New ecological halotolerant bacteria and method for enhancing cascade energy treatment of wastewater by using new ecological halotolerant bacteria

Abstract

The invention discloses a nascent-state halotolerant bacterium and a method for enhancing cascade energy treatment of wastewater by using the nascent-state halotolerant bacterium, and belongs to the technical field of organic wastewater treatment. According to the invention, a novel gradient energy regeneration reactor CEAR is designed, hydrogen and methane are respectively generated in an inner cylinder and an outer cylinder, and the CEAR is strengthened by utilizing a new ecological halotolerant bacterium Bacillus altitudinis K3, so that the treatment effect on high-salinity kitchen wastewater is superior to that of a traditional UASB (Upflow Anaerobic Sludge Blanket). According to the invention, the CEAR reactor is adopted to treat kitchen wastewater with different organic loads and salinity, and when the reaction is finished, the SCOD removal rate and methane yield are respectively improved by 15.5%-23.5% and 20.0%-25.0% compared with UASB (Upflow Anaerobic Sludge Blanket).

Description

Technical Field

[0001] The present invention relates to a new ecological halotolerant bacterium and a method for enhancing the cascade energy conversion treatment of wastewater, belonging to the technical field of organic wastewater treatment. Background Art

[0002] With the rapid development of the catering industry, the generation of kitchen waste is increasing, and the wastewater content is as high as 90%. Kitchen wastewater is the high-concentration organic wastewater after the solid-liquid separation of kitchen waste, and has become one of the important sources of water pollution.

[0003] The composition of kitchen wastewater is mainly animal and vegetable oils based on macromolecular lipids and their derivatives, as well as a large amount of biodegradable organic matters such as carbohydrates and proteins. In addition, kitchen wastewater has a high salt content and contains trace elements such as potassium, calcium, iron, and phosphorus. The rich organic substances in kitchen wastewater are prone to fermentation and deterioration, generating malodors and breeding germs. If not effectively treated, it will cause environmental pollution and endanger human health. Therefore, the treatment of kitchen wastewater has become the focus of public attention. Kitchen wastewater also has the potential value of being recycled. By anaerobic digestion, organic matters can be converted into energy gases such as H2 and CH4, which will inject strong impetus into environmental governance and economic and social development.

[0004] At present, there are still some problems in using traditional anaerobic digestion to treat kitchen wastewater. For example, it has poor impact resistance to high salt and high organic load, is difficult to achieve gradient energy conversion, has low purity of biogas, and low COD removal rate.

[0005] Therefore, the bioaugmented anaerobic digestion technology has gradually developed, and the most commonly used method is to add halotolerant bacteria. The added new ecological halotolerant bacteria can induce the formation of dominant halotolerant bacterial communities in anaerobic sludge. The generated new ecological halotolerant bacterial communities can enhance the electron transfer rate and increase the activity of metabolic enzymes, thereby improving the salt tolerance performance of the anaerobic digestion system. Further enhance the biogas production efficiency, increase the biogas yield, increase the proportion of gas components in biogas, accelerate the utilization of organic matters such as VFAs, soluble carbohydrates, and proteins, and significantly increase the COD removal rate. Summary of the Invention

[0006] In order to solve the above problems, the present invention develops a new ecological halotolerant bacterium and a method for enhancing the cascade energy conversion treatment of wastewater. In the anaerobic reactor operating for a long time, the present invention adds new ecological halotolerant bacteria to enhance the operation of the reactor, and analyzes the SCOD removal rate and biogas production of the enhanced CEAR.

[0007] The first object of the present invention is to provide a Bacillus altitudinis, which is Bacillus altitudinis K3, deposited at the China Center for Type Culture Collection on May 23, 2024, with the deposit number CCTCC NO: M 20241042.

[0008] In one embodiment, the salt tolerance of the Bacillus altitudinis K3 reaches 4.0%.

[0009] The second object of the present invention is to provide a new ecological salt-tolerant microbial community, which contains the Bacillus altitudinis as described above.

[0010] The third object of the present invention is to provide the application of the Bacillus altitudinis as described above, or the new ecological salt-tolerant microbial community as described above in the anaerobic digestion treatment of wastewater.

[0011] In one embodiment, the wastewater includes food waste wastewater.

[0012] In one embodiment, the salinity of the food waste wastewater is 1.0% - 2.0%.

[0013] The fourth object of the present invention is to provide a method for strengthening the cascade energy conversion treatment of wastewater with a new ecological salt-tolerant bacterium. This method uses a Cascade Energetic Anaerobic Reactor (CEAR) as an anaerobic reactor to perform anaerobic digestion treatment on food waste wastewater. The anaerobic digestion treatment includes: feeding the food waste wastewater into the anaerobic reactor (i.e., the cascade energy conversion reactor) and intermittently adding the new ecological salt-tolerant bacterium in a daily feeding manner. Among them,

[0014] The new ecological salt-tolerant microbial community is Bacillus altitudinis K3, with the deposit number CCTCC NO: M20241040.

[0015] In one embodiment, the cascade energy conversion reactor includes a tank body, a three-phase separator, a methane-producing phase, and a hydrogen-producing phase.

[0016] The top of the tank body is provided with a cover body, the side wall of the cover body is provided with a water outlet, the three-phase separator is arranged inside the cover body, and the bottom of the tank body is provided with a sludge discharge port.

[0017] A hydrogen-producing tank body penetrating through the cover body and the tank body is arranged inside the tank body. The inner cylinder of the hydrogen-producing tank body forms the hydrogen-producing phase. The top of the hydrogen-producing tank body is provided with a hydrogen-producing gas outlet and a water inlet.

[0018] The outer cylinder between the tank body, the hydrogen-producing tank body and the three-phase separator forms the methane-producing phase, and a methane-producing gas outlet penetrating through the top of the cover body is provided at the top of the three-phase separator;

[0019] A plurality of slot holes are provided in the lower section of the hydrogen-producing tank body along the circumferential direction to realize the communication between the hydrogen-producing phase and the methane-producing phase.

[0020] In one embodiment, a heat preservation area is provided on the outer periphery of the tank body, a hot water circulation inlet is provided at the lower part of the heat preservation area, and a hot water circulation outlet is provided at the upper part of the heat preservation area, and the stepped energy conversion reactor is heat-preserved in a way of lower inlet and upper outlet.

[0021] In one embodiment, the hot water in the heat preservation area is at 35 - 40 °C, preferably 37 °C.

[0022] In one embodiment, the tank body is a hollow cylinder, and the height-diameter ratio of the tank body is 2:1.

[0023] In one embodiment, the height-diameter ratio of the hydrogen-producing tank body is 4.9:1.

[0024] In one embodiment, the slot holes are arranged uniformly in a circle.

[0025] In one embodiment, a diversion tank body is arranged along the circumferential direction on the outside of the slot holes.

[0026] In one embodiment, the diversion tank body is fixed to the outside of the hydrogen-producing tank body through a support frame.

[0027] In one embodiment, the height-diameter ratio of the diversion tank body is 1.2:1.

[0028] In one embodiment, the volume of the stepped energy conversion reactor CEAR is 12.5 L, the volume of the inner cylinder is 2 L, and the volume of the outer cylinder is 10.5 L.

[0029] In one embodiment, the bottom diameter of the tank body is 200 mm and the height is 400 mm.

[0030] In one embodiment, the diameter of the hydrogen-producing tank body is 80 mm and the height is 395 mm.

[0031] In one embodiment, the width of the slot hole is 2 mm, the height is 32 mm, and the number is 60.

[0032] In one embodiment, the diameter of the diversion tank body is 100 mm and the height is 120 mm.

[0033] In one embodiment, the pH of the kitchen waste water is 3.95 ± 0.15, the soluble chemical oxygen demand (SCOD) is 67.2 ± 0.56 g / L, the total suspended solids (TS) is 9.17 ± 0.19%, the organic matter content (VS) is 7.74 ± 0.09%, the soluble carbohydrate (SC) is 33.01 ± 0.15 g / L, and the soluble protein (SP) is 7.84 ± 0.07 g / L.

[0034] In one embodiment, the inner cylinder of CEAR is inoculated with hydrogen-producing sludge, and the outer cylinder is inoculated with methane-producing sludge. The volume of sludge in each reactor is 20%.

[0035] In one embodiment, the organic load of the anaerobic digestion system is controlled at 2 - 4 kg TCOD / (m 3 ·d).

[0036] In one embodiment, the salinity of the anaerobic digestion system is controlled at 1.0% - 2.0%.

[0037] In one embodiment, the reaction temperature of the anaerobic digestion system is mesophilic fermentation at 35 - 37 °C.

[0038] In one embodiment, the new eco-salt-tolerant hydrogen-producing biofilm needs to be pre-cultivated by membrane attachment outside the reactor.

[0039] In one embodiment, the total cycle of the anaerobic digestion reaction is 61 days. The 0 - 15-day period is the first stage, the 16 - 28-day period is the second stage, the 29 - 40-day period is the third stage, the 41 - 50-day period is the fourth stage, and the 51 - 60-day period is the fifth stage. No new eco-salt-tolerant bacteria are added in the first two stages.

[0040] Specifically, on the 29th day, the new eco-salt-tolerant hydrogen-producing biofilm is hung into the inner cylinder of the cascade energy conversion reactor, and 1.84 g / L of new eco-salt-tolerant bacteria are added to the outer cylinder on the 29th, 41st, and 51st days respectively.

[0041] In one embodiment, the ratio of the volume of kitchen waste water fed into the anaerobic reactor every day to the total volume of the anaerobic reactor is 1:5.0 - 1:6.25, preferably 1:6.25.

[0042] In one embodiment, the fed kitchen waste water needs to be diluted 4 - 8 times.

[0043] The present invention provides the application of the method for strengthening the cascade energy conversion anaerobic reactor for wastewater by using new eco-salt-tolerant bacteria in the fields of wastewater treatment and environmental protection.

[0044] Beneficial effects:

[0045] (1) In this invention, the new ecological salt-tolerant bacterium Bacillus altitudinis K3 was added to the cascading energy conversion reactor CEAR with food waste wastewater as the substrate for biological enhancement, and compared with UASB to study the anaerobic digestion efficiency of the biologically enhanced CEAR. It was found that the anaerobic digestion performance of the CEAR with the new ecological pressure-tolerant bacteria hanging in the inner cylinder and the new ecological bacteria intermittently added to the outer cylinder was significantly improved. Especially in the fifth stage, the SCOD removal rate of CEAR was 15.5%-23.5% higher than that of UASB, the methane production increased by 20.0-25.0%, and hydrogen was produced in the inner cylinder.

[0046] (2) The gradient energy conversion reactor CEAR of this invention has a simple structure, excellent performance, and is easy to operate. The newly constructed ecological salt-tolerant bacterial community in the designed CEAR can better tolerate high salinity. Compared with the traditional UASB, both the gas production efficiency and the COD removal rate have been effectively improved. This invention has good research and application prospects. Description of the Drawings

[0047] Figure 1 Experimental device diagram for the comparative study of the anaerobic treatment effects of CEAR and UASB on food waste wastewater. Among them, Figure (a) is the design diagram of CEAR, Figure (b) is the design diagram of UASB, and Figure (c) is the physical diagram of CEAR and UASB.

[0048] Figure 2 Variation of the daily production of biogas, H2, and CH4 in the two reactors. Among them, Figure (a) shows the variation of the daily biogas production, and Figure (b) shows the variation of the daily production of H2 and CH4.

[0049] Figure 3 Variation of the SCOD removal rate and the effluent SCOD concentration in the two reactors.

[0050] Figure 4 Variation of pH and volatile fatty acids (VFAs) in the two reactors. Among them, Figure (a) shows the variation of pH, and Figure (b) shows the variation of VFAs.

[0051] Figure 5 Variation of SC and SP in the two reactors. Among them, Figure (a) shows the variation of SC, and Figure (b) shows the variation of SP.

[0052] Figure 6 Variation of the organic component in the wastewater before and after the reaction in the fifth stage in the two reactors. Among them, Figure (a) is the influent of CEAR and UASB, Figure (b) is the effluent of CEAR, and Figure (c) is the effluent of UASB.

[0053] Figure 7The composition of the microbial flora in the two reactors. Among them, Figures (a, b) show the changes in the relative abundances of bacteria at the phylum level in the inner cylinder of CEAR, Figure (c) shows the changes in the relative abundances of archaea at the genus level in CEAR, and Figure (d) shows the changes in the relative abundances of archaea at the genus level in UASB. Detailed implementation manners

[0054] For the anaerobic digestion treatment of kitchen waste wastewater with high salt content, on the one hand, the present invention newly proposes a Bacillus altitudinis K3, which has high salt tolerance and is suitable for kitchen waste wastewater with high salt content. On the other hand, the present invention also newly designs an anaerobic reactor, specifically a Cascade Energetic Anaerobic Reactor (CEAR). In order to reflect the long-term treatment effect of CEAR, it is necessary to compare it with the traditional Upflow Anaerobic Sludge Blanket Reactor (UASB), and analyze the changes in the microbial community structure of CEAR after adding the new ecological salt-tolerant bacteria.

[0055] Example 1 Source of strains

[0056] The new ecological salt-tolerant bacteria described in the present invention are screened out using the salt-tolerant sludge in the laboratory completely mixed anaerobic reactor CSTR as the chassis flora and peptone as the sole carbon and nitrogen source, specifically as follows.

[0057] (1) First, adopt the "top-down" anaerobic flora adaptive evolution strategy to domesticate an anaerobic chassis flora that can tolerate more than 3.0% salinity by gradually increasing the salt stress pressure.

[0058] (2) Directly screen out highly salt-tolerant strains using the salt-tolerant chassis flora as the source of strains.

[0059] In some embodiments of the present invention, the salt tolerance adaptation evolution of the anaerobic flora is carried out in a continuous flow completely mixed anaerobic reactor. The initial inoculum is anaerobic methanogenic activated sludge from the anaerobic digestion tank of a food processing factory, and the inoculation amount is 25 - 35 g-TS / L. Starch is used as the sole carbon and nitrogen source, and the influent concentration is 6.0 - 10.0 g / L.

[0060] In some embodiments of the present invention, the salt tolerance adaptation evolution conditions are: temperature 30 - 38 °C, pH 6.5 - 7.5, hydraulic retention time 5 - 10 days. On the basis of a COD removal rate > 70% in the steady state period, gradually increase the salt stress pressure to obtain an anaerobic chassis flora that can tolerate more than 3.0% salinity.

[0061] In some embodiments of the present invention, new ecological salt-tolerant bacteria are isolated and screened from the salt-tolerant anaerobic chassis flora. The isolation and screening process includes: isolation and purification on a 4.0% salinity peptone solid medium, primary screening for protease production ability at 4.0% salinity, and salt tolerance re-screening at 8.0% salinity.

[0062] Specifically, the specific process of screening Bacillus altitudinis K3 is as follows.

[0063] (1) Construction of salt-tolerant anaerobic chassis flora

[0064] In this example, the microbial flora adaptive evolution strategy is adopted to construct a salt-tolerant protein-degrading chassis flora. Adaptive evolution is carried out in a completely mixed anaerobic reactor, adopting a continuous flow fermentation mode, and the inoculum is anaerobic methanogenic activated sludge from the anaerobic digestion tank of a food processing factory. The components of the reactor influent are shown in Table 1, with peptone as the sole carbon and nitrogen source, the initial inoculated sludge concentration is 33.0 g-TS / L, the stirring speed is 100 rpm, the fermentation temperature is 36 °C, and the pH is controlled at 7.0. The initial salinity of the influent is 1.5%, and the hydraulic retention time is 10 days.

[0065] Table 1 Components of the influent of the adaptive evolution reactor

[0066]

[0067] (a) Changes in COD removal rate during the adaptive evolution process

[0068] During the salt stress adaptive evolution process, the change in the salt tolerance ability of the anaerobic flora is characterized by the COD removal rate. Based on the COD removal rate being higher than 70% in the steady state period, the salinity of the reactor influent is gradually increased in gradients to conduct salt tolerance adaptive evolution on the anaerobic flora, so as to obtain an anaerobic chassis flora that can tolerate a salinity exceeding 3.0%.

[0069] The inoculated anaerobic methanogenic sludge can quickly adapt under the salt stress of 1.5% (1-6 d) and 2.0% (7-14 d), and the effluent COD is between 1280-1920 mg / L, and the COD removal rate remains above 80%.

[0070] When the influent salinity is increased to 2.6% (15-21 d), the removal rate drops to 64.0% and then quickly recovers to 77.6%.

[0071] Further increase the influent salinity to 2.9% (22-51 d). Under this stress, the COD removal rate first decreases to 58.1% and then gradually recovers to 72.3%, but the tolerance response time is significantly extended.

[0072] Continue to increase the salinity to 3.2% (day 52-61), and there is no obvious change in the effluent COD concentration. The COD removal rate remains stable between 74.4%-84.0%.

[0073] This indicates that through salinity gradient acclimation, halotolerant microorganisms have become dominant. However, when the salinity was increased to 3.6% (62 - 116 d), the COD removal rate gradually decreased to about 55.0% and was difficult to recover. Therefore, the "top - down" anaerobic flora adaptive evolution strategy can be used to initially construct an anaerobic chassis flora that can tolerate a salinity of 3.2%.

[0074] (b) Changes in protein degradation rate during adaptive evolution

[0075] Detecting the protein degradation rate of the anaerobic flora during adaptive evolution, it was found that under the low - salt stress of 1.5%, the average protein degradation rate could reach 93.2%.

[0076] With the gradual increase in the influent salinity gradient and the gradual adaptation of the anaerobic flora to salt stress, the protein degradation rate first decreased and then gradually recovered. Finally, under the salt stress of 3.2%, the protein degradation rate recovered to about 90%.

[0077] However, when the influent salinity was further increased to 3.6%, the effluent protein concentration increased significantly, and the protein degradation rate decreased to about 71.2% and was difficult to recover.

[0078] This indicates that under this salt stress, the protease - secreting ability of the anaerobic flora is insufficient, and protein hydrolysis is inhibited. Therefore, in this example, the salt - tolerant adaptively evolved sludge obtained under 3.2% salt stress was used as the source to screen for high - salt - tolerant protease - producing bacteria.

[0079] Step 2: Directed screening of high - salt - tolerant protease - producing bacteria

[0080] Although the "top - down" adaptive evolution strategy can be used to initially construct a salt - tolerant chassis flora that can tolerate a salinity of 3.2%; however, under higher salt stress, the insufficient protease - secreting ability leads to the inhibition of protein hydrolysis. Therefore, in order to increase the protease - producing performance under high - salt stress, salt - tolerant protease - producing microorganisms were directionally screened from the salt - tolerant chassis flora.

[0081] The separation and screening process includes: taking the salt - tolerant adaptively evolved sludge obtained under 3.2% salt stress as a sample, placing the anaerobic flora in an anaerobic tube and adding glass beads. After shaking and dispersing, it was gradient - diluted with normal saline. Then the diluted bacterial suspension was spread on a peptone solid medium plate with a salinity of 4.0% and incubated upside - down in an anaerobic constant - temperature incubator at 36°C for 48 hours. The single colonies that grew were further purified by the plate streaking method, and then the isolated and purified strains were stored on a solid slant medium with a salinity of 4.0%.

[0082] Several strains of bacteria were co-isolated and purified at a salinity of 4.0%. They were respectively inoculated onto a 4.0% salinity transparent circle medium and a 4.0% salinity gelatin liquefaction medium by the three-point method and the stab inoculation method. The ratio of the diameter of the transparent circle to the diameter of the colony (>1.5) and the degree of gelatin liquefaction were used as evaluation criteria to screen out salt-tolerant strains with strong protease-producing ability. And the salt tolerance of the screened protease-producing bacteria was re-screened. The specific process included: after the screened strains were cultured in LB liquid medium for 48 h, the diluted bacterial suspension was respectively spread on a salt-tolerant re-screening solid medium with a salinity of 8.0%, and then protease-producing bacteria that could tolerate a salinity of 8.0% were screened out.

[0083] Step 3 Identification and preservation of Bacillus altitudinis K3

[0084] This highly salt-tolerant protease-producing strain was identified as Bacillus altitudinis. The preservation information is: Bacillus altitudinis K3, preservation number CCTCC NO: M 20241040, was preserved in the China Center for Type Culture Collection on May 23, 2024, and the preservation address is Wuhan University, China, Wuhan.

[0085] The salt tolerance of this Bacillus altitudinis K3 reaches 4.0%, and it was stored in an anaerobic roll tube at -80 °C before use.

[0086] The bacterial flora used in the following examples was obtained by mixing anaerobic sludge and newly ecological salt-tolerant bacteria, and the ratio was 7:1 - 10:1 (mass ratio).

[0087] Example 2

[0088] A schematic diagram of the experimental device is as Figure 1 shown. The experiment was carried out in CEAR and UASB reactors with a working volume of 12.5 L, and the reaction temperature was 37 ± 1 °C.

[0089] As Figure 1 (a) shows the design diagram of the CEAR reactor. In this example, the cascaded energy conversion reactor CEAR includes a tank body 4a, a three-phase separator 9a, a methanogenic phase 13a, and a hydrogen-producing phase 10a. Among them,

[0090] A cover body is provided at the top of the tank body 4a, a water outlet 2a is provided on the side wall of the cover body, a three-phase separator 9a is provided inside the cover body, and a sludge discharge port 5a is provided at the bottom of the tank body 4a;

[0091] A hydrogen-producing tank body 11a that penetrates the cover body and the tank body 4a is provided inside the tank body 4a. The inner cylinder of the hydrogen-producing tank body 11a forms a hydrogen-producing phase 10a. A hydrogen-producing gas outlet 8a and a water inlet 1a are provided at the top of the hydrogen-producing tank body 11a;

[0092] The outer cylinder between the tank body 4a, the hydrogen-producing tank body 11a and the three-phase separator 9a forms a methane-producing phase 13a. A methane-producing gas outlet 7a penetrating through the top of the cover body is provided at the top of the three-phase separator 9a;

[0093] A plurality of slot holes are circumferentially arranged on the lower section of the hydrogen-producing tank body 11a to realize the communication between the hydrogen-producing phase 10a and the methane-producing phase 13a.

[0094] In some examples, the slot holes are uniformly arranged in a circle. Optionally, in the embodiment of the present invention, the width of the slot hole is 2 mm, the height is 32 mm, and the number is 60.

[0095] In some examples, the tank body 4a is a hollow cylinder, and the height-to-diameter ratio of the tank body 4a is 2:1. In some examples, the height-to-diameter ratio of the hydrogen-producing tank body 11a is 4.9:1. Optionally, in this embodiment, the volume of the cascade energy conversion reactor is 12.5 L, the volume of the inner cylinder is 2 L, and the volume of the outer cylinder is 10.5 L. Specifically, the bottom diameter of the tank body 4a is 200 mm and the height is 400 mm; the diameter of the hydrogen-producing tank body 11a is 80 mm and the height is 395 mm.

[0096] In some examples, a heat preservation area 12a is provided on the outer periphery of the tank body 4a. A hot water circulation inlet 6a is provided at the lower part of the heat preservation area 12a, and a hot water circulation outlet 3a is provided at the upper part of the heat preservation area 12a. The cascade energy conversion reactor is heat-preserved in a way of lower inlet and upper outlet. Optionally, the hot water in the heat preservation area 12a is at 35 - 40 °C, and preferably 37 °C in this embodiment.

[0097] In some examples, a diversion tank body 14a is circumferentially arranged outside the slot hole. Optionally, the diversion tank body 14a is fixed to the outside of the hydrogen-producing tank body 11 through a support frame. In some examples, the height-to-diameter ratio of the diversion tank body 14a is 1.2:1. Optionally, in the embodiment of the present invention, the diameter of the diversion tank body 14a is 100 mm and the height is 120 mm.

[0098] As Figure 1 (b) shows the design drawing of the UASB reactor. In this embodiment, the volume of the UASB reactor is 12.5 L, and it includes a UASB tank body 4b, a UASB three-phase separator 9b, and a UASB methane-producing area 11b;

[0099] The UASB tank body 4b is a hollow cylinder. A cover body is provided at the top of the UASB tank body 4b. A water outlet 2b is provided on the side wall of the cover body. A three-phase separator 9b is provided inside the cover body. A UASB sludge discharge port 7b, a UASB water inlet 6b, and 8b are provided at the bottom of the tank body 4b;

[0100] The space formed by the UASB tank body 4b and the three-phase separator 9b constitutes the UASB methane production area 11b. The top of the three-phase separator 9b is provided with a UASB methane production gas outlet 1b;

[0101] The UASB tank body 4b is circumferentially provided with a UASB heat preservation area 10b, and a UASB hot cycle water inlet 5b and a UASB hot cycle water outlet 3b are respectively arranged on its upper and lower parts, and the UASB reactor is thermally insulated by the way of bottom-in and top-out.

[0102] In this embodiment, the volume of the UASB reactor is 12.5L, the bottom diameter of the tank body (4b) is 200mm, and the height is 400mm.

[0103] The specific test method of the embodiment of the present invention is as follows: 2L of kitchen waste wastewater diluted 4 to 8 times with water every day, the hydraulic retention time is 7d, the influent organic load is controlled to be 2 to 4 kg TCOD / (m 3 .d), the salt concentration is 1.0% to 2.0%, the specific increase in organic load and salt concentration is shown in Table 1, and the addition amount of activated sludge is 20-30% of the reactor volume.

[0104] After the biogas is collected by a gas collection bag, the gas components are measured by a gas chromatograph, and the gas volume is calculated by a gas flow meter. The concentration and composition of VFAs are measured by a gas chromatograph, and the organic components are analyzed and measured by GC-MS. The SCOD, SC, SP and microbial diversity in the reaction system are measured, and the measurement methods are all analyzed by national standard methods (Table 2).

[0105] The total anaerobic digestion reaction time is 60 days. Starting from the 15th day, every 10 days is recorded as a stage to carry out the experiment. The specific reaction stage division is shown in Table 1.

[0106] Table 1 Operating conditions of the reactor

[0107]

[0108] On the 29th day, 1.84 g / L of new ecological salt-tolerant bacteria was added to the outer cylinder of the CEAR reactor, and a new ecological salt-tolerant bacteria membrane was built on the inner cylinder.

[0109] On the 41st and 51st days, new ecological salt-tolerant bacteria were continuously added, 1.84 g / L each time.

[0110] As a control, no new ecological salt-tolerant bacteria were added to the UASB reactor.

[0111] Table 2 Analysis items and methods

[0112]

[0113]

[0114] Variation of daily production of biogas, H2 and CH4 in two reactors in Test Example 1

[0115] Figure 2 It is for the variation of daily production of biogas, H2 and CH4 in two reactors. The total biogas volume of the two reactors was relatively close in the first three stages (0 - 40 days). In the fourth - fifth stages (41 - 60 days), the load of the two reactors was maintained at 4 kg COD / (m 3 ·d). In the fourth stage, the salt concentration was increased from 1.0% to 1.5%, and in the fifth stage, it was increased to 2.0%. The gas production performance of CEAR was significantly higher than that of UASB. The total biogas volume of CEAR was increased by 8.25% and 17.51% respectively compared with that of UASB.

[0116] The performance of the reactor is affected by salts and newly - formed salt - tolerant bacteria. With the progress of the reaction, there are obvious changes in the hydrogen and methane production in the two reactors. Compared with UASB, CEAR has a higher methane production and can achieve hydrogen production. The first stage (0 - 15 days) is the startup stage. The methane production of CEAR and UASB gradually tends to be stable. The methane production of CEAR is increased by 2.12% on average compared with that of UASB. The second and third stages (16 - 40 days) are the load - increasing stages. The methane production of each reactor is increased. At the same time, hydrogen begins to be produced in the inner cylinder of CEAR on the 29th day, realizing cascaded energy utilization. The methane production of the outer cylinder of CEAR is increased by 1.86% on average compared with that of UASB. In the fourth and fifth stages, under 1.5% and 2.0% salts, the performance of CEAR is further strengthened. The methane production of CEAR is increased by 9.77% and 22.30% respectively compared with that of UASB. It can be seen that under this salinity condition, the newly - formed salt - tolerant bacteria play a strengthening role in CEAR.

[0117] Variation of SCOD removal rate and effluent SCOD concentration in two reactors in Test Example 2

[0118] Figure 3 It shows the variation of SCOD removal rate and concentration under different organic loads and salt conditions. The two reactors showed good SCOD removal efficiency at the beginning of operation, and the effluent SCOD concentration was low. In the first 15 days, the average SCOD removal rate of the two reactors exceeded 90.0%. This is mainly due to the high biodegradability of carbohydrates and proteins in food waste, as well as the high activity of the inoculated sludge and the good structural performance of the two reactors. When the organic load is increased from 2 kg COD / (m 3 ·d) to 4 kg COD / (m 3· At (d), the average SCOD removal rates of CEAR and UASB decreased from 92.38% and 90.09% to 89.39% and 83.48% respectively. Both reactors were impacted, and the SCOD removal rate decreased to a certain extent. However, the SCOD removal performance of CEAR was better than that of UASB. From 40 to 60 days was the salinity increasing stage. The SCOD removal rate of UASB decreased significantly. CEAR with newly added ecological salt-tolerant bacteria showed good performance, and its SCOD removal rate was on average 10.08% higher than that of UASB. In the later stage of the fifth-stage reaction, the operation of UASB further deteriorated. CEAR coped well with the salinity impact, and the SCOD removal rate of CEAR was 15.5% - 23.5% higher than that of UASB.

[0119] Test Example 3 Changes in pH and VFAs in the two reactors

[0120] The changes in VFAs and the corresponding pH in the two reactors are as Figure 4 shown. During the initial startup period of the reactors (0 - 15 days), the concentration of VFAs in the effluent of CEAR decreased and gradually stabilized, while UASB had a higher VFAs effluent concentration. Subsequently, with the continuous increase of the organic load and salinity, the VFAs effluent concentrations of the two reactors continuously increased. By day 35, the VFAs effluent concentrations of CEAR and UASB reached 900.0 mg / L and 1346.0 mg / L respectively. After 40 days, although the VFAs effluent concentration of UASB decreased somewhat, it was still at a relatively high level and gradually increased with time, with an average VFAs effluent concentration above 1000 mg / L. Some studies have pointed out that when the concentration of TVFAs exceeds 1000 mg / L, it will inhibit methane production. During this period, mainly due to the increase in salinity, UASB failed to cope well with the salinity impact. CEAR began to show superior performance, and its VFAs concentration decreased significantly, with an average effluent concentration of only 305 mg / L.

[0121] To ensure the consistency of reaction conditions, the influent pH of the two reactors was 7.5. After being treated by the two reactors, the effluent pH of the two reactors increased. The average pH values of CEAR and UASB were 7.98 and 7.95 respectively. The change in pH was related to VFAs. In the first 30 days of the reaction, the pH of CEAR and UASB increased. After 35 days, UASB was always lower than CEAR, which was caused by the accumulation of VFAs in UASB.

[0122] Test Example 4 Changes in SC and SP in the two reactors

[0123] Carbohydrates and proteins are the main organic matters in kitchen waste water. Therefore, it is very necessary to monitor the concentration changes of the two. The changes in soluble carbohydrates and proteins at different stages of the two reactors are as Figure 5As shown. The influent carbohydrate concentration in the two reactors was higher than the protein concentration, but the effluent was the opposite, indicating that carbohydrates were more easily degraded and utilized by microorganisms. The concentration changes of the two in the reactor were similar to the change trend of the effluent SCOD concentration. At the beginning, the concentrations of both fluctuated and then gradually stabilized. During the whole reaction process, the CEAR strengthened by the newly developed salt-tolerant bacteria showed good treatment effects, and the effluent concentrations of carbohydrates and proteins were always lower than those of UASB. Although the organic load increased in the later stage of the two reactors, the concentrations of carbohydrates and proteins decreased, mainly because the two had been hydrolyzed but not utilized, which was reflected in the change of VFAs concentration.

[0124] Test Example 5 Changes in the organic components in the wastewater before and after the fifth-stage reaction in the two reactors

[0125] Through the analysis of the positions of each peak, the specific organic components and their proportions before and after the reaction were determined ( Figure 6 ). The organic substances with relatively high proportions in the influent of CEAR and UASB were mainly long-chain fatty acids, including heptanoic acid (27.19%), octanoic acid (14.42%), hexanoic acid (16.78%), and malonic acid (4.38%). In addition, there were oil substances such as octamethylcyclotetrasiloxane in the influent, which also reflected the high-oil characteristics of kitchen waste wastewater. In the effluent, CEAR mainly consisted of oil substances such as octamethylcyclotetrasiloxane, decamethylcyclopentasiloxane, and hexamethylcyclotrisiloxane, with a proportion reaching 50.0%. The proportion of p-cresol in the effluent of UASB was the highest, reaching 28.64%, probably because it contained a benzene ring that was relatively stable and not easily degraded, and the proportion of oil substances reached 33.5%. The anaerobic microorganisms in CEAR and UASB degraded most of the organic acids in the influent, and oil substances and benzene series compounds were relatively difficult to degrade. The anaerobic microorganisms in CEAR had good degradation performance for benzene series compounds, and the proportion in the effluent was only 0.51%, indicating that good effects were produced after the addition of the newly developed salt-tolerant bacteria.

[0126] Test Example 6 Microbial flora composition in the two reactors

[0127] The changes in the bacterial phylum level in the inner cylinder of CEAR were as Figure 7As shown, the dominant bacteria with an abundance greater than 5.0% in H0 are Chloroflexi (32.95%), Bacteroidota (21.94%), Nitrospirota (15.25%), Patescibacteria (9.56%) and Firmicutes (8.63%). With the increase of organic load and salt concentration, the microbial community structure changed, and the proportions of Firmicutes, Bacteroidota and Proteobacteria became larger and reached the maximum in H6. The results indicate that these phyla in anaerobic sludge may have strong tolerance to high-salt environments. Firmicutes is the most dominant phylum in the bacterial community of H6, with its abundance increased by 29.83%, and the abundance of Bacteroidota increased to 32.13%, indicating that they play an important role in utilizing polysaccharides in food waste water. It can be seen that the microbial community existing in the system has successfully adapted to high-salt conditions, and these environmentally adapted communities also include high abundances of Thermotogota, Synergistota and Proteobacteria. Thermotogota is considered a common anaerobic treatment bacterial group in high-salt waste water and dominates in the treatment of food waste water, which may be attributed to their adaptation to extreme environments such as complex substrates and oil-containing ecosystems. It is worth noting that the abundance of Nitrospirota has decreased from 15.25% in H0 to below 2.41% because Nitrospirota is mainly abundant in fresh water and is not suitable for growth under high-salt conditions.

[0128] At the genus level of CEAR and UASB archaea, the changes of archaea in each stage of the two reactors were further revealed under the inhibition of high salinity and the enhancement of the newly ecological salt-tolerant microbial community. There were differences in the distribution of archaea at the genus level in the two reactors. From day 29 to day 60, it was the stage of increasing salt concentration in the two reactors. The abundance of Methanomassiliicoccus in F3-F5 gradually increased, while it continuously decreased in G3-G5. The newly ecological salt-tolerant microbial community added played a strengthening role in CEAR. The enrichment of Methanomassiliicoccus could accelerate the metabolism of methanol, acetic acid and H2 / CO2. At the end of the reaction, the abundance of Methanosaeta in F6 was 8.4% higher than that in F0, and that in G6 was 7.6% higher than that in G0. Methanosaeta belongs to acetate-utilizing methanogens, which mainly consume VFAs to produce methane and plays an important role in the anaerobic digestion process. Methanobacterium, Methanolinea and Methanospirillum all belong to hydrogenotrophic methanogens and are commonly found in mesophilic anaerobic digestion systems. The abundance of Methanobacterium increased significantly. At the end, F6 and G6 were 9.5% and 6.9% higher than the initial levels respectively, indicating an increase in the reduction of CO2 to CH4 during the methanogenic fermentation process. The relative abundance of Methanospirillum in each stage of CEAR and UASB ranged from 6.17% to 9.60% and from 6.23% to 9.48% respectively. It also had a certain abundance under high salinity and might establish a DIET pathway to participate in direct electron transfer. In F0-F6 and G0-G6, the relative abundance of Methanolinea decreased with the increase of salt concentration, indicating that Methanolinea showed poorer salt tolerance than other hydrogenotrophic methanogens.

[0129] Although the present invention has been disclosed above in a preferred embodiment, it is not intended to limit the present invention. Anyone familiar with this technology can make various modifications and decorations without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention should be defined by the claims.

Claims

1. A Bacillus altitudinis, characterized in that, The Bacillus altitudinis is Bacillus altitudinis K3, which was deposited at the China Center for Type Culture Collection on May 23, 2024, with the deposit number CCTCC NO: M20241042.

2. The Bacillus altitudinis according to claim 1, characterized in that, The salt tolerance of the Bacillus altitudinis K3 reaches 4.0%.

3. A new ecological salt-tolerant microbial community, characterized in that, The new ecological salt-tolerant bacterial community contains the Bacillus altitudinis as described in claim 1 or 2.

4. The application of the Bacillus altitudinis as described in claim 1 or 2, or the new ecological salt-tolerant bacterial community as described in claim 3 in the anaerobic digestion treatment of wastewater.

5. The application according to claim 4, wherein The wastewater includes food waste wastewater. Preferably, the salt content of the food waste wastewater is 1.0% - 2.0%.

6. A method for enhancing the cascaded energy utilization treatment of wastewater by using newly ecological salt-tolerant bacteria, characterized in that, This method uses a stepped energy conversion reactor as the anaerobic reactor to perform anaerobic digestion treatment on food waste wastewater. The anaerobic digestion treatment includes: feeding food waste wastewater into the stepped energy conversion reactor every day in a daily feeding manner and intermittently adding new ecological salt-tolerant bacteria, where The new ecological salt-tolerant bacteria are Bacillus altitudinis K3, with the deposit number CCTCC NO: M20241040; The stepped energy conversion reactor includes a tank body (4a), a three-phase separator (9a), a methane-producing phase (13a), and a hydrogen-producing phase (10a). The top of the tank body (4a) is provided with a cover body. The side wall of the cover body is provided with a water outlet (2a). The three-phase separator (9a) is arranged inside the cover body. The bottom of the tank body (4a) is provided with a sludge discharge port (5a). A hydrogen-producing tank body (11a) that penetrates the cover body and the tank body (4a) is arranged inside the tank body (4a). The inner cylinder of the hydrogen-producing tank body (11a) forms the hydrogen-producing phase (10a). The top of the hydrogen-producing tank body (11a) is provided with a hydrogen-producing gas outlet (8a) and a water inlet (1a). The outer cylinder between the tank body (4a) and the hydrogen-producing tank body (11a) and the three-phase separator (9a) forms the methane-producing phase (13a). The top of the three-phase separator (9a) is provided with a methane-producing gas outlet (7a) that penetrates the top of the cover body. Several slot holes are arranged circumferentially on the lower part of the hydrogen-producing tank body (11a) to realize the communication between the hydrogen-producing phase (10a) and the methane-producing phase (13a).

7. The method according to claim 6, characterized in that A heat preservation area (12a) is arranged on the outer periphery of the tank body (4a). The lower part of the heat preservation area (12a) is provided with a hot water circulation inlet (6a), and the upper part of the heat preservation area (12a) is provided with a hot water circulation outlet (3a). The stepped energy conversion reactor is heat-preserved in a way of bottom-in and top-out. Preferably, the hot water in the heat preservation area (12a) is at 35 - 40 °C, preferably 37 °C; and / or The tank body (4a) is a hollow cylinder, and the height-to-diameter ratio of the tank body (4a) is 2:1; and / or The height-to-diameter ratio of the hydrogen-producing tank body (11a) is 4.9:1; and / or The slot holes are arranged circumferentially and evenly; and / or A diversion tank body (14a) is arranged circumferentially along the outside of the slot holes. Preferably, the diversion tank body (14a) is fixed to the outside of the hydrogen production tank body (11) through a support frame; Preferably, the height-diameter ratio of the diversion tank body (14a) is 1.2:

1.

8. The method according to claim 7, characterized in that The volume of the cascade energy conversion reactor is 12.5 L, the volume of the inner cylinder is 2 L, and the volume of the outer cylinder is 10.5 L; and / or The bottom diameter of the tank body (4a) is 200 mm and the height is 400 mm; The diameter of the hydrogen production tank body (11a) is 80 mm and the height is 395 mm; and / or The width of the slot is 2 mm, the height is 32 mm, and the number is 60; and / or The diameter of the diversion tank body (14a) is 100 mm and the height is 120 mm.

9. The method according to claim 6, characterized in that, The pH of the food waste wastewater is 3.95 ± 0.15, SCOD is 67.2 ± 0.56 g / L, TS is 9.17 ± 0.19%, VS is 7.74 ± 0.09%, SC is 33.01 ± 0.15 g / L, SP is 7.84 ± 0.07 g / L; and / or The inner cylinder is inoculated with hydrogen-producing sludge, and the inoculation amount is 20% of the volume of the inner cylinder; the outer cylinder is inoculated with methane-producing sludge, and the inoculation amount is 20% of the volume of the outer cylinder; and / or The organic loading of the anaerobic digestion system in the stepped energy conversion reactor is controlled at 2 - 4 kg TCOD / (m 3 ·d); and / or The salinity of the anaerobic digestion system is controlled at 1.0% - 2.0%; and / or The reaction temperature of the anaerobic digestion system is 35 - 37 °C.

10. The method according to claim 5, characterized in that, Using the new ecological salt-tolerant bacteria, a new ecological salt-tolerant bacteria hydrogen-producing biofilm is prepared by pre-film hanging and domestication outside the cascade energy conversion reactor; The total cycle of the anaerobic digestion reaction in the cascade energy conversion reactor is 61 days. The 0 - 15th day is the first stage, the 16 - 28th day is the second stage, the 29 - 40th day is the third stage, the 41 - 50th day is the fourth stage, and the 51 - 60th day is the fifth stage. On the 29th day, a new ecological salt-tolerant bacteria hydrogen-producing biofilm is hung into the inner cylinder of the cascade energy conversion reactor, and 1.84 g / L of new ecological salt-tolerant bacteria is added to the outer cylinder on the 29th, 41st, and 51st days respectively; Preferably, the ratio of the volume of the food waste wastewater fed into the cascade energy conversion reactor every day to the total volume of the cascade energy conversion reactor is 1:5.0 - 1:6.25; Preferably, the fed food waste wastewater is diluted 4 - 8 times in advance.

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