that after low-concentration acclimation, the total nitrogen removal rate stabilized at 80% (stage N1), consistent with the results of Example 2. After adding 50 mg / L PFNA, the total nitrogen removal rate decreased from 81% to 54% within 21 days (stage N2); with continued addition of 100 mg / L PFNA, the total nitrogen removal rate continued to decline, eventually reaching almost zero (stage N3). PFNA addition was stopped to observe recovery; after one month of operation, the total nitrogen removal rate remained below 5% (stage N4). These comparative results demonstrate that although anaerobic ammonia oxidizing bacteria maintained good activity after low-concentration acclimation, they still could not tolerate the impact of high concentrations of perfluorinated compounds without low-temperature-starvation stimulation; moreover, their activity could not recover after the removal of perfluorinated compound stress.

[0083] Application examples The anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds (hereinafter referred to as tolerant bacteria) cultured using the method of the present invention are used to remediate riverbed sediment contaminated with perfluorinated compounds. The steps are as follows: The cultured tolerant bacteria (cultivated in Example 1) were added to the riverbed sediment at a dry weight ratio of 1:20, and their denitrification rate under different concentrations of perfluorinated compound exposure was tested. Five parallel sediment samples were prepared. In sediment uncontaminated with PFBA, the ammonia nitrogen removal rate was 1.111 mg / (g·d), the nitrous oxide removal rate was 0.974 mg / (g·d), and the total nitrogen removal rate was 1.987 mg / (g·d). In sediment contaminated with PFBA at concentrations of 1 ng / L, 1 μg / L, 1 mg / L, and 10 mg / L, the ammonia nitrogen removal rates were 0.736, 0.528, 0.424, and 0.319 mg / (g·d), the nitrous oxide removal rates were 0.774, 0.505, 0.413, and 0.308 mg / (g·d), and the total nitrogen removal rates were 1.408, 0.955, 0.722, and 0.608 mg / (g·d). The tolerant bacterial populations cultured using the method provided in Example 1 of this invention were added to the PFBA-contaminated sediment at mg / L for remediation. After running under PFBA exposure conditions for a period of time, the ammonia nitrogen removal rate was as follows: Figure 15 As shown, the ammonia nitrogen removal rates increased to 5.553, 5.430, 5.307, and 5.183 mg / (g·d), respectively. Compared with the contaminated sediment, the ammonia nitrogen removal rates of the remediated sediment increased to 7.5, 10.3, 12.5, and 16.2 times that of the contaminated sediment, respectively; the nitrous oxide removal rate was as follows: Figure 16 As shown, the nitrogen removal rates increased to 5.167, 5.077, 5.017, and 4.992 mg / (g·d), respectively, and the nitrogen removal rates of the remediated sediment increased to 6.7, 10.0, 12.1, and 16.2 times that of the contaminated sediment, respectively; the total nitrogen removal rates were as follows: Figure 17 As shown, the total nitrogen removal rate was stable at 9.844, 9.638, 9.617, and 9.355 mg / (g·d). Compared with the contaminated sediment, the total nitrogen removal rate of the remediated sediment was increased to 7.0, 10.1, 13.3, and 15.4 times that of the contaminated sediment, respectively.

[0084] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds, characterized in that, The steps are as follows: (1) Enrichment culture: Simulated wastewater containing ammonia nitrogen, nitrous oxide and inorganic carbon source is used as influent in the reactor. The reaction time is adjusted according to the total nitrogen removal rate. After circulation, the reactor is stably operated to obtain the enriched anaerobic ammonia-oxidizing bacteria. (2) Low concentration acclimatization: Perfluorinated compounds are added in a gradient to the anaerobic ammonia oxidation bacteria culture enriched in step (1) until the concentration of perfluorinated compounds reaches 1-10 mg / L, or the total nitrogen removal rate is always <80% and has a gradual downward trend. Then the low concentration acclimatization is ended and the anaerobic ammonia oxidation sludge after low concentration acclimatization is obtained. (3) Low temperature-starvation stimulation: The anaerobic ammonia oxidation sludge after low concentration acclimation in step (2) was left to stand, and the sediment was refrigerated and sealed to obtain anaerobic ammonia oxidation sludge under low temperature-starvation stimulation. (4) High concentration enhancement: The anaerobic ammonia oxidation sludge after low temperature-starvation stimulation in step (3) is placed in the reactor and run until the total nitrogen removal rate recovers to >80% or the change range is <5% for 7 consecutive days. Then, perfluorinated compounds are added to the influent in a gradient until the perfluorinated compound concentration reaches 50-100 mg / L. The reactor is run stably for more than 30 days to obtain anaerobic ammonia oxidation bacteria that are tolerant to perfluorinated compounds.

2. The method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds according to claim 1, characterized in that: In step (1), the molar concentration ratio of nitrous oxide to ammonia nitrogen in the influent is 1-1.05:1, and the mass concentration ratio of inorganic carbon source to ammonia nitrogen is 2-4:

1.

3. The method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds according to claim 2, characterized in that, The step (1) of adjusting the reaction time according to the total nitrogen removal rate is as follows: if the total nitrogen removal rate is >10% when the running time is >30 days, continue running; if the total nitrogen removal rate is <10% and the change range is <5% for more than 7 consecutive days, keep other conditions unchanged and adjust the influent nitrous oxide concentration to make the influent nitrous oxide / ammonia nitrogen ratio 0.45-0.55:1; continue running until the total nitrogen removal rate is >60% and the change range is <5% for more than 7 consecutive days, then restore the nitrous oxide concentration to the influent nitrous oxide / ammonia nitrogen ratio of 1-1.05:1, and continue running until the total nitrogen removal rate is >85% and stable operation is achieved for more than 30 days to reach a stable operating state.

4. The method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds according to claim 1, characterized in that: In step (2), the concentration of perfluorinated compounds added in a gradient is 1 ng / L-10 mg / L; the daily sludge discharge during the low-concentration acclimatization process is 1%-2% of the effective volume of the reactor.

5. The method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds according to claim 1, characterized in that: In step (3), the temperature for refrigeration and sealing is 2-6℃ and the time is 20-30 days.

6. The method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds according to claim 1, characterized in that: In step (4), the concentration of the perfluorinated compound added in a gradient is 10-100 mg / L.

7. The method for culturing anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds according to any one of claims 1-6, characterized in that, The step of adding the perfluorinated compound in gradients is as follows: each gradient is a stage. After a stage runs for 20-30 days, if the total nitrogen removal rate is >80%, or the total nitrogen removal rate changes by <5% for more than 7 consecutive days, then the concentration of perfluorinated compound is increased to the next gradient.

8. Anaerobic ammonia-oxidizing bacteria tolerant to perfluorinated compounds, cultured using the method of claim 7.

9. The application of the anaerobic ammonia-oxidizing bacteria community tolerant to perfluorinated compounds as described in claim 8 in the synergistic removal of total nitrogen and perfluorinated compounds from wastewater.

10. The application of the perfluorinated anaerobic ammonia-oxidizing bacteria community described in claim 8 in the remediation of riverbed sediment.

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