CN106929448B - The acinetobacter calcoaceticus CZ1701 bacterial strain and application thereof of one plant of degradation different shape nitrogen - Google Patents

The acinetobacter calcoaceticus CZ1701 bacterial strain and application thereof of one plant of degradation different shape nitrogen Download PDF

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Publication number
CN106929448B
CN106929448B CN201710186658.XA CN201710186658A CN106929448B CN 106929448 B CN106929448 B CN 106929448B CN 201710186658 A CN201710186658 A CN 201710186658A CN 106929448 B CN106929448 B CN 106929448B
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China
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bacterial strain
nitrogen
degradation
cod
acinetobacter calcoaceticus
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CN106929448A (en
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程国军
杜冬云
占伟
陈绍华
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South Central Minzu University
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South Central University for Nationalities
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • 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
    • 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
    • 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
    • C02F2101/10—Inorganic compounds
    • C02F2101/16—Nitrogen compounds, e.g. ammonia
    • 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
    • C02F2101/10—Inorganic compounds
    • C02F2101/16—Nitrogen compounds, e.g. ammonia
    • C02F2101/163—Nitrates
    • 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
    • C02F2101/10—Inorganic compounds
    • C02F2101/16—Nitrogen compounds, e.g. ammonia
    • C02F2101/166—Nitrites
    • C—CHEMISTRY; METALLURGY
    • C02—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F—TREATMENT OF WATER, WASTE WATER, SEWAGE, OR SLUDGE
    • C02F2103/00—Nature of the water, waste water, sewage or sludge to be treated
    • C02F2103/20—Nature of the water, waste water, sewage or sludge to be treated from animal husbandry

Abstract

The present invention provides a kind of denitrogenation bacterial strains of degradable different shape nitrogen, the bacterial strain is named as CZ1701, belongs to a kind of acinetobacter calcoaceticus, which is preserved in China typical culture collection center, the deposit date is on December 27th, 2016, deposit numbers are as follows: CCTCC NO:M2016788.CZ1701 bacterial strain provided by the present invention has very high degradation rate for various forms nitrogen, to NH3‑N、NO3‑N、NO2The degradation rate of-N and total N (TN) respectively reach 89.1%, 53.2%, 66.7% and 86.6%.In addition, bacterial strain also has the ability of removal COD, the degradation rate of COD is 77.6%.Therefore, CZ1701 bacterial strain provided by the present invention has the advantages that the nitrogen and COD of wide Acclimation temperature range, energy while efficient degradation various forms, in the processing especially suitable for degrade high-concentration ammonia nitrogenous wastewater such as farm's excrement and urine water.

Description

The acinetobacter calcoaceticus CZ1701 bacterial strain and application thereof of one plant of degradation different shape nitrogen
Technical field
The present invention relates to a kind of acinetobacter calcoaceticus new strains, the nitrogen of degradable different valence state, in particular to the bacterial strain is aerobic Denitrification and process performance to piggery wastewater, belong to microorganisms technical field.
Background technique
Environmental problem is more prominent since new century, and wherein the pollution of water body is such as: eutrophication, wawter bloom, red tide etc. take place frequently, It arouses widespread concern.The basic reason of water eutrophication is mainly that nitrogen and phosphorus element too high levels cause water body table Layer cyanobacteria and algae excessive propagation.Therefore, the removal of Water element for protection environment, develops the economy and ensures health All have great importance.With the large-scale development of livestock and poultry breeding industry, pig farm waste waste water causes sternly ambient enviroment Heavily contaminated, waste are discharged into lake waters, will cause water eutrophication, such as permeate the ground, can make nitrate nitrogen in underground water or Nitrite nitrogen concentration increases, and toxic component increases, and leads to water quality deterioration, jeopardizes the water quality of periphery domestic water.For pig farm excrement Stain disease has the characteristics that high ammonia nitrogen, low C/N than, the water quality that pH value is higher, biodegradability is poor, and (biology removes microorganism treatment Nitrogen) it is to remove one of polluted by nitrogen most efficient method in water.Microorganism can be the pollutant ammonium nitrogen and nitre of most serious in water body Hydrochlorate etc. is converted into the nitrogen source of own growth, gaseous state nitride (nitrogen and nitrous oxide) etc., to reduce nitrogen residue.
Therefore, separation screening can remove high ammonia nitrogen under the conditions of pH value is higher, and have one to other valence state nitrogen sources and COD Fixed removal effect, and its biological characteristics and the application in the improvement of piggery waste contaminated wastewater are studied, it can be controlled for microorganism It manages high ammonia-nitrogen wastewater and good resource is provided, be of great significance for the biological treating of the waste water such as poultry waste.
Summary of the invention
The object of the present invention is to provide a kind of denitrogenation bacterial strain of degradable different shape nitrogen, which can be used for pig farm Deng the biological treating containing high-concentration ammonia nitrogenous wastewater.
The present inventor screens one plant of novel strain, which is named as CZ1701, belongs to a kind of acinetobacter calcoaceticus (Acinetobacter sp.), which is preserved in China typical culture collection center, address: China, Wuhan, Wuhan are big It learns；Postcode 430072, the deposit date is on December 27th, 2016, deposit numbers are as follows: CCTCC NO:M2016788；The bacterial strain 16S rDNA sequence is as shown in SEQ ID NO.1.
CZ1701 bacterial strain be it is isolated from Hubei Province's pig-farm wastewater pollution discharge pond mud sample, be identified as Acinetobacter sp..The form and physiological and biochemical property of the bacterial strain are as follows: bacterium colony is observed on LB solid medium in milky white Color, neat in edge are smooth wet；It is Gram-negative, rod-shaped, no gemma that microscopy, which observes bacterial strain,.
By strain inoculated to 5ml LB culture solution (peptone 10.0g, yeast extract 5.0g, NaCl 10.0g, H2O1000mL), 37 DEG C, 200rpm/min culture 24 hours, 10000rpm/min is centrifuged 1min, collects thallus and is centrifuged in 1.5ml Guan Zhong extracts bacterial genomes using CTAB method.Use 16S rDNA universal primer (upstream primer: 5 '- AGAGTTTGATCCTGGCTCAG-3′；Downstream primer: 5 '-AAGGAGGTGATCCAGCCGCA-3 ') carry out PCR amplification, amplification Product carries out nucleotide sequence homology with the listed 16S rDNA sequence of GenBank after being sequenced and compares analysis, and motionless The 16S rDNA sequence similarity of bacillus (Acinetobacter) reaches 99%, is accredited as this category.
Compared with prior art, the invention has the following advantages that acinetobacter calcoaceticus CZ1701 bacterial strain pair provided by the present invention There is very high degradation rate in various forms nitrogen, to NH3-N、NO3-N、NO2The degradation rate of-N and total N (TN) respectively reach 89.1%, 53.2%, 66.7% and 86.6%.In addition, bacterial strain also has the ability of removal COD, compared with the control group, COD Degradation rate be 77.6%.Therefore, CZ1701 bacterial strain provided by the present invention has temperature compared with known denitrogenation acinetobacter calcoaceticus The advantages of spending the nitrogen and COD of wide adaptation range, energy while efficient degradation various forms, it is useless especially suitable for degradation ammonia nitrogen in high density In the processing of water such as farm's excrement and urine water.
Specific embodiment
Detailed specific description done to the present invention combined with specific embodiments below, but protection scope of the present invention not office It is limited to following embodiment.
The correlated performance of acinetobacter calcoaceticus CZ1701 bacterial strain provided by the present invention is studied in following embodiment.
Influence of 1 different temperatures of embodiment to CZ1701 strain growth
Using ammoniacal nitrogen as heterotrophic nitrification culture medium (the sodium citrate 5g, ammonium sulfate 1.0g, potassium dihydrogen phosphate of only nitrogen source 1g, dipotassium hydrogen phosphate 0.1g, magnesium sulfate 0.2g, ferrous sulfate 0.03g, distilled water 1000mL, pH value 8.5) in, it is inoculated with logarithmic phase CZ1701 bacterium solution makes bacterium solution initial OD600Value is 0.1, is placed in shake culture in the shaking table of different temperatures, detection growth feelings after 48h Condition, 3 repetitions of this experimental setup.CZ1701 strain growth temperature range is very wide, can grow in the range of 15 DEG C to 42 DEG C, OD in 48 hours600Value can reach 1.5 or more, and it is stronger to illustrate that CZ1701 strain adapts to ambient temperature capability.
2 CZ1701 bacterial strain of embodiment takes off ammoniacal nitrogen ability
Using ammoniacal nitrogen as heterotrophic nitrification culture medium (the sodium citrate 5g, ammonium sulfate 1.0g, potassium dihydrogen phosphate of only nitrogen source 1g, dipotassium hydrogen phosphate 0.1g, magnesium sulfate 0.2g, ferrous sulfate 0.03g, distilled water 1000mL, pH value 8.5) in, setting is not added Bacterium plus inactivated bacteria control, 3 repetitions of each processing.It is inoculated with logarithmic phase CZ1701 bacterium solution, makes bacterium solution initial OD600Value is 0.1, is connect It is cultivated 48 hours after kind in 35 DEG C, 200rpm, culture solution is centrifuged 10min through 12 000rpm, takes supernatant.Ammoniacal nitrogen is used and is received Family name's reagent photometry measures ammonia nitrogen concentration.The result shows that: after 48 hours, bacterium solution OD600Value reaches 1.8, is inoculated with CZ1701 bacterial strain Ammoniacal nitrogen degradation rate be 88.4%.
3 CZ1701 bacterial strain of embodiment takes off nitrate nitrogen ability
Using nitrate as culture medium (sodium citrate 5g, potassium nitrate 1.2g, potassium dihydrogen phosphate 1g, the phosphoric acid hydrogen of only nitrogen source Dipotassium 0.1g, magnesium sulfate 0.2g, ferrous sulfate 0.03g, distilled water 1000mL, pH value 8.5) in, bacterium plus inactivated bacteria is not added in setting Control, 3 repetitions of each processing.It is inoculated with logarithmic phase CZ1701 bacterium solution, makes bacterium solution initial OD600Value be 0.1, after inoculation 35 DEG C, 200rpm is cultivated 48 hours, and culture solution is centrifuged 10min through 12 000rpm, takes supernatant.Supernatant uses ultraviolet point of Pbenoldisulfonic Acid Light photometry measures nitrate.The result shows that: after 48 hours, bacterium solution OD600Value reaches 1.7, is inoculated with the nitre of CZ1701 bacterial strain State nitrogen concentration 170.17mg/L is down to 39.27mg/L, degradation rate 76.9%.
4 CZ1701 bacterial strain of embodiment takes off nitrite nitrogen ability
Using nitrite as culture medium (sodium citrate 5g, sodium nitrite 0.5g, the potassium dihydrogen phosphate 1g, phosphorus of only nitrogen source Sour hydrogen dipotassium 1g, magnesium sulfate 0.2g, ferrous sulfate 0.03g, distilled water 1000mL.PH value 8.5) in, bacterium plus inactivation is not added in setting Bacterium control, 3 repetitions of each processing.It is inoculated with logarithmic phase CZ1701 bacterium solution, makes bacterium solution initial OD600Value is 0.1,35 after inoculation DEG C, 200rpm cultivate 48 hours, culture solution through 12 000rpm be centrifuged 10min, take supernatant.Supernatant uses N- (1- naphthalene)- Ethylenediamine photometry measures nitrite nitrogen concentration.The result shows that: after 48 hours, bacterium solution OD600Value is 1.2, is inoculated with CZ1701 bacterial strain Nitrite nitrogen concentration be down to 19.52mg/L, degradation rate 81.8% from 106.94mg/L.
Degradation of the 5 CZ1701 bacterial strain of embodiment to high ammonia nitrogen pig-farm wastewater
The wastewater sample for acquiring the high ammonia nitrogen of certain pig breeding farm adjusts pH 8.5 with NaOH, and loading amount 1L is in the triangular flask of 3L In, 121 DEG C of sterilizing 30mins, the content of various forms nitrogen and COD COD see the table below in piggery wastewater after sterilizing:
With 5% inoculum concentration, thallus is followed by triangular flask with sterile water washing, and 35 DEG C, 200rpm culture.It is taken after 6 days Sample, sample measure NH in supernatant after 10000rpm centrifugation 5mins removes thallus3-N、NO3-N、NO2- N, total N (TN) and COD Content, and calculate degradation rate.Setting plus viable bacteria processing group, and dead bacterium control group, every group of 3 Duplicate Samples, test result is as follows Shown in table:
As seen from the above table, compared with adding dead bacterium control group, it is inoculated with CZ1701 viable bacteria processing group, is had very to various forms nitrogen High degradation rate, to NH3-N、NO3-N、NO2The degradation rate of-N and total N (TN) respectively reach 89.1%, 53.2%, 66.7% With 86.6%.In addition, bacterial strain also has the ability of removal COD, compared with the control group, the degradation rate of COD is 77.6%.
<110>: South-Center University For Nationalities
<120>: the acinetobacter calcoaceticus CZ1701 bacterial strain and application thereof of one plant of degradation different shape nitrogen
<160>: 1
<210>: 1
<211>: 1529
<212>: DNA
<213>: acinetobacter calcoaceticus (Acinetobacter sp.)
<400>: 1
agagtttgat cctggctcag attgaacgct ggcggcaggc ttaacacatg caagtcgagc 60
gggggaaggt agcttgctac cggacctagc ggcggacggg tgagtaatgc ttaggaatct 120
gcctattagt gggggacaac atctcgaaag ggatgctaat accgcatacg tcctacggga 180
gaaagcaggg gatcttcgga ccttgcgcta atagatgagc ctaagtcgga ttagctagtt 240
ggtggggtaa aggcctacca aggcgacgat ctgtagcggg tctgagagga tgatccgcca 300
cactgggact gagacacggc ccagactcct acgggaggca gcagtgggga atattggaca 360
atggggggaa ccctgatcca gccatgccgc gtgtgtgaag aaggccttat ggttgtaaag 420
cactttaagc gaggaggagg ctactttagt taatacctag agatagtgga cgttactcgc 480
agaataagca ccggctaact ctgtgccagc agccgcggta atacagaggg tgcgagcgtt 540
aatcggattt actgggcgta aagcgtgcgt aggcggctta ttaagtcgga tgtgaaatcc 600
ccgagcttaa cttgggaatt gcattcgata ctggtgagct agagtatggg agaggatggt 660
agaattccag gtgtagcggt gaaatgcgta gagatctgga ggaataccga tggcgaaggc 720
agccatctgg cctaatactg acgctgaggt acgaaagcat ggggagcaaa caggattaga 780
taccctggta gtccatgccg taaacgatgt ctactagccg ttggggcctt tgaggcttta 840
gtggcgcagc taacgcgata agtagaccgc ctggggagta cggtcgcaag actaaaactc 900
aaatgaattg acgggggccc gcacaagcgg tggagcatgt ggtttaattc gatgcaacgc 960
gaagaacctt acctggcctt gacatactag aaactttcca gagatggatt ggtgccttcg 1020
ggaatctaga tacaggtgct gcatggctgt cgtcagctcg tgtcgtgaga tgttgggtta 1080
agtcccgcaa cgagcgcaac ccttttcctt acttgccagc atttcggatg ggaactttaa 1140
ggatactgcc agtgacaaac tggaggaagg cggggacgac gtcaagtcat catggccctt 1200
acggccaggg ctacacacgt gctacaatgg tcggtacaaa gggttgctac acagcgatgt 1260
gatgctaatc tcaaaaagcc gatcgtagtc cggattggag tctgcaactc gactccatga 1320
agtcggaatc gctagtaatc gcggatcaga atgccgcggt gagtacgttc ccgggccttg 1380
tacacaccgc ccgtcacacg atgggagttt gttgcaccag aagtagctag cctaactgca 1440
aagagggcgg ttaccacggt gtggccgatg actggggtga agtcgtaaca aggtagccgt 1500
aggggaacct gcggctggat cacctcctt 1529

Claims (3)

1. acinetobacter calcoaceticus (Acinetobacter sp.) CZ1701 bacterial strain of one plant of degradation different shape nitrogen, the bacterial strain are preserved in China typical culture collection center, address: China, Wuhan, Wuhan University；Postcode 430072, the deposit date is 2016 12 The moon 27, deposit number are as follows: CCTCC NO:M2016788, the 16S rDNA sequence of the bacterial strain is as shown in SEQ ID NO.1.
2. application of the acinetobacter calcoaceticus CZ1701 bacterial strain described in claim 1 in degradation high-concentration ammonia nitrogenous wastewater.
3. purposes according to claim 2, it is characterised in that: the high-concentration ammonia nitrogenous wastewater is useless for farm's excrement pond Water.
CN201710186658.XA 2017-03-27 2017-03-27 The acinetobacter calcoaceticus CZ1701 bacterial strain and application thereof of one plant of degradation different shape nitrogen Expired - Fee Related CN106929448B (en)

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CN108148790A (en) * 2018-03-07 2018-06-12 苏州顶裕节能设备有限公司 A kind of mixed bacteria, carrier and the method for livestock and poultry biology dejection degradation
CN115261253B (en) * 2022-05-16 2023-08-22 湖南农业大学 Acinetobacter radioresistant and product and application thereof

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CN103756947B (en) * 2014-02-17 2015-09-16 武汉科技大学 The preparation method of a kind of acinetobacter calcoaceticus and microbial inoculum thereof
CN105331552B (en) * 2014-08-13 2018-09-28 广州中国科学院先进技术研究所 One plant of efficient denitrification acinetobacter calcoaceticus novel species and its application
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