CN106929448A - One plant of acinetobacter calcoaceticus CZ1701 bacterial strain and application thereof of degraded different shape nitrogen - Google Patents

One plant of acinetobacter calcoaceticus CZ1701 bacterial strain and application thereof of degraded different shape nitrogen Download PDF

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Publication number
CN106929448A
CN106929448A CN201710186658.XA CN201710186658A CN106929448A CN 106929448 A CN106929448 A CN 106929448A CN 201710186658 A CN201710186658 A CN 201710186658A CN 106929448 A CN106929448 A CN 106929448A
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bacterial strain
nitrogen
plant
acinetobacter calcoaceticus
cod
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CN106929448B (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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    • 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
    • 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
    • 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

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  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • Microbiology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Genetics & Genomics (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Biotechnology (AREA)
  • Water Supply & Treatment (AREA)
  • Biomedical Technology (AREA)
  • Virology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Medicinal Chemistry (AREA)
  • Environmental & Geological Engineering (AREA)
  • Hydrology & Water Resources (AREA)
  • Biodiversity & Conservation Biology (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Purification Treatments By Anaerobic Or Anaerobic And Aerobic Bacteria Or Animals (AREA)

Abstract

The invention provides a kind of denitrogenation bacterial strain of degradable different shape nitrogen, the bacterial strain is named as CZ1701, belongs to a kind of acinetobacter calcoaceticus, and the bacterial strain is preserved in China typical culture collection center, and preservation date is on December 27th, 2016, and deposit number is：CCTCC NO：M2016788.CZ1701 bacterial strains provided by the present invention have degradation rate very high for various form nitrogen, and it is to NH3‑N、NO3‑N、NO2The degradation rate of N and total N (TN) respectively reaches 89.1%, 53.2%, 66.7% and 86.6%.Additionally, bacterial strain also has the ability of removal COD, the degradation rate of its COD is 77.6%.Therefore, CZ1701 bacterial strains provided by the present invention have the advantages that Acclimation temperature scope it is wide, can various forms of efficient degradation simultaneously nitrogen and COD, be particularly well-suited in the treatment of degrading high concentration ammonia nitrogen waste water such as plant's excrement and urine water.

Description

One plant of acinetobacter calcoaceticus CZ1701 bacterial strain and application thereof of degraded different shape nitrogen
Technical field
The present invention relates to a kind of acinetobacter calcoaceticus new strains, the nitrogen of degradable different valence state, the more particularly to bacterial strain it is aerobic Denitrification and the process performance to piggery wastewater, belong to microbial technology field.
Background technology
Environmental problem is protruded all the more since new century, and wherein the pollution of water body is such as：Eutrophication, wawter bloom, red tide etc. take place frequently, Arouse widespread concern.The basic reason of body eutrophication is mainly that nitrogen, phosphorus element content are too high to cause water body table Layer cyanobacteria and algae excessive propagation.Therefore, the removal of Water element, for environmental protection, develops the economy and ensures health All have great importance.With the large-scale development of livestock and poultry breeding industry, pig farm excrement stain disease is caused sternly to surrounding environment Heavily contaminated, excrement dirt enters lake waters, can cause body eutrophication, such as permeates the ground, can make nitrate nitrogen in underground water or Nitrite nitrogen concentration increases, and toxic component increases, and causes water quality deterioration, jeopardizes the water quality of periphery domestic water.For pig farm excrement The features such as stain disease has ammonia nitrogen high, low C/N than, the water quality that pH value is higher, biodegradability is poor, (biology is removed microorganism treatment Nitrogen) it is to remove one of polluted by nitrogen most efficient method in water.Microorganism can be the pollutant ammonium nitrogen of most serious in water body and nitre Hydrochlorate etc. is converted into the nitrogen source of own growth, gaseous state nitride (nitrogen and nitrous oxide) etc., so as to reduce nitrogen residue.
Therefore, separation screening can remove ammonia nitrogen high, and have one to other valence state nitrogen sources and COD under the conditions of pH value is higher Fixed removal effect, and its biological characteristics and the application in the improvement of piggery waste contaminated wastewater are studied, can be controlled for microorganism Reason high ammonia-nitrogen wastewater provides the resource of high-quality, and the biological treating for waste water such as poultry-dung dirts is significant.
The content of the invention
Object of the present invention is to provide a kind of denitrogenation bacterial strain of degradable different shape nitrogen, the bacterial strain 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, and the bacterial strain is named as CZ1701, belongs to a kind of acinetobacter calcoaceticus (Acinetobacter sp.), the bacterial strain is preserved in China typical culture collection center, address：China, Wuhan, Wuhan is big Learn；Postcode 430072, preservation date is on December 27th, 2016, and deposit number is：CCTCC NO：M2016788；The bacterial strain 16S rDNA sequences are as shown in SEQ ID NO.1.
CZ1701 bacterial strains are identified as to be isolated from Hubei Province's pig-farm wastewater pollution discharge pond mud sample Acinetobacter sp..The form and physiological and biochemical property of the bacterial strain be：Bacterium colony is observed on LB solid mediums in milky white Color, neat in edge, smooth moistening；Microscopy observation bacterial strain is Gram-negative, shaft-like, without gemma.
By inoculation to 5ml LB nutrient solutions (peptone 10.0g, yeast extract 5.0g, NaCl 10.0g, H2O1000mL), 37 DEG C, 200rpm/min cultivate 24 hours, 10000rpm/min centrifugation 1min, collects thalline in 1.5ml be centrifuged Guan Zhong, bacterial genomes are extracted using CTAB methods.Use 16S rDNA universal primer (sense primers:5′- AGAGTTTGATCCTGGCTCAG-3′；Anti-sense primer:5 '-AAGGAGGTGATCCAGCCGCA-3 ') enter performing PCR amplification, amplification Product 16S rDNA sequences listed with GenBank after sequencing carry out nucleotide sequence homology and compare analysis, and motionless The 16S rDNA sequence similarities of bacillus (Acinetobacter) reach 99%, are accredited as this category.
Compared with prior art, the present invention has advantages below：Acinetobacter calcoaceticus CZ1701 bacterial strains pair provided by the present invention There is degradation rate very high in various form nitrogen, it is to NH3-N、NO3-N、NO2The degradation rate of-N and total N (TN) is respectively reached 89.1%th, 53.2%, 66.7% and 86.6%.Additionally, bacterial strain also has the ability of removal COD, compared with control group, its COD Degradation rate be 77.6%.Therefore, CZ1701 bacterial strains provided by the present invention are compared with known denitrogenation acinetobacter calcoaceticus, with temperature Degree wide adaptation range, energy are while the nitrogen and the advantage of COD of the various forms of efficient degradation, are particularly well-suited to degrading high concentration ammonia nitrogen and give up In the treatment of water such as plant's excrement and urine water.
Specific embodiment
Detailed specific description is done to the present invention with reference to specific embodiment, but protection scope of the present invention not office It is limited to following examples.
Correlated performance in following examples to acinetobacter calcoaceticus CZ1701 bacterial strains provided by the present invention is studied.
Influence of the different temperatures of embodiment 1 to CZ1701 strain growths
Heterotrophic nitrification culture medium (sodium citrate 5g, ammonium sulfate 1.0g, potassium dihydrogen phosphate with ammoniacal nitrogen as 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 solutions, make bacterium solution initial OD600It is 0.1 to be worth, and is placed in concussion and cultivate in the shaking table of different temperatures, and growth feelings are detected after 48h Condition, 3 repetitions of this Setup Experiments.CZ1701 strain growth temperature ranges are very wide, can be grown in the range of 15 DEG C to 42 DEG C, OD in 48 hours600Value can reach more than 1.5, illustrate that CZ1701 strains adapt to ambient temperature capability stronger.
The de- ammoniacal nitrogen ability of the CZ1701 bacterial strains of embodiment 2
Heterotrophic nitrification culture medium (sodium citrate 5g, ammonium sulfate 1.0g, potassium dihydrogen phosphate with ammoniacal nitrogen as 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 with Bacterium, plus inactivated bacteria control, each treatment 3 repetitions.Inoculation logarithmic phase CZ1701 bacterium solutions, make bacterium solution initial OD600It is 0.1 to be worth, and is connect Cultivated 48 hours in 35 DEG C, 200rpm after kind, nutrient solution is centrifuged 10min through 12 000rpm, takes supernatant.Ammoniacal nitrogen is used and received Family name's reagent photometry determines ammonia nitrogen concentration.Result shows：After 48 hours, bacterium solution OD600Value reaches 1.8, is inoculated with CZ1701 bacterial strains Ammoniacal nitrogen be down to 39.27mg/L from 170.17mg/L, degradation rate is 88.4%.
The de- nitrate nitrogen ability of the CZ1701 bacterial strains of embodiment 3
Culture medium (sodium citrate 5g, potassium nitrate 1.2g, potassium dihydrogen phosphate 1g, phosphoric acid hydrogen with nitrate as only nitrogen source Dipotassium 0.1g, magnesium sulfate 0.2g, ferrous sulfate 0.03g, distilled water 1000mL, pH value 8.5) in, setting is not added with bacterium, adds inactivated bacteria Control, each 3 repetition for the treatment of.Inoculation logarithmic phase CZ1701 bacterium solutions, make bacterium solution initial OD600It is 0.1 to be worth, after inoculation 35 DEG C, 200rpm is cultivated 48 hours, and nutrient solution is centrifuged 10min through 12 000rpm, takes supernatant.Supernatant uses ultraviolet point of Pbenoldisulfonic Acid Light photometry determines nitrate.Result shows：After 48 hours, bacterium solution OD600Value reaches 1.7, is inoculated with the nitre of CZ1701 bacterial strains State nitrogen concentration 170.17mg/L is down to 39.27mg/L, and degradation rate is 76.9%.
The de- nitrite nitrogen ability of the CZ1701 bacterial strains of embodiment 4
Culture medium (sodium citrate 5g, natrium nitrosum 0.5g, potassium dihydrogen phosphate 1g, phosphorus with nitrite as only nitrogen source Sour hydrogen dipotassium 1g, magnesium sulfate 0.2g, ferrous sulfate 0.03g, distilled water 1000mL.PH value 8.5) in, setting is not added with bacterium, adds inactivation Bacterium compares, each 3 repetition for the treatment of.Inoculation logarithmic phase CZ1701 bacterium solutions, make bacterium solution initial OD600It is 0.1 to be worth, 35 after inoculation DEG C, 200rpm cultivate 48 hours, nutrient solution through 12 000rpm be centrifuged 10min, take supernatant.Supernatant using N- (1- naphthyls)- Ethylenediamine photometry determines nitrite nitrogen concentration.Result shows：After 48 hours, bacterium solution OD600It is 1.2 to be worth, and is inoculated with CZ1701 bacterial strains Nitrite nitrogen concentration be down to 19.52mg/L from 106.94mg/L, degradation rate is 81.8%.
Degradation of the CZ1701 bacterial strains of embodiment 5 to ammonia nitrogen pig-farm wastewater high
The wastewater sample of certain pig breeding farm ammonia nitrogen high is gathered, pH 8.5 is adjusted with NaOH, loading amount 1L is in the triangular flask of 3L In, 121 DEG C of sterilizing 30mins, the content of various form nitrogen and COD COD see the table below in piggery wastewater after sterilizing：
With 5% inoculum concentration, thalline is followed by triangular flask with aseptic water washing, 35 DEG C, 200rpm cultures.Taken after 6 days Sample, after sample removes thalline through 10000rpm centrifugations 5mins, determines NH in supernatant3-N、NO3-N、NO2- N, total N (TN) and COD Content, and calculate degradation rate.Setting plus viable bacteria treatment group, and dead bacterium control group, every group of 3 Duplicate Samples, test result are as follows Shown in table：
As seen from the above table, compared with dead bacterium control group is added, CZ1701 viable bacteria treatment groups are inoculated with, are had very to various form nitrogen Degradation rate high, it is to NH3-N、NO3-N、NO2The degradation rate of-N and total N (TN) respectively reaches 89.1%, 53.2%, 66.7% With 86.6%.Additionally, bacterial strain also has the ability of removal COD, compared with control group, the degradation rate of its COD is 77.6%.
<110>：South-Center University For Nationalities
<120>：One plant of acinetobacter calcoaceticus CZ1701 bacterial strain and application thereof of degraded 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. one plant of acinetobacter calcoaceticus (Acinetobacter sp.) CZ1701 bacterial strain of degraded different shape nitrogen, the bacterial strain is preserved in China typical culture collection center, address：China, Wuhan, Wuhan University；Postcode 430072, preservation date is 2016 12 The moon 27, deposit number is：CCTCC NO：M2016788, the 16S rDNA sequences of the bacterial strain are as shown in SEQ ID NO.1.
2. application of the acinetobacter calcoaceticus CZ1701 bacterial strains described in claim 1 in degrading high concentration ammonia nitrogen waste water.
3. purposes according to claim 2, it is characterised in that：Described high-concentration ammonia nitrogenous wastewater is useless for plant'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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Cited By (2)

* Cited by examiner, † Cited by third party
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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
CN115261253A (en) * 2022-05-16 2022-11-01 湖南农业大学 Radiation-resistant acinetobacter, and product and application thereof

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CN105586294A (en) * 2016-01-07 2016-05-18 温州大学 Acinetobacter and application of acinetobacter in removal of nitrogen and phosphorus from wastewater

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
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
CN115261253A (en) * 2022-05-16 2022-11-01 湖南农业大学 Radiation-resistant acinetobacter, and product and application thereof
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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