CN115286118B - Method for treating sewage by using low-temperature-resistant denitrification and dephosphorization composite microbial agent - Google Patents
Method for treating sewage by using low-temperature-resistant denitrification and dephosphorization composite microbial agent Download PDFInfo
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Abstract
The invention discloses a method for treating sewage by using a low-temperature-resistant denitrification and dephosphorization composite microbial agent, and relates to a method for treating sewage by using a microbial agent. The nitrifying bacteria and the denitrifying bacteria in the low-temperature-resistant denitrification and dephosphorization composite microbial agent are matched with each other to remove total nitrogen in the sewage, and the phosphorus accumulating bacteria are used for removing total phosphorus in the sewage, so that the aims of synergistic interaction and synchronous denitrification and dephosphorization can be achieved, and the effect of removing nitrogen and phosphorus in the sewage is obviously higher than that of a single strain in a low-temperature environment. Each strain in the low-temperature-resistant denitrification and dephosphorization composite microbial agent is separated from winter sludge and river sediment in northern cold areas, and is an environment-friendly microorganism. When in use, the water is directly put in without complex engineering technology, is convenient to operate and has no secondary pollution, and has good application prospect.
Description
Technical Field
The invention relates to a method for treating sewage, in particular to a method for treating sewage by a low-temperature-resistant denitrification and dephosphorization composite microbial agent.
Background
Along with the rapid development of social economy and the continuous improvement of living standard, the continuous discharge of a large amount of urban sewage containing nitrogen and phosphorus pollutants causes excessive pollution load of water bodies, so that eutrophication and black and odorous phenomena of water bodies such as rivers, lakes and the like are caused, and serious threat is caused to ecological environment and human health. Therefore, the search for safe, efficient, simple and feasible sewage denitrification and dephosphorization technology becomes a research hotspot.
At present, the technology of denitrification and dephosphorization for sewage at home and abroad is mainly divided into a physical method (adsorption and electrodialysis), a chemical method (flocculation precipitation and ion exchange), a biological method and the like. Among them, biological methods often employ microbial agents to remove nitrogen and phosphorus. The microbial nitrogen and phosphorus removal technology has the advantages of simple operation, low running cost, good treatment effect, no secondary pollution and the like, and is the most widely applied method with the most development prospect in various sewage nitrogen and phosphorus removal technologies. However, the temperature of the urban sewage in winter in northern cold areas of China can be reduced to 8-15 ℃ or even lower than 5 ℃. Most microorganisms with nitrogen and phosphorus removal functions belong to mesophilic bacteria, the optimum growth temperature is 20-37 ℃, the treatment effect is good at 20-25 ℃ generally, the biological activity and the metabolic efficiency of the microorganisms are rapidly reduced in a low-temperature environment below 15 ℃, the nitrogen and phosphorus removal effect of the functional bacteria is seriously hindered, and the nitrogen and phosphorus removal effect on domestic sewage is poor in the low-temperature environment. Therefore, the development of the microbial agent which grows well under the low-temperature condition and can carry out high-efficiency denitrification and dephosphorization on sewage has very important social and economic benefits.
At present, researchers have proposed that low-temperature microbial agents can improve the problems of poor cell membrane fluidity, poor permeability, low enzyme activity and the like under low-temperature conditions, so that the effect of removing nitrogen and phosphorus in sewage under low-temperature environments in winter is improved. However, most of domestic related researches are still in a starting stage, most of the adopted single low-temperature strains are difficult to achieve the purification treatment effect of urban sewage, the development and construction forces of the low-temperature-resistant composite microbial agent are weak, and the method has few reports on successful application cases of water pollution treatment.
Disclosure of Invention
The invention aims to provide a method for treating sewage by using a low-temperature-resistant nitrogen and phosphorus removal compound microbial agent, which aims to solve the problems of low microbial activity and poor nitrogen and phosphorus removal effect of microorganisms on urban sewage under low-temperature conditions in winter.
The invention aims at realizing the following technical scheme:
a method for treating sewage by using a low-temperature-resistant denitrification and dephosphorization composite microbial agent, which comprises the following steps:
firstly, preparing a low-temperature-resistant denitrification and dephosphorization composite microbial agent, which comprises the following preparation steps:
(1) Preparing heterotrophic nitrification composite bacterial liquid: 3-4 loops of cold-resistant pseudomonas are selected from the inclined solid culture medium by an inoculating loop and inoculated into the heterotrophic nitrifying bacteria culture medium, and the cold-resistant pseudomonas seed liquid (the viable count is 1 multiplied by 10) is obtained by shaking culture for 24-72 hours in a full-temperature shaking incubator with the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min 7 ～2×10 9 Colony count/ml); inoculating 3-4 loops of cold-resistant pseudomonas from the inclined solid culture medium to the heterotrophic nitrifier culture medium by using an inoculating loop, and carrying out shaking culture in a full-temperature shaking incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 24-72 hours to obtain the cold-resistant pseudomonas seed liquid (the viable count is 1 multiplied by 10) 7 ～2×10 9 Colony count/ml); the pseudomonas cold-resistant grass seed liquid and the pseudomonas cold-resistant mussel seed liquid are mixed according to the volume ratio of the inoculation amount of (1-10): (1-10), and simultaneously inoculating 5% -20% of the total inoculum size into a heterotrophic nitrifying bacteria culture medium, and carrying out shake culture in a full-temperature shake incubator at a temperature of 6-15 ℃ and a rotating speed of 120-170 rpm for 48-96 hours to obtain a heterotrophic nitrifying composite bacterial liquid; the total viable count of the heterotrophic nitrification composite bacterial liquid is 1 multiplied by 10 8 ～5×10 9 Colony count/ml;
(2) And (3) preparing an aerobic denitrification compound bacterial liquid: 3-4 loops of cold-resistant pseudomonas Virroniana are selected from the inclined plane solid culture medium by an inoculating loop to be inoculated to the aerobicShaking culture is carried out in a full-temperature shaking incubator with the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 24-72 hours in a denitrifying bacteria culture medium to obtain cold-resistant verona pseudomonad seed liquid (the viable count is 1 multiplied by 10) 7 ～2×10 9 Colony count/ml); 3-4 loops of fluorescent pseudomonas are selected from the inclined plane solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium, and the fluorescent pseudomonas seed liquid (the viable count is 1 multiplied by 10) is obtained by shaking culture for 24-72 hours in a full-temperature shaking incubator with the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min 7 ～2×10 9 Colony count/ml); the pseudomonas cold-resistant veronii seed solution and the pseudomonas fluorescens are mixed according to the volume ratio of the inoculum size of (1-10): (1-10), and the total inoculum size of 5-20% is inoculated into an aerobic denitrifying bacteria culture medium, and the culture is carried out in a full-temperature shaking incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 48-96 hours, thus obtaining a well-cultured denitrifying composite bacterial liquid; the total viable count of the heterotrophic nitrification composite bacterial liquid is 1 multiplied by 10 8 ～5×10 9 Colony count/ml;
(3) Preparing phosphorus-accumulating composite bacterial liquid: inoculating 3-4 loops of Arthrobacter antarcticus from the inclined solid culture medium by using an inoculating loop, inoculating into a phosphorus accumulating bacteria culture medium, and carrying out shake culture in a full-temperature shake incubator at a temperature of 6-15 ℃ and a rotation speed of 120-170 rpm for 23-72 hours to obtain Arthrobacter antarcticus (the viable count is 1×10) 7 ～2×10 9 Colony count/ml). Inoculating 3-4 loops of salt crystal psychrophilic bacillus to the phosphorus accumulating bacteria culture medium with inoculating loop, and oscillating culturing in a full-temperature oscillating incubator at 6-15 deg.c and 120-170 rpm for 24-72 hr to obtain salt crystal psychrophilic bacillus seed liquid with viable count of 1×10 7 ～2×10 9 Colony count/ml). The seed solution of Antarctic arthrobacter and the seed solution of salt crystal psychrophilic bacillus are mixed according to the volume ratio of the inoculation amount of (1-10): (1-10), and the total inoculum size of 5-20% is inoculated into a phosphorus accumulating bacteria culture medium, and the phosphorus accumulating composite bacteria liquid is obtained by shake culture in a full-temperature shake incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 rpm for 48-96 hours; the total viable count of the phosphorus-accumulating composite bacterial liquid is 1 multiplied by 10 8 ～5×10 9 Colony count/ml;
(4) Low resistancePreparing a warm denitrification and dephosphorization compound microbial agent: the heterotrophic nitrification composite bacterial liquid, the aerobic denitrification composite bacterial liquid and the phosphorus-accumulating composite bacterial liquid are mixed according to the volume ratio of (1-10): (1-10): mixing the components (1-20) in proportion to prepare the low-temperature-resistant denitrification and dephosphorization composite microbial agent; the total viable count of the composite microbial agent is not less than 1×10 10 Colony count/ml;
the low-temperature-resistant denitrification and dephosphorization composite microbial agent is applied to low-temperature biological sewage treatment and comprises the following steps:
the additive amount in the urban sewage to be treated is 2 multiplied by 10 2 ～5×10 2 The low temperature resistant nitrogen and phosphorus removal compound microbial agent with colony count/square centimeter is aerated for 24 to 48 hours with aeration quantity of 1.0 to 3.5 cubic meters/hour, and is filtered to obtain the water after standing for 20 to 30 minutes.
The method for treating sewage by the low-temperature-resistant denitrification and dephosphorization composite microbial agent comprises the steps of preparing heterotrophic nitrification composite bacterial liquid, wherein cold-resistant pseudomonas and cold-resistant mush pseudomonas are both sewage treatment plant sedimentation tank activated sludge, and respectively identifying the cold-resistant pseudomonas and cold-resistant mush pseudomonas as pseudomonas grass (p.) by 16SrDNA sequence analysisPseudomonas poae) And Pseudomonas mussaendaPseudomonas peli)。
The method for treating sewage by using the low-temperature-resistant denitrification and dephosphorization composite microbial agent comprises the steps of preparing heterotrophic nitrification composite bacterial liquid and preparing a heterotrophic nitrifying bacterial culture medium: 1.0 to 5.0 g/liter of sodium citrate, 0.2 to 1.0 g/liter of ammonium sulfate, 0.3 to 1.5 g/liter of monopotassium phosphate, 0.1 to 1.0 g/liter of magnesium sulfate heptahydrate, 0.1 to 1.0 g/liter of sodium chloride, 0.01 to 0.1 g/liter of ferrous sulfate heptahydrate, and 0.01 to 0.1 g/liter of anhydrous calcium chloride. The pH of the heterotrophic nitrifier culture medium is adjusted to 6.5-7.5 by adopting 10% HCl and NaOH.
The method for treating sewage by the low-temperature-resistant denitrification and dephosphorization composite microbial agent comprises the steps of preparing heterotrophic nitrification composite bacterial liquid, wherein cold-resistant pseudomonas veronii and fluorescent pseudomonas are both river water frozen sediment, and respectively identifying the cold-resistant pseudomonas veronii and the fluorescent pseudomonas veronii as veronii after 16SrDNA sequence analysisPseudomonas veronii) And Pseudomonas fluorescens [ ]Pseudomonas fluorescens)。
The method for treating sewage by the low-temperature-resistant denitrification and dephosphorization composite microbial agent comprises the steps of preparing heterotrophic nitrification composite bacterial liquid, and preparing an aerobic denitrification bacterial culture medium, wherein the aerobic denitrification bacterial culture medium comprises the following components: 1.0 to 5.0 g/l sodium citrate, 0.2 to 2.0 g/l sodium nitrate, 0.3 to 1.5 g/l monopotassium phosphate, 0.1 to 1.0 g/l magnesium sulfate heptahydrate, 0.1 to 1.0 g/l sodium chloride, 0.01 to 0.1 g/l ferrous sulfate heptahydrate, 0.01 to 0.1 g/l anhydrous calcium chloride; the pH of the aerobic denitrifying bacteria culture medium is adjusted to 6.5-7.5 by adopting 10% HCl and NaOH.
The method for treating sewage by the low-temperature-resistant denitrification and dephosphorization composite microbial agent adopts a QHZ-98A type full-temperature oscillation incubator for microbial liquid culture.
The method for treating sewage by using the low-temperature-resistant denitrification and dephosphorization composite microbial agent adopts an SN210C type vertical pressure steam sterilizer for high-pressure steam sterilization.
The method for treating sewage by the low-temperature-resistant denitrification and dephosphorization composite microbial agent comprises the steps of sterilizing the culture medium by high-pressure steam, wherein the treatment condition is that the temperature is 121 ℃, the pressure is 0.105 megapascal, and the treatment time is 20-30 minutes.
The invention has the following remarkable technical advantages:
1. the low-temperature-resistant nitrogen and phosphorus removal compound microbial agent can normally grow, metabolize and reproduce in a lower temperature environment (6-15 ℃), and effectively remove nitrogen and phosphorus from sewage.
2. The strain in the low-temperature-resistant denitrification and dephosphorization composite microbial agent is separated from winter sludge and river sediment in northern cold areas, and is an environment-friendly microorganism. When in use, the water is directly put in without complex engineering technology, is convenient to operate and has no secondary pollution, and has good application prospect.
3. The nitrifying bacteria and the denitrifying bacteria in the low-temperature-resistant denitrification and dephosphorization composite microbial agent are matched with each other to remove total nitrogen in sewage, and the phosphorus accumulating bacteria are used for removing total phosphorus in sewage, so that the purposes of synergistic interaction and synchronous denitrification and dephosphorization can be achieved, and the effect of removing nitrogen and phosphorus in sewage is obviously higher than that of a single strain in a low-temperature environment.
Drawings
FIG. 1 is an electron micrograph of Pseudomonas grass of the present invention;
FIG. 2 is a photograph of a Pseudomonas mushy of the present invention;
FIG. 3 is an electron micrograph of Pseudomonas velocina of the present invention;
FIG. 4 is an electron micrograph of Pseudomonas fluorescens of the present invention;
FIG. 5 is an electron micrograph of Arthrobacter antarcticus of the invention;
FIG. 6 is a photograph of a salt crystal of the invention in a cold-bacillus acidophilus electron microscope.
Detailed Description
The present invention will be described in detail with reference to the embodiments shown in the drawings.
The invention firstly prepares the low temperature resistant denitrification and dephosphorization composite microbial agent, the total viable count of the composite microbial agent is not less than 1 multiplied by 10 10 Colony count/ml, including low temperature resistant heterotrophic nitrifying complex bacteria, low temperature resistant aerobic denitrifying complex bacteria and low temperature resistant phosphorus accumulating complex bacteria. Wherein the low temperature resistant heterotrophic nitrifying composite bacteria comprise Pseudomonas grass (P.sp.) A.MPseudomonas poae) And Pseudomonas mussaendaPseudomonas peli) The total viable count of the low temperature resistant heterotrophic nitrification composite bacteria is 1 multiplied by 10 8 ～5×10 9 Colony count/ml; the low temperature resistant aerobic denitrifying composite bacteria comprise pseudomonas VirronaPseudomonas veronii) And Pseudomonas fluorescens [ ]Pseudomonas fluorescens) The total viable count of the low temperature resistant aerobic denitrification compound bacteria is 1 multiplied by 10 8 ～5×10 9 Colony count/ml; the low temperature resistant phosphorus accumulating composite bacteria include Arthrobacter antarcticusArthrobacter antarcticus) And salt crystal psychrophilic bacillusPsychrobacter cryohalolentis) The total viable count of the low temperature resistant phosphorus accumulating composite bacteria is 1 multiplied by 10 8 ～5×10 9 Colony count/ml.
Experiments prove that the compound microbial agent disclosed by the invention is reasonable in formula, does not have antagonism among various strains of pseudomonas herbicola, pseudomonas artridge, pseudomonas veronii, pseudomonas fluorescens, arthrobacter antarctica and psychrophilium halophilum, is synergistic after being compounded, has remarkable denitrification and dephosphorization effects on urban sewage under a low-temperature condition, and has good application prospects.
Pseudomonas grassPseudomonas poae) Pseudomonas mustinePseudomonas peli) Pseudomonas veronii (Virrona)Pseudomonas veronii) Pseudomonas fluorescens @Pseudomonas fluorescens) Botrytis antarctica (L.) HemslArthrobacter antarcticus) And salt crystal psychrophilic bacillusPsychrobacter cryohalolentis) The secondary sedimentation tank activated sludge and the winter river water frozen sediment of a sewage treatment plant in Shenyang city of Liaoning province are respectively separated to be identified as known existing strains, and all large strain preservation libraries are preserved.
The preparation method of the low-temperature-resistant denitrification and dephosphorization composite microbial agent comprises the following steps:
(1) Preparing heterotrophic nitrification composite bacterial liquid: 3-4 loops of cold-resistant pseudomonas are selected from the inclined solid culture medium by an inoculating loop and inoculated into the heterotrophic nitrifying bacteria culture medium, and the cold-resistant pseudomonas seed liquid (the viable count is 1 multiplied by 10) is obtained by shaking culture for 24-72 hours in a full-temperature shaking incubator with the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min 7 ～2×10 9 Colony count/ml). Inoculating 3-4 loops of cold-resistant pseudomonas from the inclined solid culture medium to the heterotrophic nitrifier culture medium by using an inoculating loop, and carrying out shaking culture in a full-temperature shaking incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 24-72 hours to obtain the cold-resistant pseudomonas seed liquid (the viable count is 1 multiplied by 10) 7 ～2×10 9 Colony count/ml). The pseudomonas cold-resistant grass seed liquid and the pseudomonas cold-resistant mussel seed liquid are mixed according to the volume ratio of the inoculation amount of (1-10): (1-10) and 5-20% of total inoculum size are inoculated into a heterotrophic nitrifying bacteria culture medium at the same time, and the heterotrophic nitrifying composite bacterial liquid is obtained by shake culture in a full-temperature shake incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 rpm for 48-96 hours. The total viable count of the heterotrophic nitrification composite bacterial liquid is 1 multiplied by 10 8 ～5×10 9 Colony count/ml.
Wherein, the cold-resistant pseudomonas and the cold-resistant pseudomonas musae are both separated from the secondary sedimentation tank of the sewage treatment plant in the south of Shenyang city of Liaoning province in the 1 st month of 2021The sexual sludge is respectively identified as pseudomonas grass by 16SrDNA sequence analysisPseudomonas poae) And Pseudomonas mussaendaPseudomonas peli)。
The heterotrophic nitrifying bacteria culture medium comprises the following components: 1.0 to 5.0 g/liter of sodium citrate, 0.2 to 1.0 g/liter of ammonium sulfate, 0.3 to 1.5 g/liter of monopotassium phosphate, 0.1 to 1.0 g/liter of magnesium sulfate heptahydrate, 0.1 to 1.0 g/liter of sodium chloride, 0.01 to 0.1 g/liter of ferrous sulfate heptahydrate, and 0.01 to 0.1 g/liter of anhydrous calcium chloride. The pH of the heterotrophic nitrifier culture medium is adjusted to 6.5-7.5 by adopting 10% HCl and NaOH.
(2) And (3) preparing an aerobic denitrification compound bacterial liquid: 3-4 loops of cold-resistant pseudomonas Vironoidis are selected from the inclined plane solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium to be subjected to shaking culture in a full-temperature shaking incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 24-72 hours, so as to obtain cold-resistant pseudomonas Vironoidis seed liquid (the viable count is 1 multiplied by 10) 7 ～2×10 9 Colony count/ml). 3-4 loops of fluorescent pseudomonas are selected from the inclined plane solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium, and the fluorescent pseudomonas seed liquid (the viable count is 1 multiplied by 10) is obtained by shaking culture for 24-72 hours in a full-temperature shaking incubator with the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min 7 ～2×10 9 Colony count/ml). The pseudomonas cold-resistant veronii seed solution and the pseudomonas fluorescens are mixed according to the volume ratio of the inoculum size of (1-10): (1-10), and the total inoculum size of 5-20% is inoculated into an aerobic denitrifying bacteria culture medium, and the culture is carried out in a full-temperature shaking incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 48-96 hours, thus obtaining the aerobic denitrifying composite bacterial liquid. The total viable count of the heterotrophic nitrification composite bacterial liquid is 1 multiplied by 10 8 ～5×10 9 Colony count/ml.
Wherein, cold-resistant pseudomonas and fluorescent pseudomonas are both separated from the frozen sediment of North canal river in Shenyang city of Liaoning in 1 month of 2021, and are respectively identified as pseudomonas veronii (V.E.) by 16SrDNA sequence analysisPseudomonas veronii) And Pseudomonas fluorescens [ ]Pseudomonas fluorescens)。
The aerobic denitrifying bacteria culture medium comprises the following components: 1.0 to 5.0 g/liter of sodium citrate, 0.2 to 2.0 g/liter of sodium nitrate, 0.3 to 1.5 g/liter of monopotassium phosphate, 0.1 to 1.0 g/liter of magnesium sulfate heptahydrate, 0.1 to 1.0 g/liter of sodium chloride, 0.01 to 0.1 g/liter of ferrous sulfate heptahydrate, and 0.01 to 0.1 g/liter of anhydrous calcium chloride. The pH of the aerobic denitrifying bacteria culture medium is adjusted to 6.5-7.5 by adopting 10% HCl and NaOH.
(3) Preparing phosphorus-accumulating composite bacterial liquid: inoculating 3-4 loops of Arthrobacter antarcticus from the inclined solid culture medium by using an inoculating loop, inoculating into a phosphorus accumulating bacteria culture medium, and carrying out shake culture in a full-temperature shake incubator at a temperature of 6-15 ℃ and a rotation speed of 120-170 rpm for 23-72 hours to obtain Arthrobacter antarcticus (the viable count is 1×10) 7 ～2×10 9 Colony count/ml). Inoculating 3-4 loops of salt crystal psychrophilic bacillus to the phosphorus accumulating bacteria culture medium with inoculating loop, and oscillating culturing in a full-temperature oscillating incubator at 6-15 deg.c and 120-170 rpm for 24-72 hr to obtain salt crystal psychrophilic bacillus seed liquid with viable count of 1×10 7 ～2×10 9 Colony count/ml). The seed solution of Antarctic arthrobacter and the seed solution of salt crystal psychrophilic bacillus are mixed according to the volume ratio of the inoculation amount of (1-10): (1-10) and 5-20% of total inoculum size are inoculated into a phosphorus accumulating bacteria culture medium, and the phosphorus accumulating composite bacteria liquid is obtained by shaking culture in a full-temperature shaking incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 rpm for 48-96 hours. The total viable count of the phosphorus-accumulating composite bacterial liquid is 1 multiplied by 10 8 ～5×10 9 Colony count/ml.
Wherein, the Antarctic Arthrobacter and the Salmonella typhimurium are both separated from the water frozen sediment of North canal of Shenyang city of Liaoning in 2021 month, and are respectively identified as Antarctic Arthrobacter by 16SrDNA sequence analysisArthrobacter antarcticus) And salt crystal psychrophilic bacillusPsychrobacter cryohalolentis)。
The phosphorus accumulating bacteria culture medium comprises the following components: 1.0 to 5.0 g/liter of sodium acetate, 0.1 to 2.0 g/liter of ammonium sulfate, 0.01 to 1.0 g/liter of dipotassium hydrogen phosphate, 0.05 to 0.2 g/liter of magnesium sulfate heptahydrate, 0.01 to 0.1 g/liter of calcium chloride, and 7.0 to 9.0 g/liter of 4-hydroxyethyl piperazine ethane sulfonic acid. The pH of the polyphosphoric bacteria culture medium is adjusted to 6.5-7.5 by adopting 10% HCl and NaOH.
(4) Low temperature resistant strippingPreparing a nitrogen and phosphorus removal compound microbial agent: the heterotrophic nitrification composite bacterial liquid, the aerobic denitrification composite bacterial liquid and the phosphorus-accumulating composite bacterial liquid are mixed according to the volume ratio of (1-10): (1-10): mixing the components (1-20) in proportion to prepare the low-temperature-resistant denitrification and dephosphorization composite microbial agent. The total viable count of the composite microbial agent is not less than 1×10 10 Colony count/ml.
The invention relates to a low-temperature-resistant denitrification and dephosphorization composite microbial agent for treating sewage, which comprises the following steps:
the additive amount in the urban sewage to be treated is 2 multiplied by 10 2 ～5×10 2 The low temperature resistant nitrogen and phosphorus removal compound microbial agent with colony count/square centimeter is aerated for 24 to 48 hours with aeration quantity of 1.0 to 3.5 cubic meters/hour, and is filtered to obtain the water after standing for 20 to 30 minutes.
The technical scheme of the invention is further described by the following specific embodiments.
Example 1:
(1) Preparing heterotrophic nitrification composite bacterial liquid: 3 loops of cold-resistant pseudomonas are selected from the inclined solid culture medium by an inoculating loop and inoculated into a heterotrophic nitrification culture medium, and are subjected to shaking culture in a full-temperature shaking incubator at 6 ℃ and 150 revolutions per minute for 48 hours to obtain cold-resistant pseudomonas seed liquid (the viable count is 2 multiplied by 10) 7 Colony count/ml). 3 loops of cold-resistant pseudomonas mussel are selected from the inclined plane solid culture medium by an inoculating loop and inoculated into a heterotrophic nitrification culture medium, and are subjected to shaking culture in a full-temperature shaking incubator at 6 ℃ and 150 revolutions per minute for 48 hours, so as to obtain cold-resistant pseudomonas mussel seed liquid (the viable count is 3 multiplied by 10) 7 Colony count/ml). The method comprises the steps of (1) mixing a cold-resistant pseudomonas seed liquid and a cold-resistant mush pseudomonas seed liquid according to an inoculation amount volume ratio of 1: 2. simultaneously inoculating 15% of the total inoculum size into a heterotrophic nitrifying bacteria culture medium, and carrying out shaking culture in a full-temperature shaking incubator at 6 ℃ and 150 rpm for 96 hours to obtain a heterotrophic nitrifying composite bacterial liquid. The total viable count of the heterotrophic nitrification composite bacterial liquid is 1 multiplied by 10 8 Colony count/ml.
Wherein, the heterotrophic nitrifying bacteria culture medium comprises the following components: 2.5 g/l sodium citrate, 0.5 g/l ammonium sulphate, 0.3 g/l potassium dihydrogen phosphate, 0.1 g/l magnesium sulphate heptahydrate, 0.4 g/l sodium chloride, 0.04 g/l ferrous sulphate heptahydrate, 0.03 g/l anhydrous calcium chloride. The pH of the heterotrophic nitrifier medium was adjusted to 7.0 using 10% HCl and NaOH.
(2) And (3) preparing an aerobic denitrification compound bacterial liquid: 3 loops of cold-resistant pseudomonas Vena are selected from the inclined solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium, and the culture is carried out for 48 hours in a full-temperature shaking incubator at 6 ℃ and 150 revolutions per minute to obtain cold-resistant pseudomonas Vena seed liquid (the viable count is 4 multiplied by 10) 8 Colony count/ml). 3 loops of Pseudomonas fluorescens are selected from the inclined plane solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium, and the culture is carried out in a full-temperature shaking incubator at 6 ℃ and 150 revolutions per minute for 48 hours to obtain Pseudomonas fluorescens seed liquid (the viable count is 3 multiplied by 10) 7 Colony count/ml). The pseudomonas cold-resistant veronii seed solution and the pseudomonas fluorescens are mixed according to the inoculum size volume ratio of 2: 1. inoculating 15% of the total inoculum size into an aerobic denitrifying bacteria culture medium, and carrying out shaking culture in a full-temperature shaking incubator at 6 ℃ and 150 rpm for 96 hours to obtain the aerobic denitrifying composite bacterial liquid. The total viable count of the heterotrophic nitrification composite bacterial liquid is 3 multiplied by 10 8 Colony count/ml.
The aerobic denitrifying bacteria culture medium comprises the following components: 2.5 g/l sodium citrate, 1.5 g/l sodium nitrate, 0.3 g/l potassium dihydrogen phosphate, 0.1 g/l magnesium sulfate heptahydrate, 0.4 g/l sodium chloride, 0.04 g/l ferrous sulfate heptahydrate, 0.03 g/l anhydrous calcium chloride. The pH of the aerobic denitrifying bacteria culture was adjusted to 7.0 with 10% HCl and NaOH.
(3) Preparing phosphorus-accumulating composite bacterial liquid: 3-loop Antarctic bacteria are selected from the inclined solid culture medium by an inoculating loop, inoculated into a phosphorus accumulating bacteria culture medium, and subjected to shaking culture in a full-temperature shaking incubator at 6 ℃ and 150 revolutions per minute for 48 hours to obtain Antarctic bacteria (the viable count is 3 multiplied by 10) 7 Colony count/ml). Inoculating 3 loops of Salmonella typhimurium into phosphorus accumulating bacteria culture medium with inoculating loop, and shake culturing at 6deg.C in 150 rpm full-temperature shake incubator for 48 hr to obtain Salmonella typhimurium seed solution (viable count of 3×10) 7 Colony count/ml). Seed liquid and salt of Arthrobacter antarcticusThe volume ratio of the seed solution of the psychrophilic bacillus to the seed solution of the psychrophilic bacillus is 1: 1. 10% of the total inoculation amount is inoculated into a phosphorus accumulating bacteria culture medium, and the phosphorus accumulating composite bacteria liquid is obtained by shaking culture for 96 hours in a full-temperature shaking incubator at 6 ℃ and 150 revolutions per minute. The total viable count of the phosphorus-accumulating composite bacterial liquid is 2 multiplied by 10 8 Colony count/ml.
The phosphorus accumulating bacteria culture medium comprises the following components: 3.5 g/l sodium acetate, 0.8 g/l ammonium sulphate, 0.02 g/l dipotassium hydrogen phosphate, 0.1 g/l magnesium sulphate heptahydrate, 0.02 g/l calcium chloride, 8.5 g/l 4-hydroxyethyl piperazine ethane sulphonic acid. The pH of the polyphosphoric bacteria medium was adjusted to 7.0 with 10% HCl and NaOH.
(4) Preparation of low-temperature-resistant denitrification and dephosphorization composite microbial agent: the heterotrophic nitrification composite bacterial liquid, the aerobic denitrification composite bacterial liquid and the phosphorus-accumulating composite bacterial liquid are mixed according to the volume ratio of 3:3:5, mixing the materials in proportion to prepare the low-temperature-resistant denitrification and dephosphorization composite microbial agent. The total viable count of the composite microbial agent is 2 multiplied by 10 10 Colony count/ml.
Application of low-temperature-resistant denitrification and dephosphorization composite microbial agent in sewage: and adopting a low-temperature-resistant denitrification and dephosphorization composite microbial agent to carry out a domestic sewage treatment test under a low-temperature condition. Domestic sewage is taken from a sewage treatment center of a university, and water quality indexes are shown in table 1.
TABLE 1 domestic sewage Water quality index
The low temperature resistant denitrification and dephosphorization composite microbial agent, the pseudomonas grass, the pseudomonas musae, the pseudomonas veronii, the pseudomonas fluorescens, the Arthrobacter antarctica and the psychrophilic bacillus halofop described in the embodiment 1 are respectively inoculated into 1 liter of domestic sewage, and the total quantity of the composite microbial agent and the inoculation of single bacteria is the same (3 multiplied by 10 2 Colony count per square centimeter), aeration for 24 hours at 8 ℃ with aeration rate of 1.8 cubic meters per hour, standing for 20 minutes, filtering the effluent, measuring the mass concentration of ammonia nitrogen, nitrate nitrogen, total phosphorus and COD in the effluent, and adding no compound microbial agentThe domestic sewage is used as a control, and the removal rates of ammonia nitrogen, nitrate nitrogen, total phosphorus and COD are calculated, and the results are shown in Table 2.
TABLE 2 treatment effect of composite microbial inoculant and single strain on domestic sewage
The removal rates of the composite microbial agent described in the embodiment 1 on ammonia nitrogen, nitrate nitrogen, total phosphorus and COD in domestic sewage under the low-temperature condition are respectively 88.4%, 86.7%, 83.2%, 88.5% and 98.2%, and compared with the removal rate of a single strain, the removal rate of the composite microbial agent is obviously improved. Therefore, the compound microorganism strains formed after the compounding can synergistically promote the growth metabolism, and the removal efficiency of nitrogen and phosphorus in the sewage is effectively improved.
Example 2:
(1) Preparing heterotrophic nitrification composite bacterial liquid: picking 4 loops of cold-resistant pseudomonas from the inclined solid culture medium by using an inoculating loop, inoculating into a heterotrophic nitrifying bacteria culture medium, and performing shake culture in a full-temperature shake incubator at a temperature of 10 ℃ and a rotation speed of 160 revolutions per minute for 48 hours to obtain a cold-resistant pseudomonas seed solution (the viable count is 2 multiplied by 10) 8 Colony count/ml). The method comprises the steps of selecting 4 loops of cold-resistant pseudomonas from a slant solid culture medium by an inoculating loop, inoculating the cold-resistant pseudomonas into a heterotrophic nitrifying bacteria culture medium, and carrying out shaking culture in a full-temperature shaking incubator at a temperature of 10 ℃ and a rotating speed of 160 revolutions per minute for 48 hours to obtain cold-resistant pseudomonas seed liquid (the viable count is 4 multiplied by 10) 8 Colony count/ml). The pseudomonas cold-resistant grass seed liquid and the pseudomonas cold-resistant mud seed liquid are mixed according to the inoculation amount volume ratio of 2: 1. simultaneously inoculating 15% of the total inoculum size into a heterotrophic nitrifying bacteria culture medium, and carrying out shaking culture in a full-temperature shaking incubator at the temperature of 10 ℃ and the rotating speed of 160 revolutions per minute for 72 hours to obtain a heterotrophic nitrifying composite bacterial liquid. The total viable count of the heterotrophic nitrification composite bacterial liquid is 3 multiplied by 10 9 Colony count/ml.
Wherein, the heterotrophic nitrifying bacteria culture medium comprises the following components: 3.0 g/l sodium citrate, 0.8 g/l ammonium sulfate, 0.5 g/l potassium dihydrogen phosphate, 0.2 g/l magnesium sulfate heptahydrate, 0.1 g/l sodium chloride, 0.03 g/l ferrous sulfate heptahydrate, 0.02 g/l anhydrous calcium chloride. The pH of the heterotrophic nitrifier medium was adjusted to 7.0 using 10% HCl and NaOH.
(2) And (3) preparing an aerobic denitrification compound bacterial liquid: the 4-loop cold-resistant pseudomonas Veneti is selected from the inclined solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium, and is subjected to shaking culture in a full-temperature shaking incubator at the temperature of 10 ℃ and the rotating speed of 160 revolutions per minute for 48 hours to obtain cold-resistant pseudomonas Veneti seed liquid (the viable count is 2 multiplied by 10) 8 Colony count/ml). The 4-loop Pseudomonas fluorescens is selected from the inclined solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium, and is subjected to shaking culture in a full-temperature shaking incubator at the temperature of 10 ℃ and the rotating speed of 160 revolutions per minute for 48 hours to obtain a Pseudomonas fluorescens seed solution (the viable count is 3 multiplied by 10) 8 Colony count/ml). The pseudomonas cold-resistant veronii seed solution and the pseudomonas fluorescens are mixed according to the inoculum size volume ratio of 3: 2. inoculating 10% of the total inoculum size into an aerobic denitrifying bacteria culture medium, and carrying out shaking culture in a full-temperature shaking incubator at the temperature of 10 ℃ and the rotating speed of 160 revolutions per minute for 72 hours to obtain an aerobic denitrifying composite bacterial liquid. The total viable count of the aerobic denitrification compound bacterial liquid is 3 multiplied by 10 9 Colony count/ml.
The aerobic denitrifying bacteria culture medium comprises the following components: 3.0 g/l sodium citrate, 1.0 g/l sodium nitrate, 0.5 g/l potassium dihydrogen phosphate, 0.2 g/l magnesium sulfate heptahydrate, 0.1 g/l sodium chloride, 0.03 g/l ferrous sulfate heptahydrate, 0.02 g/l anhydrous calcium chloride. The pH of the aerobic denitrifying bacteria culture was adjusted to 7.0 with 10% HCl and NaOH.
(3) Preparing phosphorus-accumulating composite bacterial liquid: inoculating 4-loop Arthrobacter antarctica from slant solid culture medium with inoculating loop, shake culturing in full-temperature shake incubator at 10deg.C and 160 rpm for 48 hr to obtain Arthrobacter antarctica (viable count of 2×10) 8 Colony count/ml). Inoculating 4 loops of Salmonella typhimurium into phosphorus accumulating bacteria culture medium with inoculating loop, and shake culturing in a full-temperature shake incubator at 10deg.C and 160 rpm for 48 hr to obtain Salmonella typhimurium seed solution (viable count)Is 3X 10 8 Colony count/ml). The seed solution of the Antarctic arthrobacter and the seed solution of the salt crystal psychrophilic bacilli are mixed according to the volume ratio of the inoculation amount of 1: 1. 10% of the total inoculation amount is inoculated into a phosphorus accumulating bacteria culture medium, and the phosphorus accumulating composite bacteria liquid is obtained by shaking culture in a full-temperature shaking incubator at the temperature of 10 ℃ and the rotating speed of 160 revolutions per minute for 72 hours. The total viable count of the phosphorus-accumulating composite bacterial liquid is 2 multiplied by 10 9 Colony count/ml.
The phosphorus accumulating bacteria culture medium comprises the following components: 4.0 g/l sodium acetate, 1.0 g/l ammonium sulphate, 0.03 g/l dipotassium hydrogen phosphate, 0.05 g/l magnesium sulphate heptahydrate, 0.01 g/l calcium chloride, 7.5 g/l 4-hydroxyethyl piperazine ethane sulphonic acid. The pH of the polyphosphoric bacteria medium was adjusted to 7.0 with 10% HCl and NaOH.
(4) Preparation of low-temperature-resistant denitrification and dephosphorization composite microbial agent: the heterotrophic nitrification composite bacterial liquid, the aerobic denitrification composite bacterial liquid and the phosphorus-accumulating composite bacterial liquid are mixed according to the volume ratio of 5:5:12 to prepare the low temperature resistant denitrification and dephosphorization composite microbial inoculum. The total viable count of the composite microbial agent is 3×10 10 Colony count/ml.
Application of low-temperature-resistant denitrification and dephosphorization composite microbial inoculum in sewage: and adopting a low-temperature-resistant denitrification and dephosphorization composite microbial agent to carry out a domestic sewage treatment test under a low-temperature condition. Domestic sewage is taken from a sewage treatment center of a university, and water quality indexes are shown in table 3.
TABLE 3 domestic sewage Water quality index
The low temperature resistant denitrification and dephosphorization composite microbial agent, the pseudomonas grass, the pseudomonas musae, the pseudomonas veronii, the pseudomonas fluorescens, the Arthrobacter antarctica and the psychrophilic bacillus halofop described in the example 2 are respectively inoculated into 1 liter of domestic sewage, the total quantity of the composite microbial agent and the inoculation of single bacteria is the same (all are 5 multiplied by 10) 2 Colony count/square cm), aeration for 36 hours at 10 ℃ with aeration rate of 2.1 cubic meters/hour, standing for 20 minutes, filtering the water, and measuring ammonia nitrogen, nitrate nitrogen and total nitrogen in the waterThe mass concentration of total phosphorus and COD, and the removal rate of ammonia nitrogen, nitrate nitrogen, total phosphorus and COD were calculated by taking domestic sewage without adding the composite microbial agent as a control, and the results are shown in Table 4.
TABLE 4 treatment effect of composite microbial inoculant and single strain on domestic sewage
The removal rates of the composite microbial agent in the embodiment 2 on ammonia nitrogen, nitrate nitrogen, total phosphorus and COD in domestic sewage under the low-temperature condition are respectively 98.6%, 96.5%, 98.5%, 96.2% and 100%, and compared with the removal rate of a single strain, the removal rate of the composite microbial agent is obviously improved. Therefore, the compound microorganism strains formed after the compounding can synergistically promote the growth metabolism, and the removal efficiency of nitrogen and phosphorus in the sewage is effectively improved.
Example 3:
(1) Preparing heterotrophic nitrification composite bacterial liquid: picking 4 loops of cold-resistant pseudomonas from the inclined solid culture medium by using an inoculating loop, inoculating into a heterotrophic nitrifying bacteria culture medium, and performing shake culture in a full-temperature shake incubator at 15 ℃ and 140 revolutions per minute for 24 hours to obtain cold-resistant pseudomonas seed liquid (the viable count is 2×10) 9 Colony count/ml). The 4 loops of cold-resistant pseudomonas mussel are selected from the inclined plane solid culture medium by an inoculating loop and inoculated into the heterotrophic nitrifying bacteria culture medium, and are subjected to shaking culture in a full-temperature shaking incubator with the temperature of 15 ℃ and the rotating speed of 140 revolutions per minute for 24 hours, so as to obtain the cold-resistant pseudomonas mussel seed liquid (the viable count is 3 multiplied by 10) 9 Colony count/ml) the pseudomonas cold-resistant and the pseudomonas cold-resistant seed liquid are mixed according to the inoculum volume ratio of 1: 1. simultaneously inoculating 20% of the total inoculum size into a heterotrophic nitrifying bacteria culture medium, and carrying out shaking culture in a full-temperature shaking incubator at the temperature of 14 ℃ and the rotating speed of 140 revolutions per minute for 48 hours to obtain a heterotrophic nitrifying composite bacterial liquid. The total viable count of the heterotrophic nitrification composite bacterial liquid is 3 multiplied by 10 10 Colony count/ml.
The heterotrophic nitrifying bacteria culture medium comprises the following components: 3.5 g/l sodium citrate, 1.5 g/l ammonium sulfate, 0.5 g/l potassium dihydrogen phosphate, 0.3 g/l magnesium sulfate heptahydrate, 0.2 g/l sodium chloride, 0.05 g/l ferrous sulfate heptahydrate, 0.05 g/l anhydrous calcium chloride. The pH of the heterotrophic nitrifier medium was adjusted to 7.0 using 10% HCl and NaOH.
(2) And (3) preparing an aerobic denitrification compound bacterial liquid: the 4-loop cold-resistant pseudomonas Veneti is selected from the inclined solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium, and is subjected to shaking culture in a full-temperature shaking incubator with the temperature of 15 ℃ and the rotating speed of 140 revolutions per minute for 24 hours to obtain cold-resistant pseudomonas Veneti seed liquid (the viable count is 4 multiplied by 10) 9 Colony count/ml). The 4-loop Pseudomonas fluorescens is selected from the inclined solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium, and the culture is carried out for 24 hours in a full-temperature shaking incubator with the temperature of 15 ℃ and the rotating speed of 140 revolutions per minute, thus obtaining the Pseudomonas fluorescens seed liquid (the viable count is 2 multiplied by 10) 9 Colony count/ml). The pseudomonas cold-resistant veronii seed solution and the pseudomonas fluorescens are mixed according to the inoculum size volume ratio of 1: 1. inoculating 20% of the total inoculum size into an aerobic denitrifying bacteria culture medium, and carrying out shaking culture in a full-temperature shaking incubator at 15 ℃ and a rotating speed of 140 revolutions per minute for 48 hours to obtain an aerobic denitrifying composite bacterial liquid. The total viable count of the aerobic denitrification compound bacterial liquid is 4 multiplied by 10 10 Colony count/ml.
The aerobic denitrifying bacteria culture medium comprises the following components: 3.5 g/l sodium citrate, 1.0 g/l sodium nitrate, 0.5 g/l potassium dihydrogen phosphate, 0.3 g/l magnesium sulfate heptahydrate, 0.2 g/l sodium chloride, 0.05 g/l ferrous sulfate heptahydrate, 0.05 g/l anhydrous calcium chloride. The pH of the aerobic denitrifying bacteria culture was adjusted to 7.0 with 10% HCl and NaOH.
(3) Preparing phosphorus-accumulating composite bacterial liquid: inoculating 4-loop Antarctic bacteria to phosphorus accumulating bacteria culture medium, and shake culturing at 15deg.C and 140 rpm in a full-temperature shake incubator for 24 hr to obtain Antarctic bacteria (viable count of 2×10) 8 Colony count/ml). Inoculating 4-loop salt crystal psychrophilic bacillus to phosphorus accumulating bacteria culture medium at 10 deg.c and 140 rpm in full-temperature shaking incubatorCulturing for 24 hr to obtain seed solution of Salmonella typhimurium (viable count of 4×10) 8 Colony count/ml). The seed solution of the Antarctic arthrobacter and the seed solution of the salt crystal psychrophilic bacilli are mixed according to the volume ratio of the inoculation amount of 2: 1. inoculating 15% of the total inoculum size into a phosphorus accumulating bacteria culture medium, and carrying out shaking culture in a full-temperature shaking incubator at 15 ℃ and a rotating speed of 140 revolutions per minute for 48 hours to obtain phosphorus accumulating composite bacteria liquid. The total viable count of the phosphorus-accumulating composite bacterial liquid is 3 multiplied by 10 9 Colony count/ml.
The phosphorus accumulating bacteria culture medium comprises the following components: 4.5 g/l sodium acetate, 1.5 g/l ammonium sulphate, 1.0 g/l dipotassium hydrogen phosphate, 0.05 g/l magnesium sulphate heptahydrate, 0.03 g/l calcium chloride, 8.5 g/l 4-hydroxyethyl piperazine ethane sulphonic acid. The pH of the polyphosphoric bacteria medium was adjusted to 7.0 with 10% HCl and NaOH.
(4) Preparation of low-temperature-resistant denitrification and dephosphorization composite microbial agent: the heterotrophic nitrification composite bacterial liquid, the aerobic denitrification composite bacterial liquid and the phosphorus-accumulating composite bacterial liquid are mixed according to the volume ratio of 1:1:20 to prepare the low temperature resistant denitrification and dephosphorization composite microbial inoculum. The total viable count of the composite microbial agent is 3×10 10 Colony count/ml.
Application of low-temperature-resistant denitrification and dephosphorization composite microbial inoculum in sewage: and adopting a low-temperature-resistant denitrification and dephosphorization composite microbial agent to carry out a domestic sewage treatment test under a low-temperature condition. Domestic sewage is taken from a sewage treatment center of a university, and water quality indexes are shown in table 5.
TABLE 5 domestic sewage Water quality index
The low temperature resistant denitrification and dephosphorization composite microbial agent, the pseudomonas grass, the pseudomonas musae, the pseudomonas veronii, the pseudomonas fluorescens, the Arthrobacter antarctica and the psychrophilic bacillus halofop described in the embodiment 3 are respectively inoculated into 1 liter of domestic sewage, and the total quantity of the composite microbial agent and the inoculation of single bacteria is the same (4 multiplied by 10 2 Colony count/square cm), and aeration was carried out at 13℃for 24 hours at an aeration rate of 2.4 cubic meters/hour, followed by standing for 20 minutesThe effluent was filtered, the mass concentrations of ammonia nitrogen, nitrate nitrogen, total phosphorus and COD in the effluent were determined, and the removal rates of ammonia nitrogen, nitrate nitrogen, total phosphorus and COD were calculated by using domestic sewage without adding a composite microbial agent as a control, and the results are shown in Table 6.
Table 6 treatment effect of composite microbial agent and single strain on domestic sewage
The removal rates of the composite microbial agent in the embodiment 3 on ammonia nitrogen, nitrate nitrogen, total phosphorus and COD in domestic sewage under the low-temperature condition are respectively 99.1%, 98.8%, 99.0%, 97.3% and 100%, and compared with the removal rate of a single strain, the removal rate of the composite microbial agent is obviously improved. Therefore, the compound microorganism strains formed after the compounding can synergistically promote the growth metabolism, and the removal efficiency of nitrogen and phosphorus in the sewage is effectively improved.
Claims (8)
1. A method for treating sewage by using a low-temperature-resistant denitrification and dephosphorization composite microbial agent is characterized by comprising the following steps:
firstly, preparing a low-temperature-resistant denitrification and dephosphorization composite microbial agent, which comprises the following preparation steps:
(1) Preparing heterotrophic nitrification composite bacterial liquid: 3-4 loops of cold-resistant pseudomonas are selected from the inclined solid culture medium by an inoculating loop and inoculated into the heterotrophic nitrifying bacteria culture medium, and are subjected to shaking culture in a full-temperature shaking incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 24-72 hours to obtain the cold-resistant pseudomonas seed liquid, wherein the viable count is 1 multiplied by 10 7 ～2×10 9 Colony count/ml; inoculating 3-4 loops of cold-resistant pseudomonas from the inclined solid culture medium to the heterotrophic nitrifying bacteria culture medium by using an inoculating loop, and carrying out shaking culture in a full-temperature shaking incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 24-72 hours to obtain cold-resistant pseudomonas seed liquid with the viable count of 1X 10 7 ～2×10 9 Colony count/ml; mixing cold-resistant Pseudomonas putida seed liquid and cold-resistant mush pseudosheetThe volume ratio of the seed liquid of the cell strain is 1-10: 1-10 percent of total inoculum size is inoculated into a heterotrophic nitrifying bacteria culture medium at the same time, and the heterotrophic nitrifying composite bacterial liquid is obtained by shake culture in a full-temperature shake incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 48-96 hours; the total viable count of the heterotrophic nitrification composite bacterial liquid is 1 multiplied by 10 8 ～5×10 9 Colony count/ml;
(2) And (3) preparing an aerobic denitrification compound bacterial liquid: 3-4 loops of cold-resistant veroniona is selected from the inclined solid culture medium by an inoculating loop, inoculated into an aerobic denitrifying bacteria culture medium and subjected to shaking culture in a full-temperature shaking incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 24-72 hours, so as to obtain cold-resistant veroniona seed liquid, wherein the viable count is 1 multiplied by 10 7 ～2×10 9 Colony count/ml; 3-4 loops of fluorescent pseudomonas are selected from the inclined plane solid culture medium by an inoculating loop and inoculated into an aerobic denitrifying bacteria culture medium, and the fluorescent pseudomonas seed liquid is obtained by shaking culture for 24-72 hours in a full-temperature shaking incubator with the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min, and the viable count is 1 multiplied by 10 7 ～2×10 9 Colony count/ml; the pseudomonas cold-resistant veronii seed solution and the pseudomonas fluorescens are inoculated according to the volume ratio of 1-10: 1-10 percent of total inoculum size is inoculated into an aerobic denitrifying bacteria culture medium, and shake culture is carried out in a full-temperature shake incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 48-96 hours, thus obtaining a well-cultured denitrifying composite bacterial liquid; the total viable count of the aerobic denitrification compound bacterial liquid is 1 multiplied by 10 8 ～5×10 9 Colony count/ml;
preparing phosphorus-accumulating composite bacterial liquid: inoculating 3-4 loops of Antarctic Arthrobacter from the inclined solid culture medium into the phosphorus accumulating bacteria culture medium, and shake culturing in a full-temperature shake incubator at 6-15deg.C and 120-170 rpm for 23-72 hr to obtain Antarctic Arthrobacter with viable count of 1×10 7 ～2×10 9 Colony count/ml; inoculating 3-4 loops of salt crystal psychrophilic bacillus to the phosphorus accumulating bacteria culture medium with inoculating loop, and oscillating culturing in a full-temperature oscillating incubator at 6-15 deg.c and 120-170 rpm for 24-72 hr to obtain the productTo the seed liquid of the salt crystal psychrophilic bacillus, the viable count is 1 multiplied by 10 7 ～2×10 9 Colony count/ml; the seed solution of the Antarctic arthrobacter and the seed solution of the salt crystal psychrophilic bacilli are mixed according to the volume ratio of the inoculation amount of 1-10: 1-10 percent of total inoculum size is inoculated into a phosphorus accumulating bacteria culture medium, and the phosphorus accumulating composite bacteria liquid is obtained by shake culture in a full-temperature shake incubator at the temperature of 6-15 ℃ and the rotating speed of 120-170 r/min for 48-96 hours; the total viable count of the phosphorus-accumulating composite bacterial liquid is 1 multiplied by 10 8 ～5×10 9 Colony count/ml;
(3) Preparation of low-temperature-resistant denitrification and dephosphorization composite microbial agent: the heterotrophic nitrification composite bacterial liquid, the aerobic denitrification composite bacterial liquid and the phosphorus-accumulating composite bacterial liquid are mixed according to the volume ratio of 1-10: 1 to 10: 1-20, and preparing the low-temperature-resistant denitrification and dephosphorization composite microbial agent; the total viable count of the composite microbial agent is not less than 1×10 10 Colony count/ml;
the low-temperature-resistant denitrification and dephosphorization composite microbial agent is applied to low-temperature biological sewage treatment and comprises the following steps:
the additive amount in the urban sewage to be treated is 2 multiplied by 10 2 ～5×10 2 The low temperature resistant nitrogen and phosphorus removal compound microbial agent with colony count/square centimeter is aerated for 24 to 48 hours with aeration quantity of 1.0 to 3.5 cubic meters/hour, and is filtered to obtain the water after standing for 20 to 30 minutes.
2. The method for treating sewage by using the low-temperature-resistant denitrification and dephosphorization composite microbial agent as claimed in claim 1, wherein the heterotrophic nitrification composite microbial agent is prepared from cold-resistant pseudomonas and cold-resistant mush-resistant pseudomonas which are both sewage treatment plant sedimentation tank activated sludge, and are respectively identified as pseudomonas grass through 16SrDNA sequence analysisPseudomonas poae) And Pseudomonas mussaendaPseudomonas peli)。
3. The method for treating sewage by using the low-temperature-resistant denitrification and dephosphorization composite microbial agent according to claim 1, which is characterized in that the heterotrophic nitrification composite microbial liquid is prepared, and a heterotrophic nitrifying bacteria culture medium comprises the following components: 1.0 to 5.0 g/liter of sodium citrate, 0.2 to 1.0 g/liter of ammonium sulfate, 0.3 to 1.5 g/liter of monopotassium phosphate, 0.1 to 1.0 g/liter of magnesium sulfate heptahydrate, 0.1 to 1.0 g/liter of sodium chloride, 0.01 to 0.1 g/liter of ferrous sulfate heptahydrate, 0.01 to 0.1 g/liter of anhydrous calcium chloride; the pH of the heterotrophic nitrifier culture medium is adjusted to 6.5-7.5 by adopting 10% HCl and NaOH.
4. The method for treating sewage by using the low-temperature-resistant denitrification and dephosphorization composite microbial agent according to claim 1, which is characterized in that the heterotrophic nitrification composite microbial agent is prepared from cold-resistant pseudomonas and fluorescent pseudomonas which are both river water frozen sediment, and are respectively identified as pseudomonas veronii by 16SrDNA sequence analysisPseudomonas veronii) And Pseudomonas fluorescens [ ]Pseudomonas fluorescens)。
5. The method for treating sewage by using the low-temperature-resistant denitrification and dephosphorization composite microbial agent according to claim 1, which is characterized in that the preparation of the aerobic denitrification composite microbial agent is characterized in that an aerobic denitrification bacterial culture medium comprises the following components: 1.0 to 5.0 g/l sodium citrate, 0.2 to 2.0 g/l sodium nitrate, 0.3 to 1.5 g/l monopotassium phosphate, 0.1 to 1.0 g/l magnesium sulfate heptahydrate, 0.1 to 1.0 g/l sodium chloride, 0.01 to 0.1 g/l ferrous sulfate heptahydrate, 0.01 to 0.1 g/l anhydrous calcium chloride; the pH of the aerobic denitrifying bacteria culture medium is adjusted to 6.5-7.5 by adopting 10% HCl and NaOH.
6. The method for treating sewage by using the low-temperature-resistant denitrification and dephosphorization composite microbial agent according to claim 1, which is characterized in that a QHZ-98A type full-temperature shaking incubator is adopted for microbial liquid culture.
7. The method for treating sewage by using the low-temperature-resistant denitrification and dephosphorization composite microbial agent according to claim 1, which is characterized in that the heterotrophic nitrifying bacteria culture medium, the aerobic denitrifying bacteria culture medium and the phosphorus accumulating bacteria culture medium are subjected to high-pressure steam sterilization treatment under the treatment condition that the temperature is 121 ℃ and the pressure is 0.105 megapascal, and the treatment time is 20-30 minutes.
8. The method for treating sewage by using the low-temperature-resistant denitrification and dephosphorization composite microbial agent according to claim 7, wherein the high-pressure steam sterilization adopts an SN210C type vertical pressure steam sterilizer.
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| CN105296398A (en) * | 2015-11-12 | 2016-02-03 | 天津大学 | Low-temperature-resistant composite microbial agent used for treating contaminated water, and preparation method and application thereof |
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| US20170166466A1 (en) * | 2015-12-09 | 2017-06-15 | Microbial Discovery Group Llc | Cold temperature-resistant microbials and methods of their use |
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