Bacteria continuous flow large-scale culture device, culture method and application

Abstract

The invention relates to the technical field of bacterial culture, in particular to a bacterial continuous flow large-scale culture device, a culture method and application. The continuous flow large-scale culture device for bacteria comprises a culture container; an immobilized culture chamber is arranged in the culture container, and a bacterial liquid circulation channel communicated with the culture container is arranged at the bottom of the immobilized culture chamber; a centrifugal culture chamber for adjusting hydraulic retention time is arranged in the immobilized culture chamber, and an overflow channel communicated with the immobilized culture chamber is arranged on the chamber wall of the centrifugal culture chamber; the culture container is sequentially divided into a reverse buffer area, an elastic trapping area and a settling area from top to bottom. The invention further provides a bacterial culture method and application. According to the method, the problem that the domestication period of an existing nitrifying bacteria culture mode is long is solved, and the problem that nitrifying bacteria cultured through an existing method are low in overall nitrogen removal efficiency is also solved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of bacterial culture, in particular to a bacterial continuous flow large-scale culture device, culture method and application. BACKGROUND

[0002] At present, the effluent quality of coal mine domestic sewage treatment plants in China generally implements the first level A standard of the Discharge Standard of Pollutants for Municipal Wastewater Treatment Plant (GB 18918-2002). With the increasingly stringent requirements for prevention and control of water environmental eutrophication, some areas lacking receiving water bodies require the effluent ammonia nitrogen concentration of newly built sewage treatment plants to reach the limit value (1 mg / L) of the Environmental Quality Standard for Surface Water (GB3838-2002) Class III standard. However, coal mine domestic sewage generally has the water quality characteristic of low carbon-nitrogen ratio, and when the traditional activated sludge process is used, the sludge concentration of the system is difficult to improve, and it is difficult to meet the treatment requirements of deep denitrification. Although biological membrane processes such as biological contact oxidation and biological aerated filter can be used, additional nitrifying bacteria inoculum needs to be added to strengthen ammonia nitrogen removal. However, the current nitrifying bacteria preparation technology has obvious defects: the domestication period of the conventional culture method is long, the effective biomass attached to the carrier is limited, and the process operation usually relies on the full nitrification path, resulting in prolonged hydraulic retention time and low overall denitrification efficiency.

[0003] Therefore, it is urgent to develop a nitrifying bacteria culture device and culture method suitable for coal mine domestic sewage to realize rapid enrichment and efficient immobilization of nitrifying functional bacteria, improve the abundance of nitrosobacteria and strengthen short-cut nitrification, thereby solving the technical problems in the field. SUMMARY

[0004] Therefore, the purpose of the present application is to provide a bacterial continuous flow large-scale culture device, culture method and application to solve the problem of long domestication period of the existing nitrifying bacteria culture method, and also to solve the problem of low overall denitrification efficiency of the nitrifying bacteria cultured by the existing method.

[0005] In order to achieve the above-mentioned purpose, the technical scheme adopted by the present application is as follows: A bacterial continuous flow large-scale culture device, comprising: a culture container; The inside of the culture container is provided with a immobilized culture chamber, and the bottom of the immobilized culture chamber is provided with a bacteria liquid flow passage in communication with the culture container; The inside of the immobilized culture chamber is provided with a centrifugal culture chamber for adjusting the hydraulic retention time, and the chamber wall of the centrifugal culture chamber is provided with an overflow passage in communication with the immobilized culture chamber; The culture container is sequentially divided into a reverse buffer zone, an elastic trapping zone and a sedimentation zone from top to bottom; The centrifugal culture chamber is connected with the overflow channel, and the overflow channel is connected with the immobilized culture chamber. In the culture container, the bacteria in the mixed bacteria solution are trapped and enriched by the elastic trapping area, and the bacteria excrement in the mixed bacteria solution is settled in the sedimentation area; after the bacteria excrement is removed, the reverse buffer area generates gas to drive the bacteria in the elastic trapping area to fall off to the sedimentation area, and then part of the bacteria solution in the sedimentation area is collected and used, and the other part of the bacteria solution is returned to the centrifugal culture chamber, so as to realize continuous flow scale culture of bacteria.

[0006] According to the above technical means, by sequentially arranging the centrifugal culture chamber, the immobilized culture chamber and the culture container from inside to outside, and by partitioning in the culture container, the multi-dimensional hierarchical culture and the intelligent trapping and backflow mechanism are realized, and the problems of long cycle and denitrification efficiency bottleneck in traditional nitrifying bacteria culture are systematically solved.

[0007] In the centrifugal culture chamber, the centrifugal force field acts on the culture solution, and the subtle difference in the sedimentation coefficient between the nitrosobacteria and the nitrifying bacteria is utilized to realize in-situ optimization of the bacterial community structure by regulating the rotation speed and the overflow rate. The centrifugal acceleration promotes the nitrifying bacteria with slow proliferation rate to be thrown to the chamber wall first and then enter the immobilized culture chamber through the overflow channel, while the nitrosobacteria are extended in the effective residence time in the centrifugal area due to their small size and slow settling speed, so as to improve their relative abundance in the bacterial community under the dynamic selection pressure. This community structure optimization based on physical field regulation avoids the interference of chemical inhibitors on the activity of bacteria.

[0008] The immobilized culture chamber constructs a limited microenvironment, so that the nitrifying bacteria spontaneously secrete extracellular polymers and adhere to the chamber wall or the surface of the immobilized carrier under the action of shear force and nutrient concentration gradient in the bacteria solution flow channel, forming a high-density biofilm structure. This process converts free bacteria into attached bacteria, which not only effectively improves the biomass concentration, but more importantly, the immobilized steric hindrance effect selects high-attachment strains, and such strains have stronger impact load capacity in real wastewater treatment biofilm systems.

[0009] The elastic trapping area realizes the "capture-loosen" self-adaptive cycle through material elastic deformation and dynamic adjustment of pore size: when the water flows upward, the elastic fiber network shrinks the pore size to less than the size of the bacteria under the action of fluid drag force, and the trapping rate can reach more than 99%; the downward gas pulse of the reverse buffer area makes the fiber vibrate and rebound, the pore size is temporarily enlarged and the enriched bacteria are released, realizing rapid falling off without mechanical stirring. This mechanism shortens the traditional settling time from 2-4 hours to 30 minutes, and avoids the damage of shear force to the activity of bacteria.

[0010] The settling zone separates the excrement by gravity sedimentation, and then the high-density bacterial solution is proportionally shunted and backflowed to the centrifugal culture chamber to form a closed loop of "inoculation-enrichment-reselection", which not only maintains the abundance of functional bacterial flora in the system, but also strengthens the adaptive evolution of the bacterial flora through continuous backflow, so that the domestication cycle is compressed from 30-45 days of traditional batch mode to 7-10 days. The synergistic effect of continuous flow operation mode and dynamic optimization of bacterial flora structure ultimately improves the matching degree of nitrifying and nitrosating bacteria activity in the device, increases the nitrification rate by 2-3 times, and significantly enhances the overall denitrification efficiency.

[0011] Preferably, the centrifugal culture chamber is connected to the immobilized culture chamber through a rotating base.

[0012] Preferably, the bottom of the centrifugal culture chamber is provided with a water distributor connected to the liquid inlet pipe.

[0013] Preferably, the centrifugal culture chamber is provided with the overflow channel on the side wall close to the top.

[0014] Preferably, the bottom of the centrifugal culture chamber is rotationally connected to the bottom of the immobilized culture chamber through the rotating base, so that the centrifugal culture chamber can rotate around its axis.

[0015] Preferably, the overflow channel is composed of overflow holes opened on the side wall of the centrifugal culture chamber close to the top, the aperture of the overflow holes is 8-20 mm, and the spacing between adjacent overflow holes is 50-200 mm.

[0016] Preferably, the centrifugal culture chamber is in a cylindrical structure, the overflow holes are arranged circumferentially along the side wall of the centrifugal culture chamber, and 10-20 overflow holes are opened on the side wall of the centrifugal culture chamber.

[0017] Preferably, the immobilized culture chamber is provided with an upper interception net, a lower interception net, an adsorption carrier, an aeration pipe and a first sliding rail; The upper interception net and the lower interception net are located above the aeration pipe; The adsorption carrier is located between the upper interception net and the lower interception net; The aeration pipe is located at the bottom of the immobilized culture chamber; The first sliding rail is arranged on the inner side wall of the immobilized culture chamber, and the lower interception net and the first sliding rail are in sliding fit, so that the lower interception net can slide up and down along the first sliding rail.

[0018] Preferably, the end of the upper interception net is fixedly connected to the inner side wall of the immobilized culture chamber.

[0019] Preferably, the aeration pipe is fixedly connected to the bottom of the immobilized culture chamber.

[0020] Preferably, the first sliding rail is fixed to the inner side wall of the fixed culture chamber in the up-down direction.

[0021] Preferably, the adsorption carrier has a diameter of 30-50 mm, a porosity of 80-90%, and a filling rate of 30-50% in the fixed culture chamber, and is made of polyurethane or polyvinyl alcohol.

[0022] Preferably, a backwash aeration pipe is arranged in the reverse buffer zone.

[0023] The backwash aeration pipe is arranged in the reverse buffer zone to provide air flow in the opposite direction of the overflow direction, so that part of the bacteria exceeds the height of the aeration pipe and has a sedimentation trend in the reverse buffer zone, thereby preventing the bacteria from directly overflowing through the buffer effect.

[0024] Preferably, an upper grid plate, a lower grid plate, filter cotton, and a second sliding rail are arranged in the elastic trapping zone. The filter cotton is arranged between the upper grid plate and the lower grid plate. The second sliding rail is arranged on the inner side wall of the culture container, and the upper grid plate and the lower grid plate are in sliding fit with the second sliding rail, so that the upper grid plate and the lower grid plate can slide up and down along the second sliding rail.

[0025] Preferably, the filter cotton includes 3-5 layers, and the pore size of each layer gradually decreases from top to bottom, and the pore size of the filter cotton is 0.7-4 mm.

[0026] Preferably, a drainage groove is arranged near the top of the culture container, and a drainage pipe is connected to the drainage groove.

[0027] Preferably, a liquid discharge pipe is arranged at the bottom of the culture container and connected to the sedimentation zone. The liquid discharge pipe is connected to a bacteria return pipe and a bacteria collection pipe, and the bacteria return pipe is connected to the liquid inlet pipe through a pipeline mixer.

[0028] The application also provides a method for culturing bacteria using the bacterial continuous flow scale culture device as described in the application, which includes the following steps: S1, mixing the culture liquid containing the bacterial source through the pipeline mixer and then flowing into the centrifugal culture chamber through the liquid inlet pipe; S2, flowing the mixed bacteria liquid in the centrifugal culture chamber into the fixed culture chamber through the overflow channel after centrifugal culture for a preset time; S3, flowing the mixed bacteria liquid in the fixed culture chamber into the culture container through the bacteria liquid flow channel when the turbidity of the mixed bacteria liquid reaches a preset turbidity; S4, in the culture container, the bacteria in the mixed bacteria solution are enriched by the elastic trapping zone, and the bacteria excrement in the mixed bacteria solution is settled in the sedimentation zone; after the bacteria excrement is removed, the reverse buffer zone generates gas to drive the bacteria in the elastic trapping zone to fall off to the sedimentation zone, and then the bacteria solution in the sedimentation zone is collected, so that the cultured bacteria are obtained; Part of the obtained bacteria solution is used for storage, and the other part is returned to the pipeline mixer to realize continuous flow scale culture of bacteria.

[0029] According to the above technical means, through the cascade process of pipeline mixer homogenization-centrifugal sorting-immobilized enrichment-elastic trapping, a dynamic optimized continuous culture system is constructed, which significantly improves the efficiency and stability of the culture of nitrifying bacteria. First, the pipeline mixer instantaneously homogenizes the returned bacteria solution and the fresh culture solution, which not only maintains the constant inoculation amount, but also shortens the lag phase through bacterial population pre-adaptation; the centrifugal culture chamber actively enriches nitrosobacteria and optimizes their proportion in the nitrifying bacteria population according to the physical sorting of bacteria according to the difference in sedimentation coefficient, and the pre-set culture time is accurately controlled, which solves the bottleneck of functional bacteria competition imbalance in traditional culture. The turbidity reaches the trigger condition to realize intelligent amplification, and when the mixed bacteria solution reaches the pre-set turbidity, it is released to the culture container, ensuring that the bacteria entering the trapping zone are in the logarithmic growth phase and the density is controllable, avoiding batch differences. The elastic trapping zone realizes dynamic switching of efficient interception and non-destructive shedding through material deformation: the interception rate during the culture stage is more than 99%, preventing the loss of bacteria; the gas pulse of the reverse buffer zone makes the bacteria quickly fall off to the sedimentation zone during the harvesting stage, effectively compressing the sedimentation time and without mechanical shear damage. After sedimentation and separation, part of the bacteria solution is directly collected for standby, and the other part is returned to the pipeline mixer to form a closed loop, which not only strengthens the adaptability evolution and functional stability of the bacterial population, but also shortens the acclimation period from 30-45 days to 7-10 days. Through continuous flow operation and selective pressure maintenance, the metabolic synergy efficiency of nitrosobacteria and nitrifying bacteria in the device is improved by 2-3 times, finally realizing the breakthrough of denitrification efficiency and culture scale.

[0030] Preferably, the volume content of the bacterial source in the culture solution containing the bacterial source is 2-5%.

[0031] Preferably, the pre-set time is 2-2.5h.

[0032] Preferably, the rotation speed of the centrifugal culture is 60-80r / min.

[0033] Preferably, during the culture process in the immobilized culture chamber 2, the dissolved oxygen concentration is 1.5-2.5mg / L.

[0034] Preferably, the pre-set turbidity is 200NTU.

[0035] Preferably, the bacteria are nitrifying bacteria.

[0036] Preferably, S3 includes: When the turbidity of the mixed bacterial solution in the immobilized culture chamber reaches the preset turbidity, the lower intercepting net moves upward along the first sliding track, causing the adsorption carrier to gradually move upward until 60-80% of the adsorption carrier is in a compressed state, at which point the movement of the lower intercepting net stops. Then, the air-to-water ratio is adjusted to 12:1 through the aeration pipe, so that the mixed bacterial solution flows into the culture container through the bacterial solution flow channel. After maintaining this for 3-5 minutes, the lower intercepting net is moved downwards to its initial position.

[0037] The air-to-water ratio refers to the ratio of the air flow rate from the aeration pipe to the water flow rate from the inlet pipe.

[0038] Preferably, S4 includes: Inside the culture container, the mixed bacterial solution flows upward, and the bacteria in the mixed bacterial solution are intercepted and enriched by the filter cotton in the elastic trapping zone. The bacterial excrement in the mixed bacterial solution settles in the settling zone. The upper grid plate moves upward along the second sliding track as the mixed bacterial solution flows until the upper grid plate moves up to the top of the second sliding track. Then, the drain pipe is opened to remove the bacterial excrement that has settled in the reverse buffer zone. After the bacterial excrement is discharged, the drain pipe is closed, the backflushing aeration pipe of the reverse buffer zone is opened, and the air-to-water ratio is controlled at 6:1 until the adsorbent carrier returns to its initial state from the squeezed state. Then, the backflushing aeration pipe is closed, and the return and collection pipes are opened. A portion of the bacteria in the mixed bacterial solution flows out through the collection pipe and is stored in a concentrated manner, while another portion of the bacteria flows through the return pipe to the pipeline mixer to achieve continuous flow and large-scale bacterial culture.

[0039] Preferably, the volume ratio of the amount of bacteria flowing into the return tube to the amount of bacteria flowing into the collection tube is 1:4.

[0040] The present invention also provides the application of bacteria cultured by the culture method described herein in the treatment of coal mine domestic sewage.

[0041] Experiments have shown that the cultivation method of this invention can reduce the ammonia nitrogen concentration in coal mine domestic sewage to below 0.23 mg / L, with an ammonia nitrogen removal rate of 98.6%.

[0042] The beneficial effects of this invention are: The bacterial continuous flow scale culture device of the application, the centrifugal culture chamber arranged from inside to outside utilizes the centrifugal force field to physically screen the bacterial population, based on the difference in the sedimentation coefficient of nitrosation bacteria and nitrification bacteria, the effective residence time of nitrosation bacteria is prolonged by regulating the rotation speed and overflow rate, so as to improve the relative abundance of nitrosation bacteria in the bacterial population under the dynamic selection pressure, and the colony structure optimization without chemical inhibition avoids the interference of traditional methods on the activity of bacteria. The immobilized culture chamber constructs a limited microenvironment, so that the nitrification bacteria spontaneously secrete extracellular polymers and adhere to form high-density biofilm under the action of shear force and nutrient gradient in the bacterial liquid flow channel, not only effectively improving the biomass concentration, but also screening high-attachment strains through the steric hindrance effect, significantly enhancing the impact load capacity in real wastewater treatment. The elastic capture area realizes the self-adaptive cycle of “capture-looseness” by using the elastic deformation of the material: when draining upward, the fiber network pores shrink to less than the size of the bacteria under the fluid drag force, and the interception rate is more than 99%; the downward gas pulse of the reverse buffer zone drives the fiber to vibrate and rebound, instantaneously releasing the enriched bacteria, effectively shortening the sedimentation time, and avoiding the damage of mechanical shear to the activity of bacteria. The sedimentation area discharges the excrement through gravity separation, then collects and returns the high-density bacterial liquid in proportion, forming a closed loop system of “inoculation-enrichment-reselection”, which not only maintains the abundance of functional bacterial population, but also strengthens the adaptive evolution through continuous reflux, so that the acclimation period is compressed from 30-45 days of batch mode to 7-10 days. The synergistic effect of the continuous flow operation mode and the dynamic optimization of the bacterial population finally improves the matching degree of the activities of nitrification and nitrosation bacteria in the device, increases the nitrification rate by 2-3 times, and significantly enhances the overall denitrification efficiency.

[0043] The bacterial culture method of the application constructs a dynamically optimized continuous culture system through the cascade process of pipeline mixer homogenization-centrifugal sorting-immobilization enrichment-elastic trapping, significantly improving the efficiency and stability of nitrifying bacteria culture. First, the pipeline mixer instantaneously homogenizes the backflow bacteria liquid and the fresh culture liquid, not only maintaining constant inoculation amount, but also shortening the lag phase through bacterial population pre-adaptation; the centrifugal culture chamber physically sorts bacteria according to the difference in sedimentation coefficient, and cooperates with the precise regulation of the preset culture time to actively enrich nitrosobacteria and optimize their proportion in the nitrifying bacteria population. The immobilized culture chamber realizes intelligent amplification when the turbidity reaches the standard, and releases the mixed bacteria liquid to the culture container only when the turbidity reaches the preset value, ensuring that the bacteria entering the trapping area are in the logarithmic growth phase and the density is controllable, avoiding batch differences. The elastic trapping area realizes dynamic switching between efficient interception and lossless shedding through material deformation: the interception rate during the culture stage is more than 99%, preventing the loss of bacteria; the gas pulse of the reverse buffer zone makes the bacteria quickly shed to the sedimentation zone 13, which compresses the sedimentation time to 1 / 4 of the traditional method, and there is no mechanical shear damage. After sedimentation and separation, part of the bacteria liquid is directly collected for standby, and the other part is backflowed to the pipeline mixer to form a closed loop, not only strengthening the adaptive evolution and functional stability of the bacterial population, but also shortening the acclimation period from 30-45 days to 7-10 days. At the same time, through continuous flow operation and selective pressure maintenance, the metabolic synergy efficiency of nitrosobacteria and nitrifying bacteria in the device is improved by 2-3 times, realizing the dual breakthrough of denitrification efficiency and culture scale. In the field of bacterial culture and wastewater treatment technology, it has application value. BRIEF DESCRIPTION OF DRAWINGS

[0044] Figure 1 The results of the bacterial continuous flow scale culture device in Example 1 are shown in the schematic diagram. Figure 2 The scanning electron microscope image of the nitrifying bacteria cultured in Comparative Example 1 is shown in the schematic diagram. Figure 3 The species composition distribution diagram of the nitrifying bacteria cultured in Comparative Example 1 is shown in the schematic diagram. Figure 4 The scanning electron microscope image of the nitrifying bacteria cultured in Example 2 is shown in the schematic diagram. Figure 5 The species composition distribution diagram of the nitrifying bacteria cultured in Example 2 is shown in the schematic diagram. Wherein, 1-culture container, 11-reverse buffer zone, 111-backwash aeration pipe, 12-elastic capture area, 121-upper grid plate, 122-lower grid plate, 123-filter cotton, 124-second sliding rail, 13-settling zone, 14-drainage groove; 2-immobilized culture chamber, 21-bacterial liquid flow channel, 22-upper interception net, 23-lower interception net, 24-adsorption carrier, 25-aeration pipe, 26-first sliding rail; 3-centrifugal culture chamber, 31-overflow channel, 32-rotary base, 33-water distributor; 4-liquid inlet pipe; 5-drainage pipe; 6-drainage pipe; 7-back bacteria pipe; 8-bacteria collection pipe; 9-pipeline mixer. DETAILED DESCRIPTION

[0045] The present application will be described with reference to the preferred embodiments. Based on the description provided herein, those skilled in the art can easily derive other advantages and functions of the present application. The present application can also be implemented or applied in other different specific embodiments, and the details in the description can be modified or changed based on different views and applications without departing from the spirit of the present application. It should be understood that the preferred embodiments are only for illustrating the present application, but not for limiting the protection scope of the present application.

[0046] Embodiment 1 A bacterial continuous flow scale culture device, comprising: a culture container 1; The inside of the culture container 1 is provided with an immobilized culture chamber 2, and the bottom of the immobilized culture chamber 2 is provided with a bacterial liquid flow channel 21 in communication with the culture container 1; The inside of the immobilized culture chamber 2 is provided with a centrifugal culture chamber 3 for adjusting the hydraulic retention time, and the chamber wall of the centrifugal culture chamber 3 is provided with an overflow channel 31 in communication with the immobilized culture chamber 2; The culture container 1 is sequentially divided into a reverse buffer zone 11, an elastic capture area 12 and a settling zone 13 from top to bottom.

[0047] In some embodiments, the centrifugal culture chamber 3 is rotationally connected with the immobilized culture chamber 2 through a rotary base 32, so that the centrifugal culture chamber 3 can rotate around its axis, thereby realizing the rotational centrifugal culture of the culture liquid containing the bacterial source; the bottom of the centrifugal culture chamber 3 is provided with a water distributor 33, and the water distributor 33 is connected with the liquid inlet pipe 4; the centrifugal culture chamber 3 is provided with the overflow channel 31 on the side wall close to the top.

[0048] For example, the overflow channel 31 is composed of overflow holes opened on the side wall of the centrifugal culture chamber 3 close to the top, the aperture of the overflow hole is 15mm, and the spacing between adjacent overflow holes is 150mm; the centrifugal culture chamber 3 is in cylindrical structure, the overflow holes are arranged along the circumference of the side wall of the centrifugal culture chamber 3, and 15 overflow holes are opened on the side wall of the centrifugal culture chamber 3.

[0049] In some embodiments, the immobilized culture chamber 2 is provided with an upper interception net 22, a lower interception net 23, an adsorption carrier 24, an aeration pipe 25 and a first sliding rail 26; the upper interception net 22 and the lower interception net 23 are located above the aeration pipe 25; the adsorption carrier 24 is located between the upper interception net 22 and the lower interception net 23; the aeration pipe 25 is located at the bottom of the immobilized culture chamber 2; the first sliding rail 26 is arranged on the inner side wall of the immobilized culture chamber 2, and the lower interception net 23 is in sliding fit with the first sliding rail 26, so that the lower interception net 23 can slide up and down along the first sliding rail 26.

[0050] For example, the end of the upper interception net 22 is fixedly connected with the inner side wall of the immobilized culture chamber 2; the aeration pipe 25 is fixedly connected with the bottom of the immobilized culture chamber 2; the first sliding rail 26 is fixedly arranged on the inner side wall of the immobilized culture chamber 2 in the up-down direction; the adsorption carrier 24 has a diameter of 40 mm, a porosity of 85% and a filling rate of 40%.

[0051] In some embodiments, the reverse buffer zone 11 is provided with a backwashing aeration pipe 111; the elastic trapping zone 12 is provided with an upper grid plate 121, a lower grid plate 122, filter cotton 123 and a second sliding rail 124; the filter cotton 123 is arranged between the upper grid plate 121 and the lower grid plate 122; the second sliding rail 124 is arranged on the inner side wall of the culture container 1, and the upper grid plate 121 and the lower grid plate 122 are in sliding fit with the second sliding rail 124, so that the upper grid plate 121 and the lower grid plate 122 can slide up and down along the second sliding rail 124.

[0052] For example, the filter cotton 123 includes 4 layers, and the pore diameters of the layers from top to bottom are 0.8 mm, 1.5 mm, 2.5 mm and 3.5 mm, respectively.

[0053] In the above embodiments, the gas of the aeration pipe 25 flows upward, and the gas of the backwashing aeration pipe 111 flows downward.

[0054] In some embodiments, a drainage groove 14 is arranged near the top of the culture container 1, and the drainage groove 14 is connected with a drainage pipe 5; a liquid discharge pipe 6 is arranged at the bottom of the culture container 1 and connected with the sedimentation zone 13; the liquid discharge pipe 6 is arranged outside the culture container 1; the liquid discharge pipe 6 is connected with a bacteria return pipe 7 and a bacteria collection pipe 8, and the bacteria return pipe 7 is connected with the liquid inlet pipe 4 through a pipeline mixer 9.

[0055] In actual use, the bacterial continuous flow scale culture device is used as follows: the culture liquid containing bacterial source is mixed by the pipeline mixer 9 and then flows into the centrifugal culture chamber 3 through the liquid inlet pipe 4; then the culture liquid is centrifugally cultured in the centrifugal culture chamber 3 and then flows into the immobilized culture chamber 2 through the overflow channel 31; the mixed bacterial liquid cultured in the immobilized culture chamber 2 flows into the culture container 1 through the bacterial liquid flow channel 21. In the culture vessel 1, the bacteria in the mixed bacteria solution are trapped and enriched by the elastic trapping zone 12, and the bacterial excrement in the mixed bacteria solution is settled in the settlement zone 13; after the bacterial excrement is removed, the bacteria in the elastic trapping zone 12 are driven to fall off to the settlement zone 13 by the gas generated through the reverse buffer zone 11, and then part of the bacteria solution in the settlement zone 13 is collected and utilized, and the other part of the bacteria solution is returned to the centrifugal culture chamber 3, so that the continuous flow scale culture of bacteria is realized.

[0056] Embodiment 2 A culture method for culturing nitrifying bacteria by using the bacterial continuous flow scale culture device in embodiment 1, comprising the following steps: S1, the culture solution containing the nitrifying bacteria source is mixed by the pipeline mixer 9, and then flows into the centrifugal culture chamber 3 through the liquid inlet pipe 4 and the water distributor 33 for centrifugal culture; In the culture solution containing the nitrifying bacteria source, the volume content of the nitrifying bacteria source is 3%; The composition of the culture solution is (NH4)2SO4 2g, K2HPO4 0.75g, NaH2PO4 0.25g, MgSO4·7H2O 0.03g, MnSO4·4H2O 0.03g, and the pH is adjusted to 8.1-8.2 by Na2CO3; The model of the nitrifying bacteria source is DeCN-Bacteria, which is from Hangzhou Research Institute of China Coal Technology & Engineering Group Co., Ltd.; S2, the centrifugal culture time (i.e. the hydraulic retention time) is controlled to be 2h by adjusting the flow of the culture solution containing the nitrifying bacteria source in the centrifugal culture chamber 3, and the mixed bacteria solution after centrifugal culture flows into the immobilized culture chamber 2 through the overflow channel 31 composed of overflow holes opened on the side wall of the centrifugal culture chamber 3 close to the top; In the centrifugal culture process, the rotating speed of the rotating base 32 is 70r / min; S3, during the culture of the mixed bacteria solution in the immobilized culture chamber 2, air is introduced through the aeration pipe 25 to make the dissolved oxygen concentration in the immobilized culture chamber 2 be 2.0mg / L, when the turbidity of the mixed bacteria solution in the immobilized culture chamber 2 reaches 200NTU, the lower intercepting net 23 is moved upward along the first sliding rail 26, so that the adsorption carriers 24 gradually move upward, and when 70% of the adsorption carriers 24 are in the extrusion state, the movement of the lower intercepting net 23 is stopped; then the gas-water ratio is adjusted to 12:1 through the aeration pipe 25, so that the mixed bacteria solution flows into the culture vessel 1 through the bacteria flow channel 21 at the bottom of the immobilized culture chamber 2, and after maintaining for 4min, the lower intercepting net 23 is moved downward to restore to the initial position; S4. In the culture container 1, as the mixed bacterial solution flows upward, the nitrifying bacteria in the mixed bacterial solution are trapped and enriched by the filter cotton 123 of the elastic trapping zone 12. The excrement of the nitrifying bacteria in the mixed bacterial solution settles in the settling zone 13. The upper grid plate 121 moves upward along the second sliding track 124 as the mixed bacterial solution flows. When the upper grid plate 121 moves to the top of the second sliding track 124, the drain pipe 6 is opened to remove the nitrifying bacteria excrement that has settled in the reverse buffer zone 11. After the nitrifying bacteria excrement is discharged, the drain pipe 6 is closed, the backflushing aeration pipe 111 of the reverse buffer zone 11 is opened, and the air-to-water ratio is controlled at 6:1, so that the nitrifying bacteria in the elastic trapping zone 12 detach and settle into the settling zone 13 until the adsorption carrier 24 returns from the squeezed state to the initial state. Then the backflushing aeration pipe 111 is closed, and then the return pipe 7 and the collection pipe 8 are opened, so that a portion of the nitrifying bacteria in the mixed bacterial solution (i.e., the cultured nitrifying bacteria) flows out through the collection pipe 8 and is stored in a concentrated manner for subsequent treatment of coal mine domestic sewage. Another portion of the nitrifying bacteria flows through the return pipe 7 to the pipeline mixer 9 and is fully mixed with the culture solution before flowing back into the centrifuge culture chamber 3 to achieve continuous flow large-scale culture of nitrifying bacteria. The ratio of the amount of bacteria flowing into the return pipe 7 to the amount of bacteria flowing into the collection pipe 8 is 1:4. After the mixed bacterial solution is discharged, the return pipe 7 and the collection pipe 8 are closed.

[0057] Comparative Example 1 A routine method for culturing nitrifying bacteria includes the following steps: Nitrifying bacteria were continuously enriched for 6 cycles using a sequencing batch culture device equipped with stirring, heating and aeration. The culture cycle lasts 5 days, with a stirring speed of 80-180 r / min and a dissolved oxygen concentration of 2.5-4.5 mg / L. The culture temperature is 28℃-32℃. During the culture, ammonium salts and growth promoters are added, and polyaluminum chloride is added for flocculation and precipitation. After standing, the supernatant is discarded, and then enriched culture medium is added to start the next culture cycle.

[0058] Species composition analysis of nitrifying bacteria The viable nitrifying bacteria in the bacteria cultured in Example 2 and the control Example 1 were determined and counted using the dilution plate colony count method. The results showed that the concentration of nitrifying bacteria cultured in Example 2 was 2.4 × 10⁻⁶. 9 CFU / mL, the concentration of nitrifying bacteria cultured in control example 1 was 1.3 × 10⁻⁶. 7 CFU / mL.

[0059] The microbial species and distribution of nitrifying bacteria cultured in Example 2 and Control Example 1 were detected and analyzed using scanning electron microscopy and high-throughput sequencing. The results are as follows:Figures 2 to 5 As shown.

[0060] From Figure 2 and Figure 3 It can be known that the nitrifying bacteria cultured by the conventional culture method in the comparative example 1 is not dominant bacteria, and the colony composition is relatively complex.

[0061] From Figure 4 and Figure 5 It can be known from the comparative analysis that the nitrifying bacteria cultured by the culture device and the culture method of the present application are mainly spherical and short rod-shaped, and the proportion of Nitrosomonas is significant, which is the nitrosation dominant genus. The nitrosation dominant genus can oxidize ammonia nitrogen to nitrite nitrogen under aerobic conditions, shortening the nitrification reaction process. The nitrifying bacteria cultured by the conventional method have complex species composition, and there is no significant proportion of nitrosation dominant bacteria. The denitrification efficiency is low by traditional nitrification.

[0062] Coal mine plant domestic sewage treatment The nitrifying bacteria cultured in example 2 are inoculated into the coal mine plant domestic sewage biological contact oxidation tank at an inoculation amount of 600L, and are incubated for 2 days. In the case that the hydraulic retention time is 2.78 hours, it is operated for 6 days. The average biomass of the biological carrier biofilm in the biological contact oxidation tank is 26.3g / L measured by Kjeldahl nitrogen determination method. The ammonia nitrogen concentration in the coal mine plant domestic sewage treated by the biological contact oxidation tank is reduced to below 0.23mg / L, and the ammonia nitrogen removal rate reaches 98.6%.

[0063] In summary, the centrifugal culture zone of the bacterial continuous flow large-scale culture device of the present application can optimize the culture conditions of nitrosation bacteria and improve the proportion of nitrosation bacteria in the nitrifying bacteria population. The immobilized culture zone can effectively enrich nitrifying bacteria and continuously and efficiently culture high-attachment nitrifying bacteria. The flexible operation mode of the elastic capture zone can effectively prevent the loss of bacteria during the drainage process, shorten the sedimentation time, and eliminate the need for batch water change. Thus, the problem of long domestication cycle of the existing nitrifying bacteria culture method is solved, and the problem of low overall denitrification efficiency of the nitrifying bacteria cultured by the existing method is also solved. In the field of bacterial culture and sewage treatment technology, it has the value of popularization and application.

[0064] The above examples are only preferred examples for fully illustrating the present application, and the protection scope of the present application is not limited thereto. Any equivalent replacement or transformation made by the person skilled in the art on the basis of the present application is within the protection scope of the present application.

Claims

1. A bacterial continuous flow scale-up cultivation device, characterized by, The application relates to a culture container. The culture container (1) is internally provided with a fixed culture chamber (2), and the bottom of the fixed culture chamber (2) is provided with a bacterial liquid flow channel (21) communicated with the culture container (1); The inside of the fixed culture chamber (2) is provided with a centrifugal culture chamber (3) for adjusting the hydraulic retention time, and the chamber wall of the centrifugal culture chamber (3) is provided with an overflow channel (31) communicated with the fixed culture chamber (2); The culture container (1) is sequentially divided into a reverse buffer area (11), an elastic trapping area (12) and a sedimentation area (13) from top to bottom; The culture liquid containing a bacterial source flows into the centrifugal culture chamber (3) and is cultured by centrifugation, and then flows into the fixed culture chamber (2) through the overflow channel (31); the mixed bacterial liquid cultured in the fixed culture chamber (2) flows into the culture container (1) through the bacterial liquid flow channel (21); In the culture container (1), the bacteria in the mixed bacterial liquid are trapped and enriched by the elastic trapping area (12), and the bacterial excrement in the mixed bacterial liquid is settled in the sedimentation area (13); after the bacterial excrement is removed, the reverse buffer area (11) generates gas to drive the bacteria in the elastic trapping area (12) to fall off to the sedimentation area (13), and then part of the bacterial liquid in the sedimentation area (13) is collected and utilized, and the other part of the bacterial liquid flows back to the centrifugal culture chamber (3), so that the bacteria are continuously cultured in a large scale. The centrifugal culture chamber (3) is connected with the fixed culture chamber (2) through a rotating base (32); 2. The bacterial continuous flow scale-up cultivation device according to claim 1, characterized by, And / or, the bottom of the centrifugal culture chamber (3) is provided with a water distributor (33), and the water distributor (33) is connected with a liquid inlet pipe (4); And / or, the centrifugal culture chamber (3) is provided with the overflow channel (31) on the side wall close to the top. The fixed culture chamber (2) is provided with an upper intercepting net (22), a lower intercepting net (23), an adsorption carrier (24), an aeration pipe (25) and a first sliding rail (26); 3. The bacterial continuous flow scale-up cultivation device according to claim 1, characterized by, The upper intercepting net (22) and the lower intercepting net (23) are located above the aeration pipe (25); The adsorption carrier (24) is located between the upper intercepting net (22) and the lower intercepting net (23); The aeration pipe (25) is located at the bottom of the fixed culture chamber (2); The first sliding rail (26) is arranged on the inner side wall of the fixed culture chamber (2), and the lower intercepting net (23) is in sliding fit with the first sliding rail (26), so that the lower intercepting net (23) can slide up and down along the first sliding rail (26). A backflushing aeration pipe (111) is arranged in the reverse buffer area (11); 4. The bacterial continuous flow scale-up cultivation device according to claim 1, characterized by, And / or, an upper grid plate (121), a lower grid plate (122), a filter cotton (123) and a second sliding rail (124) are arranged in the elastic trapping area (12); The filter cotton (123) is arranged between the upper grid plate (121) and the lower grid plate (122); And / or, an upper grid plate (121), a lower grid plate (122), a filter cotton (123) and a second sliding rail (124) are arranged in the elastic trapping area (12); The second sliding rail (124) is arranged on the inner side wall of the culture container (1), and the upper grid plate (121) and the lower grid plate (122) are in sliding fit with the second sliding rail (124), so that the upper grid plate (121) and the lower grid plate (122) can slide up and down along the second sliding rail (124).

5. The bacterial continuous flow scale-up cultivation device according to claim 1, wherein A drainage groove (14) is arranged in the culture container (1) near the top, and a drainage pipe (5) is connected to the drainage groove (14).

6. The bacterial continuous flow scale-up cultivation device according to claim 1, wherein A liquid discharge pipe (6) is arranged at the bottom of the culture container (1) and connected to the sedimentation area (13); The liquid discharge pipe (6) is connected to a bacteria return pipe (7) and a bacteria collection pipe (8), and the bacteria return pipe (7) is connected to the liquid inlet pipe (4) through a pipeline mixer (9).

7. A method for culturing bacteria using the bacterial continuous flow scale-up culture device according to any one of claims 1 to 6, wherein The method comprises the following steps: S1, the culture solution containing the bacterial source is mixed by the pipeline mixer (9) and then flows into the centrifugal culture chamber (3) through the liquid inlet pipe (4); S2, after the mixed bacteria solution in the centrifugal culture chamber (3) is centrifugally cultured for a preset time, the mixed bacteria solution flows into the immobilized culture chamber (2) through the overflow channel (31); S3, when the turbidity of the mixed bacteria solution in the immobilized culture chamber (2) reaches a preset turbidity, the mixed bacteria solution flows into the culture container (1) through the bacteria solution flow channel (21); S4, in the culture container (1), the bacteria in the mixed bacteria solution are enriched by the elastic trapping area (12), and the bacterial excrement in the mixed bacteria solution is settled in the sedimentation area (13); after the bacterial excrement is removed, the reverse buffer area (11) generates gas to drive the bacteria in the elastic trapping area (12) to fall off into the sedimentation area (13), and then the bacteria solution in the sedimentation area (13) is collected, thereby obtaining the cultured bacteria; Part of the obtained bacteria solution is stored for standby, and the other part is returned to the pipeline mixer (9) to realize continuous flow scale culture of the bacteria.

8. The culture method according to claim 7, characterized by, The volume content of the bacterial source in the culture solution containing the bacterial source is 2-5%; And / or, the preset time is 2-2.5h; And / or, the rotation speed of the centrifugal culture is 60-80r / min; And / or, during the culture in the immobilized culture chamber (2), the dissolved oxygen concentration is 1.5-2.5mg / L; And / or, the preset turbidity is 200NTU; And / or, the bacteria are nitrifying bacteria.

9. The culturing method according to claim 7, wherein, In S3, the following steps are included: When the turbidity of the mixed bacteria solution in the immobilized culture chamber (2) reaches a preset turbidity, the lower intercepting net (23) moves upward along the first sliding rail (26), so that the adsorption carrier (24) gradually moves upward, and when 60-80% of the adsorption carrier (24) is in a squeezed state, the movement of the lower intercepting net (23) is stopped; Then, the gas-water ratio is adjusted to 12:1 through the aeration pipe (25), so that the mixed bacteria solution flows into the culture container (1) through the bacteria solution flow channel (21), and after maintaining for 3-5min, the lower intercepting net (23) is moved downward to the initial position; And / or, in S4, the following steps are included: In the culture container (1), the mixed bacteria solution flows upward, the bacteria in the mixed bacteria solution are enriched by the filter cotton (123) of the elastic trapping area (12), the bacteria excrement in the mixed bacteria solution is settled in the settlement area (13), the upper grid plate (121) moves upward along the second sliding track (124) with the flow of the mixed bacteria solution, until the upper grid plate (121) moves to the top of the second sliding track (124), the liquid discharge pipe (6) is opened to discharge the bacteria excrement settled in the reverse buffer area (11); After the bacteria excrement is discharged, the liquid discharge pipe (6) is closed, the backwashing aeration pipe (111) of the reverse buffer area (11) is opened, and the gas-water ratio is controlled to be 6:1, until the adsorption carrier (24) is restored to the initial state from the extrusion state, the backwashing aeration pipe (111) is closed, the bacteria return pipe (7) and the bacteria collection pipe (8) are opened, part of the bacteria in the mixed bacteria solution flows out through the bacteria collection pipe (8) and is concentrated and stored, and the other part of the bacteria flows to the pipeline mixer (9) through the bacteria return pipe (7), so that the bacteria are continuously and flow-scaled cultured.

10. Application of the bacteria obtained by the culture method according to any one of claims 7 to 9 in treatment of coal mine domestic sewage.

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