Disease infection factor of wheat scab and application of disease infection factor

Abstract

The invention discloses a susceptible factor of wheat scab and application thereof, and belongs to the technical field of prevention and treatment of the wheat scab. Experiments prove that LA and PC (-- / 18: 2) belong to susceptible factors of wheat scab, and can promote field planting of pathogenic bacteria of the wheat scab, accelerate mycelial growth, increase the diffusion range and the like, and further deepen the infection degree of the wheat scab. Furthermore, LA and PC (-- / 18: 2) are utilized to develop a novel prevention and treatment agent for the wheat scab, it is found through a large number of screening that glycerin or tetrandrine can effectively prevent and treat the wheat scab caused by LA or PC (-- / 18: 2) induction, a preparation method of tetrandrine is provided, and the novel prevention and treatment agent for the wheat scab can be used for preventing and treating the wheat scab caused by LA or PC (-- / 18: 2) induction. The extracting solution containing the pachytetrandrine, which is prepared by the method disclosed by the invention, has a better prevention and treatment effect. Meanwhile, since the tetrandrine is derived from a natural plant, the tetrandrine has the characteristics of no residue, no pollution and the like, and is more beneficial to environmental protection.

Description

Technical Field

[0001] The present invention relates to the technical field of prevention and control of Fusarium head blight, in particular to susceptibility factors of wheat Fusarium head blight and their applications. Background Art

[0002] Wheat (Triticum aestivum) is an important food crop in the world, while Fusarium head blight seriously threatens the yield and quality of wheat. Fungi of the genus Fusarium are the main pathogens of Fusarium head blight. After the wheat ear is infected with the pathogen of Fusarium head blight, it will not only cause poor development of wheat grains, reduce wheat yield and deteriorate flour quality. At the same time, it accumulates a variety of mycotoxins, causing diseases such as anorexia, diarrhea and vomiting in humans and animals, seriously threatening the health of humans and animals. In addition, about 83.2% of the grain samples collected from the areas affected by Fusarium head blight are contaminated with toxins, and more than 31.2% of the samples exceed the toxin content standard (GB 13078-2017: ≤1 mg / kg). Fusarium head blight has gradually become the biggest threat to wheat production. Conducting research on wheat resistance to Fusarium head blight is of great significance for ensuring food security.

[0003] The prevention and control of wheat Fusarium head blight mainly depends on environmental conditions, pathogen characteristics and wheat variety characteristics during the growing season, and mainly reduces yield loss and toxin accumulation by taking corresponding measures before, during and after wheat flowering. These measures include chemical control, planting resistant varieties, biological control, improving tillage and irrigation conditions, etc. Limited by the diversity of Fusarium head blight pathogen strains, tenacious vitality, fast mutation rate, and the interaction between wheat resistance to Fusarium head blight and genotype and environment, it is still unknown whether the current prevention and control measures of Fusarium head blight can be effective in the long term. Continuing to deeply study the pathogenesis of Fusarium head blight pathogens, excavating susceptible factors and disease-resistant factors among them, and continuously developing new prevention and control measures and breeding strategies based on them are important measures to promote breeding for resistance to Fusarium head blight. Summary of the Invention

[0004] The purpose of the present invention is to provide susceptibility factors of wheat Fusarium head blight and their applications to solve the problems existing in the above-mentioned prior art. The present invention develops a new type of fungicide for preventing and controlling wheat Fusarium head blight by using LA and PC(-- / 18:2). After a large number of screenings, it is found that tetrandrine and glycerol can effectively prevent and control wheat Fusarium head blight induced by PC(-- / 18:2).

[0005] To achieve the above purpose, the present invention provides the following solutions:

[0006] The present invention provides the application of LA and / or PC(-- / 18:2) in any one of the following:

[0007] Application in inducing wheat Fusarium head blight;

[0008] Application in optimizing the test system for wheat scab resistance;

[0009] Application in screening wheat scab control agents.

[0010] Optionally, the LA promotes the colonization of the scab pathogen;

[0011] The PC(-- / 18:2) promotes the growth rate and spread range of the mycelia of the scab pathogen.

[0012] The present invention also provides the application of tetrandrine and / or glycerol in controlling wheat scab.

[0013] Optionally, the tetrandrine and / or glycerol play a role in controlling wheat scab by inhibiting LA and / or PC(-- / 18:2).

[0014] Optionally, the preparation method of the tetrandrine comprises the following steps:

[0015] Take tetrandra root and add it to water, heat and boil, then add an equal volume of chloroform, extract overnight, and then remove the chloroform to obtain it.

[0016] Optionally, the mass-volume ratio of the tetrandra root to water is 10 g: 0.3 L;

[0017] The boiling is to boil until the volume is 0.12 times the original volume.

[0018] The present invention also provides a wheat scab control agent, and the active ingredient of the agent comprises tetrandrine and / or glycerol.

[0019] Optionally, the preparation method of the tetrandrine comprises the following steps:

[0020] Take tetrandra root and add it to water, heat and boil, then add an equal volume of chloroform, extract overnight, and then remove the chloroform to obtain it.

[0021] Optionally, the mass-volume ratio of the tetrandra root to water is 10 g: 0.3 L;

[0022] The boiling is to boil until the volume is 0.12 times the original volume.

[0023] The present invention discloses the following technical effects:

[0024] It has been experimentally verified that LA and PC(-- / 18:2) are susceptibility factors for Fusarium head blight of wheat. They can deepen the infection degree of Fusarium head blight of wheat by promoting the colonization of pathogenic bacteria of Fusarium head blight, accelerating the growth and spreading range of hyphae, etc. Further, the present invention uses LA and PC(-- / 18:2) to develop a new type of control agent for Fusarium head blight of wheat. Through a large number of screenings, it is found that glycerol or tetrandrine can effectively control Fusarium head blight of wheat induced by LA or PC(-- / 18:2) respectively, and a preparation method of tetrandrine is provided. The extract containing tetrandrine prepared by the method of the present invention has a better control effect. At the same time, since tetrandrine is derived from natural plants and has the characteristics of no residue and no pollution, it is more beneficial to environmental protection.

[0025] Moreover, LA and PC(-- / 18:2) can be further used to optimize the field resistance test system for Fusarium head blight of wheat, providing technical conditions for exploring resistance genes of Fusarium head blight of wheat. Brief Description of the Drawings

[0026] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required to be used in the embodiments. Obviously, the drawings in the following description are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can be obtained based on these drawings.

[0027] Figure 1 To screen differential metabolites of anthers under different treatments; A: Analysis of differential metabolites of different anther treatments by non-target metabolomics, B: Relative quantification of LA and PC(-- / 18:2) in anthers;

[0028] Figure 2 For the effects of LA and PC(-- / 18:2) on Fusarium; A: Effects of adding 1 mM LA and / or 1 mM PC(-- / 18:2) (PC) on the growth of Fusarium graminearum; B: Statistical analysis of growth rate; C: Number of conidia of Fusarium graminearum germinated in liquid medium after inoculation for 12 h;

[0029] Figure 3 For the effect of LA on the colonization of Fusarium;

[0030] Figure 4 For the phenotype after co-culturing different spike tissues at the flowering stage of wheat with 1×10 5 compound conidia of Fusarium for 48 h;

[0031] Figure 5Effect of tetrandrine on Fusarium; A: Inhibitory effect of tetrandrine on Fusarium; B: Phenotype of Stephania tetrandra plants; C: Extraction of tetrandrine from Stephania tetrandra by different methods and its content; D: Inhibitory effect of tetrandrine extracted by different methods on Fusarium

[0032] Figure 6 Incidence of scab in different groups; A: Induction of scab in different treatments of wheat ears; B: Number of scabbed florets in different treatments of wheat ears; C: Detection of linoleic acid content in wheat ears after spraying 3% glycerol Detailed implementation mode

[0033] Now, various exemplary embodiments of the present invention will be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, characteristics, and implementation schemes of the present invention

[0034] It should be understood that the terms used in the present invention are only for describing specific embodiments and are not used to limit the present invention. Additionally, for the numerical ranges in the present invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Any intermediate value within any stated value or stated range, as well as each smaller range between any other stated value or intermediate value within the stated range, is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded from the range

[0035] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the present invention pertains. Although the present invention only describes preferred methods and materials, any methods and materials similar or equivalent to those described herein can also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the documents. In case of conflict with any incorporated document, the content of this specification shall prevail

[0036] Without departing from the scope or spirit of the present invention, various improvements and changes can be made to the specific implementation modes of the present invention specification, which are obvious to those skilled in the art. Other implementation modes obtained from the present invention specification are obvious to those skilled in the art. The present invention specification and examples are only exemplary

[0037] Regarding the use of "comprising", "including", "having", "containing", etc. in this article, they are all open-ended terms, meaning including but not limited to

[0038] The infection process of the Gibberella zeae pathogen in wheat begins with ascospores on plant residues. After maturation, they eject conidia, and this process is affected by temperature, the falling position, and the water potential of wheat tissues at the corresponding position. These conidia are dispersed by air currents to the ears of wheat during the flowering stage, enter through the withered pollen tubes or the gaps formed after the lemma and palea unfold, and rely on the internal anthers or the remains of anthers or pollen grains on the lemma surface as substrates to continuously proliferate, thereby infecting the entire ear. Thus, it can be seen that anthers play an important role in inducing the occurrence of Fusarium head blight.

[0039] In previous studies, to deeply analyze the potential metabolic components promoting the growth of Fusarium graminearum in anthers, anthers treated differently were selected for non-targeted metabolome detection. The research results showed that there were differences in the linoleic acid metabolic pathway, and the relative contents of LA (Linoleic acid) and PC (-- / 18:2) (1-unknown acyl-2-linoleoyl-sn-glycero-3-phosphocholine) in sterile anthers were lower than those in fertile anthers, and further decreased with the infection of Fusarium ( Figure 1 ). Therefore, it is speculated that LA and PC may be the key susceptibility factors promoting the infection of the Gibberella zeae pathogen in wheat anthers.

[0040] The LA and PC (-- / 18:2) used in this invention are from sigma, with the corresponding product numbers being LA (#L1376) and PC (-- / 18:2) (#429415); the compound Fusarium used is a highly pathogenic compound strain of Fusarium head blight in Sichuan Province, provided and preserved by the Wheat Research Institute of Sichuan Agricultural University.

[0041] Special medium for Gibberella zeae pathogen (Modified sperzieller Nahrstoffarmer agar, mSNA): KH2PO4 1g, KNO3 1g, MgSO4·7H2O 0.5g, KCl 0.5g, glucose 1g, sucrose 1g, and agar powder 20g. Add double-distilled water to 1L and sterilize at 121°C for 15 min.

[0042] Example 1 LA and PC are susceptibility factors for Fusarium head blight in wheat

[0043] The compound Fusarium was cultured using the special medium for Gibberella zeae pathogen, and 1 mM LA and PC (-- / 18:2) were added respectively for comparative experiments. Phenotypic analysis showed that LA significantly enhanced the conidia germination, aerial mycelium growth, and hyphal colonization ability of Fusarium, but also significantly inhibited the colony radial growth rate. In contrast, PC (-- / 18:2) significantly promoted the radial growth of Fusarium on the mSNA medium but decreased the conidia germination ( Figure 2) Thus, LA and PC(-- / 18:2) have differential regulatory effects on the growth of Fusarium graminearum and play important roles as key candidate susceptibility factors.

[0044] Furthermore, 1 mM LA was sprayed onto wheat leaves, and at the same time, 1×10 5 conidia of compound Fusarium were inoculated, and the leaves were kept moist for 48 hours. Seven days after inoculation, obvious lesions appeared on the leaves sprayed with LA ( Figure 3 ). Thus, it can be seen that LA can significantly promote the colonization of the pathogenic bacteria causing Fusarium head blight.

[0045] Exogenous pollen and 1 mM PC(-- / 18:2) were used to treat different wheat tissues, with H2O as the control. The results of microscopic observation are as Figure 4 shown. It can be seen that, compared with the control group, the treatment group added with 1 mM PC(-- / 18:2) and pollen showed significant acceleration of hyphal growth and expansion of the diffusion range in each wheat tissue.

[0046] Based on the above experimental results, LA and PC are susceptibility factors of wheat Fusarium head blight, which can deepen the degree of wheat infection with Fusarium head blight by promoting the colonization of the pathogenic bacteria causing Fusarium head blight, accelerating hyphal growth and expanding the diffusion range, etc.

[0047] Example 2 Screening of new control agents for wheat Fusarium head blight

[0048] Based on the discovery of the susceptibility factors in Example 1, to verify the possibility in its practical application. The present invention utilizes them to develop new control agents for wheat Fusarium head blight. Through a large number of screening tests, tetrandrine and glycerol can effectively control wheat Fusarium head blight caused by Fusarium induced by PC(-- / 18:2).

[0049] 1. Tetrandrine (Te)

[0050] A co-culture experiment was carried out using tetrandrine (source: Solarbio, #ST8130) and compound Fusarium (with 1 mM PC as the control). The results showed that tetrandrine could significantly relieve the promoting effect of PC(-- / 18:2) on the growth of Fusarium ( Figure 5 A in), and the EC 50 of tetrandrine was 311.38 mg / L, and the EC 95 was 1245.50 mg / L.

[0051] 1.1 Extraction of tetrandrine

[0052] Due to the water-insoluble property of tetrandrine, its practical application in production is limited. Therefore, the present invention further utilizes as Figure 5Extract Te using the two extraction methods (MeSt and TriSt) shown in C. The details are as follows.

[0053] MeSt extraction method: Take 50 g of Stephania tetrandra (plant phenotype shown in Figure 5 C), add 180 mL of methanol and extract overnight. After filtering through gauze, the MeSt extraction product is obtained, which is the Te extracted by MeSt.

[0054] TriSt extraction method: Take 50 g of Stephania tetrandra, add 1.5 L of H2O, boil in a boiling water bath until 180 mL (0.12 times of 1.5 L), then add an equal volume of chloroform and extract overnight. Then use a rotary evaporator to remove chloroform to obtain the TriSt extraction product, which is the Te extracted by TriSt.

[0055] The HPLC results show that the concentration of Te extracted by the MeSt method is 6010 ± 2750 mg L -1 , and the concentration of Te extracted by the TriSt method is 5.38 ± 1.88 mg L -1 . The concentration of Te extracted by MeSt is significantly higher than that extracted by TriSt, but both can significantly inhibit the growth of Fusarium, and the effective inhibitory concentration is far lower than the inhibitory concentration of tetrandrine against Fusarium head blight recorded in Patent CN113197214A ( Figure 5 D).

[0056] To further verify the effects of the above two extracts on the prevention and control of Fusarium head blight, the present invention conducts a spraying test on the ears of wheat at the flowering stage. First, select the ears of wheat at the flowering stage and spray 1×10 6 compound conidia of Fusarium, keep the humidity > 80%, and cultivate at 25°C for 48 hours. Then spray 1 mM PC (-- / 18:2) for induction. After 12 hours, spray Te extracted by MeSt (dilute the above extract with water to 5.01 ± 3.68 mg L -1 , dosage is 500 mL / m 2 ) and Te extracted by TriSt (concentration is 5.38 ± 1.88 mg L -1 , dosage is 500 mL / m 2 ), keep the humidity > 80%, and observe the phenotype after cultivating at 25°C for 20 days.

[0057] The results are as Figure 6 shown. It can be seen that the Te obtained by the TriSt extraction method has a significant effect on preventing and controlling Fusarium head blight, while the Te extracted by the MeSt method cannot effectively prevent and control Fusarium head blight, indicating that the extract containing tetrandrine prepared by the TriSt extraction method provided by the present invention has a better prevention and control effect.

[0058] 2. Glycerol

[0059] Carry out a spraying test on the ears of wheat at the flowering stage to verify whether glycerol has a control effect on Fusarium head blight. First, select the ears of wheat at the flowering stage and spray 1×10 6 conidia of Fusarium complex, maintain the humidity > 80%, and incubate at 25°C for 48 hours. Then spray glycerol with a volume fraction of 3% (dosage: 500 mL / m 2 ), maintain the humidity > 80%, and observe the phenotype after incubating at 25°C for 20 days.

[0060] The results are as Figure 6 shown. It can be seen that 3% glycerol can significantly reduce the content of LA in the ears of wheat ( Figure 6 C in the figure), and further reduce the occurrence of Fusarium head blight ( Figure 6 A - B).

[0061] In summary, LA and PC(-- / 18:2) as susceptible factors for the occurrence of Fusarium head blight can be used to develop corresponding control agents and measures based on them.

[0062] Example 3 Optimize the field resistance test system for wheat Fusarium head blight

[0063] In addition, in the aspect of breeding and selection of wheat offspring resistant to Fusarium head blight: Given that significant progress has been made in the mining of Fusarium head blight resistance genes (for example: Fhb1 - Fhb9, etc.), but it is difficult to induce Fusarium head blight in the field of wheat, and the traditional single - flower drip - injection is time - consuming and laborious, which has led to the slow progress of the identification work of resistant offspring.

[0064] The present invention selects the ears of wheat at the flowering stage and sprays 1×10 6 conidia of Fusarium complex, maintain the humidity > 80%, and incubate at 25°C for 48 hours. Then spray 1 mM PC(-- / 18:2) for induction, maintain the humidity > 80%, and observe the phenotype after incubating at 25°C for 20 days. The phenotype identification results show that compared with the normal induction of wheat Fusarium head blight, the incidence of Fusarium head blight in the ears of wheat after adding PC(-- / 18:2) treatment is significantly increased ( Figure 6 A - B in the figure).

[0065] Spray 1 mM LA onto the wheat leaves, and at the same time inoculate 1×10 5 conidia of Fusarium complex, and keep the humidity for 48 hours. On the 7th day after inoculation, the incidence of Fusarium head blight in the wheat leaves after spraying LA treatment is significantly increased ( Figure 3 ), It can be seen that the discovery of the susceptible factors in the present invention provides an important basis for further developing the optimization of the field resistance test system for wheat Fusarium head blight.

[0066] The embodiments described above are only descriptions of the preferred embodiments of the present invention, and do not limit the scope of the present invention. Without departing from the design spirit of the present invention, various deformations and improvements made by those of ordinary skill in the art to the technical solutions of the present invention shall fall within the protection scope determined by the claims of the present invention.

Claims

1. Use of LA and / or PC(-- / 18:2) in any of the following: Use in inducing Fusarium head blight of wheat; Use in optimizing the test system for resistance to Fusarium head blight of wheat; Use in screening agents for controlling Fusarium head blight of wheat.

2. The application according to claim 1, characterized in that The said LA promotes the colonization of the pathogenic bacteria causing Fusarium head blight; The said PC(-- / 18:2) promotes the growth rate and spreading range of the hyphae of the pathogenic bacteria causing Fusarium head blight.

3. Use of tetrandrine and / or glycerol in controlling Fusarium head blight of wheat.

4. The application according to claim 3, characterized in that, The said tetrandrine and / or glycerol play a role in controlling Fusarium head blight of wheat by inhibiting LA and / or PC(-- / 18:2).

5. The application according to claim 3, wherein The preparation method of the said tetrandrine comprises the following steps: Take tetrandra root, add it to water, heat and boil, then add an equal volume of chloroform, extract overnight, and then remove the chloroform to obtain it.

6. The application according to claim 5, characterized in that, The mass-volume ratio of the said tetrandra root to water is 10 g: 0.3 L; The said boiling is to boil until the volume is 0.12 times the original volume.

7. A medicament for preventing and treating wheat scab, characterized in that, The active ingredient of the said agent comprises tetrandrine and / or glycerol.

8. The medicament according to claim 7, wherein The preparation method of the said tetrandrine comprises the following steps: Take tetrandra root, add it to water, heat and boil, then add an equal volume of chloroform, extract overnight, and then remove the chloroform to obtain it.

9. The medicament according to claim 7, characterized in that, The mass-volume ratio of the said tetrandra root to water is 10 g: 0.3 L; The said boiling is to boil until the volume is 0.12 times the original volume.

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