A kind of deltamethrin-degrading bacteria and its application
By isolating and identifying Serratia marcescens X011 strain, the problem of difficulty in efficiently removing deltamethrin residue pollution in the prior art is solved, and the effect of efficient deltamethrin is achieved, and good application prospects are provided.
Patent Information
- Application Number
- CN202310051952.5
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-02-02
- Publication Date
- 2025-03-25
- Estimated Expiration
- 2043-02-02
AI Technical Summary
The prior art is difficult to efficiently remove residual contamination of deltamethrin in aquaculture environment, and Serratia slime has poor degradation effect on it.
A strain of Serratia marcescens X011 was isolated and identified. The degradation rate of 45 mg/L deltamethrin reached 95.2% under aerobic conditions and was cultured and applied through specific culture media and conditions.
It has achieved efficient degradation of deltamethrin, with better degradation effect than the deltamethrin strains in the prior art and has good application prospects, especially in the restoration of the bottom mud of river crab ponds.
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Figure CN116333918B_ABST
Abstract
Description
Technical Field
[0001] The invention belongs to the field of microbial degradation, and particularly relates to a cypermethrin-degrading bacterium and an application thereof. Background Art
[0002] Pyrethroid pesticides are a class of cyclopropane ester synthetic insecticides containing phenoxy groups. They are highly effective, broad-spectrum, low-toxic and rapidly metabolized. They are not only widely used in crop pest control and sanitary pest control, but also used to kill a variety of external parasites in livestock and poultry. Deltamethrin is a commonly used pyrethroid insecticide with contact and stomach killing effects and high toxicity to arthropods. In aquaculture, it is used to kill catfish, Chinese catfish and fish lice that parasitize on the surface of fish. In crab farming pond cleaning, it is often used to kill remaining crustaceans such as Procambarus clarkii and river crabs. Deltamethrin is almost insoluble in water, stable to light and heat, relatively stable in acidic media, unstable in alkaline media, and persistent in the environment. Deltamethrin is low toxic to mammals, highly toxic to fish, and highly toxic to arthropods such as cladocerans, copepods, rotifers, shrimps and crabs, which can cause changes in the structure and function of the aquatic ecosystem. Therefore, eliminating the residual pollution of cypermethrin in the breeding environment has become an urgent problem to be solved.
[0003] Microbial degradation is an important way to eliminate pesticide residue pollution in aquaculture environments. It has the advantages of high efficiency, low cost, mild conditions, etc., and the microbial strains isolated and strengthened from the crab breeding environment have no secondary pollution problems, and have broad application prospects. At present, the reported pyrethroid pesticide degradation bacteria include Enterobacter cloacae, Pseudomonas, Bacillus, Serratia, Micrococcus, Klebsiella pneumoniae, Rhodococcus, Sphingobacterium multivorum, Alcaligenes, Aspergillus niger, Rhizobium, Sphingobacterium, etc., and there has been no report on the efficient degradation of deltamethrin by Serratia marcescens. Summary of the invention
[0004] The technical problem to be solved by the present invention is to provide a cypermethrin-degrading bacterium and application thereof. The bacterium can achieve a degradation rate of 95.2% for 45 mg / L cypermethrin in 5 days under aerobic conditions, and the degradation effect is higher than that of the cypermethrin strain in the prior art, and has good application prospects.
[0005] The invention provides a deltamethrin-degrading bacterium. The deltamethrin-degrading bacterium is Serratia marcescens X011, the preservation number of which is CCTCC M 20221992, and the GenBank accession number of 16S rRNA of the strain is OP984150.1.
[0006] The colonies of the strain are round, red / pink, with slightly lighter edges, raised, smooth surfaces, neat edges, and moist; the strain is a Gram-negative short bacillus; it produces red pigment when cultured at 28°C, but does not produce red pigment when cultured at 37°C.
[0007] The strain is derived from the bottom mud of river crab ponds, and is obtained through cypermethrin concentration gradient domestication, separation, purification and screening, and will be mainly used for the restoration of river crab pond mud contaminated by cypermethrin.
[0008] The present invention also provides an application of deltamethrin-degrading bacteria in the bottom mud of river crab ponds.
[0009] The culture medium of the cypermethrin-degrading bacteria comprises: 10-20 g / L glucose, 5-10 g / L yeast powder, 0.1-1 g / L NaCl, 1.0-5.0 g / L NH4SO4, 1.0-2.0 g / L NaH2PO4, 0.1-0.5 g / L K2HPO4, 0.1-0.5 g / L MgSO4 and 0.01-0.05 g / L FeSO4.
[0010] The culture conditions of the deltamethrin-degrading bacteria are as follows: culture temperature 28-30° C., pH 7.4-7.8, inoculation amount 3-5%, and culture time 120-130 hours.
[0011] The Serratia marcescens X011 strain of the present invention has a deposit number of CCTCC M20221992 and was deposited in the China Center for Type Culture Collection CCTCC on December 19, 2022. The deposit address is Wuhan University, No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province.
[0012] Beneficial Effects
[0013] The strain of the invention can achieve a degradation rate of 95.2% for 45 mg / L deltamethrin in 5 days under aerobic conditions, and the degradation effect is higher than that of the deltamethrin strain in the prior art, and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS
[0014] Figure 1 The inorganic salt agar plate bacterial lawn is the Serratia marcescens X011 strain of the present invention.
[0015] Figure 2 The degradation of deltamethrin in soil by the Serratia marcescens X011 strain of the present invention is shown. DETAILED DESCRIPTION
[0016] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and are not intended to limit the scope of the present invention. In addition, it should be understood that after reading the content taught by the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms fall within the scope limited by the appended claims of the application equally.
[0017] Example 1
[0018] Screening, isolation and identification of deltamethrin-degrading bacteria:
[0019] (1) Collect the bottom mud of a river crab pond where deltamethrin has been used for pond cleaning for many years.
[0020] (2) The environmental samples were enriched and cultured in multiple rounds using an inorganic salt culture medium (NaCl 0.5 g / L, NH4NO3 1.0 g / L, K2HPO4 1.5 g / L, KH2PO4 0.5 g / L, MgSO4·H2O 0.5 g / L) with cypermethrin as the carbon source.
[0021] (3) Add 100 mg / L of deltamethrin to the inorganic salt medium and culture at 30°C and 150 r / min on a shaker for 7 days; then transfer to the next round of liquid enrichment medium at a volume ratio of 10%, increase the deltamethrin concentration to 200 mg / L, and culture for 7 days in the same way. Repeat this cycle, increasing the deltamethrin concentration by 100 mg / L each time until the deltamethrin concentration is increased to 500 mg / L.
[0022] (4) After the enrichment and acclimation culture, the bacterial liquid was spread on a basic inorganic salt agar medium containing 300 mg / L of deltamethrin. After culturing at 30°C for 3 days, individual colonies of different morphologies were picked out for separation and purification on LB plates. The obtained single strains were inoculated into glycerol liquid culture medium and stored in a -80°C refrigerator.
[0023] (5) The DNA of the preserved strain was extracted and used as a template. 16S rDNA universal primers (F: 5'-AGAGTTTGATCCTGGCTCAG-3', R: 5'-AAGGAGGTGATCCAGCCGC-3') were used for PCR amplification. The amplified fragment was sequenced. Homology comparison analysis showed that the preserved strain X011 had the highest homology with Serratia marcescens. The GenBank accession number of the strain's 16S rDNA is OP984150.1.
[0024] Toxicity test:
[0025] At a water temperature of 23±2℃, Chinese mitten crab (10-12g / crab) was immersed in 2×10 7Among the strains of Serratia marcescens X011 with a content of 1.5 cfu / ml, the survival rate after 5 days was 100%, indicating that strain X011 had no toxicity to crabs.
[0026] Deltamethrin-degrading bacteria degrade deltamethrin in liquid:
[0027] Liquid culture medium: glucose 15g / L, yeast powder 8g / L, NaCl 0.5g / L, NH4SO4 3.0g / L, NaH2PO41.5g / L, K2HPO4 0.2g / L, MgSO4 0.3g / L, FeSO4 0.05g / L; culture conditions: culture temperature 28-30°C, pH 7.4-7.8, inoculation amount 5%, cypermethrin is added to the above liquid culture medium to a concentration of 45.06mg / L, after 120h of culture, the concentration of cypermethrin in the culture medium is 2.17mg / L, and the degradation rate of cypermethrin is 95.2%.
[0028] Inoculation of deltamethrin-degrading bacteria into pond mud to degrade deltamethrin:
[0029] Deltamethrin was added to the pond sediment that was tested to be free of deltamethrin. Two experimental groups with added degradation bacteria and one control group without added degradation bacteria were set up. The inoculation amount of deltamethrin-degrading bacteria was 3% and 5%, and the initial deltamethrin content was 1.905 mg / kg and 1.912 mg / kg, respectively. After 24 days under natural conditions, the deltamethrin content in the sediment of the two experimental groups was 0.120 mg / kg and 0.078 mg / kg, respectively, and the degradation rate of deltamethrin in the sediment was 87.9% and 92.2% (see Figure 2 ).
Claims
1. An application of deltamethrin-degrading bacteria in degrading deltamethrin in crab pond sediment, characterized in that: The cypermethrin-degrading bacteria is Serratia marcescens ( Serratia marcescens )X011, the deposit number is CCTCC M20221992, and the GenBank accession number of the strain 16S rRNA is OP984150.
1.
2. The use according to claim 1, characterized in that: The culture medium of the cypermethrin-degrading bacteria comprises: 10-20 g / L glucose, 5-10 g / L yeast powder, 0.1-1 g / L NaCl, 1.0-5.0 g / L NH4SO4, 1.0-2.0 g / L NaH2PO4, 0.1-0.5 g / L K2HPO4, 0.1-0.5 g / L MgSO4 and 0.01-0.05 g / L FeSO4.
3. The use according to claim 2, characterized in that: The culture conditions of the cypermethrin-degrading bacteria are as follows: culture temperature 28-30° C., pH 7.4-7.8, inoculation amount 3-5%, and culture time 120-130 h.