CN113846020A - Aspergillus flavus XZCY1805 without producing toxicity and application thereof - Google Patents
Aspergillus flavus XZCY1805 without producing toxicity and application thereof Download PDFInfo
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Abstract
The invention relates to a non-toxigenic aspergillus flavus strain and application thereof. The non-toxigenic aspergillus flavus XZCY1805 does not produce a plurality of toxins such as Aflatoxin (AFT), cyclopiazonic acid (CPA), aflatoxin (Aflatrem) and aflatoxin (ST), and is a biocontrol strain with high safety and strong competitive advantage. The non-toxigenic aspergillus flavus XZCY1805 has a strong toxigenic inhibition effect on toxigenic aspergillus flavus separated from a standard toxigenic aspergillus flavus strain 3.4408 and different peanut producing areas, and can remarkably reduce the contents of aflatoxin, cyclopiazonic acid, aflatoxin and variegated aspergillus chromous toxin in agricultural products such as peanuts and the like.
Description
Technical Field
The invention belongs to the technical field of microorganisms, and particularly relates to non-toxigenic aspergillus flavus XZCY1805 and application thereof.
Background
Aflatoxins (AFT) are secondary metabolites produced by Aflatoxins (Aspergillus flavus) and Aspergillus parasitism (Aspergillus parasiticus) and other fungi after infecting host plants, are mycotoxins discovered so far and have the strongest toxicity to polluted agricultural products, have acute and chronic toxicity, and can cause cancers, teratogenesis and mutation. The common aflatoxin species is mainly aflatoxin B1(AFB1) Aflatoxins B2(AFB2) Aflatoxin G1(AFG1) Aflatoxin G2(AFG2). Aflatoxin B with the highest toxicity and the strongest carcinogenicity1The toxicity of the traditional Chinese medicine is 10 times that of potassium cyanide and 68 times that of arsenic, the carcinogenicity of the traditional Chinese medicine is 10000 times that of sixty-six, and the traditional Chinese medicine is listed as a class I carcinogen by the International Agency for Research on Cancer (IARC) of World Health Organization (WHO). Besides aflatoxin, aspergillus flavus can also produce toxic secondary metabolites such as Cyclopiazonic acid (CPA), aflatoxin (Aflatrem), variegated aflatoxin (ST) and the like, thus threatening consumption safety and life health of people. CPA is an indole derivative that has toxic effects on humans and various animals, including liver degeneration and necrosis, myocardial injury, kidney injury, and the like. Aflatoxin is an indole diterpenoid mycotoxin, has tumorigenic properties and acute neurotoxic effects, and can cause animal tremor. The aflatoxin is an intermediate product in the later stage of the process of synthesizing the aflatoxin by the aspergillus flavus, and the toxicity and the carcinogenicity of the aflatoxin are highly regarded by countries in the world due to the similarity of the structure of the aflatoxin. China is in a region with serious aflatoxin pollution, agricultural products such as peanuts, corns, rice, nuts and the like and food are reported and detected, and the phenomenon of mixed pollution with cyclopiazonic acid, aflatoxin and the like exists, so that the consumption safety of agricultural products in China is threatened, and the industrial development and export trade are limited. Among them, peanuts and corns are the easiest to produceIs polluted by aflatoxin infection and aflatoxin and other mycotoxins. Therefore, the research on field prevention and control of the aspergillus flavus pollution of agricultural products can reduce the aspergillus flavus infection and the mycotoxin pollution generated by the aspergillus flavus infection from the source, and the method has important significance for ensuring the quality safety of agricultural products in China and improving the international competitiveness of agricultural products in China.
The biological prevention and control has the advantages of no damage to the original quality of agricultural products, safety, high efficiency, environmental protection and the like, and is an effective way for green control of aspergillus flavus and aflatoxin. The method utilizes the aspergillus flavus which does not produce toxin to compete and inhibit the growth and colonization of the aspergillus flavus which produces toxin in the field, thereby reducing the number of bacteria colony of the toxin producing bacteria and reducing the pollution level of the aflatoxin, and is a very effective biological control method for controlling the aflatoxin pollution in agricultural products such as peanuts, corns and the like from the source at present. The research of the aspergillus flavus biocontrol bacteria which do not produce the toxin in China is in an initial stage, and no case exists for popularization and application in fields. The prior art discloses non-toxigenic aspergillus flavus used for biological control and application thereof (such as CN 110157626A, CN 110305796A, CN 110129212A, CN 107177516 and CN 104789480A), and only reports on biocontrol bacteria with high safety aiming at various toxic metabolites such as non-aflatoxin, cyclopiazonic acid, aflatoxin, heteroauxin and the like, and the strains can bring mycotoxin pollution risks such as cyclopiazonic acid, aflatoxin and the like after field application. Researches show that after aspergillus flavus AF36 (Afla-/CPA +) which does not produce toxicity is used as a biocontrol bacterium, the content of cyclopiazonic acid in corn and peanut is increased, and new risk potential is brought. Therefore, the high-safety aspergillus flavus which does not produce a plurality of toxins such as aflatoxin, cyclopiazonic acid, aflatoxin and the like is screened, developed and utilized, toxin pollution prevention and control are carried out before crops are harvested, the risk that aflatoxin, cyclopiazonic acid, aflatoxin and the like enter a food chain is effectively reduced, and the wide application prospect is realized in the aspect of improving the quality safety level of agricultural products. According to the invention, through a large amount of experimental work, 1 strain of aspergillus flavus XZCY1805 which does not produce secondary metabolites such as aflatoxin, cyclopiazonic acid, aflatoxin, variegated aspergillus toxin and the like is screened out from 97 strains which do not produce aflatoxin, has strong competitive advantage, and can remarkably reduce the mycotoxin pollution risk such as aflatoxin, cyclopiazonic acid, aflatoxin, variegated aspergillus toxin and the like in agricultural products such as peanuts and the like.
Disclosure of Invention
The invention aims to solve the technical problem of providing a non-toxigenic aspergillus flavus strain which does not produce a plurality of toxins such as aflatoxin, cyclopiazonic acid, aflatoxin, variegated aspergillus toxin and the like and application thereof.
Provides a non-toxigenic Aspergillus flavus strain XZCY1805(Aspergillus flavus), which is preserved in China Center for Type Culture Collection (CCTCC) at 9/18 of 2020, with a preservation number of CCTCC NO: M2020521, a preservation address: china, wuhan university.
The non-toxigenic aspergillus flavus strain XZCY1805 was isolated from the west corner peanut plantation in tibet.
And (3) screening: the aspergillus flavus XZCY1805 is screened from 1600 aspergillus flavus by strain toxin production capability evaluation and biocontrol inhibition effect test, and the process is as follows: collecting more than 2000 parts of peanuts and soil samples from northeast, northern and Yangtze river watersheds and southern peanut main producing areas of China, and separating, purifying and identifying to obtain 1600 strains of aspergillus flavus; after the aflatoxin toxin production capacity of the strain is measured, 97 non-toxigenic aflatoxins (mainly separated from 15 peanut planting fields such as Henan, Shandong, Hubei, Jiangxi, Tibet and the like) are selected to be co-cultured on peanut powder and respectively subjected to toxigenic standard strains 3.4408 and evaluation of toxigenic inhibition effect, and 11 strains with remarkable inhibition effect on toxigenic aflatoxins are preliminarily screened; further co-culturing 11 strains of non-toxigenic aspergillus flavus and toxigenic aspergillus flavus separated from different regions (Liaoning funxin, Hebei Tangshan, Sichuan paean, Tibet corner, Hubei hong' an, Hubei yang logical, Jiangxi camphor tree) on peanut powder and peanut seeds, and screening out 3 strains with remarkable competitive inhibition effect on toxigenic strains from different geographical sources after evaluating the toxigenic inhibition effect; and (3) further measuring toxic secondary metabolites such as aflatoxin, cyclopiazonic acid, aflatoxin and variegated aspergillus toxin on 3 strains of the aspergillus flavus strain which does not produce the toxin, screening 1 strain XZCY1805 which does not produce the aflatoxin, cyclopiazonic acid, aflatoxin and variegated aspergillus toxin, and remarkably competing and inhibiting the growth and reproduction capacity of the toxin-producing aspergillus flavus, and reducing the contents of the aflatoxin, the cyclopiazonic acid, the aflatoxin and the variegated aspergillus toxin in agricultural products such as peanuts and the like.
The non-toxigenic aspergillus flavus strain XZCY1805 has the characteristics of rapid growth and reproduction, has strong competitive advantage, can remarkably compete to inhibit the growth and reproduction of toxigenic aspergillus flavus, and can reduce the content of mycotoxins such as aflatoxin, cyclopiazonic acid, aflatoxin, variegated aspergillus toxin and the like in agricultural products such as peanuts and the like. In peanut powder, the non-toxigenic bacterium XZCY1805 is against AFB of toxigenic bacterium 3.44081The toxin production inhibition rate is 99.86 percent, and the toxin production inhibition rate is applied to toxin production strains AFT and AFB which are separated from different producing areas1The toxin production inhibition rates of the composition are respectively 81.5-94.4% and 84.5-98.2%; the total aflatoxin content in the peanut seeds is reduced by 51.5-97.8 percent, and the aflatoxin B1The content is reduced by 51.2 to 100.0 percent; field pot experiment of peanut for AFT and AFB1The toxin production inhibition effect is obvious, and the inhibition rates are respectively 76.5% and 68.0%. The result shows that the non-toxigenic bacterium XZCY1805 has the capability of efficiently competing to inhibit the growth and reproduction of toxigenic bacteria and reduce the probability of infecting crops by the toxigenic bacteria, so that the content of mycotoxins such as aflatoxin, cyclopiazonic acid, aflatoxin, variegated aspergillus toxin and the like in agricultural products is reduced. Compared with other aspergillus flavus biocontrol bacteria which do not produce aflatoxin, the product does not produce a plurality of toxins such as aflatoxin, cyclopiazonic acid, aflatoxin, variegated aflatoxin and the like, and has higher safety.
The colony morphology of the non-toxigenic aspergillus flavus strain XZCY1805 is as follows: the strain produces white hypha and yellow-green spores on a PDA culture medium, produces yellow-green spores on a DG-18 culture medium, and presents characteristic bacterial colonies with bright orange color on the back of an AFPA culture medium. After 5 days of culture in DG-18 medium at 28 + -1 deg.C, the colony diameter is 7.73cm, and the spore yield is 5.5X 108Compared with the standard strain 3.4408 (the diameter of the colony is 7.0cm, the spore yield is 4.25 multiplied by 10) of the aspergillus flavus strain which produces the toxin under the same condition82) has better growth and reproductive capacityIs strong.
The method for inhibiting the generation of aflatoxin by using the non-toxigenic aspergillus flavus strain XZCY1805 is provided, and the non-toxigenic aspergillus flavus strain XZCY1805 is inoculated on a biological sample to inhibit the generation of mycotoxin.
The biological sample can be peanut powder or peanut kernel.
According to the scheme, the mycotoxin comprises aflatoxin, cyclopiazonic acid, aflatoxin, variegated aspergillus toxin and the like, and the aflatoxin comprises aflatoxin B1Aflatoxins B2Aflatoxin G1And aflatoxin G2。
Specifically, after activating the non-toxigenic aspergillus flavus strain XZCY1805 in a DG-18 solid medium and culturing for 5 days at 28 +/-1 ℃ in the dark, washing conidia on the surface of the medium by using sterilized 0.1% Tween 80, and diluting the conidia in sterile water to obtain a conidia suspension for use.
According to the above scheme, the concentration of the spore suspension can be 1 × 106One per ml.
Provides a method for preventing and reducing the pollution risk of aflatoxin, cyclopiazonic acid, aflatoxin and variegated aspergillin in agricultural products in fields. The Aspergillus flavus XZCY1805 conidium suspension is used for being scattered at the rhizosphere of crops.
According to the scheme, the concentration of spores of the Aspergillus flavus XZCY1805 suspension is preferably 1 x 106The dosage is preferably 80L/mu.
According to the scheme, the agricultural product is peanuts, and the application time is preferably the flowering and fruiting period of the peanuts.
The invention has the beneficial effects that:
(1) the non-toxigenic aspergillus flavus XZCY1805 has a strong effect of inhibiting toxigenic aspergillus flavus produced by a standard toxigenic aspergillus flavus strain 3.4408 and different peanut producing areas and separated, is a biocontrol strain with high safety and strong competitive advantage, can obviously inhibit the toxigenic aspergillus flavus from infecting crops, and can reduce mycotoxin pollution of aflatoxin, cyclopiazonic acid, aflatoxin, variegated aspergillus chromous toxin and the like in agricultural products such as peanuts and the like. Has obvious effect of inhibiting the production of toxicity on peanut powder, peanut seeds and peanut seedlings.
(2) The non-toxigenic aspergillus flavus XZCY1805 disclosed by the invention does not produce Aflatoxin (AFT) and does not produce various toxins such as Cyclopianile (CPA), variegated aflatoxin (ST), aflatoxin (Aflatrem) and the like, and is a biocontrol strain with high safety. The compound has the characteristics of rapid growth and reproduction, has strong competitive inhibition capability, can remarkably competitively inhibit the growth and reproduction of toxin-producing aspergillus flavus, and reduces the content of mycotoxins such as aflatoxin, cyclopiazonic acid, aflatoxin, variegated aflatoxin and the like.
Drawings
Table 1: the toxigenicity of the yellow aspergillus flavus XZCY1805 is not toxigenic;
table 2: the aspergillus flavus XZCY1805 without producing the toxin has the effect of inhibiting the production of the toxin of the aspergillus flavus on the peanut powder;
table 3: aspergillus flavus XZCY1805 without producing toxin produces AFT and AFB on peanut seed1An inhibitory effect;
table 4: aspergillus flavus XZCY1805 without producing toxin produces AFG on peanut seed for producing aspergillus flavus with toxin1、AFG2Inhibiting effect
Table 5: the XZCY1805 with different concentrations has the function of inhibiting the aspergillus flavus from producing toxin;
table 6: the aspergillus flavus XZCY1805 without generating toxin has the effect of inhibiting the content of aflatoxin in potted flowers;
FIG. 1: the aspergillus flavus XZCY1805 without producing toxin is in colony morphology on the back of a DG-18 culture medium and an AFPA culture medium, the strain XZCY1805 of the invention grows yellow-green spores on the DG-18 culture medium, and produces bright orange color reaction on the AFPA culture medium;
FIG. 2: aspergillus flavus XZCY1805 with different spore concentrations for producing toxin to Aspergillus flavus AFT and AFB1The inhibition rate.
Detailed Description
Example 1: separation, purification and identification of strain XZCY1805
Collecting more than 2000 parts of peanuts and soil samples from northeast, northern and Yangtze river watersheds and southern peanut main producing areas of China, and separating, purifying and identifying to obtain 1600 strains of aspergillus flavus; after the aflatoxin toxin production capacity of the strain is measured, 97 strains of non-toxin-producing aflatoxins (mainly separated from 15 peanut planting fields such as Henan, Shandong, Hubei, Jiangxi, Tibet and the like) are selected for competitive inhibition effect research and toxin production capacity evaluation: co-culturing the peanut powder and a toxigenic standard strain 3.4408 respectively, and evaluating the toxigenic inhibition effect, and primarily screening 11 strains with remarkable inhibition effect on toxigenic aspergillus flavus; further co-culturing 11 strains of non-toxigenic aspergillus flavus and toxigenic aspergillus flavus separated from different regions (Liaoning funxin, Hebei Tangshan, Sichuan paean, Tibet corner, Hubei hong' an, Hubei yang logical, Jiangxi camphor tree) on peanut powder and peanut seeds, and screening 3 strains with remarkable competitive inhibition effect on toxigenic strains from different geographical sources after evaluating the toxigenic inhibition effect; and (3) further carrying out the determination of toxic secondary metabolites such as aflatoxin, cyclopiazonic acid, aflatoxin, variegated aspergillus toxin and the like on 3 strains of the aspergillus flavus which do not produce the toxin, screening out 1 strain XZCY1805 which does not produce the aflatoxin, cyclopiazonic acid, aflatoxin and variegated aspergillus toxin, and being capable of remarkably competing and inhibiting the growth and reproductive capacity of the toxin-producing aspergillus flavus and reducing the contents of the aflatoxin, the cyclopiazonic acid, the aflatoxin and the variegated aspergillus toxin in agricultural products such as peanuts. The strain XZCY1805 which does not produce aflatoxin, cyclopiazonic acid, aflatoxin and variegated aspergillus toxin is separated from the peanut plantation in the Tibet Tanqua.
For the soil sample, the separation process is as follows: and (3) removing plant residues and large-particle stones from the soil sample, grinding the soil, uniformly mixing and the like. The ground 10.0g of soil sample is weighed and added into 90mL of sterilized water, and the mixture is put into a shaking table to shake for 2h, so as to prepare 100mL of sample suspension. 50 μ L of the suspension was added to a DG-18 medium plate and spread evenly with a spreading bar. Then placing the coated flat plate into a constant temperature incubator, and culturing at 28 +/-1 ℃ under the relative humidity of 90% and under the dark condition. After 5 days, colonies with yellow-green spores are picked and transferred to a new DG-18 medium plate for secondary streak purification culture until single colonies are obtained for later use.
For peanut samples, the separation process was: pulverizing peanut to obtain peanut powder, preparing sample suspension, and separating and purifying by the above method to obtain single colony for use. And (3) identification:
morphological identification
The strain XZCY1805 of the invention grows yellow green spores on a DG-18 culture medium, the diameter of a colony cultured for 5 days at the temperature of 28 +/-1 ℃ is 7.73cm, and the spore yield is 5.5 multiplied by 108The colony diameter of the standard Aspergillus flavus strain 3.4408 under the same conditions is 7.0cm, and the spore yield is 4.25X 108And (4) respectively. Therefore, the aspergillus flavus XZCY1805 provided by the invention has the advantages of rapid growth and propagation.
Colonies with yellowish green spores were picked from DG-18 medium and transferred to AFPA (Aspergillus flavus and Aspergillus parasiticus Agar) as a selective medium and cultured at 28. + -. 1 ℃ under 90% relative humidity and dark conditions. The colony back color was observed after 5 days, producing a bright orange color reaction on AFPA medium (fig. 1), initially identified as aspergillus flavus or aspergillus parasiticus.
Molecular biological identification
The strain XZCY1805 is subjected to molecular identification through an ITS gene sequence and a calmodulin gene sequence of the fungus.
Primers used for amplification of Aspergillus flavus genome ITS were:
ITS1:5′-TCCGTAGGTGAACCTGCGG-3′
ITS4:5′-TCCTCCGCTTATTGATATGC-3′
the primers used for amplifying the calmodulin coding gene of the aspergillus flavus genome are as follows:
CF1:5′-AGGCCGAYTCTYTGACYGA-3′
CF4:5′-TTTYTGCATCATRAGYTGGAC-3′
the PCR amplification conditions of the ITS gene sequence are as follows: pre-denaturation at 94 ℃ for 5 min; denaturation at 94 ℃ for 30s, annealing at 58 ℃ for 30s, and extension at 72 ℃ for 30s for 35 cycles; final extension at 72 ℃ for 10 min. After amplification, the product was stored at 4 ℃. Sending to Wuhan division of Biotechnology Limited, Beijing Ongchongke, for sequencing, and performing comparative analysis on the sequencing result on NCBI website (https:// blast.
The PCR amplification conditions of the calmodulin coding gene sequence are as follows: pre-denaturation at 94 ℃ for 5 min; denaturation at 94 ℃ for 30s, annealing at 57 ℃ for 30s, and extension at 72 ℃ for 1min for 35 cycles; final extension at 72 ℃ for 10 min. After amplification, the product was stored at 4 ℃. Sending to Wuhan division of Biotechnology Limited, Beijing Ongko for sequencing, and obtaining the sequencing result on NCBI website
(https:// blast. ncbi. nlm. nih. gov/blast. cgi) was performed.
Sequencing shows that the ITS sequence of the strain XZCY1805 is shown as SEQ ID No. 1; the calmodulin sequence is shown in SEQ ID No. 2.
The ITS amplified sequence is compared with a BLASTN database at an NCBI website, so that the similarity between the ITS gene sequence of the strain XZCY1805 and the RNA gene sequence of the ribosome small subunit of Aspergillus flavus (Aspergillus flavus) is 99.67 percent. The comparison of the calmodulin coding gene amplification sequence in NCBI website through BLASTN database shows that the similarity between the calmodulin coding gene sequence of the strain XZCY1805 and the calmodulin A gene sequence of Aspergillus flavus is 98.86%.
The strain XZCY1805 of the invention is identified as Aspergillus flavus (Aspergillus flavus) by combining the identification results of morphology and molecular biology.
Example 2: analysis of the toxigenicity of Strain XZCY1805
1. Strain toxin-producing culture
Inoculating the non-toxigenic aspergillus flavus strain XZCY1805 to DG-18 solid culture medium for activation. After culturing for 5 days at 28 +/-1 ℃ in the dark, the aspergillus flavus conidia on the obtained plate is washed by sterilized 0.1% Tween 80 to obtain a spore suspension. Spore suspension concentrations were determined using a blood-ball counting plate. Taking a certain amount of spore liquid, injecting into a triangular flask containing sterilized 30ml liquid Sabouraud's medium to make its final concentration be 4 × 105One per ml. The flask was placed on a shaker at 28. + -. 1 ℃ and cultured in the dark at 200 rpm.
2. Determination of toxin content in bacterial strains
5 days later, filtering mycelium pellet culture solution with sterilized gauze, discarding mycelium pellet, collecting toxin-producing culture solution, and storing in centrifuge tubeStoring, standing for 1h, transferring 500 μ l of the toxin-producing culture solution into a 2ml centrifuge tube, adding 500 μ l of methanol, uniformly vortexing, centrifuging for 10min at 20000rmp, carefully sucking out supernatant in the tube by using a 1ml syringe (preventing a needle from contacting with the bottom of the tube for precipitation), filtering toxin extract in the syringe through a 0.22 μm organic filter membrane, filling the tube into a sample injection bottle, and measuring the content of toxic metabolites such as aflatoxin, cyclopiazonic acid, aflatoxin, mottle toxin and the like by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS), wherein the detection result is shown in Table 1. Chromatographic conditions are as follows: the chromatographic column is C18(100 mm. times.2.1 mm, 3 μm); the column temperature is 40 ℃, the sample injection volume is 2 mu L, and the sample tray temperature is 10 ℃; mobile phase a was methanol to water (V: V ═ 5:95), mobile phase B methanol to water (V: V ═ 5:95), each containing 0.1% formic acid and 10mM ammonium formate at a flow rate of 0.3 ml/min; gradient elution procedure: 0-1min:85A, 1-3 min: 85-50% of A, 3-5 min: 50-30% of A, 5-10 min: 30-0% of A, 10-13 min: 0% A, 13-15 min: 0-85% of A, 15-20 min: 85% of A.
Mass spectrum conditions: ESI (+) mode: 20mL/min of air curtain air; the temperature of the atomizer is 450 ℃; 20mL/min of an atomizer; the auxiliary heating gas pressure is 40 mL/min. The quantitative mode is selected ion detection (SIM); analysis software Analyst 1.14. A chromatographic column: waters Symmetry C18 column (2.1X 150mm, 3.5 μm); mobile phase A: 10mM acetic acid in water; mobile phase B: 100% methanol; the column temperature is 40 ℃; the flow rate is 0.3 mL/min; gradient program: b is 50% in 0min, is increased to 90% in 0-10 min, and is reduced from 90% to 50% in 10-15 min.
As can be seen from Table 1, strain XZCY1805 does not produce aflatoxin, cyclopiazonic acid, aflatoxin, and tremolin.
TABLE 1 toxigenic Properties of non-toxigenic Aspergillus flavus XZCY1805
aAFT is an abbreviation for aflatoxin (aflatoxin B)1Aflatoxins B2Aflatoxin G1And Aspergillus flavus G2Total amount of (c);
bCPA is cyclopiazonic acid;cST is aflatoxin; dAflatrem is aflatoxin;
eND means not detected.
Example 3: inhibition effect of strain XZCY1805 on toxigenic aspergillus flavus on peanut powder
1. Spore suspension preparation
The non-toxigenic Aspergillus flavus strain XZCY1805, the toxigenic Aspergillus flavus standard strain 3.4408 and the toxigenic Aspergillus flavus (Aspergillus flavus) separated from different regions, wherein the numbers of the strain are SCPA-32-12 (Sichuan pavon), XZCY-24-6 (Tibet Kwang), LNFX-25-1 (Liaoning funxin), HBHA-129-1 (Hubei Hongan), HBYL-12-7 (Hubei yang logical), HBTS-94-2 (Hebei Tangshan) and JXZS-118-9 (Jiangxi camphor tree) are respectively inoculated to DG-18 solid medium for activation. After culturing for 5 days at 28 +/-1 ℃ in the dark, the aspergillus flavus conidia on the flat plate are washed by sterilized 0.1% Tween 80 in a 10ml centrifuge tube to obtain a spore suspension. The spore suspension concentration was determined under a microscope using a hemocytometer, and the spore concentration was diluted to 1X 10 with sterile water6And each ml is ready for use.
2. Competitive inhibition culture
Weigh 10.0g of peanut powder in a sterile petri dish (9cm), set control and treatment groups: respectively preparing toxigenic aspergillus flavus (spore concentration is 1 multiplied by 10)6One/ml) spore suspension (control group) mixed with equal volume of sterile water (250. mu.l each), and non-toxigenic Aspergillus flavus XZCY1805 (spore concentration 1X 10)6One/ml) and standard Aspergillus flavus strain 3.4408 or Aspergillus flavus producing strain isolated from different peanut producing areas (spore concentration 1X 10)6Pieces/ml) of spore suspension (treatment group) mixed in equal volumes (250. mu.l each), the mixed spore suspension was inoculated uniformly onto peanut powder for 3 biological replicates per treatment, and the samples were placed in a constant temperature incubator and incubated in the dark at 28 ℃. + -. 1 ℃ and 90% relative humidity for 14 days.
3. Determination of aflatoxin content
After the incubation period, the peanut powder in the petri dish was transferred to a 50ml centrifuge tube and 15ml of 70% methanol solution (containing4% NaCl), evenly mixing by vortex, placing on an oscillator to shake for 2h, centrifuging at 4500r/min, taking 1ml of supernatant, filtering through an organic filter membrane, and detecting aflatoxin B by High Performance Liquid Chromatography (HPLC)1、B2、G1、G2The content of (a). The total aflatoxin content (AFT) is the sum of the above 4 aflatoxins.
HPLC conditions C18 column (4.6 mm. times.150 mm,5 μm); the column temperature was 35 ℃; mobile phase: methanol: water (V: V ═ 45: 55); the flow rate is 0.8 mL/min; post-column photochemical derivatization method, photochemical derivatization device 254 nm; fluorescence detector (excitation wavelength 360nm, emission wavelength 440nm), sample volume 10 u l, determination time 22 min.
The toxicity generation inhibition rate calculation formula is as follows: the inhibition rate (%) [1- (treatment group aflatoxin content)/(control group aflatoxin content) ] × 100, formula I.
As can be seen from Table 2, the peanut powder is used as the matrix, and the non-toxigenic bacteria XZCY1805 is used for AFB of toxigenic bacteria 3.44081The toxin production inhibition rate is 99.86 percent, and the toxin production inhibition rate is applied to toxin production strains AFT and AFB which are separated from different producing areas1、AFB2The toxin production inhibition rates of the strain are respectively 81.5-94.4%, 84.5-98.2% and 39.8-93.7%, which shows that the non-toxigenic bacterium XZCY1805 has broad-spectrum and high-efficiency toxin production capacity of competitive inhibition of toxin-producing aspergillus flavus, thereby reducing the content of aflatoxin in peanuts.
TABLE 2 inhibition of non-toxigenic Aspergillus flavus XZCY1805 on peanut powder
Wherein AFT is total aflatoxin (aflatoxin B)1Aflatoxins B2Aflatoxin G1And Aspergillus flavus G2Composition of
Example 4: inhibition effect of strain XZCY1805 on producing toxin aspergillus flavus on peanut grains
1. Spore suspension preparation
The same as in example 3.
2. Competitive inhibition culture
Selecting mature, full, healthy and normal peanut seeds without insect mouth disease spots and with complete and uniform seed coats. Soaking peanut seeds in 70% ethanol solution for 2min for surface disinfection, washing with sterile distilled water for 3 times to wash off residual ethanol on the peanut surface, and ensuring that the moisture of the peanut seeds is about 20% and within 13.0 min.
Setting a control group and a treatment group, and respectively preparing the aspergillus flavus producing toxin (the spore concentration is 11 multiplied by 10)6One/ml) spore suspensions (12.5 ml each) mixed with equal volumes of sterile water (control group), and non-toxigenic aspergillus flavus XZCY1805 (spore concentration 1 × 10)6One/ml) and standard strain 3.4408 of Aspergillus flavus and Aspergillus flavus producing Aspergillus flavus (spore concentration 1X 10) separated in different regions6Pieces/ml) of spore suspension (treatment group) mixed in equal volume (12.5 ml each), 10 peanut kernels were placed in it and soaked for 5min, then picked up with sterile forceps and placed in a sterile petri dish. 3 biological replicates of each treatment were performed and the samples were placed in a constant temperature incubator and incubated continuously in the dark at 28. + -. 1 ℃ and 90% relative humidity for 7 days.
3. Determination of aflatoxin content
Sterilizing the cultured peanut seeds at high temperature and high pressure (121 ℃, 30min), drying the peanut seeds in a constant-temperature drying oven (110 ℃, 1h), cooling, grinding the peanut seeds into peanut powder by using a coffee machine, weighing 1.0g of peanut powder sample, putting the peanut powder sample into a centrifugal tube, adding 5ml of 70% methanol solution (containing 4% NaCl), uniformly mixing in a vortex manner, placing the peanut powder sample on an oscillator to vibrate for 2h, centrifuging at 4500r/min, passing through an immunoaffinity column and an organic filter membrane, and detecting aflatoxin B by using High Performance Liquid Chromatography (HPLC)1、B2、G1、 G2The content of (a). The total aflatoxin content (AFT) is the sum of the above 4 aflatoxins.
Chromatographic conditions are as described in example 3.
The formula for calculating the toxicity inhibiting rate is the same as that described in example 3, i.e. formula I. As can be seen from tables 3 and 4, the peanut seeds are used as the culture medium, and the non-toxigenic bacteria XZCY1805 are used for AFT and AFB of toxigenic bacteria 3.44081、AFB2The toxin production inhibition rate is respectively 97.8 percent100.0 percent and 80.7 percent of the total weight of the peanut oil, and the strains AFT and AFB which are separated from different peanut producing areas and produce toxicity1、AFB2、AFG1、 AFG2The toxin production inhibition rates of the strain are respectively 51.5-88.8%, 51.2-100.0%, 17.4-89.6%, 72.8-100.0% and 40.9-100.0%, which shows that the non-toxigenic strain XZCY1805 can efficiently compete and inhibit the toxin production capability of the toxin-producing aspergillus flavus and remarkably reduce the aflatoxin pollution level in peanuts.
TABLE 3 inhibition effect of non-toxigenic Aspergillus flavus XZCY1805 on AFT and AFB production of toxigenic Aspergillus flavus on peanut seeds
AFT is total aflatoxin (from aflatoxin B)1Aflatoxins B2Aflatoxin G1And aflatoxin G2Composition of
TABLE 4 Aflatopsis flavus XZCY1805 production without toxigenicity on peanut seeds for AFG production by toxigenicity Aspergillus flavus1And AFG2Inhibiting effect
Example 5: inhibition experiments with different concentration ratios
Separately preparing the spore at a concentration of 1 × 103、1×104、1×105、1×106Aspergillus flavus XZCY1805 and 1X 10 of non-toxigenic aspergillus flavus per ml5The preparation method of the spore suspension of yellow-producing Aspergillus flavus 3.4408 is the same as that of example 3.
Setting a control group and a treatment group:
preparation of Aspergillus flavus Standard Strain 3.4408 (1X 10)5) Mixing with equal volume of sterile water (12.5 ml each) to obtain spore suspension containing spore with number of 5 × 104A/ml standard strain of Aspergillus flavus 3.4408 (control).
The Aspergillus flavus XZCY1805 and the Aspergillus flavus standard strain 3.4408 with the volume ratio of 1:1 are added in seriesMixing to obtain spore concentration ratio of 5 × 102:5×104(1:100)、5×103:5×104(1:10)、5×104:5×104(1:1)、 5×105:5×104A mixture of Aspergillus flavus SX0104 (10:1) and Aspergillus flavus standard strain 3.4408.
Soaking the sterilized and washed peanut seeds in the control group and the treatment group for 5min respectively, clamping the peanut seeds by using sterile forceps, placing the peanut seeds in a sterile culture dish, placing the culture dish in a constant-temperature incubator, and continuously culturing the peanut seeds in the dark for 7 days at the temperature of 28 ℃ and the relative humidity of 90%.
Sterilizing the cultured peanut seeds at high temperature and high pressure (121 ℃, 30min), drying the peanut seeds in a constant-temperature drying oven (110 ℃, 1h), cooling, grinding the peanut seeds into peanut powder by using a coffee machine, weighing 1g of peanut powder sample, putting the peanut powder sample into a centrifuge tube, adding 5ml of 70% methanol solution (containing 4% NaCl), uniformly mixing the peanut powder sample in a vortex manner, placing the peanut powder sample on an oscillator for shaking for 2h, centrifuging at 4500r/min, passing through an immunoaffinity column and an organic filter membrane, and detecting aflatoxin B by using high performance liquid chromatography1、B2、G1、G2The aflatoxin inhibition was calculated according to example 3, and the results are shown in table 5 and fig. 2, as the chromatographic conditions were as described in example 3.
As can be seen from table 5 and fig. 2, the non-toxigenic aspergillus flavus with different concentration ratios has an inhibitory effect on the production of mold and mildew, to a certain extent, the inhibition rate of the non-toxigenic aspergillus flavus on the production of toxigenic bacteria increases with the increase of the concentration of spores thereof, and the ratio of the number of the spores of the non-toxigenic aspergillus flavus to the number of the spores of toxigenic bacteria 3.4408 is 1:100 to 10:1, the AFT toxin production inhibition rate of the aspergillus flavus XZCY1805 on the toxin-producing strain 3.4408 in peanut grains is 45.4-96.3%. Lower concentration ratio of 5X 102:5×104(1:100), the inhibition rate of aflatoxin total amount AFT is 45.4%. When the ratio of the spore concentration of the aspergillus flavus XZCY1805 to the aspergillus flavus standard strain 3.4408 is 5 x 103:5×104(1:10), the inhibition rate of the total aflatoxin amount is 66.8%; concentration ratio of 5X 104:5×104(1:1), the inhibition ratio is 76.1%; at a concentration ratio of 5X 105:5×104The highest inhibition rate was 96.3% in the case of (10: 1).
TABLE 5 inhibition of Aspergillus flavus production by XZCY1805 at different concentrations
Example 6 Effect of Strain XZCY1805 on Aflatoxin inhibition of potted peanuts
1. Aflatopsis flavus XZCY1805 potting application method without producing toxin
Peanut seedlings are transplanted into flowerpots (the area of each flowerpot is 0.3 m) from peanut planting land24 peanuts in each pot, no damage to peanut seedlings and soil at roots). 9mL of a conidia suspension of non-toxigenic Aspergillus flavus XZCY1805 (concentration 1X 10)6one/mL) was scattered at the rhizosphere of each peanut (test group), the blank control group was scattered with an equal volume of sterile water, and the other daily management test groups were identical to the blank control group. The assay was set up for 3 biological replicates.
2. Collection of peanut samples at harvest stage
Collecting peanut samples in the peanut harvesting period, airing and drying the peanut samples, and detecting the aflatoxin content after the samples are subjected to condensation separation by a quartering method.
3. Cultivation of peanut pod toxigenicity
In order to reduce the risk of aflatoxin pollution caused by improper storage environment, an aflatoxin pollution experiment under extreme environmental conditions (toxin production and culture conditions) is carried out. Harvested and dried peanut pods were placed in petri dishes with 5 pods per petri dish and 3 replicates per sample. Placing into an incubator, culturing at 28 + -1 deg.C and relative humidity of 90%, and detecting aflatoxin content after 20 days.
4. Detection of aflatoxin content in peanuts and toxin production inhibition effect
Drying peanut pods in a constant temperature drying oven (110 deg.C, 1h), cooling, grinding into peanut powder with a coffee machine, weighing 1.0g peanut powder sample, placing into a centrifuge tube, adding 5ml 70% methanol solution (containing 4% NaCl), vortex mixing, placing on an oscillator, shaking for 2h, and centrifuging at 4500r/minDetecting aflatoxin B by High Performance Liquid Chromatography (HPLC) after passing through immunoaffinity column and organic filter membrane1、B2、G1、G2The content of (a). The total aflatoxin content (AFT) is the sum of the above 4 aflatoxins.
The chromatographic conditions were as described in example 3.
The formula for calculating the toxicity inhibiting rate is the same as that described in example 3, i.e. formula I. The results are shown in Table 6
The toxin detection result shows that aflatoxin is not detected in the harvested and dried peanut control group and the experimental group.
As can be seen from Table 6, under the toxin-producing culture conditions, the aflatoxin content in the peanut pods of the placebo group was 69.3. mu.g/kg, and the aflatoxin B was1The content of (A) is 40.3 mu g/kg; the content of aflatoxin in the peanut pods of the test group is 16.3 mug/kg, and the aflatoxin B1The content was 12.9. mu.g/kg. The non-toxigenic aspergillus flavus XZCY1805 has the capabilities of efficiently competing and inhibiting the growth and reproduction of toxigenic bacteria and reducing the proportion of the toxigenic bacteria infecting crops, thereby reducing the mycotoxin pollution content in agricultural products and treating AFT and AFB in peanuts1The inhibition ratios of (A) and (B) were 76.5% and 68.0%, respectively.
TABLE 6 non-toxigenic Aspergillus flavus XZCY1805 inhibitory Effect on potted peanut aflatoxin content
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Claims (8)
1. A non-toxigenic Aspergillus flavus strain XZCY1805(Aspergillus flavus) is characterized in that: the non-toxigenic aspergillus flavus strain is preserved in China Center for Type Culture Collection (CCTCC) NO: M2020521 in 9-18 2020.
2. The aspergillus flavus XZCY1805 according to claim 1, wherein the non-toxigenic aspergillus flavus XZCY1805 does not produce aflatoxin, cyclopiazonic acid, aflatoxin and variegated aflatoxin.
3. A method of inhibiting aflatoxin production by the non-toxigenic aspergillus flavus strain XZCY1805 of claim 1, wherein the non-toxigenic aspergillus flavus strain XZCY1805 is inoculated onto a biological sample to inhibit mycotoxin production.
4. The method of claim 3, wherein: the biological sample is peanut powder or peanut kernel.
5. The method of claim 3, wherein: the mycotoxin comprises aflatoxin, cyclopiazonic acid, aflatoxin and variegated aflatoxin, and the aflatoxin comprises aflatoxin B1Aflatoxins B2Aflatoxin G1And aflatoxin G2。
6. The method of claim 3, wherein: the non-toxigenic Aspergillus flavus strain XZCY1805 was used as a conidium suspension.
7. A method for preventing and reducing the pollution risk of aflatoxin, cyclopiazonic acid, aflatoxin and aflatoxin in agricultural products in fields, wherein the aspergillus flavus XZCY1805 conidia suspension of claim 1 is applied to the rhizosphere of peanuts.
8. The method of claim 7, wherein: the agricultural product is peanuts, and the application time is the flowering and injection period of the peanuts.
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Citations (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6027724A (en) * | 1998-07-06 | 2000-02-22 | The United States Of America, As Represented By The Secretary Of Agriculture | Non-toxigenic strain of Aspergillus oryzae and Aspergillus sojae for biocontrol of toxigenic fungi |
US7361499B1 (en) * | 2005-01-11 | 2008-04-22 | The United States Of America, As Represented By The Secretary Of Agriculute | Non-aflatoxigenic Aspergillus flavus isolates |
US20120183507A1 (en) * | 2011-01-19 | 2012-07-19 | Dorner Joe W | Non-Toxigenic Strains of Aspergillus Flavus for Control of Aflatoxin Contamination in Crops |
WO2014191917A1 (en) * | 2013-05-27 | 2014-12-04 | Universita' Cattolica Del Sacro Cuore | Non-toxigenic strain of aspergillus flavus |
CN107177516A (en) * | 2017-07-07 | 2017-09-19 | 福建农林大学 | A kind of aspergillus flavus avirulent strain and its application of preventing and treating aspergillus flavus pollution |
CN107245453A (en) * | 2017-06-02 | 2017-10-13 | 中国农业科学院农产品加工研究所 | One plant is not produced malicious aspergillus flavus and its application in terms of aflatoxin degradation |
CN107279686A (en) * | 2017-06-02 | 2017-10-24 | 中国农业科学院农产品加工研究所 | Application of the malicious aspergillus flavus in terms of aflatoxin degradation is not produced |
CN107828694A (en) * | 2017-11-29 | 2018-03-23 | 中国农业科学院油料作物研究所 | One plant height effect suppresses serratia marcescens biocontrol microorganisms and its application of aspergillus flavus synthesis aflatoxin |
CN108531407A (en) * | 2018-04-27 | 2018-09-14 | 福建农林大学 | One plant of method that structure of aflatoxigenic strain and prevention aspergillus flavus do not pollute |
CN109913375A (en) * | 2019-04-03 | 2019-06-21 | 吉林大学 | A kind of Aspergillus flavus of not toxin producing, the composition containing it and its application |
-
2021
- 2021-06-24 CN CN202110701929.7A patent/CN113846020B/en active Active
Patent Citations (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6027724A (en) * | 1998-07-06 | 2000-02-22 | The United States Of America, As Represented By The Secretary Of Agriculture | Non-toxigenic strain of Aspergillus oryzae and Aspergillus sojae for biocontrol of toxigenic fungi |
US7361499B1 (en) * | 2005-01-11 | 2008-04-22 | The United States Of America, As Represented By The Secretary Of Agriculute | Non-aflatoxigenic Aspergillus flavus isolates |
US20120183507A1 (en) * | 2011-01-19 | 2012-07-19 | Dorner Joe W | Non-Toxigenic Strains of Aspergillus Flavus for Control of Aflatoxin Contamination in Crops |
WO2014191917A1 (en) * | 2013-05-27 | 2014-12-04 | Universita' Cattolica Del Sacro Cuore | Non-toxigenic strain of aspergillus flavus |
CN107245453A (en) * | 2017-06-02 | 2017-10-13 | 中国农业科学院农产品加工研究所 | One plant is not produced malicious aspergillus flavus and its application in terms of aflatoxin degradation |
CN107279686A (en) * | 2017-06-02 | 2017-10-24 | 中国农业科学院农产品加工研究所 | Application of the malicious aspergillus flavus in terms of aflatoxin degradation is not produced |
CN107177516A (en) * | 2017-07-07 | 2017-09-19 | 福建农林大学 | A kind of aspergillus flavus avirulent strain and its application of preventing and treating aspergillus flavus pollution |
CN107828694A (en) * | 2017-11-29 | 2018-03-23 | 中国农业科学院油料作物研究所 | One plant height effect suppresses serratia marcescens biocontrol microorganisms and its application of aspergillus flavus synthesis aflatoxin |
CN108531407A (en) * | 2018-04-27 | 2018-09-14 | 福建农林大学 | One plant of method that structure of aflatoxigenic strain and prevention aspergillus flavus do not pollute |
CN109913375A (en) * | 2019-04-03 | 2019-06-21 | 吉林大学 | A kind of Aspergillus flavus of not toxin producing, the composition containing it and its application |
Non-Patent Citations (5)
Title |
---|
FRISVAD, J.C.等: "Taxonomy of Aspergillus section Flavi and their production of aflatoxins, ochratoxins and other mycotoxins", STUDIES IN MYCOLOGY * |
VISHWAMBAR NAVALE等: "Aspergillus derived mycotoxins in food and the environment: Prevalence, detection, and toxicity", TOXICOLOGY REPORTS * |
刘俊等: "不产毒黄曲霉菌株的筛选鉴定及分子机理研究", 花生学报 * |
惠明等: "酒曲中一株不产毒黄曲霉的分离与鉴定", 中国酿造 * |
朱婷婷等: "湖北省典型花生种植区土壤中黄曲霉菌分布及产毒力研究", 中国油料作物学报 * |
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN113322189A (en) * | 2021-06-16 | 2021-08-31 | 中国农业科学院油料作物研究所 | Aspergillus flavus HuBXY33 without producing toxicity and application thereof |
CN113322189B (en) * | 2021-06-16 | 2022-03-11 | 中国农业科学院油料作物研究所 | Aspergillus flavus HuBXY33 without producing toxicity and application thereof |
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