CN115992058A - Raoultella ornithinolytica PN-1, and culture method and application thereof - Google Patents
Raoultella ornithinolytica PN-1, and culture method and application thereof Download PDFInfo
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- 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
- Y02W10/00—Technologies for wastewater treatment
- Y02W10/10—Biological treatment of water, waste water, or sewage
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Abstract
The invention provides a Raoultella ornithinolytica PN-1, a culture method and application thereof, wherein the Raoultella ornithinolytica PN-1 is Raoultella ornithinolytica @Raoultella ornithinolytica) PN-1 is preserved in China center for type culture Collection (China, university of Wuhan, china, with a preservation number of CCTCC NO: M20211429) at 11 and 16 of 2021; the Raoultella ornithinolytica PN-1 is continuously cultured in LB liquid medium under the conditions of 20 ℃ and 180 r/minCulturing for 36h; the application of the Raoultella ornithinolytica PN-1 in the regulation of the culture water body at the low temperature of 10-20 ℃. The Raoultella ornithinolytica PN-1 can efficiently degrade total phosphorus and nitrate in a culture water body under a low-temperature and aerobic environment, has no toxic or harmful effect on aquatic economic animals, can be recycled for agricultural production links such as fertilization, and can be applied to mass production to efficiently improve the environment of the culture water body under a low-temperature condition.
Description
Technical Field
The invention relates to the technical field of microorganisms, in particular to a Raoultella ornithinolytica PN-1, a culture method and application thereof.
Background
The water environment is an ecological environment in which aquatic economic animals live. The too high nitrogen and phosphorus content in the culture water body not only can cause a large number of death of aquatic economic animals in the culture process, but also has a relatively close relationship with the too high nitrogen and phosphorus content of the culture tail water due to the high eutrophication of the natural water body environment.
The traditional biological denitrification and dephosphorization technology needs to consider the influence of the competition and intergrowth relation of environmental factors and microbial communities on the growth of microorganisms, and has great restriction on the environment. The denitrifying phosphorus accumulating bacteria are an emerging compound nitrogen and phosphorus reduction functional flora, and can improve the utilization efficiency of carbon sources in the nitrogen and phosphorus reduction process of sewage and wastewater. In addition, the latitude span of the land in China is larger, the average water temperature in winter in many areas can only be maintained at about 10 ℃, and the nitrogen and phosphorus removal efficiency of the traditional biological nitrogen and phosphorus removal technology applied to the low-temperature condition can be obviously reduced. Therefore, the denitrification and dephosphorization technology in the low-temperature environment is very important to solve the problem of deterioration of the aquaculture water environment in China.
In recent years, the denitrification phosphorus accumulating bacteria denitrification and phosphorus reduction technology based on normal temperature conditions has good application prospect in low-carbon sewage treatment, but has fewer research and development applications on cold-philic denitrification phosphorus accumulating bacteria for aquaculture. Therefore, there is a strong need to develop a psychrophilic denitrifying phosphorus accumulating bacterium for aquaculture that is harmless to aquatic economic animals.
The main differences and advantages of the ornithinolytic Raoultella PN-1 disclosed by the present published patent (patent numbers CN201210158648.2, CN201210158468.4 and 201910759615.5) are as follows:
(1) The inoculum size of the ornithinolytic Raoultella PN-1 is obviously lower than that of the existing strain, and the strain expansion and culture use cost can be saved.
(2) The ornithinolytic Raoult bacteria PN-1 is separated from the aquatic economic animal breeding environment, is applied to the aquatic economic animal breeding water body, has no toxic or harmful effect on the aquatic economic animal, and can be widely applied to more aquatic economic animal breeding scenes.
Disclosure of Invention
The invention aims to solve the technical problem of providing the ornithine-degrading Raoult bacteria PN-1, the culture method and the application thereof, wherein the strain belongs to aerobic heterotrophic bacteria, can degrade total phosphorus and nitrate in a culture water body in a low-temperature environment, has no toxic or harmful effect on aquatic economic animals, and has the advantages of wide application conditions, good degradation effect, high efficiency, no secondary pollution and the like.
The invention is realized in the following way:
the first object of the invention is to provide a Raoultella ornithinolytica PN-1, wherein the Raoultella ornithinolytica PN-1 is Raoultella ornithinolytica @Raoultella ornithinolytica) PN-1 is preserved in China center for type culture Collection (China, university of Wuhan, china, with a preservation number of CCTCC NO: M20211429) at 11 and 16 of 2021.
The second object of the invention is to provide a culture method of the Raoultella ornithinolytica PN-1, wherein the Raoultella ornithinolytica is prepared by the methodRaoultella ornithinolytica) PN-1 is continuously cultured in LB liquid medium for 36h under the conditions of 20 ℃ and 180 r/min.
The third object of the invention is to provide one of the applications of the Raoultella ornithinolytica PN-1, which is characterized in that: the Raoultella ornithinolytica is preparedRaoultella ornithinolytica) The application of PN-1 in the regulation of aquaculture water at the low temperature of 10-20 ℃.
Further, the Raoultella ornithinolytica is preparedRaoultella ornithinolytica) PN-1 can degrade the concentration of total phosphorus and nitrate in the culture water body at the low temperature of 10-20 ℃ and in an aerobic environment.
Further, the Raoultella ornithinolytica is preparedRaoultella ornithinolytica) PN-1 degrades total phosphorus and total phosphorus in aquaculture water under low temperature of 10-20 ℃ and aerobic environmentThe inoculation amount of the Raoultella ornithinolytica PN-1 is 0.25% when nitrate is used.
The fourth object of the invention is to provide two applications of the Raoultella ornithinolytica PN-1, wherein the Raoultella ornithinolytica is prepared by the steps ofRaoultella ornithinolytica) PN-1 can be used for preparing a psychrophilic microecological degradation preparation of total phosphorus and nitrate in a culture water body.
The invention has the following advantages:
the ornithinolytic Raoulet bacteria PN-1 provided by the invention can degrade total phosphorus and nitrate in aquaculture wastewater under a low temperature condition, belongs to aerobic heterotrophic bacteria, can degrade total phosphorus and nitrate in aquaculture water under a low temperature environment, has no toxic or harmful effect on aquatic economic animals, and has the advantages of wide application condition, good degradation effect, high efficiency, no secondary pollution and the like. The recycling can be used for agricultural production links such as fertilization, and the large-scale production can be applied to improving the environment of the culture water body under the low-temperature condition.
Drawings
The invention will be further described with reference to examples of embodiments with reference to the accompanying drawings.
FIG. 1 is a phylogenetic tree based on Raoultella ornithinolytica PN-1 16S rDNA according to the present invention;
FIG. 2 shows the growth curve of Raoultella ornithinolytica PN-1 according to the present invention.
Detailed Description
The invention relates to Raoultella ornithinolytica PN-1, which is prepared from Raoultella ornithinolyticaRaoultella ornithinolytica) PN-1 is preserved in China center for type culture Collection (China, university of Wuhan, china, with a preservation number of CCTCC NO: M20211429) at 11 and 16 of 2021.
The invention also relates to a culture method of the Raoultella ornithinolytica PN-1, which is prepared by the following steps ofRaoultella ornithinolytica) PN-1 is continuously cultured in LB liquid medium for 36h under the conditions of 20 ℃ and 180 r/min.
The invention also relates to application of the Raoultella ornithinolytica PN-1, the solutionOrnithine Raoult fungusRaoultella ornithinolytica) The application of PN-1 in the regulation of aquaculture water at the low temperature of 10-20 ℃. Specifically, the Raoultella ornithinolytica PN-1 can degrade the total phosphorus and nitrate concentration in a culture water body at the low temperature of 10-20 ℃ and in an aerobic environment.
The Raoultella ornithinolytica is preparedRaoultella ornithinolytica) When PN-1 degrades total phosphorus and nitrate in the culture water body at the low temperature of 10-20 ℃ and in an aerobic environment, the inoculation amount of the Raoultella ornithinolytica PN-1 is 0.25%.
The invention relates to two applications of Raoultella ornithinolytica PN-1, which are prepared by the following steps ofRaoultella ornithinolytica) PN-1 can be used for preparing a psychrophilic microecological degradation preparation of total phosphorus and nitrate in a culture water body.
The technical scheme of the present invention will be clearly and completely described in connection with the following detailed description. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
Example 1 isolation screening of Raoultella ornithinolytica PN-1
1. Sampling: biological filler is collected from a tail water treatment pool of eel farms in the area of the Yanping of south Ping City of Fujian province and is stored at the temperature of-4 ℃ for standby. Placing into sterilized bottle containing sterile water, and treating with ultrasonic vibrator.
Enrichment culture: 10 mL sample pretreatment liquid is sucked and put into 100mL of EM culture medium (sterilized), a bottle mouth is sealed by cotton, and enrichment culture is carried out for 1-2d in a constant temperature shaking table under the condition of 12 ℃ and 150 r/min.
EM medium: beef extract 3g, peptone 10g, naCl 5g, H 2 O1L, pH 7.0-7.4, sterilizing at 121deg.C for 30min.
And (3) separating and purifying: after enrichment culture is finished, suckingTaking 150 mu L of enrichment culture solution, putting into 1.5 mL sterile water, uniformly mixing, and the like to prepare 10 -1 、10 -2 、10 -3 、10 -4 、10 -5 、10 -6 、10 -7 Bacterial suspensions with different dilution gradients are obtained. Three dilution factors (10) after 200. Mu.L were aspirated respectively -5 、10 -6 、10 -7 ) The gradient bacterial suspension was spread evenly on YG medium with sterile triangular glass spatula, each dilution factor (10 -5 、10 -6 、10 -7 ) Is repeatedly coated 3 times. The diluted and coated YG culture medium is placed in a biochemical incubator at 12 ℃ in an inverted mode, after 3d culture, colonies with obvious differences in characterization aspects such as colony morphology, colony size and the like are selected from the YG culture medium with different dilution factors, and streak separation is started on a new YG culture medium. Care was taken not to scratch the surface of the medium until a pure culture developed from individual microbial cells was obtained.
YG medium: yeast extract 1g, anhydrous glucose 1g, K 2 HPO 4 •3H 2 O 0.3931g，KH 2 PO 4 0.25g, agar 20g, mgSO 4 •7H 2 O 0.4094g，H 2 O1L, pH 7.0-7.2, sterilizing at 121deg.C for 30min.
Screening and preserving: and taking pure cultures obtained by separation and purification, respectively inoculating the pure cultures on a phosphorus-passing/phosphorus-limiting SM culture medium, culturing the pure cultures in an inversion manner for 3d at 12 ℃ in a biochemical incubator, and observing the color development condition of the strain to be detected on the phosphorus-passing/phosphorus-limiting SM culture medium. The strain which can show blue bacterial plaque on the phosphorus-passing/phosphorus-limiting SM culture medium is a primary screening strain. The obtained primary screening strain is subjected to liquid expansion culture, inoculated in DM culture medium with an inoculum size of 2 percent, a bottle mouth is sealed by cotton (an aerobic environment is kept), and the primary screening strain is cultured for 2-3d in a constant-temperature oscillator at the temperature of 12 ℃ and 180 r/min, and then subjected to low-temperature aerobic denitrification phosphorus-reduction domestication. Determination of OD of domesticated bacterial suspension 600 Value, select OD 600 Strains with larger values were subjected to secondary screening. Taking a proper amount of bacterial suspension acclimatized by the strain to be tested, centrifuging in a centrifuge tube at 12 ℃ and 10000 rpm for 10min in a high-speed centrifuge, and measuring according to ultraviolet spectrophotometry and ammonium molybdate spectrophotometryAnd finally selecting the strain with high efficiency as a target strain.
MOPS-Glu superphosphorus/phosphorus limited SM Medium combination:
(1) MOPS-Glu mixture (100 mL): MOPS 8.372g and Tricine 0.717g dissolved in distilled water<38 mL), 10 mol.L -1 KOH was used to adjust the pH of the solution to 7.4 overall, distilled water was added to bring the total volume of the solution to 44 mL, freshly prepared 0.0183 mol.L -1 FeSO 4 •7H 2 O solution 1 mL, 1.9 mol.L were added sequentially -1 NH 4 Cl solution 5mL, 0.276 mol.L -1 K 2 SO 4 Solution 1 mL, 5×10 -4 mol•L -1 CaCl 2 Solution 1 mL, 1.127mol. L -1 MgCl 2 •6H 2 O solution 1 mL, 5mol -1 NaCl solution 10 mL, trace element solution 1 mL, distilled water 36 mL, anhydrous glucose 0.1g.
(2) Phosphorus/phosphorus limited SM medium (500 mL each of phosphorus/phosphorus limited): 50mL of Glu-MOPS mixture was taken in a 500mL Erlenmeyer flask. When preparing the phosphorus SM culture medium, K is added in sequence 2 HPO 4 •3H 2 O 0.2282g、1% VB 1 5mL of solution, 0.025g of X-Pi, 440mL of distilled water, 10g of agar and pH 7.2; when preparing the phosphorus-limited SM culture medium, sequentially adding 1% VB 1 5mL of solution, 0.025g of X-Pi, 440mL of distilled water, 10g of agar and pH 7.2. After autoclaving at 121 ℃ for 30min, taking out, cooling to 60 ℃, shaking uniformly and pouring into a flat plate.
DM medium: anhydrous glucose 1.2851g, KH 2 PO 4 0.0659g，KNO 3 0.722g，MgSO 4 •7H 2 O 0.2g，H 2 O1L, 2mL of trace element solution.
Trace element solution: (NH) 4 ) 6 Mo 7 O 24 •4H 2 O 0.0956g，H 3 BO 3 0.62g，CaCl 2 0.18g，CuSO 4 0.06g，MnCl 2 •4H 2 O 0.6289g，ZnSO 4 •7H 2 O 0.1246g，H 2 O 1L。
Example 2 identification of Raoultella ornithinolytica PN-1
The purified bacteria were used to extract genomic DNA from the strain using TaKaRa MiniBEST Bacteria Genomic DNA Extraction Kit Ver.3.0. The PCR amplification was performed using a 2X TsingKE Master Mix system with the universal primers 27F (5'-AGAGTTTGATCCTGGCTCAG-3') of the 16S rDNA sequence (SEQ ID NO: 1) and 1492R (5'-GGTTACCTTGTTACGACTT-3') (SEQ ID NO: 2) as PCR reaction primers. The PCR reaction system is as follows: 1. Mu.L of genomic DNA, 2X TsingKE Master Mix. Mu.L, 1. Mu.L of 27F Primer (10. Mu.M), 1. Mu.L of 1492R Primer (10. Mu.M), dH 2 O22. Mu.L. The PCR reaction conditions were: pre-denaturation at 94℃for 10min,30 cycles (94℃for 30s, 55℃for 30s, 72℃for 1.5 min) and extension at 72℃for 10min. The PCR product obtained was purified and then sent to the engineering and bioengineering (Shanghai) Co.Ltd for sequencing.
The 16S rDNA sequence of strain PN-1 (shown as SEQ ID No: 3) was uploaded to the NCBI database and BLAST homologous sequence alignment (https:// BLAST. NCBI. Lm. Nih. Gov/BLAST. Cgi) was performed, and the results showed that strain PN-1 was identical to that of strainRaoultella sp. Strain GZ2 (GenBank accession number: MK 999961.1) andRaoultella ornithinolyticathe similarity of the strain FJG3 (GenBank accession number: MG 516115.1) is up to 99.93%. The construction of phylogenetic tree (Neighbor-training method) using MEGA7 is shown in FIG. 1, and the results show that the strain PN-1 and NCBI database are availableRaoultella ornithinolyticaThe strain SY15 16S rDNA sequence (GenBank accession number: MH 259979.1) is in the same branch of the tree and is closest to it. The 16S rDNA sequence of strain PN-1 was uploaded to the EzBioCloud database (https:// www.ezbiocloud.net /), and the results showed that the strains PN-1 and 16S rRNA sequence were in the databaseRaoultella ornithinolyticaThe similarity of the strain JCM 6096 (Access: AJ 251467) is up to 99.03%. And judging that the bacterial strain PN-1 is Raoultella ornithinolytica by combining the identification and analysis resultsRaoultella ornithinolytica) And named as Raoultella ornithinolytica @ and the likeRaoultella ornithinolytica）PN-1。
Example 3 growth curve of Raoultella ornithinolytica PN-1
Picking sheetPure strain PN-1 is inoculated in sterilized LB liquid medium, and cultured overnight in a constant temperature shaker at 20 ℃ and 180 r/min to prepare seed liquid for later use. 18X 180 test tubes were prepared, each tube was filled with 15mL of LB liquid medium, and autoclaved at 121℃for 30min after sealing with cotton stoppers. Transferring seed solution into sub-packaged test tubes (200 μL of seed solution per tube), shake culturing the inoculated test tubes at 20deg.C and 180 r/min, and culturing target strains to 0, 1, 2.5, 4, 6, 8, 10, 12.5, 15, 17.5, 20, 22.5, 25, 29, 33, 37, 41, 47 and 53 hr, taking one LB culture solution, immediately measuring absorbance (OD) of the LB culture solution at 600nm wavelength with unvaccinated LB liquid culture medium as reference 600 ). The data at each time was repeated for 3 measurements on the samples to reduce test errors. At OD 600 The growth curve of strain PN-1 was plotted on the ordinate and the incubation time on the abscissa (FIG. 2).
As can be seen from FIG. 2, the strain PN-1 is in a slow growth period within 0-3 hours and grows more slowly; after 3 hours, the curve grows in a logarithmic form, and the growth is quicker; after 25 hours, the growth rate gradually decreases, the growth period starts to enter a stable growth period, and reaches the peak value of the growth curve of the bacterial strain PN-1 in 36 hours, so that the total number of bacterial cells tends to be stable; after 47h growth enters the decay phase. The results showed that the optimal cultivation time for strain PN-1 was 36h. Therefore, when the strain PN-1 seed solution is cultured and prepared, the selection of the culture time should be paid attention to, and the culture is preserved in time.
Example 4 determination of Total phosphorus and nitrate degradation Effect of Raoultella ornithinolytica PN-1 on aquaculture Water quality at different inoculum sizes
Single pure bacterial strain PN-1 is selected and inoculated into sterilized LB liquid medium, and amplified and cultured for 36 hours at 20 ℃ and 180 r/min in a constant temperature oscillator to prepare seed liquid. 8 treatment groups were set, the inoculation amount was adjusted to be 0.25%, 0.5%, 0.75%, 1%, 2%, 3%, 4%, 5% by changing the amount of seed solution of the strain PN-1 inoculated into 1L eel culture water, and the culture was carried out in a constant temperature shaker at 15℃with the addition of an anhydrous glucose adjusting solution C/N ratio of 4 and 180 r/min, with 3 identical replicates for each treatment group. During the test, samples of each treatment group were collected immediately after 48 hours of culture, centrifuged at 15℃and 10000 rpm in a high-speed centrifuge for 10 minutes, and the supernatant was taken to measure the degradation rate of total phosphorus and nitrate, and the measurement results are shown in Table 1 and Table 2.
According to the measurement result, when the inoculation amount interval is 0.25% -2%, the degradation rate of the bacterial strain PN-1 on the total phosphorus in the culture water quality is above 70%; when the inoculation amount is 0.25% and 0.5%, the degradation rate of the bacterial strain PN-1 on nitrate nitrogen in the culture water quality is more than 40%. Wherein, the degradation effect of the bacterial strain PN-1 on the total phosphorus is optimal when the inoculation amount is 0.75%, and the degradation rate can reach 77.03%; the degradation effect of the bacterial strain PN-1 on nitrate nitrogen is optimal when the inoculation amount is 0.25%, and the degradation rate can reach 60.71%.
The result shows that the higher inoculation amount can inhibit the process of degrading total phosphorus and nitrate nitrogen in the culture water by the bacterial strain PN-1. In terms of inoculum size selection, the degradation effect of the strain PN-1 on total phosphorus was optimal at an inoculum size of 0.75%, but the degradation rate was not much different from that at an inoculum size of 0.25%. Therefore, the bacterial strain PN-1 is comprehensively considered to be applied to synchronous denitrification and phosphorus reduction of the aquaculture water quality, and the optimal inoculation amount is preferably selected to be 0.25%.
TABLE 1 Total phosphorus degradation effects of Raoultella ornithinolytica PN-1 on aquaculture water quality at different inoculum sizes
TABLE 2 degradation effects of Raoultella ornithinolytica PN-1 on nitrate nitrogen in aquaculture Water at different inoculum sizes
Example 5 determination of Total phosphorus and nitrate degradation Effect of Raoultella ornithinolytica PN-1 on quality of cultured Water at different temperatures and verification of psychrophilic Property
Single pure bacterial strain PN-1 is selected and inoculated into sterilized LB liquid medium, and amplified and cultured for 36 hours at 20 ℃ and 180 r/min in a constant temperature oscillator to prepare seed liquid. Setting 6 treatment groups, respectively changing the environmental temperature to 10 ℃,15 ℃,20 ℃,25 ℃,30 ℃ and 35 ℃, adding an anhydrous glucose regulating solution with a C/N ratio of 4, taking 2.5mL of seed liquid of the bacterial strain PN-1, inoculating the seed liquid into 1L of eel culture water quality, culturing in a constant temperature oscillator at 180 r/min, and carrying out 3 identical repetitions on each treatment group. During the test, samples of each treatment group were collected immediately after 48 hours of culture, centrifuged at 15℃and 10000 rpm in a high-speed centrifuge for 10 minutes, and the supernatant was taken to measure the degradation rate of total phosphorus and nitrate, and the measurement results are shown in Table 3 and Table 4.
According to the measurement result, when the temperature range is 10-25 ℃, the degradation rate of the bacterial strain PN-1 on the total phosphorus in the aquaculture water is over 80 percent, and the degradation rate of the total phosphorus after 25 ℃ is obviously attenuated; when the temperature range is 10-20 ℃, the degradation rate of the bacterial strain PN-1 on nitrate nitrogen in the culture water quality is over 30 percent. Wherein, the degradation effect of the bacterial strain PN-1 on the total phosphorus and the nitrate nitrogen is optimal at the temperature of 15 ℃, and the degradation rate can reach 86.88 percent and 43.92 percent respectively.
The results show that the strain PN-1 can have a certain degradation effect on total phosphorus and nitrate nitrogen in aquaculture water under the low temperature condition (10-20 ℃), and the strain PN-1 also proves that the ornithinolytic Raoult strain PN-1 is a psychrophilic strain and has a certain application value in the aspect of low-temperature synchronous denitrification and phosphorus reduction of aquaculture wastewater.
TABLE 3 Total phosphorus degradation effects of Raoultella ornithinolytica PN-1 on aquaculture water at different temperatures
TABLE 4 degradation effects of Raoultella ornithinolytica PN-1 on nitrate nitrogen in aquaculture Water at different temperatures
Example 6 safety detection of Raoultella ornithinolytica PN-1 in aquaculture applications
Single pure bacterial strain PN-1 is selected and inoculated into sterilized LB liquid medium, and amplified and cultured for 36 hours at 20 ℃ and 180 r/min in a constant temperature oscillator to prepare seed liquid. At 2.53×10 4 Inoculating cfu/mL to a culture wastewater treatment subsystem of a circulating water Australia eel culture system (two moving bed biomembrane reaction tanks are provided, and the effective water volume of each tank for treating culture wastewater is 4.8 m) 3 ) Anhydrous glucose is added as a carbon source, and the circulating water Australian eel culture system normally feeds and manages the feed according to the feed rate of 2.0 percent for 60 days.
The circulating water can ensure that the Anguilla anguilla grows healthily during the culture period of the Anguilla anguilla, and no morbidity and mortality exist. The results show that the ornithinolytic Raoult bacteria PN-1 has relatively stable safety in the denitrification and phosphorus reduction process of the culture of the Australian eel.
Ornithine-decomposing Raoult fungusRaoultella ornithinolytica) Although it has no toxic effect on aquatic economic animals. But of high concentration (greater than 10 7 cfu/mL) may convert histidine in the breeding water to histamine, and if applied at high concentrations to a factory high density breeding water, may cause a very small number of fish highly sensitive to histamine concentration such as mackerel @Scombridae) And the bamboo-sword fish family%Scomberesocidae) Mackerel syndrome (scombroid syndrome) occurs in the fish species. Therefore, if the strain PN-1 provided by the invention is applied to industrial high-density culture water bodies of mackerel and saury, the concentration of the strain PN-1 does not exceed the normal release concentration (lower than 10 5 cfu/mL).
In conclusion, the ornithinolytic Raouli bacteria PN-1 capable of degrading total phosphorus and nitrate in aquaculture wastewater under low temperature conditions belongs to aerobic heterotrophic bacteria, can degrade total phosphorus and nitrate in aquaculture water under low temperature conditions, has no toxic or harmful effects on aquatic economic animals, and has the advantages of wide application conditions, good degradation effect, high efficiency, no secondary pollution and the like. The recycling can be used for agricultural production links such as fertilization, and the large-scale production can be applied to improving the environment of the culture water body under the low-temperature condition.
While specific embodiments of the invention have been described above, it will be appreciated by those skilled in the art that the specific embodiments described are illustrative only and not intended to limit the scope of the invention, and that equivalent modifications and variations of the invention in light of the spirit of the invention will be covered by the claims of the present invention.
Claims (6)
1. An ornithinolytic Raoultella PN-1, characterized in that: the Raoultella ornithinolytica PN-1 is Raoultella ornithinolytica @Raoultella ornithinolytica) PN-1 is preserved in China center for type culture Collection (China, university of Wuhan, china, with a preservation number of CCTCC NO: M20211429) at 11 and 16 of 2021.
2. The culture method of the Raoultella ornithinolytica PN-1, according to claim 1, wherein the culture method comprises the following steps: the Raoultella ornithinolytica is preparedRaoultella ornithinolytica) PN-1 is continuously cultured in LB liquid medium for 36h under the conditions of 20 ℃ and 180 r/min.
3. The use of raleigh-degrading bacterium PN-1 according to claim 1, characterized in that: the Raoultella ornithinolytica is preparedRaoultella ornithinolytica) The application of PN-1 in the regulation of aquaculture water at the low temperature of 10-20 ℃.
4. The use of a Raoultella ornithinolytica PN-1 as claimed in claim 3, wherein: the Raoultella ornithinolytica is preparedRaoultella ornithinolytica) PN-1 can degrade the concentration of total phosphorus and nitrate in the culture water body at the low temperature of 10-20 ℃ and in an aerobic environment.
5. The use of the Raoultella ornithinolytica PN-1 according to claim 4, wherein the use comprises the following steps: the Raoultella ornithinolytica is preparedRaoultella ornithinolytica) When PN-1 degrades total phosphorus and nitrate in the culture water body at the low temperature of 10-20 ℃ and in an aerobic environment, the inoculation amount of the Raoultella ornithinolytica PN-1 is 0.25%.
6. The use of raleigh-degrading bacterium PN-1 according to claim 1, characterized in that: the Raoultella ornithinolytica is preparedRaoultella ornithinolytica) PN-1 can be used for preparing a psychrophilic microecological degradation preparation of total phosphorus and nitrate in a culture water body.
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| CN102391961A (en) * | 2011-11-02 | 2012-03-28 | 东华大学 | Raoultella ornithinolytica N-4 bacterial strains and preparation and application thereof |
| CN105861356A (en) * | 2016-03-31 | 2016-08-17 | 浙江工业大学 | Raoultella sp. pan22x and application thereof |
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