CN111269860A - Microbial strain for degrading kitchen waste and application thereof - Google Patents
Microbial strain for degrading kitchen waste and application thereof Download PDFInfo
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- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
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- Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
- Y02W30/00—Technologies for solid waste management
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Abstract
The invention belongs to the technical field of environmental protection biology, and particularly relates to a microbial strain for degrading kitchen waste and application thereof. The strain is named as Geobacillus thermocatenulatus GT-C1, and the preservation number is CCTCC M2019814. The strain GT-C1 can efficiently and quickly degrade various organic matters in the kitchen waste under the high-temperature condition, can be used for quickly reducing the kitchen waste, and realizes quick biological reduction and stabilization of the kitchen waste. The strain has obvious aerobic biological treatment effect on the kitchen waste, can obviously reduce the treatment cost, and has higher application value in the field of kitchen waste treatment.
Description
Technical Field
The invention relates to the technical field of environmental protection biology, in particular to a microbial strain for degrading kitchen waste and application thereof.
Background
The catering industry creates great benefits for the economy of China and simultaneously generates a large amount of kitchen waste, thereby bringing great pressure to the living environment. The kitchen waste mainly comprises three parts: the food processing leftovers (kitchen waste), the food residues (swill) and the farmer market residues (vegetable market) have complex components and are the mixture of various substances such as water, oil, vegetable peels, rice flour, meat, bone residues, plastic tableware, glass, paper towels and the like. After impurities are removed, the materials are classified according to chemical components and can be classified into four major categories of starch, cellulose, protein and fat. Because of different sources, the proportions of the components are greatly different, and the common points are high water content, easy decay, rich nutrient elements and less harmful substances. The conventional treatment methods for kitchen waste include incineration, landfill, feed treatment and biological treatment. The first three methods have many disadvantages, and the kitchen waste can not only reduce the heat value of the waste when burning, but also can generate substances such as dioxin and the like due to insufficient burning, so that the leaching rate of heavy metals in fly ash is increased, and secondary pollution is caused to the environment; if the kitchen waste and other household waste are landfilled hygienically, leachate generated by landfilling contains a large amount of high-concentration organic pollutants, so that secondary pollution is caused, and a large amount of organic matters, nitrogen, phosphorus, potassium and other nutrient elements in the kitchen waste are wasted; the kitchen waste is delivered to suburban farms as livestock feed, is usually deteriorated quickly, is easy to breed bacteria and mosquitoes, is harmful to the health of livestock and poultry, and can also indirectly harm the health of human beings by accumulating toxic substances in the livestock and poultry bodies to cause cross infection among people and livestock. The kitchen waste is treated by using the biotechnology, so that the secondary pollution is avoided, the kitchen waste can be effectively recycled, and the treatment rate can reach over 80 percent. Therefore, the treatment of the kitchen waste by utilizing the microbial technology becomes a main treatment method.
The microbial treatment of the kitchen waste is a treatment method for degrading organic components in the waste into simple inorganic matters by utilizing the growth and metabolism of microorganisms, and has the advantages of high efficiency, environmental protection and energy conservation. The microbial kitchen waste treatment method mainly comprises three methods, namely anaerobic digestion, aerobic composting and aerobic decrement. At present, the treatment target of the kitchen waste is mainly reduction and resource. Wherein, the aerobic decrement method has convenient use and high garbage decrement rate, and is very suitable for centralized treatment of a large amount of urban kitchen garbage. The core technology of aerobic decrement is the screening and application of microbial strains with special environment adaptation and special degradation functions, so that suitable high-efficiency microbial strains are searched according to the characteristics of the dietary habits in China and are imperatively applied to the treatment of kitchen waste.
Disclosure of Invention
In view of the above technical problems, a first object of the present invention is to provide an aerobic biological treatment strain for kitchen waste, so as to achieve rapid biological reduction and stabilization of kitchen waste.
In order to achieve the purpose, the strain GT-C1 for efficiently degrading the kitchen waste is identified as Geobacillus thermocatenulatus (Geobacillus thermocatenulatus), is preserved in the China Center for Type Culture Collection (CCTCC), has the preservation date of 2019, 10 and 12 days, and has the preservation number of CCTCC M2019814.
The Geobacillus thermocatenulatus GT-C1 contains a gene sequence shown as SEQID NO. 1.
The Geobacillus thermocatenulatus GT-C1 is a gram-negative bacterium and is short rod-shaped, the colony form is milky white and round, the edge is neat, and the surface is smooth and moist.
The second purpose of the invention is to provide a preparation method of a Geobacillus thermocatenulatus GT-C1 liquid microbial inoculum, which is characterized in that the Geobacillus thermocatenulatus GT-C1 pure strain is inoculated in a liquid culture medium to be cultured for 12-20 h to obtain the liquid microbial inoculum.
As a further proposal, the concentration of Geobacillus herminiophilus GT-C1 in the liquid microbial inoculum is at least 1 x 107cfu/mL。
The third purpose of the invention is to provide a preparation method of a solid microbial inoculum of Geobacillus thermocatenulatus GT-C1, which is characterized in that the solid microbial inoculum is prepared by mixing and drying a liquid microbial inoculum of Geobacillus thermocatenulatus GT-C1 with carriers such as wood dust and wheat bran, wherein the mass ratio of the carriers to the liquid microbial inoculum is 1: 1-10.
The fourth purpose of the invention is to disclose the application of geobacillus caldolyticus GT-C1 in the treatment of kitchen waste.
As a further scheme, the kitchen waste comprises perishable organic waste such as food and beverage waste, kitchen waste, vegetable yard waste and the like.
As a further scheme, the application comprises the following steps: (1) sorting the kitchen waste to be treated, and removing impurities such as plastics, bones, chopsticks and the like; (2) adding the sorted kitchen waste into the geobacillus thermocellus solid microbial inoculum, wherein the mass ratio of the solid microbial inoculum to the kitchen waste is 1: 1-5; (3) the mixed materials are uniformly stirred, aerobic fermentation is carried out at the temperature of 30-60 ℃, and continuous ventilation stirring is needed in the process so as to ensure that the materials are uniformly heated and are carried out under aerobic conditions.
Compared with the prior art, the invention has the following beneficial effects:
1. the geobacillus thermocatenulatus disclosed by the invention is wide in application temperature range, and can be used for quickly degrading kitchen waste at the temperature of 30-60 ℃;
2. the geobacillus thermocatenulatus disclosed by the invention has the capability of producing protease, amylase and lipase at high yield under a high temperature condition (50 ℃), can efficiently degrade organic matters such as protein, starch, grease and the like in kitchen waste, and has a kitchen waste reduction rate of 86% at a temperature of 50 ℃.
By combining the two characteristics, the Geobacillus thermocatenulatus GT-C1 can be applied to kitchen waste treatment, so that the rapid and efficient biological reduction and stabilization of the kitchen waste can be realized.
Drawings
FIG. 1 is a phylogenetic tree of the strain GT-C1 of the present invention;
FIG. 2 shows the wet weight loss rate of the kitchen waste by the strain GT-C1 of the present invention;
FIG. 3 shows the dry weight reduction rate of the kitchen waste by the strain GT-C1 of the present invention;
FIG. 4 is the wet weight loss rate of the inventive strain GT-C1 on mixed perishable waste.
Detailed description of the preferred embodiments
The invention is further illustrated with reference to the following figures and examples.
The strain capable of efficiently degrading the kitchen waste is obtained by primary screening of a separation and purification culture medium, secondary screening of an enzyme production culture medium and experimental verification of high-temperature aerobic biological treatment of the kitchen waste, and is named as GT-C1. The taxonomic status of the strain is identified by comprehensively analyzing data such as colony and cell morphology, physiological and biochemical characteristics, 16S rRNA gene sequence determination and the like: bacillus thermocatenulatus (Geobacillus thermocatenulatus). The strain is preserved in China Center for Type Culture Collection (CCTCC), the preservation date is 10 months and 12 days in 2019, the preservation number is CCTCC M2019814, and the preservation address is China center for type culture Collection of Wuhan university in Bayilu in Wuhan city, Hubei province.
The experiment result of the GT-C1 strain for degrading the kitchen waste shows that the strain can improve the decrement rate of the kitchen waste under the aerobic condition, achieves the purpose of kitchen waste decrement and stabilization, can be used for degrading perishable organic wastes such as the kitchen waste, vegetable yard waste and the like, and has high application value.
The following are specific examples of the present invention, in which the following media are used:
separating and purifying the culture medium: 10g/L of peptone and 5g/L, NaCl 10g/L, MgCl of yeast extract20.2 g/L、CaCl20.2 g/L、FeSO4·7H20.01g/L of O and 15g/L of agar.
Liquid culture medium: 10g/L of peptone and 5g/L, NaCl 10g/L, MgCl of yeast extract20.2 g/L、CaCl20.2g/L、FeSO4·7H2O 0.01g/L。
Protease screening medium: casein 10g/L, peptone 5g/L, yeast extract 2.5g/L, NaCl5g/L, MgCl g20.2 g/L、CaCl20.2 g/L、FeSO4·7H20.01g/L of O and 15g/L of agar.
Amylase screening culture medium: 10g/L of soluble starch, 5g/L of peptone and 2.5g/L, NaCl5g/L, MgCl of yeast extract20.2 g/L、CaCl20.2 g/L、FeSO4·7H20.01g/L of O and 15g/L of agar.
Lipase screening culture medium: emulsified olive oil 10mL/L, peptone 5g/L, yeast extract 2.5g/L, NaCl5g/L, MgCl20.2 g/L、CaCl20.2 g/L、FeSO4·7H20.01g/L of O, 20g/L of agar and the pH value is adjusted to 7-7.5.
Example 1: separation screening and performance determination of strain GT-C1
Primary screening: the test sample is collected from an aerobic digestion reactor of a kitchen waste treatment plant, a proper amount of sample is added with sterile water and mixed uniformly, gradient dilution is carried out, 100 mu L of diluent with different gradients is taken and coated on a separation and purification culture medium plate, after culture is carried out in a constant temperature incubator at 50 ℃ for 24h, single bacterial colonies with different shapes and sizes are selected, the single bacterial colonies are subjected to scribing purification and numbering preservation, and 9 bacterial strains are obtained through primary screening.
Re-screening: inoculating the pure strains obtained by primary screening to a protease, amylase and lipase screening culture medium by adopting a point grafting method, and measuring the enzyme production capacity of the strains so as to perform secondary screening. And (3) culturing the point-connected protease, amylase and lipase screening plate at 50 ℃ for 48-72 h, measuring the diameter (D) of each bacterial colony and the diameter (D) of a transparent ring, calculating D/D, and selecting strains with strong protein and fat degradation capability at the same time to obtain 2 re-screened strains.
Preliminary verification: respectively inoculating 2 strains obtained by re-screening into 100mL of liquid culture medium, culturing for 12h at 37 ℃ and 200r/min, taking 50mL of cultured bacterial liquid, washing with sterile normal saline after centrifugation, suspending to 20mL, attaching to 60g of sawdust, inoculating the bacterial liquid into 120g of kitchen waste, uniformly mixing, and continuously ventilating at 50 ℃ for preliminary verification, wherein the result shows that the inoculated strain GT-C1 has the maximum wet weight reduction rate on the kitchen waste and reaches 86% at 48 h.
The main biological characteristics of the strain GT-C1 are gram-negative bacteria and short rod-shaped, the size of a bacterial colony cultured on a solid plate for 16 hours is 4-5 mm, and the bacterial colony is milky white and round, has neat edges and smooth and moist surface. The 16S rDNA sequence of the strain GT-C1 is shown as SEQ ID NO.1, the sequence length is 1437bp, the 16S rRNA sequence is submitted in NCBI, the homology sequence comparison analysis is carried out by software and GenBank, and MEGA 7 software is used for constructing a phylogenetic tree of the strain (figure 1). Based on the above information, the strain GT-C1 was identified as Geobacillus thermocatenulatus (Geobacillus thermophilus).
Example 2: preparation of microbial inoculum of strain GT-C1
Preparing a liquid microbial inoculum: inoculating GT-C1 pure bacteria into 100mL of liquid culture medium, culturing for 12-20 h in a constant temperature shaking table at 37 ℃ and 200r/min, then inoculating the GT-C1 pure bacteria into the same liquid culture medium by 1-5% of inoculum size, and carrying out amplification culture under the same culture condition to obtain the liquid microbial inoculum of the strain.
Preparing a solid microbial inoculum: and mixing and drying the liquid microbial inoculum of the strain with carriers such as sawdust and wheat bran to prepare the solid microbial inoculum, wherein the mass ratio of the carriers to the liquid microbial inoculum is 1: 1-10.
Example 3: treatment effect of strain GT-C1 on kitchen waste
Sorting the kitchen waste to be treated, removing impurities such as plastics, bones, chopsticks and the like, adding a geobacillus thermocatenulatus solid microbial inoculum into the sorted kitchen waste, wherein the mass ratio of the solid microbial inoculum to the kitchen waste is 1: 3, carrying out aerobic treatment at 55 ℃ after uniformly stirring, wherein uninterrupted ventilation and stirring are needed in the process to ensure that the mixed material is uniformly heated and is carried out under aerobic conditions. Weighing the treatment system every 12h to obtain the decrement rate of the kitchen waste, wherein the decrement effect is shown in figures 2 and 3. As can be seen from FIG. 2, the experimental group inoculated with the strain GT-C1 always showed higher reduction rate than the control group without inoculated strain, the wet weight reduction rate of the inoculated strain reached 83.2% at 48h, which was 22.8% higher than 67.6% of the control group at 48 h. At the end of the experiment, the wet weight loss rate of the 72h experimental group reached 84.5%. Further, as can be seen from the dry weight reduction rate in fig. 3, the experiment group inoculated with geobacillus caldus almost reached the highest value of 43.5% even in 48 hours, and the control group reached 27.3% dry weight reduction rate. Therefore, the bacillus thermocatenulatus GT-C1 is inoculated, so that the reduction effect of the kitchen waste can be obviously improved, and the rapid biological reduction and stabilization of the kitchen waste are realized.
Example 4: treatment effect of strain GT-C1 on mixed perishable organic garbage
Mixing three kinds of perishable organic garbage of kitchen garbage, vegetable field garbage and kitchen garbage to be treated according to the ratio of 1:1:1, sorting, and removing impurities such as plastics, bones, chopsticks and the like. Adding a bacillus tenuissima solid microbial inoculum into the mixed perishable organic waste, wherein the mass ratio of the solid microbial inoculum to the kitchen waste is 1: 2, after being uniformly stirred, aerobic fermentation is carried out at 60 ℃, and continuous ventilation and stirring are needed in the process so as to ensure that the mixed materials are uniformly heated and are carried out under aerobic conditions. Weighing the treatment system every 12h to obtain the decrement rate of the kitchen waste, wherein the decrement effect is shown in figure 4. The results show that the wet weight loss of litter in the experimental group inoculated with the strain GT-C1 at 48h was as high as 90.2% and was already close to the maximum value after 24h, whereas the wet weight loss of the control group inoculated with no strain was as high as 81.1% and the strain GT-C1 increased the wet weight loss of litter by 11.2% compared to the control group. Therefore, the bacillus thermocatenulatus can improve the decrement rate of the kitchen waste under the aerobic condition, can be used for degrading mixed perishable organic waste such as the kitchen waste, vegetable yard waste and the like, and has high application value and wide application prospect.
Sequence listing
<110> Senno technologies, Inc., Senno technologies, Shandong Yushang energy-saving and environmental-protection technologies, Inc
<120> microbial strain for degrading kitchen waste and application cggctataatgcagtcgagcggaccggattggggcttgctcttgtttggtcagcggcggacgggtgagtaacacgtgggcaacctgcccgcaagaccgggataactccgggaaaccggagctaataccggataacaccgaagaccgcatggtcttcggttgaaaggcggcctttgggctgtcacttgcggatgggcccgcggcgcattagctagttggtgaggtaacggctcaccaaggcgacgatgcgtagccggcctgagagggtgaccggccacactgggactgagacacggcccagactcctacgggaggcagcagtagggaatcttccgcaatgggcgaaagcctgacggagcgacgccgcgtgagcgaagaaggccttcgggtcgtaaagctctgttgtgagggacgaaggagcgccgttcgaagagggcggcgcggtgacggtacctcacgagaaagccccggctaactacgtgccagcagccgcggtaatacgtagggggcgagcgttgtccggaattattgggcgtaaagcgcgcgcaggcggtctcttaagtctgatgtgaaagcccacggctcaaccgtggagggtcattggaaactgggggacttgagggcaggagaggagagcggaattccacgtgtagcggtgaaatgcgtagagatgtggaggaacaccagtggcgaaggcggctctctggcctgcacctgacgctgaggcgcgaaagcgtggggagcaaacaggattagataccctggtagtccacgccgtaaacgatgagtgctaagtgttagaggggtcacaccctttagtgctgcagctaacgcgataagcactccgcctggggagtacggccgcaaggctgaaactcaaaggaattgacgggggcccgcacaagcggtggagcatgtggtttaattcgaagcaacgcgaagaaccttaccaggtcttgacatcccctgacaacccaagagattgggcgttcccccttcggggggacagggtgacaggtggtgcatggttgtcgtcagctcgtgtcgtgagatgttgggttaagtcccgcaacgagcgcaaccctcgcctctagttgccagcattcagttgggcactctagagggactgccggcgacaagtcggaggaaggtggggatgacgtcaaatcatcatgccccttatgacctgggctacacacgtgctacaatgggcggtacaaagggctgcgaacccgcgagggggagcgaatcccaaaaagccgctctcagttcggattgcaggctgcaactcgcctgcatgaagccggaatcgctagtaatcgcggatcagcatgccgcggtgaatacgttcccgggccttgtacacaccgcccgtcacaccacgagagcttgcaacacccgaagtcggtgaggtaaccgcacggagccagccgccga thereof
<160>0
<170>SIPOSequenceListing 1.0
Claims (10)
1. Geobacillus thermocatenulatus GT-C1 is preserved in China Center for Type Culture Collection (CCTCC) with the preservation date of 2019, 10 months and 12 days and the preservation number of CCTCC M2019814.
2. The Geobacillus thermocatenulatus GT-C1 according to claim 1, wherein the Geobacillus thermocatenulatus GT-C1 has the gene sequence shown in SEQ ID NO. 1.
3. The Geobacillus thermocatenulatus GT-C1 according to claim 1, wherein the Geobacillus thermocatenulatus GT-C1 is a gram-negative bacterium, has a short rod shape, and has a milky white round colony shape, neat edges and a smooth and moist surface.
4. A preparation method of a Geobacillus thermocatenulatus GT-C1 liquid microbial inoculum is characterized in that the Geobacillus thermocatenulatus GT-C1 pure strain of any one of claims 1 to 3 is inoculated in a liquid culture medium to be cultured for 12 to 20 hours to obtain the liquid microbial inoculum.
5. The method for preparing the Geobacillus thermocatenulatus GT-C1 liquid microbial inoculum according to claim 4, wherein the liquid culture medium comprises the following components: 10g/L of peptone and 5g/L, NaCl 10g/L, MgCl of yeast extract20.2 g/L、CaCl20.2 g/L、FeSO4·7H2O 0.01g/L。
6. The method for preparing Geobacillus thermocatenulatus GT-C1 liquid bacterial agent according to claim 4, wherein the concentration of Geobacillus thermocatenulatus GT-C1 in the liquid bacterial agent is at least 1 x 107cfu/mL。
7. The preparation method of the Geobacillus thermocatenulatus GT-C1 solid microbial inoculum is characterized in that the solid microbial inoculum is prepared by mixing and drying a liquid microbial inoculum, sawdust and a rice hull carrier, wherein the mass ratio of the carrier to the liquid microbial inoculum is 1: 1-10.
8. The method for preparing the Geobacillus thermocatenulatus GT-C1 solid microbial inoculum of claim 7, wherein the method for preparing the liquid microbial inoculum according to any one of claims 4 to 6 comprises the following steps.
9. A method for using Geobacillus thermocatenulatus GT-C1 in the treatment of kitchen waste according to any one of claims 1 to 3, comprising the steps of:
(1) preparing a Geobacillus thermocatenulatus GT-C1 solid microbial inoculum;
(2) sorting the kitchen waste to be treated, and removing impurities such as plastics, bones and chopsticks;
(3) adding the sorted kitchen waste into a Geobacillus thermocatenulatus GT-C1 solid microbial inoculum, wherein the mass ratio of the solid microbial inoculum to the kitchen waste is 1: 1-5;
(4) the mixed materials are uniformly stirred, aerobic fermentation is carried out at the temperature of 30-60 ℃, and continuous ventilation stirring is needed in the process so as to ensure that the materials are uniformly heated and are carried out under aerobic conditions.
10. The use method according to claim 9, wherein the solid microbial inoculum is prepared according to the steps of the preparation method of the solid microbial inoculum according to any one of claims 7 to 8.
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Cited By (6)
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CN114437976A (en) * | 2022-01-28 | 2022-05-06 | 上海康鸿凯科技有限公司 | Compound microbial agent and application thereof in biological reduction of kitchen waste |
CN114525220A (en) * | 2022-01-28 | 2022-05-24 | 上海康鸿凯科技有限公司 | Biological reduction strain for kitchen waste and application thereof |
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CN114525220B (en) * | 2022-01-28 | 2023-09-19 | 上海康鸿凯科技有限公司 | Kitchen waste biological decrement strain and application thereof |
CN115786206A (en) * | 2022-12-02 | 2023-03-14 | 森诺技术有限公司 | Self-heat-production high-temperature aerobic strain and application thereof in biological treatment of kitchen waste |
CN115786206B (en) * | 2022-12-02 | 2023-12-19 | 森诺技术有限公司 | Self-heat-generating high-temperature aerobic strain and application thereof in biological treatment of kitchen waste |
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