CN112877244B - Ultraviolet-resistant immobile bacterium and application thereof - Google Patents
Ultraviolet-resistant immobile bacterium and application thereof Download PDFInfo
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F1/00—Treatment of water, waste water, or sewage
- C02F1/30—Treatment of water, waste water, or sewage by irradiation
- C02F1/32—Treatment of water, waste water, or sewage by irradiation with ultraviolet light
-
- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F3/00—Biological treatment of water, waste water, or sewage
- C02F3/34—Biological treatment of water, waste water, or sewage characterised by the microorganisms used
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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
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- C—CHEMISTRY; METALLURGY
- C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
- C02F2101/00—Nature of the contaminant
- C02F2101/30—Organic compounds
- C02F2101/34—Organic compounds containing oxygen
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- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
- Y02W10/00—Technologies for wastewater treatment
- Y02W10/10—Biological treatment of water, waste water, or sewage
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Abstract
An ultraviolet-resistant immobile bacterium and application thereof, relating to the field of microorganisms, in particular to an ultraviolet-resistant immobile bacterium and application thereof. The problem that a biological membrane is easily damaged by ultraviolet light in the conventional ICPB process is solved. The bacterium is Acinetobacter (Acinetobacter enhantopacatal) with strong light catalysisysis) F3, which is preserved in the China general microbiological culture Collection center, the preservation date is 11 months and 20 days in 2019, and the preservation number is CGMCC No. 18985. The strong light catalysis acinetobacter F3 can be used for catalyzing the Acinetobacter under ultraviolet light UV254Growth under irradiation, can enhance the degradation effect of ultraviolet light on humic acid, and can be applied to ultraviolet light UV254The humic acid removal rate can reach 65.77 percent after 10 hours of treatment under the irradiation condition. The invention is applied to the field of wastewater treatment.
Description
Technical Field
The invention relates to the field of microorganisms, in particular to an ultraviolet-resistant immobile bacterium and application thereof.
Background
The photocatalytic oxidation technology utilizes the characteristic that the surface of a semiconductor oxide material can be activated under the illumination condition, so that organic matters are effectively oxidized and decomposed, and the photocatalytic oxidation technology has the advantages of high efficiency, low energy consumption, low pollution, low price, wide application range and the like. TiO 22The material has the advantages of good stability, convenient use, no toxicity and the like, and is the most widely used semiconductor photocatalyst at present. Loading the biological membrane on TiO2The porous carrier skeleton of (2) can form a photocatalytic Biodegradation direct Coupling technology (ICPB) to make TiO2The biodegradable part of the photocatalytic degradation product can be immediately degraded by microorganisms, so that the degradation time is shortened, and the degradation efficiency is improved. At present, most of research focuses on how to increase the specific surface area and porosity of the photocatalytic material to increase the attachment rate of the biofilm on the photocatalytic material and increase the photocatalytic efficiency.
TiO2The ultraviolet light is a commonly used sterilization tool and is excited only when the radiation light source irradiates ultraviolet light with the wavelength of less than 387nm, under the irradiation of the ultraviolet light, a biological film attached to the outside of the carrier is greatly damaged, and the ultraviolet light can also have adverse effects on the biological film inside the carrier, such as causing the biological film to fall off, damaging microbial cells, generating soluble microbial products and the like. Screening to obtainThe ultraviolet light resistant dominant bacteria have important significance for improving the treatment efficiency of the photocatalytic biodegradation direct coupling technology.
Disclosure of Invention
The invention provides an ultraviolet-resistant immobile bacterium and application thereof, aiming at solving the problem that a biological membrane is easily damaged by ultraviolet light in the conventional ICPB (intensive care hardening) process.
The ultraviolet-resistant immobile bacterium is high light catalytic Acinetobacter (Acinetobacter enhantopacatalysis) F3, is preserved in the common microorganism center of China Committee for culture Collection of microorganisms, has a preservation address of No. 3 Xilu No.1 Beijing, Chaoyang, and has a preservation date of 11 and 20 days in 2019 and a preservation number of CGMCC No. 18985.
The morphological characteristics of Acinetobacter strongly catalyzing (Acinetobacter enhantopacatalysis) F3 are as follows: the bacillus cocci have the length of 0.7-1.3 mu m and the width of 1.4-1.5 mu m and do not have flagella; gram stain negative, but not easy to decolorize; no spore exists; the bacterial colony is round, smooth and convex.
The invention has the physiological and biochemical characteristics of Acinetobacter sinensis (Acinetobacter enhanophotocatalysis) F3: is an obligate aerobic bacterium and can grow under the conditions of 15-45 ℃ and pH 6-8; the optimum growth temperature is 35 deg.C, the optimum growth pH is 7.2, and the growth can be performed under ultraviolet light UV254Survival under the condition; catalase-positive and oxidase-positive, can grow by taking sodium acetate as a single carbon source, does not hydrolyze glucose, and can utilize L-alanine, alpha-ketoglutaric acid, D-malic acid, L-apple and 1% sodium lactate; has stronger resistance to lincomycin, vancomycin, rifamycin and aztreonam.
The invention discloses a molecular biological identification result of Acinetobacter strongly catalyzing F3: through 16S rDNA sequence comparison analysis, the similarity of the Acinetobacter bouvatii (Acinetobacter bouvatii) and the similar species thereof is only 97 percent, the G + C content is 44.23 percent, and the strong light catalysis Acinetobacter F3 is determined by combining the morphological characteristics of thalli, growth conditions and physiological and biochemical identification results and is a new bacterium.
The invention discloses application of Acinetobacter enhantophyllus F3 in wastewater treatment.
The invention discloses application of Acinetobacter blazei (Acinetobacter enhanthocatalysis) F3 in photocatalytic oxidation.
Further, the application of the strong light catalytic Acinetobacter (Acinetobacter enhantopacatalysis) F3 in an ICPB system.
Further, the strong light catalytic Acinetobacter (Acinetobacter enhantopacatalysis) F3 is used for forming a biofilm supported on TiO2Within the porous carrier backbone.
The invention has the beneficial effects that:
the invention relates to an ultraviolet light resistant immobile bacterium, which is a new bacterium named Acinetobacter angusticeps (Acinetobacter enhantophyllum) F3 and can be used for ultraviolet light UV254Growing under irradiation condition, and the generation time under ultraviolet condition at 35 deg.C is 3.58 min. The strong light catalytic acinetobacter F3 can enhance the degradation effect of ultraviolet light on humic acid, and the degradation effect is enhanced under the action of ultraviolet light UV254The humic acid removal rate can reach 65.77 percent after 10 hours of treatment under the irradiation condition.
Drawings
FIG. 1 shows the form of Acinetobacter F3 in the present invention;
FIG. 2 is a colony morphology of the strongly photocatalytic A.acinetobacter F3 according to the present invention;
FIG. 3 is a graph showing the growth of Acinetobacter furiosus F3 under UV light and dark culture conditions.
Detailed Description
The technical solution of the present invention is not limited to the following specific embodiments, but includes any combination of the specific embodiments.
The first embodiment is as follows: the ultraviolet-resistant immobilized bacteria is Acinetobacter enhanthocathalysis F3, is preserved in the common microorganism center of China Committee for culture Collection of microorganisms, has the preservation address of No. 3 Xilu No.1 of Beijing republic of Chaoyang, the preservation date of 11 and 20 days in 2019 and the preservation number of CGMCC No. 18985.
The second embodiment is as follows: the screening method of the ultraviolet-resistant immobile bacteria F3 of the embodiment comprises the following steps: from the university of Heilongjiang, ChinaTaking 300mg of surface soil in the jar, adding distilled water to 1L, stirring uniformly and standing. And standing to obtain supernatant as soil leaching solution. Adding 3 parts of 300mL soil leachate into 500mL of screening solution respectively, and performing UV treatment at 35 deg.C254Culturing for 3 days under the irradiation of ultraviolet lamp and magnetic stirrer. After gradient dilution, UV at 35 ℃254Streaking separation is carried out under the irradiation of an ultraviolet lamp, and a typical single colony is selected for purification culture, namely the strain F3 of the embodiment.
The screening liquid comprises the following components: 1g/L carbon source, NH4Cl 0.1～0.5g/L，MgSO4·7H2O 0.01～0.05g/L，K2HPO4 0.05～0.2g/L，NaCl 0.05～0.12g/L，MnSO4·4H2O 0.01g/L，FeSO40.01g/L and 0.5-1 g/L of catalyst for screening, wherein the carbon source is humic acid, glucose, starch or sodium acetate, and the catalyst for screening can be TiO2Iron powder, ferrous sulfate, and the like.
In the embodiment, the ultraviolet-resistant immobilized bacteria F3 is subjected to physiological and biochemical identification by referring to Bergey's Manual of bacteria identification, eighth edition and Manual of identification of common bacteria systems:
the morphological characteristics of the strain are as follows: coccobacillus, as shown in FIG. 1, has a length of 0.7-1.3 μm and a width of 1.4-1.5 μm, and has no flagella; gram stain negative, but not easy to decolorize; no spore exists; the colonies were round, smooth, and convex, as shown in FIG. 2.
The physiological and biochemical characteristics are as follows: the obligate aerobic bacteria can grow under the conditions of 15-45 ℃ and pH 6-8; the optimum growth temperature is 35 deg.C, the optimum growth pH is 7.2, and the growth can be performed under ultraviolet light UV254Survival under the condition; catalase-positive and oxidase-positive, can grow by taking sodium acetate as a single carbon source, does not hydrolyze glucose, and can utilize L-alanine, alpha-ketoglutaric acid, D-malic acid, L-apple and 1% sodium lactate; has stronger resistance to lincomycin, vancomycin, rifamycin and aztreonam.
Performing molecular identification on the strain F3, extracting bacterial genome DNA, and performing 16S rDNA sequence amplification, wherein PCR reaction primers for amplification are general primers: a forward primer 5'-GAGCGGATAACAATTTCACACAGG-3'; the reverse primer 5'-CGCCAGGGTTTTCCCAGTCACGAC-3'. The PCR reaction system is as follows: 50-100 ng of template DNA, 25 mu L of Taq enzyme mixed liquor, 0.5 mu L forward primer, 0.5 mu L reverse primer and sterile deionized water are added to 50 mu L. PCR amplification conditions: pre-deforming at 94 ℃ for 5min, then denaturing at 94 ℃ for 1min, amplifying at 50-55 ℃ for 1min, extending at 72 ℃ for 1.5min, and extending at 72 ℃ for 5min after 30 cycles. The fragments obtained were used for sequencing. Obtaining 1473bp sequence length, submitting to GenBank registration number MN587978, wherein the 16SrDNA sequence is shown as SEQ ID NO: 1 is shown.
The similarity of the strain to the similar species of the Acinetobacter buuveitis bacteria is only 97 percent, the G + C content is 44.23 percent, and the strain F3 is determined by combining morphological characteristics, growth conditions and physiological and biochemical identification results of bacteria, and is named as Acinetobacter blazei (Acinetobacter enhantopacatalysis) F3.
The third concrete implementation mode: the strong light catalysis of the embodiment is Acinetobacter (Acinetobacter enhantopacatalysis) F3 in ultraviolet light UV254Growth test under irradiation conditions:
254nm ultraviolet light with a light intensity of 27.8 μ w/cm2The strain Acinetobacter strongly catalized F3 was cultured in 100mL of a standard LB medium (tryptone 10g/L, yeast extract 5g/L, NaCl 10g/L) at 35 ℃ by measuring the absorbance value (OD) of the solution at 600nm600) The growth curve of the strain was plotted and compared with the growth curve of strain F3 cultured at 35 ℃ in the absence of light, and the results are shown in FIG. 3, in which ■ indicates dark culture conditions and ● indicates ultraviolet light conditions. UV lamp254Under the irradiation condition, the acclimation period for the growth of the strain F3 is shortened, which is related to that organic matters in water are decomposed by ultraviolet light, so that the organic matters are more easily utilized by the strain. After culturing for 24 hours under the ultraviolet condition, the growth of the strain F3 shows a decline trend, which indicates that the strain is still damaged by long-time ultraviolet irradiation, and the ultraviolet light intensity is 27.8 mu w/cm2The irradiation culture for 2-11 h under the condition is most beneficial to the growth of the strain F3.
The strong light catalytic Acinetobacter (Acinetobacter enhantopacatalysis) F3 can be used for ultraviolet light UV254Growing under irradiation condition, and the generation time under ultraviolet condition at 35 deg.C is 3.58 min.
The fourth concrete implementation mode: the strong light catalysis of the embodiment is Acinetobacter (Acinetobacter enhantopacatalysis) F3 in ultraviolet light UV254Test for removing humic acid from water under irradiation conditions:
taking 0.3-0.5 mg of newly cultured F3 thallus, inoculating into 100mL of humic acid solution, and culturing at 35 deg.C under 254nm ultraviolet light and 27.8 μ w/cm ultraviolet light intensity2The treatment was carried out at 160rpm, the Total Organic Carbon (TOC) concentration in water was measured after the humic acid treatment for 10 hours, and the effect of removing humic acid by the strain F3 was analyzed with reference to a humic acid solution without inoculated cells, and the results are shown in Table 1.
TABLE 1 removal of humic acid by Acinetobacter F3 under catalysis of ultraviolet light
As a result, the TOC removal effect was as low as less than 10% in the treatment without inoculation of bacterial cells. And 65.77 percent of humic acid in water can be removed by treating for 10 hours through the strain F3.
The ultraviolet irradiation can decompose humic acid into biodegradable micromolecular organic matter, and the bacterial strain F3 can resist ultraviolet irradiation and utilize biodegradable organic matter to carry out growth and metabolism to oxidize the biodegradable organic matter thoroughly, so that the organic matter in water can be decomposed thoroughly. Therefore, the strain F3 can enhance the degradation effect of ultraviolet light on humic acid.
Sequence listing
<110> university of Heilongjiang
<120> ultraviolet-resistant immobile bacterium and application thereof
<160> 3
<210> 1
<211> 1473
<212> DNA
<213> Acinetobacter strain (Acinetobacter enhanophoatalysis)
<220>
<223> strong light catalysis acinetobacter F3
<400> 1
ggctcagatt gaacgctggc ggcaggctta acacatgcaa gtcgagcgga gttgtggtgc 60
ttgcaccata acttagcggc ggacgggtga gtaatgctta ggaatctgcc tattagtggg 120
ggacaacatc tcgaaaggga tgctaatacc gcatacgtcc tacgggagaa agcaggggac 180
cttcgggcct tgcgctaata gatgagccta agtcggatta gctagttggt ggggtaaagg 240
cctaccaagg cgacgatctg tagcgggtct gagaggatga tccgccacac tgggactgag 300
acacggccca gactcctacg ggaggcagca gtggggaata ttggacaatg gggggaaccc 360
tgatccagcc atgccgcgtg tgtgaagaag gccttttggt tgtaaagcac tttaagcgag 420
gaggaggcta ctggtattaa tactaccgga tagtggacgt tactcgcaga ataagcaccg 480
gctaactctg tgccagcagc cgcggtaata cagagggtgc gagcgttaat cggatttact 540
gggcgtaaag cgtgcgtagg cggcttttta agtcggatgt gaaatccctg agcttaactt 600
aggaattgca ttcgatactg ggaggctaga gtatgggaga ggatggtaga attccaggtg 660
tagcggtgaa atgcgtagag atctggagga ataccgatgg cgaaggcagc catctggcct 720
aatactgacg ctgaggtacg aaagcatggg gagcaaacag gattagatac cctggtagtc 780
catgccgtaa acgatgtcta ctagccgttg gggcctttga ggctttagtg gcgcagctaa 840
cgcgataagt agaccgcctg gggagtacgg tcgcaagact aaaactcaaa tgaattgacg 900
ggggcccgca caagcggtgg agcatgtggt ttaattcgat gcaacgcgaa gaaccttacc 960
tggtcttgac atacagagaa ctttccagag atggattggt gccttcggga actctgatac 1020
aggtgctgca tggctgtcgt cagctcgtgt cgtgagatgt tgggttaagt cccgcaacga 1080
gcgcaaccct tttccttact tgccatcggg tcatgccggg aactttaagg atactgccag 1140
tgacaaactg gaggaaggcg gggacgacgt caagtcatca tggcccttac gaccagggct 1200
acacacgtgc tacaatggtc ggtacaaagg gttgctacct agcgatagga tgctaatctc 1260
aaaaagccga tcgtagtccg gattggagtc tgcaactcga ctccatgaag tcggaatcgc 1320
tagtaatcgc ggatcagaat gccgcggtga atacgttccc gggccttgta cacaccgccc 1380
gtcacaccat gggagtttgt tgcaccagaa gtaggtagtc taaccgcaag gaggacgctt 1440
accacggtgt ggccgatgac tggggtgaag tcg 1473
<210> 2
<211> 24
<212> DNA
<213> Artificial sequence
<220>
<223> Forward primer
<400> 2
gagcggataacaatttcacacagg 24
<210> 3
<211> 24
<212> DNA
<213> Artificial sequence
<220>
<223> reverse primer
<400> 3
cgccagggttttcccagtcacgac 24
Claims (5)
1. A strain of ultraviolet-resistant immobile bacterium is characterized in that the bacterium is strong light catalytic acinetobacter (A), (B) and (C)Acinetobacter enhanphotocatalysis) F3, deposited in China general microbiological culture Collection center, with the deposit address of No. 3 Xilu No.1 on North Chen of the south-facing-Yang district, Beijing, the date of deposit being 11 months and 20 days in 2019, and the number of deposit being CGMCC No. 18985.
2. Use of the UV-resistant immobilized bacteria of claim 1 in the treatment of wastewater.
3. Use of the UV-resistant immobilized bacteria of claim 1 in photocatalytic oxidation.
4. Use according to claim 3, characterized in that the UV-resistant immobile bacteria are used in the ICPB system.
5. Use according to claim 4, characterized in that the UV-resistant immobile bacteria are used to form a biofilm supported on TiO2Within the porous carrier backbone.
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