CN111595975B - Method for rapidly inhibiting activated sludge non-filamentous bacteria expansion - Google Patents
Method for rapidly inhibiting activated sludge non-filamentous bacteria expansion Download PDFInfo
- Publication number
- CN111595975B CN111595975B CN202010482120.5A CN202010482120A CN111595975B CN 111595975 B CN111595975 B CN 111595975B CN 202010482120 A CN202010482120 A CN 202010482120A CN 111595975 B CN111595975 B CN 111595975B
- Authority
- CN
- China
- Prior art keywords
- activated sludge
- filamentous bacteria
- sludge
- sewage
- microorganisms
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F11/00—Treatment of sludge; Devices therefor
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6869—Methods for sequencing
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/645—Specially adapted constructive features of fluorimeters
- G01N21/6456—Spatial resolved fluorescence measurements; Imaging
- G01N21/6458—Fluorescence microscopy
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/62—Detectors specially adapted therefor
- G01N30/72—Mass spectrometers
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
- G01N2021/6439—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" with indicators, stains, dyes, tags, labels, marks
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Immunology (AREA)
- Physics & Mathematics (AREA)
- Organic Chemistry (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Pathology (AREA)
- Engineering & Computer Science (AREA)
- General Physics & Mathematics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Microbiology (AREA)
- Hydrology & Water Resources (AREA)
- Water Supply & Treatment (AREA)
- Environmental & Geological Engineering (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Optics & Photonics (AREA)
- Biodiversity & Conservation Biology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Molecular Biology (AREA)
- Biophysics (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention provides a method for rapidly inhibiting the non-filamentous bacteria swelling of activated sludge, belonging to the technical field of sewage treatment, and the treatment method specifically comprises the following steps: sampling activated sludge, determining the type of sludge bulking, determining the type and content of signal molecules in sewage, screening reinforced microorganisms, fermenting reinforced microorganisms, and finally adding fermented microorganism fermentation liquor into a target tank body. The method of the invention is based on the microbionosis effect, promotes the secretion of extracellular polymers of microorganisms, further improves the specific gravity of sludge, and achieves the purpose of rapidly inhibiting the non-filamentous bacteria expansion of activated sludge. The method provided by the invention adopts a specific strengthening means on the basis of defining the wastewater treatment microorganisms, and has the advantages of strong pertinence, good gain effect and high sludge bulking inhibition speed.
Description
Technical Field
The invention belongs to the technical field of sewage treatment by an activated sludge process.
Background
The activated sludge process is taken as a mainstream process of a sewage treatment plant, and the sludge bulking is a phenomenon frequently generated in the activated sludge process of the sewage treatment plant and has high performance, universality and harmfulness. At present, more than 30 percent of domestic sewage treatment plants adopting the traditional activated sludge process suffer from the sludge bulking problem every year, so an effective means is urgently needed to solve the activated sludge bulking problem and ensure the efficient and stable operation of an activated sludge system. Typical sludge bulking includes filamentous bacteria bulking and non-filamentous bacteria bulking, and the number of filamentous bacteria can be generally divided into 0-5 grades, wherein: FI is 0: almost no filamentous bacteria exist in the activated sludge; FI 1: a small amount of filamentous bacteria was observed in the activated sludge; FI is 2: a moderate number of filamentous bacteria were observed in the activated sludge, but the total amount was lower than that of zoogloea bacteria; and FI is 3: a large number of filamentous bacteria were observed in the activated sludge, the total amount being approximately equal to zoogloea bacteria; FI 4: the number of filamentous bacteria in the activated sludge exceeds that of zoogloea bacteria; FI 5: the number of filamentous bacteria in the activated sludge is absolutely predominant, the zoogloea bacteria are few, and when FI is less than 3, the activated sludge bulking can be regarded as non-filamentous bacteria bulking.
At present, a great deal of research is carried out on the expansion of activated sludge filamentous bacteria, and the control means comprises specific control means mainly for supplementing nutrient substances, adjusting process parameters and the like and non-specific control means mainly for adding an oxidant, a weighting agent, a flocculating agent and the like. But the means of controlling the expansion of non-filamentous bacteria in activated sludge is relatively limited.
Chinese invention patent CN201710611895.6, published as 2019, 10.25.A broad-spectrum biochemical sludge bulking control agent and a preparation method thereof are disclosed, wherein the agent is prepared by mixing an iron ion salt solution, activated carbon and graphene and then reacting the mixture with an alkali solution, the final component is a mixture of iron oxide, activated carbon and graphene, and the agent combines the high-efficiency electron conduction function of graphene, the biological affinity of activated carbon, the electron neutralization of iron ions and the flocculation effect. Further, as disclosed in chinese invention patent CN201810913887.1, published as 2018, 12 and 18 months, a sludge bulking eliminating agent and a method for using the same are disclosed, which comprises: 25-35 parts of Portland cement clinker and 65-75 parts of fly ash dry ash, wherein the usage method of the remover comprises the step of pouring the sludge bulking remover into the sewage, and the pouring amount is 50-100mg/L, so that the method can cause activated sludge hardening and reduce the sewage treatment efficiency.
The methods all belong to methods for treating by adding chemical reagents, not only can increase the burden of subsequent wastewater treatment, but also have difficult and durable action, so that a new, safer and more effective method is urgently needed to be developed aiming at the sludge bulking caused by the activated sludge non-filamentous bacteria.
Disclosure of Invention
1. Problems to be solved
Aiming at the problems that the sludge bulking caused by non-filamentous bacteria in the activated sludge is difficult to completely solve by adding a chemical reagent and the problem of wastewater treatment burden caused by adding the chemical reagent, the invention develops a biological treatment method based on the sludge property in the wastewater, can be used for pertinently treating the sludge bulking caused by the non-filamentous bacteria, does not need to add the chemical reagent, does not increase the subsequent wastewater treatment burden, and is more effective for the sludge bulking treatment for a long time.
2. Technical scheme
In order to solve the problems, the technical scheme adopted by the invention is as follows:
a method for rapidly inhibiting the non-filamentous bacterium bulking of activated sludge comprises the following steps:
1) sampling
Dividing the target tank body into regions, sampling each region, respectively collecting samples (including water and activated sludge) at different depth positions away from the liquid level, and mixing the collected samples to obtain a mixed sample;
2) sludge bulking species determination
Taking the mixed sample obtained in the step 1), separating out the activated sludge, and identifying the filamentous fungus index in the activated sludge within 0-2 h, wherein when the filamentous fungus index is less than 3, the identification of the microbial community structure of the activated sludge is carried out;
3) identification of activated sludge microbial community structure
Analyzing by using a 16S rRNA gene sequencing technology to obtain microbial community structure information of the activated sludge, and screening out microorganisms with quorum sensing according to the obtained microbial community structure information;
4) determination of signal molecule type and content in sewage
Taking the mixed sample in the step 1), adjusting the pH value to 4.0-5.0, performing centrifugal separation to obtain sewage, and determining the content and the type of signal molecules in the sewage by using a liquid chromatography-mass spectrometer to obtain the main signal molecule type; the purpose of adjusting the pH to 4.0-5.0 is to prevent degradation of the signal molecule by acidification.
5) Screening of the enhanced microorganisms: comparing the main signal molecule type in the step 4) with the signal molecule type secreted by the microorganism with quorum sensing screened in the step 3), and further screening out the microorganism capable of secreting the signal molecule type same as that in the step 4), wherein the microorganism is taken as an enhanced microorganism;
6) strengthening the fermentation and addition of microorganisms
Performing fermentation culture on the enhanced microorganisms obtained in the step 5), and adding fermentation liquor to the target tank body.
Preferably, the main signal molecule species comprises AHLs signal molecules.
Preferably, the scheme is as follows: in the step 1), the target pool body is divided into 3-5 areas, and sampling is carried out on each area; respectively collecting samples with the depths of 0-0.5 m, 1-1.5 m and 2.5-3 m from the liquid level during sampling, and mixing 9-15 collected samples; the specific method during sampling is as follows: sampling is carried out by using a constant-flow peristaltic pump, a sample inlet of a pump pipe of the constant-flow peristaltic pump is placed at a preset target position during sampling, the sampling is carried out for 1-3min, the sampling flow is 10-100 mL/min, and after each sampling, the peristaltic pump is started to pump 500-1500mL sampling point sewage to rinse the pump pipe before next sampling.
Preferably, in the step 2), the specific method for identifying the filamentous Fungus Index (FI) in the activated sludge is as follows: taking 3-5mL of activated sludge, dyeing an activated sludge sample (diluted by 2 times) by using 10% ammonium oxalate crystal violet dyeing solution, observing and measuring the number of filamentous bacteria in the sludge sample by using a fluorescence microscope to further determine a filamentous bacteria index (FI), and determining the filamentous bacteria index (FI) according to the ratio of the filamentous bacteria and zoogloea bacteria observed in a visual field by using a visual evaluation method.
Preferably, in the step 3), the specific steps of obtaining the microbial community structure information of the activated sludge by using 16S rRNA gene sequencing technology analysis are as follows:
adding equal volume of ethanol into 10-20 mL of the activated sludge obtained in the step 1), preserving at-20 ℃ until DNA is extracted, preferably performing PCR amplification on a V1-V2 region of the extracted DNA, and then sequencing 16S rRNA to obtain community structure information of microorganisms; wherein, the FastDNA SPIN Kit is preferably selected for extracting the DNA of the activated sludge, and the specific operation steps refer to a Kit operation manual; wherein the preferable times for extracting the DNA sample is 3-5 times;
the V1-V2 region amplification primers are as follows: the forward primer is 5 '-AGAGTTTGATYMTGGCTCAG-3' and the reverse primer is 5'-TGCTGCCTCCCGTAGGAGT-3';
purifying the PCR amplification product by adopting an E.Z.N.A.TM Cycle-Pure kit; high throughput sequencing can be directly determined using the Illumina Miseq platform.
Preferably, in the step 3), the microorganisms having quorum sensing in the microorganisms are selected according to the acquired microbial community structure information, and the steps are as follows: inputting the acquired microbial community structure information into an environmental quorum sensing microbe retrieval platform (http:// www.njuqsb.com/microbe/f), clicking to accurately search and compare, and directly outputting whether the activated sludge microbes have a quorum sensing function or not by the retrieval platform.
Preferably, in the step 4), the mixed sample in the step 1) is taken, HCl is adopted to adjust the pH value to 4.0-5.0, then the mixed sample is centrifuged for 5-10 min under the centrifugal force of 3000-5000 Xg, and the centrifuged sewage is enriched by using a solid phase extraction column (preferably HLB).
Preferably, in the 5), the strengthening microorganism further needs to satisfy the following conditions: the pure culture can be carried out under the conditions that the culture temperature is 5-35 ℃ and the culture medium is LB culture solution.
In a preferred embodiment, in 6), the conditions for culturing the enhanced microorganism are as follows: the culture temperature is the same as the temperature of the sewage in the target tank, and the culture solution is a mixture of the sewage in the target tank and LB culture solution (prepared according to molecular cloning experimental guidelines (J. SammBruker D.W. Lassel.).
Preferably, in the culture solution, the volume ratio of the added sewage to the LB culture solution is (300-100): 1.
preferably, in the step 6), the OD value of the microorganism is 0.6-0.8, and the fermentation can be completed in the logarithmic growth phase.
Preferably, in the 6), the volume ratio of the adding amount of the fermentation liquid to the target tank body is 1: (5000 to 20000).
In the preferable scheme, in the step 6), after the fermentation liquid is added into the target tank body, the sludge reflux ratio of the target tank body needs to be increased to 1.2-1.5 times of the original reflux ratio.
3. Advantageous effects
Compared with the prior art, the invention has the beneficial effects that:
(1) the invention provides a method for rapidly inhibiting activated sludge non-filamentous bacteria expansion, which comprises the steps of firstly defining waste water treatment microorganisms, then adopting a specific inhibition method, wherein the non-filamentous bacteria expansion is viscous sludge expansion caused by high-hydrophilicity viscous substances adhered to the sludge, and is difficult to directly and effectively treat by adding chemical reagents relative to the filamentous bacteria expansion. Thereby further improving the specific gravity of the sludge and achieving the purpose of quickly inhibiting the non-filamentous bacteria expansion of the activated sludge; compared with the existing method for inhibiting sludge bulking, the method does not need to add chemical reagents and does not increase the burden of wastewater treatment; moreover, the effect is durable and the repetition is not easy.
(2) In the process of directionally screening the enhanced microorganism strains, firstly, a large-range microorganism variety is obtained based on the microbial community structure information obtained by the activated sludge in the wastewater, the large-range microorganism variety is compared with an environmental quorum sensing microorganism retrieval platform to obtain a small-range microorganism variety with quorum sensing in the wastewater, on the basis, a main signal molecule variety obtained from the wastewater is compared with a signal molecule variety secreted by the screened microorganism to screen the enhanced strains capable of secreting the same signal molecule variety, and a layer-by-layer screening mode from a large range to a small range is adopted in the steps, so that the obtained enhanced strain quorum sensing effect is stronger, and the sludge bulking problem generated in the wastewater can be more specifically inhibited.
(3) According to the method for rapidly inhibiting the non-filamentous bacteria expansion of the activated sludge, the target pool body is divided into regions in the sampling process, each region is sampled, samples at different depths from the liquid level are respectively collected, the collected samples are mixed, a mixed sample is obtained and processed, and the obtained sample is more uniform and complete; the sludge expansion level of the target tank body can be reflected to the maximum extent, and the misjudgment probability of the traditional random sampling method is overcome to the maximum extent.
(4) According to the method for rapidly inhibiting the expansion of the activated sludge non-filamentous bacteria, provided by the invention, the microorganisms are cultured and fermented based on the simulation of the target tank body environment, the adaptability of the microorganisms to the target tank body environment is improved to the greatest extent on the basis of ensuring the survival of the microorganisms, and the risk of screening and applying strains is reduced.
Drawings
FIG. 1 is a diagram showing the microbial community structure in the sludge of the aerobic tank in example 1;
FIG. 2 shows Sphingomonas rubra growth curves and secretory signal molecule species.
Detailed Description
The invention is further described with reference to specific examples.
Example 1
The sedimentation performance of the activated sludge in the aerobic pool in the AAO process of a certain municipal sewage plant is poor, the sedimentation time of the sludge is higher than the design time, and the sludge bulking is presumed to occur and is analyzed. The method comprises the following specific steps:
1) sampling
The aerobic pool is divided into areas, sampling is carried out on each area, samples (including water and biological sludge) at different depth positions from the liquid level are respectively collected, and the collected samples are mixed to obtain a mixed sample; specifically, a target pool body is divided into 3 areas, and sampling is carried out on each area; respectively collecting samples with the depth of 0m, 1m and 2.5m from the liquid level during sampling, and mixing 9 collected samples; the specific method during sampling is as follows: sampling is carried out by using a constant-flow peristaltic pump, a pump pipe sample inlet of the constant-flow peristaltic pump is placed at a preset target position during sampling, the sampling is carried out for 3min, the sampling flow is 10mL/min, and after each sampling, the peristaltic pump is required to be started to pump 500mL of sampling point sewage to rinse the pump pipe before next sampling.
2) Sludge bulking species determination
Taking the mixed sample in the step 1), separating out activated sludge, and identifying the filamentous fungus index in the activated sludge within 2 hours; identifying the microbial community structure of the activated sludge; the specific method for identifying the filamentous Fungus Index (FI) in the activated sludge is as follows: taking 3mL of activated sludge, dyeing an activated sludge sample (diluted by 2 times) by using 10% ammonium oxalate crystal violet dyeing liquid, observing and measuring the number of filamentous bacteria in the sludge sample by adopting a fluorescence microscope to further determine a filamentous bacteria index (FI), and determining the filamentous bacteria index (FI) according to the ratio of the filamentous bacteria and zoogloea bacteria observed in a visual field by adopting a visual evaluation method; the filamentous Fungus Index (FI) in this example was 3.
3) Identification of activated sludge microbial community structure
The method comprises the following steps of utilizing 16S rRNA gene sequencing technology to analyze and obtain microbial community structure information of activated sludge: adding equal volume of ethanol into 10mL of the activated sludge obtained in the step 1), preserving at-20 ℃ until DNA is extracted, preferably performing PCR amplification on V1-V2 region of the extracted DNA, and then sequencing 16S rRNA to obtain community structure information of microorganisms; wherein, the FastDNA SPIN Kit is preferentially recommended for the extraction of the DNA of the activated sludge, and the specific operation steps refer to a Kit operation manual; wherein the number of times of extracting the DNA sample is 3;
the V1-V2 region amplification primers are as follows: the forward primer is 5 '-AGAGTTTGATYMTGGCTCAG-3' and the reverse primer is 5'-TGCTGCCTCCCGTAGGAGT-3';
the amplification procedure is preferably selected as follows: preheating (98 ℃,5min) → amplification procedure [ denaturation (98 ℃,30s) → annealing (50 ℃; 30s) → extension (2 ℃; 40s) → extension (72 ℃; 10min) → cycles (20 times) ];
purifying the PCR amplification product by adopting an E.Z.N.A.TM Cycle-Pure kit; high throughput sequencing was directly determined using the Illumina Miseq platform.
The microbial community structure information obtained in this example is shown in fig. 1.
Inputting the acquired microbial community structure information into an environmental quorum sensing microbial retrieval platform (http:// www.njuqsb.com/microbe/f), clicking for accurate search, comparing, and screening out the microbial results with quorum sensing, wherein QS shows that the microbe has quorum sensing function, and QQ shows that the microbe has quorum sensing quenching function, and the results are shown in Table 1; the top 5 types of microorganisms in table 1 are potentially applicable in this example by bacteria detailed information comparison.
Table 1 environmental group-sensitive microorganism search platform output results
4) Determination of signal molecule type and content in sewage
Taking the mixed sample in the step 1), adjusting the pH value to 4.0 by adopting HCl, then centrifuging for 10min under the centrifugal force of 3000 Xg, and enriching the signal molecules in the centrifuged sewage by using an HLB solid-phase extraction column, wherein the specific enrichment conditions are as follows: the volume of the wastewater is 500mL, 3mL of methanol is used for activation, 3mL of water is used for balancing, the sample is loaded, 5:95(v/v) methanol-water solution (2% ammonium hydroxide) is used for cleaning, 5:95(v/v) methanol-water solution (2% acetic acid) is used for cleaning, and methanol (2% acetic acid) is used for eluting;
the content and the type of signal molecules in the sewage are measured by adopting a liquid chromatography-mass spectrometer (triple quadrupole liquid chromatography-mass spectrometer), and the specific conditions are as follows:
the chromatographic determination conditions are as follows: flow rate: 0.2mL/min, sample size: 10 μ L, assay time: 12min, column temperature: the mobile phase A is methanol at 35 ℃, and contains ammonium acetate with the molar concentration of 2mmoI/L and formic acid with the mass concentration of 0.1 percent; b is water, containing 2mmoI/L ammonium acetate and 0.1% formic acid;
the mass spectrometry conditions are as follows: electrospray ion source (ESI), ion source temperature 120 deg.C, desolventizing temperature 350 deg.C; desolventizing gas and taper hole gas N2The flow rate of the desolventizing agent is 500L/h, the flow rate of the taper hole is 50L/h, and the collision gas is argon; the capillary voltage is 3.50 kV; the scanning mode is positive ion scanning, and the residence time is 50 ms; the monitoring mode is the MRM mode. The main AHLs signal molecule in the wastewater is determined to be N decanoyl homoserine lactone (C)10HSL) and N-dodecylhomoserine lactone (C)12-HSL)。
5) Enhanced screening of microorganisms
Comparing the types of the main signal molecules obtained in the step 4) with the types of the signal molecules secreted by the microorganisms screened in the step 3), selecting the microorganisms with the types of the signal molecules secreted in the step 3) being the same as the types of the main signal molecules in the step 4), and performing pure culture at the culture temperature of 30 ℃ by using the microorganisms of an LB culture solution (prepared according to molecular cloning experimental guidance (J. SammBruker D.W. Lassel) as an enhanced microorganism; the microorganism of the present example is Sphingomonas rubra, and the growth curve and the type of signal molecule for secretion are shown in FIG. 2, which is N-butylhomoserine lactone (C)4-HSL), N decanoyl homoserine lactone (C)10HSL), N-dodecylhomoserine lactone (C)12-HSL), N-tetradecylhomoserine lactone (C)14-HSL)。
6) Strengthening the fermentation and addition of microorganisms
Carrying out fermentation culture on the enhanced microorganisms obtained in the step 5), wherein the specific fermentation conditions are as follows:
the culture temperature is as follows: the temperature of the water is 28 ℃ which is the same as that of the water in the target pool body;
culture solution: the adding volume ratio of the sewage to the LB culture solution is 300: 1;
the microorganism is continuously stirred and cultured, the OD value of the microorganism is 0.8, and the fermentation can be completed when the microorganism is in the logarithmic phase.
Adding the fermentation liquor into a target tank body, wherein the adding ratio is 1: 20000, and simultaneously, the sludge reflux ratio of the target tank body is increased to 1.2 times of the original reflux ratio.
And simultaneously detecting the expansion condition of the sludge in the target tank body before and after the fermentation liquor is added, wherein the detected SVI of the target tank body before the fermentation liquor is added is as high as about 350mL/g, the detected SV30 is as high as more than 85%, the SVI represents the volume index of the sludge, the good SVI of the activated sludge is usually 50-120 mL/g, the detected SV30 represents the volume percentage of the sludge after the mixed liquor in the aeration tank is statically settled for 30min, and the data show that the sludge expansion generated by the biochemical system is serious.
After the fermentation liquor is continuously added for 3 days, the SV30 value is sharply reduced, and the SVI value is reduced to 65mL/g within 3 days, which shows that the method has a particularly remarkable effect of inhibiting sludge bulking.
Example 2
The sedimentation performance of the activated sludge in the aerobic pool in the SBR process of a certain municipal sewage plant is poor, the sedimentation time of the sludge is higher than the design time, and the sludge bulking is presumed to possibly occur and is analyzed. The method comprises the following specific steps:
1) activated sludge sampling
Dividing SBR, sampling each region, respectively collecting samples at different depth positions from the liquid level, and mixing the collected samples to obtain a mixed sample; specifically, a target pool body is divided into 5 areas, and sampling is carried out on each area; respectively collecting samples with the depths of 0.5m, 1.5m and 3m from the liquid level during sampling, and mixing the 15 collected samples; the specific method during sampling is as follows: sampling is carried out by using a constant-flow peristaltic pump, a sample inlet of a pump pipe of the constant-flow peristaltic pump is placed at a preset target position during sampling, the sampling is carried out for 1min, the sampling flow is 100mL/min, and after each sampling, the peristaltic pump is required to be started to pump 1500mL sampling point sewage to rinse the pump pipe before next sampling.
2) Sludge bulking species determination
Taking the mixed sample in the step 1), separating out activated sludge, and identifying the filamentous fungus index in the activated sludge within 2 hours; identifying the microbial community structure of the activated sludge; the specific method for identifying the filamentous Fungus Index (FI) in the activated sludge is as follows: taking 5mL of activated sludge, dyeing an activated sludge sample (diluted by 2 times) by using 10% ammonium oxalate crystal violet dyeing liquid, observing and measuring the number of filamentous bacteria in the sludge sample by adopting a fluorescence microscope to further determine a filamentous bacteria index (FI), and determining the filamentous bacteria index (FI) according to the ratio of the filamentous bacteria and zoogloea bacteria observed in a visual field by adopting a visual evaluation method; the filamentous Fungus Index (FI) in this example was 2.
3) Identification of activated sludge microbial community structure
The method comprises the following steps of utilizing 16S rRNA gene sequencing technology to analyze and obtain microbial community structure information of activated sludge: adding equal volume of ethanol into 20mL of the activated sludge obtained in the step 1), preserving at-20 ℃ until DNA is extracted, preferably performing PCR amplification on a V1-V2 region of the extracted DNA, and then sequencing 16S rRNA to obtain community structure information of microorganisms; wherein, the FastDNA SPIN Kit is preferentially recommended for the extraction of the DNA of the activated sludge, and the specific operation steps refer to a Kit operation manual; wherein the number of times of extracting the DNA sample is 5;
the V1-V2 region amplification primers are as follows: the forward primer is 5 '-AGAGTTTGATYMTGGCTCAG-3' and the reverse primer is 5'-TGCTGCCTCCCGTAGGAGT-3';
the amplification procedure is preferably selected as follows: preheating (98 ℃,5min) → amplification procedure [ denaturation (98 ℃,30s) → annealing (50 ℃; 30s) → extension (2 ℃; 40s) → extension (72 ℃; 10min) → cycles (20 times) ];
purifying the PCR amplification product by adopting an E.Z.N.A.TM Cycle-Pure kit; high throughput sequencing was performed directly using Illumina Miseq platform to obtain information on microbial community structure in this example.
The obtained microbial community structure information is input into an environmental quorum sensing microbial retrieval platform (http:// www.njuqsb.com/microbe/f) to click an accurate search for comparison, and microbes having quorum sensing therein are screened as potential microbes to be applied in the embodiment.
4) Determination of signal molecule type and content in sewage
Taking the mixed sample in the step 1), adjusting the pH value to 5.0 by adopting HCl, then centrifuging for 5min under the centrifugal force of 5000 Xg, and enriching the signal molecules in the centrifuged sewage by using an HLB solid-phase extraction column, wherein the specific enrichment conditions are as follows: the volume of the wastewater is 500mL, 3mL of methanol is used for activation, 3mL of water is used for balancing, the sample is loaded, 5:95(v/v) methanol-water solution (2% ammonium hydroxide) is used for cleaning, 5:95(v/v) methanol-water solution (2% acetic acid) is used for cleaning, and methanol (2% acetic acid) is used for eluting;
the content and the type of signal molecules in the sewage are measured by adopting a liquid chromatography-mass spectrometer (triple quadrupole liquid chromatography-mass spectrometer), and the specific conditions are as follows:
the chromatographic determination conditions are as follows: flow rate: 0.2mL/min, sample size: 10 μ L, assay time: 12min, column temperature: the mobile phase A is methanol containing ammonium acetate with the molar number of 2mmoI/L and formic acid with the mass number of 0.1 percent at 35 ℃; b is water containing 2mmoI/L ammonium acetate and 0.1 mass percent of formic acid;
the mass spectrometry conditions are as follows: electrospray ion source (ESI), ion source temperature 120 deg.C, desolventizing temperature 350 deg.C; desolventizing gas and taper hole gas N2, desolventizing gas flow rate of 500L/h, taper hole gas flow rate of 50L/h, and collision gas of argon; the capillary voltage is 3.50 kV; the scanning mode is positive ion scanning, and the residence time is 50 ms; the monitoring mode is the MRM mode. Determining the main AHLs signal molecule in the wastewater as C10HSL and C12-HSL。
5) Enhanced screening of microorganisms
Comparing the types of the main signal molecules obtained in the step 4) with the types of the signal molecules secreted by the microorganisms screened in the step 3), selecting the microorganisms with the types of the signal molecules secreted in the step 3) being the same as the types of the main signal molecules in the step 4), and performing pure culture at the culture temperature of 5-35 ℃ by using the microorganisms of an LB culture solution (prepared according to molecular cloning experimental guidelines (J. SammBruke D.W. Lassel) as an enhanced microorganism; the microorganism of the present example is Bacillus cereus.
6) Strengthening the fermentation and addition of microorganisms
Carrying out fermentation culture on the enhanced microorganisms obtained in the step 5), wherein the specific fermentation conditions are as follows:
the culture temperature is as follows: the temperature of the water is 30 ℃ which is the same as that of the water in the target pool body;
culture solution: the adding volume ratio of the sewage to the LB culture solution is 300: 1;
the microorganism is continuously stirred and cultured, the OD value of the microorganism is 0.6, and the fermentation can be completed when the microorganism is in the logarithmic phase.
And (3) adding the fermentation liquor into the target tank body at the adding ratio of 1:5000, and simultaneously increasing the sludge reflux ratio of the target tank body to be 1.5 times of the original reflux ratio.
After the continuous addition for 4 days, the sludge sedimentation performance is obviously improved, and the inhibition of sludge bulking is completed.
And simultaneously detecting the expansion condition of the sludge in the target tank body before and after the fermentation liquor is added, wherein the detected SVI of the target tank body before the fermentation liquor is added is as high as about 295mg/L, the detected SV30 is as high as more than 82%, the SVI represents the volume index of the sludge, the good SVI of the activated sludge is usually 50-120 mL/g, the detected SV30 represents the volume percentage of the sludge after the mixed liquor in the aeration tank is statically settled for 30min, and the data shows that the sludge expansion generated by the biochemical system is serious.
After the fermentation liquor is continuously added for 4 days, the SV30 value is sharply reduced, and the SVI value is reduced to 55mL/g within 4 days, which shows that the method has a particularly remarkable effect of inhibiting sludge bulking.
Example 3
The sedimentation performance of the activated sludge in the aerobic pool in the CASS process of a certain municipal sewage plant is poor, the sedimentation time of the sludge is higher than the design time, and the sludge bulking is presumed to possibly occur and is analyzed. The method comprises the following specific steps:
1) activated sludge sampling
Dividing CASS, sampling for each region, respectively collecting samples at different depth positions from the liquid level, and mixing the collected samples to obtain a mixed sample; specifically, a target pool body is divided into 4 areas, and sampling is carried out on each area; respectively collecting samples with the depths of 0.5m, 1.5m and 2.5m from the liquid level during sampling, and mixing the 12 collected samples; the specific method during sampling is as follows: sampling is carried out by using a constant-flow peristaltic pump, a sample inlet of a pump pipe of the constant-flow peristaltic pump is placed at a preset target position during sampling, the sampling is carried out for 2min, the sampling flow is 60mL/min, and after each sampling, the peristaltic pump is required to be started to pump 800mL of sampling point sewage to rinse the pump pipe before next sampling.
2) Sludge bulking species determination
Taking the mixed sample in the step 1), separating out activated sludge, and identifying the filamentous fungus index in the activated sludge within 2 hours; identifying the microbial community structure of the activated sludge; the specific method for identifying the filamentous Fungus Index (FI) in the activated sludge is as follows: taking 4mL of activated sludge, dyeing an activated sludge sample (diluted by 2 times) by using 10% ammonium oxalate crystal violet dyeing liquid, observing and measuring the number of filamentous bacteria in the sludge sample by adopting a fluorescence microscope to further determine a filamentous bacteria index (FI), and determining the filamentous bacteria index (FI) according to the ratio of the filamentous bacteria and zoogloea bacteria observed in a visual field by adopting a visual evaluation method; the filamentous Fungus Index (FI) in this example was 5.
Therefore, the sludge bulking species in this example is considered to belong to filamentous bacteria bulking and is not considered to fall within the scope of the patent for inhibiting bulking.
The invention and its embodiments have been described in detail, without limitation, and it is possible to implement the invention in other specific forms without departing from the spirit or essential characteristics thereof.
Claims (10)
1. A method for rapidly inhibiting the non-filamentous bacteria swelling of activated sludge is characterized by comprising the following steps: the method comprises the following steps:
1) sampling: collecting a target cell body sample;
2) determining the sludge bulking type: taking the sample collected in the step 1), separating out activated sludge, and identifying the filamentous fungus index in the activated sludge, wherein when the filamentous fungus index is less than 3, the identification of the microbial community structure of the activated sludge is carried out;
3) identification of the microbial community structure of activated sludge: acquiring the structure information of the microbial community of the activated sludge, and screening out microorganisms with quorum sensing according to the acquired structure information of the microbial community;
4) determining the types and the contents of signal molecules in the sewage: taking the sample collected in the step 1), performing centrifugal separation to obtain sewage, and determining the content and the type of signal molecules in the sewage to obtain the type of main signal molecules;
5) screening of the enhanced microorganisms: comparing the main signal molecule type in the step 4) with the signal molecule type secreted by the microorganism with quorum sensing screened in the step 3), and further screening out the microorganism capable of secreting the signal molecule type same as that in the step 4), wherein the microorganism is taken as an enhanced microorganism;
6) fermentation and addition of the reinforced microorganisms: performing fermentation culture on the enhanced microorganisms obtained in the step 5), and adding fermentation liquor to the target tank body.
2. The method for rapidly inhibiting activated sludge non-filamentous bacteria bulking according to claim 1, wherein: in the step 1), the target pool body is divided into regions, each region is sampled, samples at different depth positions from the liquid level are collected respectively, and the collected samples are mixed to obtain a mixed sample.
3. The method for rapidly inhibiting activated sludge non-filamentous bacteria bulking according to claim 2, wherein: in the step 1), the target pool body is divided into 3-5 areas, and sampling is carried out on each area; and respectively collecting samples with the depths of 0-0.5 m, 1-1.5 m and 2.5-3 m from the liquid level during sampling, and mixing the collected samples.
4. The method for rapidly inhibiting activated sludge non-filamentous bacteria bulking according to claim 1 or 2, wherein: and 4) enriching the centrifuged sewage by using an HLB (hydrophile-lipophile balance) solid-phase extraction column, and/or identifying the filamentous fungus index in the activated sludge within 0-2 h after the activated sludge is separated in the step 2).
5. The method for rapidly inhibiting activated sludge non-filamentous bacteria bulking according to claim 3, wherein: in the 5), the microorganism enhancement also needs to satisfy the following conditions: the pure culture can be carried out under the conditions that the culture temperature is 5-35 ℃ and the culture medium is LB culture solution.
6. The method for rapidly inhibiting activated sludge non-filamentous bacteria bulking according to claim 5, wherein: in the 6), the fermentation culture conditions of the enhanced microorganisms are as follows: the culture temperature is the same as the temperature of the sewage in the target tank body, and the culture solution is a mixture of the sewage in the target tank body and the LB culture solution.
7. The method for rapidly inhibiting activated sludge non-filamentous bacteria bulking according to claim 6, wherein: and 6), culturing until the microorganism grows logarithmically, and/or the OD value of the microorganism is 0.6-0.8, and finishing fermentation.
8. The method for rapidly inhibiting activated sludge non-filamentous bacteria bulking according to claim 6, wherein: the volume ratio of the adding amount of the fermentation liquor to the target tank body is 1: (5000-20000), and/or the volume ratio of the added sewage to the LB culture solution in the culture solution is (300-100): 1.
9. the method for rapidly inhibiting activated sludge non-filamentous bacteria bulking according to claim 3, wherein: and 3) analyzing by using a 16S rRNA gene sequencing technology to obtain the microbial community structure information of the activated sludge, and/or 4) adjusting the pH value of the mixed sample to 4.0-5.0 and then performing centrifugal separation treatment.
10. The method for rapidly inhibiting activated sludge non-filamentous bacteria bulking according to claim 5, wherein: after the fermentation liquor is added into the target tank body, the sludge reflux ratio of the target tank body needs to be increased to 1.2-1.5 times of the original reflux ratio.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010482120.5A CN111595975B (en) | 2020-05-29 | 2020-05-29 | Method for rapidly inhibiting activated sludge non-filamentous bacteria expansion |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010482120.5A CN111595975B (en) | 2020-05-29 | 2020-05-29 | Method for rapidly inhibiting activated sludge non-filamentous bacteria expansion |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN111595975A CN111595975A (en) | 2020-08-28 |
| CN111595975B true CN111595975B (en) | 2020-11-24 |
Family
ID=72182073
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010482120.5A Active CN111595975B (en) | 2020-05-29 | 2020-05-29 | Method for rapidly inhibiting activated sludge non-filamentous bacteria expansion |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111595975B (en) |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2001129583A (en) * | 1999-11-09 | 2001-05-15 | Nippon Haisui Giken Kk | Batch type sludge treating installation using both pressure floatation and oxygen addition for pre- treatment |
| CN101993146A (en) * | 2009-08-27 | 2011-03-30 | 中国科学院沈阳自动化研究所 | Sludge bulking predetermining method by activated sludge process in sewage treating process |
| CN103011419A (en) * | 2012-12-03 | 2013-04-03 | 北京工业大学 | Method for rapidly solving problem of filamentous bacteria sludge bulking through hunger treatment |
| CN105084552A (en) * | 2015-07-31 | 2015-11-25 | 北京师范大学 | Method for enhancing aerobiotic ammonia oxidizing bacterium gathering through N-acylated homoserine lactones |
| CN110117078A (en) * | 2019-04-03 | 2019-08-13 | 东北师范大学 | A kind of filamentous fungi property sludge bulking inhibitor and its application method |
| CN111003816A (en) * | 2019-12-31 | 2020-04-14 | 江苏南大华兴环保科技股份公司 | Biochemical tail water biological denitrification method for inhibiting non-filamentous bacterium expansion |
-
2020
- 2020-05-29 CN CN202010482120.5A patent/CN111595975B/en active Active
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2001129583A (en) * | 1999-11-09 | 2001-05-15 | Nippon Haisui Giken Kk | Batch type sludge treating installation using both pressure floatation and oxygen addition for pre- treatment |
| CN101993146A (en) * | 2009-08-27 | 2011-03-30 | 中国科学院沈阳自动化研究所 | Sludge bulking predetermining method by activated sludge process in sewage treating process |
| CN103011419A (en) * | 2012-12-03 | 2013-04-03 | 北京工业大学 | Method for rapidly solving problem of filamentous bacteria sludge bulking through hunger treatment |
| CN105084552A (en) * | 2015-07-31 | 2015-11-25 | 北京师范大学 | Method for enhancing aerobiotic ammonia oxidizing bacterium gathering through N-acylated homoserine lactones |
| CN110117078A (en) * | 2019-04-03 | 2019-08-13 | 东北师范大学 | A kind of filamentous fungi property sludge bulking inhibitor and its application method |
| CN111003816A (en) * | 2019-12-31 | 2020-04-14 | 江苏南大华兴环保科技股份公司 | Biochemical tail water biological denitrification method for inhibiting non-filamentous bacterium expansion |
Non-Patent Citations (2)
| Title |
|---|
| SBR法处理生活污水时非丝状菌污泥膨胀的发生与控制;陈滢 等;《环境科学学报》;20050131;第105-108页 * |
| The regulation of N-acyl-homoserine lactones (AHLs)-based quorum sensing on EPS secretion via ATP synthetic for the stability of aerobic granular sludge;Zhiming Zhang et al.;《Science of the Total Environment》;20190405;第83-91页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN111595975A (en) | 2020-08-28 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106755424B (en) | Escherichia coli ST131 strain detection primer, kit and detection method based on CRISPR | |
| CN103993007B (en) | The simple and easy method of a kind of high efficiency extraction DNA from pedotheque | |
| CN103361439B (en) | Method for integrated detection of nine beer-spoilage bacteria | |
| CN101717815B (en) | Legionnella rapid detecting and parting method | |
| CN110241240B (en) | Detection primer, kit and quantitative method for Candidatus Electron | |
| CN109778321B (en) | Database building method, kit and sequencing method for metagenome sequencing | |
| CN102329874B (en) | Real-time fluorescent quantitative PCR (polymerase chain reaction) detection method of ammonia-oxidizing bacteria in active sludge | |
| CN106929578A (en) | The evaluation method of planktonic bacteria group in a kind of Taihu Lake water body | |
| CN111595975B (en) | Method for rapidly inhibiting activated sludge non-filamentous bacteria expansion | |
| CN107447023B (en) | Method for detecting phosphorus accumulating bacteria in sewage treatment system by combining fluorescence in-situ hybridization with flow cytometry | |
| Seder-Colomina et al. | Moderate oxygen depletion as a factor favouring the filamentous growth of Sphaerotilus natans | |
| CN113528614B (en) | Plant phyllosphere surface microorganism metagenome detection method | |
| CN108486238B (en) | High-throughput sequencing-based bioaugmentation functional flora analysis method | |
| Li et al. | Dynamics of dewaterability and bacterial populations in activated sludge | |
| CN101768632B (en) | Method for detecting aspergillus by polymerase chain reaction | |
| CN102242206A (en) | Method for studying microflora diversity in fermentation process of white spirit | |
| CN110055343B (en) | Method for monitoring rapid-acting phosphorus load pollution of sediment by using specific primers | |
| CN103333883B (en) | A kind of high efficiency extraction is used for the method for the groundwater microbial DNA of pcr amplification | |
| CN111154836A (en) | Targeted nucleic acid capture and detection methods | |
| CN105132561A (en) | Quick detection method capable of simultaneously identifying three beer-spoilage bacteria megasphaera, pectinatus and pediococcus | |
| CN112813142B (en) | MicroRNA capture magnetic bead, preparation method and detection method of MicroRNA | |
| AU2021103963A4 (en) | Paper-based chromatographic viral nucleic acid extraction kit | |
| CN113444826B (en) | Primer, probe and kit for identifying micrococcus based on isothermal amplification technology and application of primer, probe and kit | |
| CN116694807A (en) | Kit and method for detecting wheat blast bacteria based on RPA-lateral flow chromatography | |
| CN101168768A (en) | Alexandrium constant temperature gene amplification fast detecting kit and method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |