WO2021244573A1 - 可降解聚对苯二甲酸乙二醇酯的嗜麦芽寡养单胞菌 - Google Patents

可降解聚对苯二甲酸乙二醇酯的嗜麦芽寡养单胞菌 Download PDF

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WO2021244573A1
WO2021244573A1 PCT/CN2021/097924 CN2021097924W WO2021244573A1 WO 2021244573 A1 WO2021244573 A1 WO 2021244573A1 CN 2021097924 W CN2021097924 W CN 2021097924W WO 2021244573 A1 WO2021244573 A1 WO 2021244573A1
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polyethylene terephthalate
stenotrophomonas
pet
jwg
product
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吴敬
颜正飞
黄青松
顾冷涛
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江南大学
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    • C08J11/00—Recovery or working-up of waste materials
    • C08J11/04—Recovery or working-up of waste materials of polymers
    • C08J11/10—Recovery or working-up of waste materials of polymers by chemically breaking down the molecular chains of polymers or breaking of crosslinks, e.g. devulcanisation
    • C08J11/105—Recovery or working-up of waste materials of polymers by chemically breaking down the molecular chains of polymers or breaking of crosslinks, e.g. devulcanisation by treatment with enzymes
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    • 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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    • C08J11/00—Recovery or working-up of waste materials
    • C08J11/04—Recovery or working-up of waste materials of polymers
    • C08J11/10—Recovery or working-up of waste materials of polymers by chemically breaking down the molecular chains of polymers or breaking of crosslinks, e.g. devulcanisation
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    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
    • C12P7/44—Polycarboxylic acids
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    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/62—Carboxylic acid esters
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2367/00—Characterised by the use of polyesters obtained by reactions forming a carboxylic ester link in the main chain; Derivatives of such polymers
    • C08J2367/02—Polyesters derived from dicarboxylic acids and dihydroxy compounds
    • C—CHEMISTRY; METALLURGY
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    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
    • Y02W30/00—Technologies for solid waste management
    • Y02W30/50—Reuse, recycling or recovery technologies
    • Y02W30/62—Plastics recycling; Rubber recycling

Definitions

  • the invention relates to a degradable polyethylene terephthalate-degradable Stenotrophomonas maltophilia, and belongs to the technical field of microorganisms.
  • PET Polyethylene terephthalate
  • EG ethylene glycol
  • TPA terephthalic acid
  • PET polyethylene terephthalate
  • PET plastic products account for about 60% of all plastic products.
  • PET polyethylene terephthalate
  • PET plastic waste also accounts for a relatively large proportion of all plastic waste. high. Therefore, the degradation of polyethylene terephthalate (PET) is very important for the treatment of plastic waste.
  • PET polyethylene terephthalate
  • the chemical degradation method requires the use of a large amount of chemicals, and a large amount of toxic and harmful substances are produced in the process, which has a serious negative impact on the ecological environment.
  • the physical degradation law requires high temperature and high pressure equipment, which greatly increases the treatment cost of polyethylene terephthalate (PET) plastic waste. Therefore, governments of various countries are actively exploring new technologies for degrading polyethylene terephthalate (PET).
  • Biodegradation technology is a technology that directly degrades plastics by strains that can degrade plastics. Because of its green, pollution-free and low cost advantages, it has gradually become a research hotspot in the field of plastic degradation.
  • the strain Ideonella sakaiensis 201-F6 can use polyethylene terephthalate (PET) as the only nutrient source, and the degradation rate is 0.13 mg/d (Science, 2016, 351(6278): 1196-1199); Paleococcus strains that can degrade polylactic acid (PLA) (publication number CN102639690A); Penicillium strains that can degrade polyhydroxyalkanoates (PHA) (Polymer-plastics Technology and Engineering, 2009, 48(1): 58 -63).
  • polyethylene terephthalate (PET) molecular chain contains a lot of aromatic groups , Resulting in a large steric hindrance of the molecular chain and a more hydrophobic surface, which is difficult to be degraded by microorganisms. Therefore, obtaining a degradable polyethylene terephthalate (PET) strain is still a difficult point.
  • PHA polyhydroxyalkanoate
  • PLA polylactic acid
  • PET polyethylene terephthalate
  • the technical problem to be solved by the present invention is to provide a stenotrophomonas pavanii which can degrade polyethylene terephthalate and its application.
  • the present invention provides a strain of Stenotrophomonas pavanii JWG-G1, which is deposited in the China Type Culture Collection with the deposit number CCTCC NO:M 2019415, and the preservation date is June 03, 2019.
  • the maltophilic oligotrophomonas (Stenotrophomonas pavanii) JWG-G1 was isolated from a soil sample derived from the Taohuashan landfill in Wuxi. The strain was sequenced and analyzed, and its 16S rDNA sequence is as SEQ ID NO.
  • the sequence obtained by sequencing is compared with the nucleic acid sequence in Genbank, and the result shows that the similarity to the nucleic acid sequence of Stenotrophomonas is as high as 99%, and the phylogenetic tree will be constructed with the strain with high similarity (see details Figure 1), the results show that the strain belongs to Stenotrophomonas maltophilia (Stenotrophomonas pavanii) JWG-G1.
  • the colony of the Stenotrophomonas pavanii JWG-G1 on the LB solid medium is round and convex, light yellow, opaque, moist and shiny, with flagella on the edges (see Figure 2- Figure 3).
  • the present invention also provides the application of the above-mentioned Stenotrophomonas pavanii JWG-G1 in the degradation of polyethylene terephthalate, gelatin or esculin.
  • the present invention also provides a method for degrading and degrading polyethylene terephthalate (PET).
  • PET polyethylene terephthalate
  • the seed liquid of JWG-G1 is inoculated into a liquid medium containing polyethylene terephthalate for cultivation.
  • the seed solution of the above-mentioned Stenotrophomonas pavanii JWG-G1 contains polyethylene terephthalate at an inoculum of not less than 10% (v/v). Inoculate the glycol ester in a liquid medium.
  • the concentration of the seed solution of Stenotrophomonas pavanii JWG-G1 in the culture medium is not less than 1 ⁇ 10 8 CFU/mL.
  • the medium is an inorganic salt medium.
  • the present invention also provides a product that can be used to degrade polyethylene terephthalate (PET), which contains the above-mentioned Stenotrophomonas maltophilia (Stenotrophomonas). pavanii) JWG-G1.
  • PET polyethylene terephthalate
  • the present invention also provides a method for hydrolyzing gelatin.
  • the method is to inoculate the above-mentioned Stenotrophomonas pavanii JWG-G1 into a plate medium containing gelatin for cultivation.
  • the present invention also provides a product that can be used to hydrolyze gelatin, and the product contains the above-mentioned Stenotrophomonas pavanii JWG-G1.
  • the present invention also provides a method for hydrolyzing esculin, which comprises inoculating the above-mentioned Stenotrophomonas pavanii JWG-G1 into a plate medium containing gelatin for cultivation.
  • the present invention also provides a product that can be used to hydrolyze esculin, which contains the above-mentioned Stenotrophomonas pavanii JWG-G1.
  • the present invention provides a strain of Stenotrophomonas pavanii JWG-G1 that can degrade polyethylene terephthalate (PET). pavanii)
  • the seed solution of JWG-G1 is inoculated into an inorganic salt liquid medium containing 2g/L polyethylene terephthalate (PET) at an inoculum of 10% (v/v).
  • PET polyethylene terephthalate
  • MHET hydroxyethyl terephthalate
  • TPA terephthalic acid
  • the ester bond functional groups on the surface of polyethylene terephthalate (PET) plastic particles can be reduced, and the polyethylene terephthalate (PET) plastic
  • PET polyethylene terephthalate
  • the weight loss rate of the particles reaches 9.4%. Therefore, the Stenotrophomonas pavanii JWG-G1 of the present invention has extremely high application prospects in the degradation of polyethylene terephthalate (PET).
  • the Stenotrophomonas pavanii JWG-G1 of the present invention has excellent salt tolerance and can grow vigorously in an LB liquid medium containing 1-9 g/L NaCl.
  • the Stenotrophomonas pavanii JWG-G1 of the present invention can hydrolyze gelatin and esculin.
  • Figure 1 Phylogenetic tree of Stenotrophomonas pavanii JWG-G1.
  • FIG. 1 The colony of Stenotrophomonas pavanii JWG-G1.
  • Figure 3 The cell morphology of Stenotrophomonas pavanii JWG-G1.
  • Figure 4 Growth curve of Stenotrophomonas pavanii JWG-G1 at different pH.
  • Figure 5 Growth curve of Stenotrophomonas pavanii JWG-G1 at different temperatures.
  • Figure 7 Gelatin hydrolysis efficiency of Stenotrophomonas pavanii JWG-G1 JWG-G1.
  • Figure 8 Hydrolysis efficiency of Stenotrophomonas pavanii JWG-G1JWG-G1 to esculin.
  • the dimethyl terephthalate (DET), polyethylene terephthalate (PET) plastic particles, and ethylene phthalate (BHET) involved in the following examples were purchased from Sigma, TPA, The standard products of MHET and BHET were purchased from Sigma Company.
  • LB solid medium g/L: peptone 10, yeast powder 5, sodium chloride 10, agar 13, pH 7.0.
  • LB liquid medium g/L: peptone 10, yeast powder 5, sodium chloride 10, pH 7.0.
  • Inorganic salt liquid medium (g/L): KH 2 PO 4 0.7, K 2 HPO 4 ⁇ 3H 2 O 0.5, NH 4 Cl 2, MgSO 4 ⁇ 7H 2 O 0.6, NaCl 0.005, FeSO 4 ⁇ 7H 2 O 0.001 , ZnSO 4 ⁇ 7H 2 O 0.002, MnSO 4 ⁇ H 2 O 0.001.
  • Inorganic salt solid medium (g/L): KH 2 PO 4 0.7, K 2 HPO 4 ⁇ 3H 2 O 0.5, NH 4 Cl 2, MgSO 4 ⁇ 7H 2 O 0.6, NaCl 0.005, FeSO 4 ⁇ 7H 2 O 0.001 , ZnSO 4 ⁇ 7H 2 O 0.002, MnSO 4 ⁇ H 2 O 0.001, agar powder 13.
  • PET polyethylene terephthalate
  • the polyethylene terephthalate (PET) plastic particles treated by the strains are repeatedly washed with deionized water for 3 to 4 times; the washed polyethylene terephthalate (PET) plastic particles are Ultrasound for 15 minutes under the condition of power 200W and frequency 58KHz; put the polyethylene terephthalate (PET) plastic particles after ultrasound into an oven at 60°C for 6h; use untreated polyethylene terephthalate Alcohol ester (PET) plastic particles are used as a control.
  • Fourier transform infrared spectrometer is used to detect the surface of untreated polyethylene terephthalate (PET) plastic particles and the polyethylene terephthalate after treatment with strains. Changes in the surface functional groups of ester (PET) plastic particles.
  • Standard treatment Weigh the standard products of TPA, MHET, and BHET respectively and dissolve them in dimethyl sulfoxide (DMSO) to make mother liquor. Use sterile water to dilute the mother liquor into a 0.1mg/mL standard solution, and filter with 0.22 ⁇ M. Head filtration, and inject the liquid phase bottle with a syringe for HPLC detection;
  • DMSO dimethyl sulfoxide
  • Sample processing Let the culture solution stand for 10 min, take 5 mL of the supernatant, centrifuge at 12000 rpm for 8 min, filter with a 0.22 ⁇ M filter, and inject the liquid phase bottle with a syringe for HPLC detection.
  • the PET plastic particles treated by the strains are repeatedly washed with deionized water for 3 to 4 times.
  • the cleaned PET plastic particles are sonicated for 15 minutes at a power of 200W and a frequency of 58KHz, and then placed in an oven at 60°C for 6 hours and weighed. Heavy;
  • the PET plastic pellets before the strain treatment are repeatedly washed with deionized water for 3 to 4 times.
  • the cleaned PET plastic pellets are sonicated for 15 minutes at a power of 200W and a frequency of 58KHz, and then placed in an oven at 60°C for 6h and weighed. Heavy.
  • D1 On the nth day, the diameter of the transparent circle on the detection plate (cm); n: the number of days of culture on the detection plate (day).
  • PET polyethylene terephthalic acid
  • DET diethyl terephthalate
  • BHET bishydroxyethyl terephthalate
  • PET polyethylene terephthalate
  • PET poly(ethylene terephthalate)
  • strain JWG-G1 The total DNA of strain JWG-G1 was extracted for 16S rDNA amplification and sequencing (completed by Wuxi Tianlin Biotechnology Co., Ltd.), and the sequence obtained by sequencing was compared in Genbank for nucleic acid sequence comparison.
  • the 16S rDNA sequence homology of the genus Monas is greater than 99%, and the similarity rate of the 16S rDNA sequence of Stenotrophomonas pavanii DSM 25135 is 99.5%. It can be seen that the strain JWG-G1 belongs to the genus Stenotrophomonas.
  • strain JWG-G1 The 16S rDNA sequence of strain JWG-G1 (sequence shown in SEQ ID NO.1) and the strain with high similarity were constructed to construct a phylogenetic tree (the phylogenetic tree constructed by strain JWG-G1 can be seen in Figure 1), and the results show that strain JWG -G1 and Stenotrophomonas pavanii DSM 25135 belong to the same branch. It can be seen that the strain JWG-G1 belongs to the genus Stenotrophomonas and is named Stenotrophomonas pavanii JWG-G1.
  • the Stenotrophomonas pavanii JWG-G1 obtained in Example 1 was gram-stained and observed under a microscope, and it was found to be a gram-positive bacteria.
  • Example 4 Degradability of different Stenotrophomonas and Stenotrophomonas pavanii to polyethylene terephthalate (PET) plastic particles
  • Stenotrophomonas pavanii JWG-G1 belongs to the genus Stenotrophomonas, the genus Stenotrophomonas may be one of the potential sources of PET plastic particle degrading strains, so the collection is related to the genus of Stenotrophomonas pavanii Stenotrophomonas pavanii (Stenotrophomonas pavanii) JWG-G1 closely related 7 strains of Stenotrophomonas pavanii DSM 25135; Stenotrophomonas bentonitica DSM 103927; Stenotrophomonas chelatiphaga DSM lactithomonas lactithomonas lachomophila DSM 21508; Stenotrophomonas lactrophonas 152; Stenotrophomonas pavanii DSM 25135; DSM 14405; Stenotrophomonas tumulicola NCIMB 15009) and Steno
  • the single colonies of Stenotrophomonas pavanii JWG-G1 and 7 strains of Stenotrophomonas pavanii obtained in Example 1 were selected, and they were respectively connected to 100 mL of LB liquid medium at 35°C and 180 rpm. Cultivate with shaking for 24 hours to obtain seed liquid A; transfer the seed liquid A to 100 mL of fresh LB liquid medium at an inoculum amount of 10% (v/v), and culture with shaking for 24 hours at 35°C and 180 rpm.
  • the inorganic salt liquid medium containing 2g/L polyethylene terephthalate (PET) was inoculated with seed liquid B as the control group, and seed liquid B was inoculated with an inoculum amount of 10% (v/v) to contain 2g /L Polyethylene terephthalate (PET) inorganic salt liquid medium (at this time, the bacterial concentration of seed solution B in the PET inorganic salt liquid medium is 1 ⁇ 10 9 CFU/mL), cultured with shaking at 35°C and 180rpm for 5 days to obtain culture medium B.
  • PET polyethylene terephthalate
  • the change of OD 600 before and after the measurement of culture medium B can be seen in Table 1.
  • Take out the PET plastic particles in culture medium B To detect the changes in the structure of the surface functional groups (the surface of the polyethylene terephthalate (PET) plastic particles in the culture solution B obtained from the cultivation of the maltophilic Stenotrophomonas JWG-G1 and 7 strains of Stenotrophomonas
  • the changes in the functional group structure of the ester bond can be seen in Table 1.
  • the changes in the surface functional group structure of the PET plastic particles in the culture solution B obtained from the culture of Stenotrophomonas maltophilia JWG-G1 can be seen in Figure 6) and the weight loss rate (maltophilic oligotrophic
  • the weight loss rate of the polyethylene terephthalate (PET) plastic particles in the culture medium B obtained by culturing 7 strains of Stenotrophomonas JWG-G1 and 7 strains can be seen in Table 1).
  • Stenotrophomonas pavanii JWG-G1 when PET plastic pellets are used as the only nutrient source, Stenotrophomonas pavanii JWG-G1 can grow; while the OD 600 of the 7 strains of Stenotrophomonas pavanii is not significant Change (within error ⁇ 0.04). It can be seen that only Stenotrophomonas pavanii JWG-G1 can grow and multiply with PET plastic particles as the only nutrient source.
  • the OD 600 changes of JWG-G1 and 7 strains of Stenotrophomonas maltophilia before and after being cultured in an inorganic salt liquid medium containing 2g/L polyethylene terephthalate (PET) are:
  • PET polyethylene terephthalate
  • the OD 600 value of Stenotrophomonas malt JWG-G1 and 7 strains of Stenotrophomonas before being cultured in an inorganic salt liquid medium containing 2g/L polyethylene terephthalate (PET) "+” : The test is positive; "-": The test is negative.
  • Example 5 Degradability of Stenotrophomonas pavanii JWG-G1 to different content of polyethylene terephthalate (PET) plastic particles
  • the specific implementation mode is the same as in Example 4, the difference is that the seed solution B of Stenotrophomonas pavanii JWG-G1 was inoculated with an inoculum amount of 10% (v/v) to contain 2.5g/L, 3.0g/L, 3.5g/L and 4.0g/L polyethylene terephthalate (PET) inorganic salt liquid culture medium;
  • PET polyethylene terephthalate
  • Example 6 Degradation ability of different addition amounts of Stenotrophomonas pavanii to polyethylene terephthalate (PET) plastic particles
  • the specific implementation mode is the same as in Example 4, the difference is that the seed solution B of Stenotrophomonas pavanii JWG-G1 is 15%, 20%, 25%, 30% (v/v) inoculum Inoculate into a liquid medium containing 2g/L polyethylene terephthalate (PET) in inorganic salts;
  • PET polyethylene terephthalate
  • the results are shown in Table 3.
  • the content of degradation product terephthalic acid (TPA) is 250, 300, 330, 360 mg/L, and the content of hydroxyethyl terephthalate (MHET) is 8.1, 10.6, 12.3, respectively. , 15.2mg/L, the weight loss rate of polyethylene terephthalate (PET) particles reached 10.3%, 11.6%, 12.3%, and 13.9%, respectively.
  • the results showed that as the inoculation amount of Stenotrophomonas pavanii JWG-G1 increased, the degradation effect of polyethylene terephthalate (PET) became more significant.
  • Example 8 The ability of Stenotrophomonas pavanii to hydrolyze gelatin and esculin
  • Stenotrophomonas pavanii grows on the test plates containing gelatin and esculin, and there is a clear hydrolysis circle around the colony. After culturing for 5 days, the diameter of the gelatin hydrolysis transparent circle reached 1.3cm, and the gelatin hydrolysis efficiency of its strain JWG-G1 was 0.26cm/day ( Figure 7). The diameter of the esculin hydrolysis transparent circle is 0.9cm, and the hydrolysis efficiency of the strain JWG-G1 to esculin is 0.18cm/day ( Figure 8). It can be seen that Stenotrophomonas pavanii JWG-G1 has the ability to hydrolyze gelatin and esculin.

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Abstract

提供一种可降解聚对苯二甲酸乙二醇酯的嗜麦芽寡养单胞菌，属于微生物技术领域。一株可降解聚对苯二甲酸乙二醇酯的嗜麦芽寡养单胞菌JWG-G1，将此嗜麦芽寡养单胞菌的种子液以10％(v/v)的接种量接种至含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基中培养5d后，可使聚对苯二甲酸乙二醇酯颗粒部分被降解为羟乙基对苯二甲酸酯和可直接回收利用的对苯二甲酸，可使聚对苯二甲酸乙二醇酯塑料颗粒表面的酯键官能团减少，且使聚对苯二甲酸乙二醇酯塑料颗粒的失重率达9.4％，因此，嗜麦芽寡养单胞菌JWG-G1在降解聚对苯二甲酸乙二醇酯中具备极高的应用前景。

Description

可降解聚对苯二甲酸乙二醇酯的嗜麦芽寡养单胞菌 技术领域
本发明涉及可降解聚对苯二甲酸乙二醇酯的嗜麦芽寡养单胞菌，属于微生物技术领域。
背景技术
随着经济的高速发展，人们对塑料制品的消耗水平显著提高，全球每年的塑料消耗量超3.2亿吨，并且消耗量每年以4～6％的速度增长。由于塑料难以降解，因此，全球每年的塑料制品回收利用率仅有14％，这使得塑料垃圾在环境中持续积累，对生态构成了严重的威胁。
聚对苯二甲酸乙二醇酯(PET)是乙二醇(Ethylene glycol,EG)和对苯二甲酸(Terephthalic acid,TPA)以酯键依次连接形成的线性大分子。目前，聚对苯二甲酸乙二醇酯(PET)塑料制品约占所有塑料制品的60％，相应的，聚对苯二甲酸乙二醇酯(PET)塑料垃圾在所有塑料垃圾中也占比较高。因此，降解聚对苯二甲酸乙二醇酯(PET)对治理塑料垃圾十分关键。
目前，降解聚对苯二甲酸乙二醇酯(PET)依旧停留在使用酸、碱、醇、热等化学物理降解法的阶段。化学降解法需要使用大量的化学品，且过程中会产生大量有毒有害物质，对生态环境产生较为严重的负面影响。物理降解法则需要高温和高压的设备，这大大增加了聚对苯二甲酸乙二醇酯(PET)塑料垃圾的治理成本。因此，各国政府积极探索降解聚对苯二甲酸乙二醇酯(PET)新技术。
生物降解技术是直接通过可降解塑料的菌株降解塑料的技术，由于其具有绿色无污染以及成本低的优势，逐渐成为塑料降解领域的研究热点。例如，菌株Ideonella sakaiensis 201-F6，能够以聚对苯二甲酸乙二醇酯(PET)为唯一营养源，降解速率为0.13mg/d(Science,2016,351(6278):1196-1199)；可降解聚乳酸(PLA)的苍白杆菌属菌株(公开号为CN102639690A)；可降解聚羟基脂肪酸酯(PHA)的青霉属菌株(Polymer-plastics Technology and Engineering,2009,48(1):58-63)。
然而，相比于同样是C-O键连接的聚羟基脂肪酸酯(PHA)和聚乳酸(PLA)等生物基塑料，聚对苯二甲酸乙二醇酯(PET)分子链中含有大量芳香基团，导致分子链空间位阻大且表面更加疏水，难以被微生物降解。因此，获得可降解聚对苯二甲酸乙二醇酯(PET)的菌株仍是一个难点。
发明内容
[技术问题]
本发明要解决的技术问题是提供一种可降解聚对苯二甲酸乙二醇酯的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)及其应用。
[技术方案]
为解决上述技术问题，本发明提供了一株嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1，该菌株保藏于中国典型培养物保藏中心，保藏编号为CCTCC NO:M 2019415，保藏日期为2019年06月03日。
所述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1是从来源于无锡桃花山垃圾填埋场的土壤样本中分离得到的，该菌株经测序分析，其16S rDNA序列如SEQ ID NO.1所示，将测序得到的序列在Genbank中进行核酸序列比对，结果显示与寡养单胞菌属的核酸序列相似度高达99％，将与其相似度高的菌株构建系统进化树(具体可见图1)，结果显示菌株属于寡养单胞菌属的嗜麦芽寡养单胞菌，将其命名为嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1。
所述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1在LB固体培养基上的菌落呈圆隆形凸起，淡黄色，不透明，湿润有光泽，边缘有鞭毛(具体可见图2-图3)。
本发明还提供了上述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1在降解聚对苯二甲酸乙二醇酯、明胶或七叶苷中的应用。
在本发明的一种实施方式中，本发明还提供了一种降解解聚对苯二甲酸乙二醇酯(PET)的方法，所述方法为将上述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的种子液接种至含有聚对苯二甲酸乙二醇酯的液体培养基中进行培养。
在本发明的一种实施方式中，上述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的种子液以不低于10％(v/v)的接种量在含有聚对苯二甲酸乙二醇酯的液体培养基中接种。
在本发明的一种实施方式中，上述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的种子液在培养基中的浓度不低于1×10 8CFU/mL。
在本发明的一种实施方式中，所述培养基为无机盐培养基。
在本发明的一种实施方式中，本发明还提供了一种可用于降解聚对苯二甲酸乙二醇酯(PET)的产品，所述产品中含有上述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1。
本发明还提供了一种水解明胶的方法，所述方法为将上述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1接种至含有明胶的平板培养基中进行培养。
本发明还提供了一种可用于水解明胶的产品，所述产品中含有上述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1。
本发明还提供了一种水解七叶苷的方法，所述方法为将上述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1接种至含有明胶的平板培养基中进行培养。
本发明还提供了一种可用于水解七叶苷的产品，所述产品中含有上述嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1。
[有益效果]
(1)本发明提供了一株可降解聚对苯二甲酸乙二醇酯(PET)的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1，将此嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的种子液以10％(v/v)的接种量接种至含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基中培养5d后，可使聚对苯二甲酸乙二醇酯(PET)颗粒部分被降解为羟乙基对苯二甲酸酯(MHET)和可直接回收利用的对苯二甲酸(TPA)，并且，在本发明嗜麦芽寡养单胞菌JWG-G1的作用下，可使聚对苯二甲酸乙二醇酯(PET)塑料颗粒表面的酯键官能团减少，且使聚对苯二甲酸乙二醇酯(PET)塑料颗粒的失重率达9.4％，因此，本发明的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1在降解聚对苯二甲酸乙二醇酯(PET)中具备极高的应用前景。
(2)本发明的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1具有优良的耐盐性能，可在含有1～9g/L NaCl的LB液体培养基中旺盛生长。
(3)本发明的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1能够水解明胶和七叶苷。
生物材料保藏
一株嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1，分类学命名为Stenotrophomonas pavanii JWG-G1，已于2019年06月03日保藏于中国典型培养物保藏中心，保藏编号为CCTCC NO:M 2019415，保藏地址为中国武汉，武汉大学。
附图说明
图1：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的系统进化树。
图2：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的菌落。
图3：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的菌体形态。
图4：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1在不同pH下生长曲线。
图5：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1在不同温度下生长曲线。
图6：聚对苯二甲酸乙二醇酯(PET)塑料颗粒表面经嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1处理后的酯键官能团变化情况。
图7：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1JWG-G1对明胶的水解效率。
图8：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1JWG-G1对七叶苷的水解效率。
具体实施方式
下面结合具体实施例对本发明进行进一步的阐述。
下述实施例中涉及的对苯二酸二甲酯(DET)、聚对苯二甲酸乙二醇酯(PET)塑料颗粒、苯二甲酸乙二醇酯(BHET)购自Sigma公司，TPA、MHET、BHET的标准品购自Sigma公司。
下述实施例中涉及的培养基如下：
LB固体培养基(g/L)：蛋白胨10、酵母粉5、氯化钠10、琼脂13，pH 7.0。
LB液体培养基(g/L)：蛋白胨10、酵母粉5、氯化钠10，pH 7.0。
无机盐液体培养基(g/L)：KH 2PO 4 0.7、K 2HPO 4·3H 2O 0.5、NH 4Cl 2、MgSO 4·7H 2O 0.6、NaCl 0.005、FeSO 4·7H 2O 0.001、ZnSO 4·7H 2O 0.002、MnSO 4·H 2O 0.001。
无机盐固体培养基(g/L)：KH 2PO 4 0.7、K 2HPO 4·3H 2O 0.5、NH 4Cl 2、MgSO 4·7H 2O 0.6、NaCl 0.005、FeSO 4·7H 2O 0.001、ZnSO 4·7H 2O 0.002、MnSO 4·H 2O 0.001、琼脂粉13。
下述实施例中涉及的检测方法如下：
聚对苯二甲酸乙二醇酯(PET)塑料颗粒表面官能团变化的检测方法：
将经菌株处理后的聚对苯二甲酸乙二醇酯(PET)塑料颗粒用去离子水反复清洗3～4次；将清洗完的聚对苯二甲酸乙二醇酯(PET)塑料颗粒于功率200W、频率58KHz的条件下超声15min；将超声后的聚对苯二甲酸乙二醇酯(PET)塑料颗粒放入烘箱60℃烘干6h；以未经处理的聚对苯二甲酸乙二醇酯(PET)塑料颗粒为对照，使用傅里叶变换红外光谱仪检测未经处理的聚对苯二甲酸乙二醇酯(PET)塑料颗粒表面以及经菌株处理后聚对苯二甲酸乙二醇酯(PET)塑料颗粒表面官能团的变化。
降解产物及其含量的检测方法：
标准品处理：分别称取TPA、MHET、BHET的标准品溶于二甲亚砜(DMSO)中制成母液，利用无菌水将母液稀释成0.1mg/mL标准品溶液，用0.22μM的滤头过滤，用注射器注入液相瓶进行HPLC检测；
样品处理：将培养液静置10min，取上清5mL，12000rpm离心8min，用0.22μM的滤头过滤，用注射器注入液相瓶，进行HPLC检测。
失重率的检测方法：
PET塑料颗粒的失重率(％)＝[(m2-m1)÷m2]×100；
m1：经菌株处理后的PET塑料颗粒用去离子水反复清洗3～4次，将清洗完的PET塑料颗粒于功率200W、频率58KHz的条件下超声15min，放入烘箱60℃烘干6h后称重；
m2：经菌株处理前的PET塑料颗粒用去离子水反复清洗3～4次，将清洗完的PET塑料颗粒于功率200W、频率58KHz的条件下超声15min，放入烘箱60℃烘干6h后称重。
明胶和七叶苷水解效率：
水解效率(cm/day)＝D1/n
D1：第n天，检测平板上透明圈直径(cm)；n：检测平板培养天数(day)。
实施例1：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)的筛选
具体步骤如下：
1、普通筛选
以来源于无锡市桃花山垃圾填埋场土壤为样本，取1g土壤加入9mL生理盐水，于35℃、180rpm的条件下振荡富集培养30min；随后取1mL上清液依次稀释至10 -4、10 -5、10 -6，分别取200μL稀释至10 -4、10 -5、10 -6的稀释液均匀涂布于含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐固体培养基上，置于35℃培养箱中恒温培养至长出菌落；实验结果显示经多日培养，分离平板未出现菌落，该方法并不能有效分离出具有降解聚对苯二甲酸乙二醇酯(PET)能力的菌株。
2、“PET诱导培养”筛选
(1)以来源于无锡市桃花山垃圾填埋场土壤为样本，取1g土壤加入9mL含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基，于35℃、180rpm的条件下振荡富集培养48h；接着吸取上述1mL富集液加入新的9mL2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基，相同条件培养10个循环；
(2)随后取步骤(1)培养10个循环后获得的培养液上清1mL依次稀释至10 -4、10 -5、10 -6，取200μL稀释至10 -4、10 -5、10 -6的稀释液均匀涂布于含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐固体培养基上，置于35℃培养箱中恒温培养至长出菌落；实验结果显示经多日培养，分离平板出现的菌落稀疏(5-6个/板)；
(3)作为对照，取步骤(1)培养10个循环后获得的培养液上清1mL依次稀释至10 -4、10 -5、10 -6，取200μL稀释至10 -4、10 -5、10 -6的稀释液转接到不含聚对苯二甲酸乙二醇酯(PET) 的无机盐固体培养基，35℃下进行培养，上述菌落均能够在不含聚对苯二甲酸乙二醇酯(PET)的无机盐固体培养基上生长；
可见，上述菌落均为自养型的纯化菌株，并非是以PET为唯一营养源的非自养型的菌株。该方法并不能有效分离出具有降解聚对苯二甲酸乙二醇酯(PET)能力的菌株。
3、“逐级筛选”策略
(1)以来源于无锡市桃花山垃圾填埋场土壤为样本，取1g土壤加入9mL含有10g/L对苯二甲酸二乙酯(DET)的无机盐液体培养基，于35℃、180rpm的条件下振荡富集培养48h，接着吸取上述1mL富集液加入新的9mL含有10g/L对苯二甲酸二乙酯(DET)的无机盐液体培养基，相同条件培养10个循环；
(2)吸取上述步骤(1)培养10个循环后获得的培养液上清1mL加入新的9mL含有对苯二甲酸双羟乙酯(BHET)的无机盐液体培养基，相同条件培养10个循环；
(3)吸取上述步骤(2)培养10个循环后获得的培养液上清1mL加入新的9mL含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基，相同条件培养10个循环；
(4)吸取上述步骤(3)培养10个循环后获得的培养液上清1mL依次稀释至10 -4、10 -5、10 -6，取200μL稀释至10 -4、10 -5、10 -6的稀释液均匀涂布于含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐固体培养基上，置于35℃培养箱中恒温培养至长出菌落(180-200个/板)；以无营养源的无机盐固体培养基作为对照，得到非自养型的纯化菌株10个，将长势最好的纯化菌株分别命名为菌株JWG-G1。
基于上述的3中不同的分离筛选方法的结果，可发现需要对苯二甲酸二乙酯(DET)和对苯二甲酸双羟乙酯(BHET)等可溶、结构简单的聚对苯二甲酸乙二醇酯(PET)中间体作为初步底物，对聚对苯二甲酸乙二醇酯(PET)降解菌株进行富集，使其菌株适应聚对苯二甲酸乙二醇酯(PET)中间体为唯一营养物。随后就以聚对苯二甲酸乙二醇酯(PET)为唯一底物再次进行降解菌株富集，使其聚对苯二甲酸乙二醇酯(PET)中间体降解菌株中衍生出能够降解聚对苯二甲酸乙二醇酯(PET)的菌株，并使其浓度进一步提升。故“逐级筛选”策略能够有效筛选出具有PET降解能力的菌株。
实施例2：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)鉴定
提取菌株JWG-G1的总DNA进行16S rDNA的扩增和测序(由无锡天霖生物科技有限公司完成)，将测序得到的序列在Genbank中进行核酸序列比对，发现菌株JWG-G1与寡养单胞菌属的16S rDNA序列同源性大于99％，其中，与嗜麦芽寡养单胞菌(Stenotrophomonas  pavanii)DSM 25135的16S rDNA序列相似率达99.5％。可见，菌株JWG-G1属于寡养单胞菌属。
将菌株JWG-G1的16S rDNA序列(序列如SEQ ID NO.1所示)与其相似度高的菌株构建系统进化树(菌株JWG-G1构建得到的系统进化树可见图1)，结果显示菌株JWG-G1与Stenotrophomonas pavanii DSM 25135同属于一个分支，可见，菌株JWG-G1属于寡养单胞菌属，命名为嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1。
实施例3：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)的培养
具体步骤如下：
刮取一环实施例1获得的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1接种至LB固体培养基划线培养，35℃培养36h后，观察其菌落，发现其菌落呈圆隆形凸起，浅黄色，不透明，湿润有光泽，边缘有鞭毛(具体可见图2-图3)。
将实施例1获得的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1经革兰氏染色后于显微镜下观察，发现为革兰氏阳性菌。
刮取一环实施例1获得的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1接种至pH分别为7～11的LB液体培养基中培养，于35℃培养，通过酶标仪检测培养液中的OD 600值，发现最适生长pH为7(具体可见图4)。
刮取一环实施例1获得的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1接种至pH为7的LB液体培养基中培养，分别于25～40℃培养，通过酶标仪检测培养液中的OD 600值，发现最适生长温度为25℃(具体可见图5)。
实施例4：不同寡养单胞菌及嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)对聚对苯二甲酸乙二醇酯(PET)塑料颗粒的降解能力
具体步骤如下：
由于嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1属于寡养单胞菌属，寡养单胞菌属可能是潜在的PET塑料颗粒降解菌株来源属之一，故收集与嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1亲缘关系近的7株寡养单胞属菌(Stenotrophomonas pavanii DSM 25135；Stenotrophomonas bentonitica DSM 103927；Stenotrophomonas chelatiphaga DSM 21508；Stenotrophomonas ginsengisoli KCTC 12539；Stenotrophomonas lactitubi DSM 104152；Stenotrophomonas rhizophila DSM 14405；Stenotrophomonas tumulicola NCIMB 15009)与嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1株作为共同测试菌株。
挑取实施例1获得的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1以及7株寡养单胞菌的单菌落，分别接入100mL的LB液体培养基中，于35℃、180rpm的条件下震荡培养24h，得到种子液A；将种子液A以10％(v/v)的接种量转接到100mL新鲜的LB液体培养基中，于35℃、180rpm的条件下震荡培养24h，得到培养液A；将培养液A于8000rpm的条件下离心10min，收集菌体；将菌体用无机盐培养基洗涤2次后制成OD 600＝1.0的菌悬液作为种子液B；以未接种种子液B的含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基为对照组，将种子液B以10％(v/v)的接种量接种至含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基中(此时，种子液B在PET无机盐液体培养基中嗜麦芽寡养单胞菌JWG-G1的菌浓度为1×10 9CFU/mL)，于35℃、180rpm的条件下震荡培养5d，得到培养液B，测定培养液B前后的OD 600的变化可见表1；将培养液B中的PET塑料颗粒取出，检测其表面官能团结构的变化(嗜麦芽寡养单胞菌JWG-G1以及7株寡养单胞菌培养获得的培养液B中的聚对苯二甲酸乙二醇酯(PET)塑料颗粒表面酯键官能团结构的变化可见表1，嗜麦芽寡养单胞菌JWG-G1培养获得的培养液B中的PET塑料颗粒表面官能团结构的变化可见图6)并检测其失重率(嗜麦芽寡养单胞菌JWG-G1以及7株寡养单胞菌培养获得的培养液B中的聚对苯二甲酸乙二醇酯(PET)塑料颗粒的失重率可见表1)，同时，检测培养液B中聚对苯二甲酸乙二醇酯(PET)塑料颗粒降解产物羟乙基对苯二甲酸酯(MHET)和对苯二甲酸(TPA)的含量(嗜麦芽寡养单胞菌JWG-G1以及7株寡养单胞菌培养获得的培养液B中聚对苯二甲酸乙二醇酯(PET)塑料颗粒降解产物羟乙基对苯二甲酸酯(MHET)和对苯二甲酸(TPA)的含量可见表1)。
由表1和图6可以看出，以PET塑料颗粒为唯一营养源时，嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1能够生长；而7株寡养单胞菌的OD 600无显著变化(误差范围内±0.04)。可见，仅有嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1可以PET塑料颗粒为唯一营养源生长繁殖。
由表1和图6可以看出，经嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1处理5d后，聚对苯二甲酸乙二醇酯(PET)塑料颗粒部分被降解为羟乙基对苯二甲酸酯(MHET)和对苯二甲酸(TPA)，聚对苯二甲酸乙二醇酯(PET)塑料颗粒表面的酯键官能团被破坏(在1000～1300cm -1之间有两个特征峰，在1700～1750cm -1之间有一个特征峰)，且聚对苯二甲酸乙二醇酯(PET)塑料颗粒失重9.4％；而经7株寡养单胞菌处理5d后，聚对苯二甲酸乙二醇酯(PET)塑料颗粒未见明显变化。可见，仅有嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1能够降解聚对苯二甲酸乙二醇酯(PET)塑料颗粒。
表1嗜麦芽寡养单胞菌JWG-G1以及7株寡养单胞菌经含有PET的培养基培养前后的性能检测
Figure PCTCN2021097924-appb-000001
其中，嗜麦芽寡养单胞菌JWG-G1以及7株寡养单胞菌经含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基培养前后的OD 600变化即嗜麦芽寡养单胞菌JWG-G1以及7株寡养单胞菌经含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基培养后的OD 600值减去嗜麦芽寡养单胞菌JWG-G1以及7株寡养单胞菌经含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基培养前的OD 600值，“+”：检测为阳性；“-”：检测为阴性。
实施例5：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1对不同含量的聚对苯二甲酸乙二醇酯(PET)塑料颗粒的降解能力
具体实施方式同实施例4，区别在于，将嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的种子液B以10％(v/v)的接种量分别接种至含有2.5g/L、3.0g/L、3.5g/L和4.0g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基中；
结果如表2所示，其中，降解产物中对苯二甲酸(TPA)含量分别为203、206、205、205mg/L，羟乙基对苯二甲酸酯(MHET)含量分别为8.1、8.0、8.3、8.2mg/L，而聚对苯二甲酸乙二醇酯(PET)颗粒失重率分别达到9.8％、9.7％、9.6％、9.7％。结果显示嗜麦芽寡养单胞菌 (Stenotrophomonas pavanii)JWG-G1对聚对苯二甲酸乙二醇酯(PET)的降解效果，不随聚对苯二甲酸乙二醇酯含量的增加而增加。
表2嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)对不同含量的聚对苯二甲酸乙二醇酯(PET)塑料颗粒的降解能力
Figure PCTCN2021097924-appb-000002
实施例6：不同添加量的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)对聚对苯二甲酸乙二醇酯(PET)塑料颗粒的降解能力
具体实施方式同实施例4，区别在于，将嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的种子液B以15％、20％、25％、30％(v/v)的接种量接种至含有2g/L聚对苯二甲酸乙二醇酯(PET)的无机盐液体培养基中；
结果如表3所示，其中降解产物对苯二甲酸(TPA)含量分别为250、300、330、360mg/L，羟乙基对苯二甲酸酯(MHET)含量分别为8.1、10.6、12.3、15.2mg/L，聚对苯二甲酸乙二醇酯(PET)颗粒失重率分别达到10.3％、11.6％、12.3％、13.9％。结果显示随着嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1接种量的增加，聚对苯二甲酸乙二醇酯(PET)的降解效果越显著。
表3不同添加量的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)对聚对苯二甲酸乙二醇酯(PET)塑料颗粒的降解能力
Figure PCTCN2021097924-appb-000003
实施例7：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)的耐盐能力
具体步骤如下：
挑取实施例1获得的嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1的单菌落接入100mL的LB液体培养基中，于35℃、180rpm条件下震荡培养24h，得到种子液A；将 种子液A以10％(v/v)的接种量转接到100mL新鲜的LB液体培养基中，于35℃、180rpm的条件下震荡培养72h，得到培养液A；将培养液A于8000rpm的条件下离心10min，收集菌体；将菌体用无机盐培养基洗涤2次后制成OD 600＝1.0的菌悬液作为种子液B；将种子液B以10％(v/v)的接种量分别接种至含有不同浓度NaCl(1、2、3、4、5、6、7、8、9、10g/L)的LB液体培养基中，于35℃、180rpm的条件下震荡培养5d，获得培养液B。
由测定培养液B的OD 600结果发现，嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1在含有1～9g/L NaCl的LB液体培养基中生长5d后获得的培养液B中的OD 600增加量分别为0.20、10.31、0.35、0.45、0.35、0.32、0.31、0.29、0.24、0.21。可见，嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1具有优良的耐盐性能。
实施例8：嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)水解明胶和七叶苷的能力
具体步骤如下：
依据参考文献“Bergey's Manual of Systematic Bacteriology.New York:Springer,2012”采用平板透明圈法检测嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1对明胶、七叶苷水解能力。
由检测结果可知，嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)在分别含有明胶、七叶苷的检查平板上生长，菌落周围出现明显的水解透明圈。培养5d后，明胶水解透明圈直径达1.3cm，其菌株JWG-G1对明胶水解效率为0.26cm/day(图7)。七叶苷水解透明圈直径达0.9cm，其菌株JWG-G1对七叶苷水解效率为0.18cm/day(图8)。可见，嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)JWG-G1具备水解明胶、七叶苷的能力。
虽然本发明已以较佳实施例公开如上，但其并非用以限定本发明，任何熟悉此技术的人，在不脱离本发明的精神和范围内，都可做各种的改动与修饰，因此本发明的保护范围应该以权利要求书所界定的为准。

Claims (15)

  1. 一株嗜麦芽寡养单胞菌(Stenotrophomonas pavanii)，其特征在于，所述嗜麦芽寡养单胞菌保藏于中国典型培养物保藏中心，保藏编号为CCTCC NO:M 2019415，保藏日期为2019年06月03日。
  2. 权利要求1所述的嗜麦芽寡养单胞菌在降解聚对苯二甲酸乙二醇酯、明胶或七叶苷中的应用。
  3. 一种降解聚对苯二甲酸乙二醇酯的方法，其特征在于，所述方法为将权利要求1所述嗜麦芽寡养单胞菌接种至含有聚对苯二甲酸乙二醇酯的液体培养基中进行培养。
  4. 如权利要求3所述的方法，其特征在于，权利要求1所述嗜麦芽寡养单胞菌以种子液的形式接种至含有聚对苯二甲酸乙二醇酯的液体培养基中；所述种子液的体积占液体培养基总体积的比例不低于10％。
  5. 如权利要求4所述的方法，其特征在于，所述种子液中，嗜麦芽寡养单胞菌的浓度不低于1×10 8CFU/mL。
  6. 如权利要求5所述的方法，其特征在于，所述培养基为无机盐液体培养基。
  7. 如权利要求6所述的方法，其特征在于，所述无机盐液体培养基的成分包括：KH 2PO 40.7g/L、K 2HPO 4·3H 2O 0.5g/L、NH 4Cl 2、MgSO 4·7H 2O 0.6g/L、NaCl 0.005g/L、FeSO 4·7H 2O 0.001g/L、ZnSO 4·7H 2O 0.002g/L、MnSO 4·H 2O 0.001g/L。
  8. 一种可用于降解聚对苯二甲酸乙二醇酯的产品，其特征在于，所述产品中含有权利要求1所述嗜麦芽寡养单胞菌。
  9. 如权利要求8所述的产品，其特征在于，所述产品中，嗜麦芽寡养单胞菌的浓度不低于1×10 8CFU/mL。
  10. 一种水解明胶的方法，其特征在于，所述方法为将权利要求1所述嗜麦芽寡养单胞菌接种至含有明胶的培养基中进行培养。
  11. 一种可用于水解明胶的产品，其特征在于，所述产品中含有权利要求1所述嗜麦芽寡养单胞菌。
  12. 如权利要求11所述的产品，其特征在于，所述产品中，嗜麦芽寡养单胞菌的浓度不低于1×10 8CFU/mL。
  13. 一种水解七叶苷的方法，其特征在于，所述方法为将权利要求1所述嗜麦芽寡养单胞菌接种至含有七叶苷的培养基中进行培养。
  14. 一种可用于水解七叶苷的产品，其特征在于，所述产品中含有权利要求1所述嗜麦芽寡养单胞菌。
  15. 如权利要求14所述的产品，其特征在于，所述产品中，嗜麦芽寡养单胞菌的浓度不低于1×10 8CFU/mL。
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