Application of nicotinamide in prevention and treatment of southern root-knot nematode

Abstract

The application provides an application of nicotinamide in prevention and treatment of southern root-knot nematode. The nicotinamide has a strong inhibiting and killing effect on the second instar larvae (J2s) of the southern root-knot nematode and can inhibit the hatching of eggs. When the concentration of the nicotinamide is 15-25 mg / mL, the mortality of the J2s of the southern root-knot nematode and the inhibition rate of egg hatching are both above 90%, and the nicotinamide shows significant nematicidal activity. The nicotinamide used in the application has abundant sources, is harmless to human beings, livestock and environment, and meets the needs of green plant protection and sustainable development of agriculture.

Description

Technical Field

[0001] This invention belongs to the field of agricultural technology, specifically, it relates to the application of nicotinamide in the control of southern root-knot nematodes. Background Technology

[0002] Plant root-knot nematode (RKN; Meloidogyne spp.) diseases are the second largest category of agricultural diseases worldwide, after fungal diseases, with the southern root-knot nematode (Meloidogyne incognita) being the primary pathogen. The southern root-knot nematode has over 1700 host species and mainly damages vegetable and cash crops such as those in the Solanaceae, Cucurbitaceae, and Brassicaceae families. Globally, the economic losses caused by southern root-knot nematodes reach $78 billion annually. Research on green and efficient control technologies for southern root-knot nematodes has become a bottleneck problem urgently needing to be solved in my country's efforts to reduce chemical fertilizers and pesticides.

[0003] Currently, the main methods for controlling root-knot nematodes include physical methods such as chemical fumigation, high-temperature greenhouse treatment, and flooding, as well as agricultural cultivation methods such as crop rotation. The use of highly toxic chemical agents is the primary control measure, but the environmental and soil pollution caused by these agents is increasingly serious, and long-term application leads to increased resistance in root-knot nematodes (RKNs). Currently, the active ingredients in registered nematicides in my country mainly include abamectin, thiamethoxam, methyl isofenphos, ethoprophos, methyl bromide, aldicarb, amino oligosaccharides, dazomet, fenitrothion, carbofuran, and thiocarbofuran, most of which are organophosphates with high toxicity. Most of these carbamate, organophosphate pesticides, and fumigants are restricted or banned. Therefore, exploring highly effective, low-toxicity, and environmentally friendly nematicides is of great significance for the safe and green control of root-knot nematodes in southern China.

[0004] Nicotinamide, also known as nicotinamide, is an amide compound of nicotinic acid. It is widely distributed in many organisms, generally extracted from tobacco, and can also be synthesized by dehydration after nicotinic acid reacts with ammonia to form a salt. Nicotinamide participates in lipid metabolism, tissue respiration oxidation, and glycogenolysis within cells, and is an essential substance; deficiency will affect normal cellular respiration and metabolism. In clinical medicine, nicotinamide is often used to prevent and treat some nicotinic acid deficiency diseases such as pellagra, but there are no publicly available reports on its application in controlling southern root-knot nematodes. Summary of the Invention

[0005] The purpose of this invention is to provide the application of nicotinamide in the control of southern root-knot nematodes.

[0006] In order to achieve the purpose of this invention, in a first aspect, this invention provides the application of nicotinamide in the control of southern root-knot nematodes.

[0007] Secondly, the present invention provides the application of nicotinamide in the preparation of an insecticide for southern root-knot nematodes.

[0008] Furthermore, the active ingredient of the insecticide is nicotinamide, and the concentration of nicotinamide is 15-25 mg / mL when the insecticide is applied.

[0009] Furthermore, the insecticide exists in solution form.

[0010] Preferably, the insecticide is prepared with water. That is, the application involves dissolving nicotinamide in sterile water, which can be used as a nematicide.

[0011] Furthermore, the application includes:

[0012] (1) Nicotinamide has an inhibitory effect on the second-stage juveniles (J2s) of the southern root-knot nematode;

[0013] (2) Nicotinamide inhibits the hatching of Southern Root-knot Nematode eggs.

[0014] By employing the above technical solution, the present invention has at least the following advantages and beneficial effects:

[0015] (I) This invention is the first to discover that nicotinamide can produce a lethal effect on second-instar larvae of the southern root-knot nematode and inhibit the hatching of its eggs within a certain concentration range, and the effect is significant. When the concentration of nicotinamide is 15-25 mg / mL, the lethality rate and egg hatching inhibition rate of the southern root-knot nematode are both above 90%, showing significant nematicidal activity.

[0016] (II) Nicotinamide is derived from plants and microorganisms, and compared with other nematicides, it has advantages such as high efficiency, low toxicity, and low residue. Its use in the control of root-knot nematode disease is in line with the needs of green plant protection and sustainable agricultural development. Attached Figure Description

[0017] Figure 1 The effect of nicotinamide solution concentration on the hatching inhibition rate of *Strombus heterophylla* eggs in a preferred embodiment of the present invention. Different lowercase letters indicate significant differences. Detailed Implementation

[0018] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials used are all commercially available products.

[0019] Example 1

[0020] Example 1: Effect of different concentrations of nicotinamide on the lethality of J2s nematode of southern root-knot nematode

[0021] 1. Sample preparation

[0022] The nicotinamide (CAS: 98-92-0) used in this embodiment was purchased from Shanghai Maclean Biochemical Technology Co., Ltd.

[0023] Dissolve 5–25 mg of high-purity (≥98%) nicotinamide in sterile distilled water to prepare the test solution. The concentrations of each solution are 25 mg / mL, 20 mg / mL, 15 mg / mL, 10 mg / mL and 5 mg / mL, respectively.

[0024] 2. Hatching of Southern Root-Knot Nematodes

[0025] Tomato roots infected with southern root-knot nematodes were collected and washed to remove soil particles adhering to the surface. Mature egg sacs were picked from the roots using sterile forceps under a stereomicroscope. The picked egg sacs were thoroughly washed with a 2% sodium hypochlorite solution for 3 minutes, and then rinsed with sterile water to remove any remaining sodium hypochlorite solution. The washed egg sacs were placed in a 500-mesh sieve in a petri dish containing sterile water and incubated in a dark incubator at 27°C for 3-4 days. The hatched southern root-knot nematodes J2s were collected for experimental use.

[0026] 3. Experimental Methods

[0027] Approximately 100 J2s suspensions were added to the wells of a 48-well cell culture plate. The number of J2s in each well was observed and recorded under a stereomicroscope. Then, 100 μL of sample solutions at concentrations of 25 mg / mL, 20 mg / mL, 15 mg / mL, 10 mg / mL, and 5 mg / mL were added to each well, with an equal volume of sterile water as a blank control. Each experimental group was performed in triplicate. The 48-well cell culture plates containing J2s were placed in a dark environment at 27°C. The number of dead J2s from *Strombus heterophyllum* was observed and counted under a stereomicroscope at 6 h, 12 h, 18 h, 24 h, and 30 h. The nematodes were considered dead when they were stiff and immobile. The J2s lethality rate was calculated.

[0028] The corrected mortality rate is calculated as follows: Corrected mortality rate % = (Nematode mortality rate in the treatment group - Nematode mortality rate in the control group) / (1 - Nematode mortality rate in the control group) × 100%

[0029] 4. Experimental Results

[0030] The toxic effects of nicotinamide on the southern root-knot nematode J2s are shown in Tables 1 and 2.

[0031] Table 1. The toxicity (%) of nicotinamide against the southern root-knot nematode J2s.

[0032]

[0033] Table 2. Calculation results of nicotinamide toxicity determination against Southern root-knot nematode.

[0034] SPSS 25.0 6 y = 2.473x - 2.965 15.802 14.320～21.313 SPSS 25.0 12 y = 3.459x - 3.601 10.993 8.758～13.849 SPSS 25.0 18 y = 3.738x - 3.095 6.733 5.677～10.011

[0035] The results showed that the mortality rate of *J2s*, a southern root-knot nematode, increased with increasing nicotinamide concentration and treatment time (Table 1). When the treatment time was 24 h, the corrected mortality rates of *J2s* for nicotinamide solutions of 25 mg / mL, 20 mg / mL, 15 mg / mL, 10 mg / mL, and 5 mg / mL were 99.32%, 98.63%, 92.44%, 86.25%, and 38.14%, respectively. Furthermore, the corrected mortality rates exceeded 90% for concentrations ranging from 15 to 25 mg / mL.

[0036] Table 2 shows that when the treatment time is 6h, 12h, and 18h, the LC50 of nicotinamide for *Symplocos spp.* is significantly higher. 50 The concentrations were 15.802 mg / mL, 10.993 mg / mL, and 6.733 mg / mL, respectively. This demonstrates that nicotinamide has a strong toxic effect on the southern root-knot nematode J2s.

[0037] Example 2: Effect of different concentrations of nicotinamide on the hatching inhibition rate of southern root-knot nematodes.

[0038] 1. Sample preparation

[0039] Same as Example 1.

[0040] 2. Obtaining Southern Root-knot Nematode Eggs

[0041] Collect tomato roots infected with southern root-knot nematodes and wash away any soil particles adhering to the surface. Cut the roots into 2-4 cm sections with scissors and place them in a beaker. Add 10% sodium hypochlorite and shake thoroughly for three minutes. Pour the contents of the shaken beaker evenly into a sieve (sieve mesh sizes from top to bottom: 80 mesh, 140 mesh, 180 mesh, 200 mesh, 500 mesh). Rinse with sterile water 5-7 times to obtain sufficient nematode eggs and to wash away excess sodium hypochlorite. Rinse the 500 mesh sieve with a wash bottle and collect the rinsing liquid in a beaker to obtain an egg suspension.

[0042] 3. Experimental Methods

[0043] A fresh egg suspension of approximately 100 eggs was added to the wells of a 48-well cell culture plate. The number of eggs in each well was observed and recorded under a stereomicroscope. Then, 100 μL of sample solutions at concentrations of 25 mg / mL, 20 mg / mL, 15 mg / mL, 10 mg / mL, and 5 mg / mL were added to each well, with an equal volume of sterile water as a blank control. Each experimental group was performed in triplicate. The 48-well cell culture plates containing eggs were placed in the dark at 27°C. The hatching number of *Symplocos spp.* was observed under a stereomicroscope every 12 hours. After 5 days, the hatching inhibition rate of *Symplocos spp.* eggs was calculated.

[0044] The relative inhibition rate of egg hatching is calculated as follows: Relative inhibition rate % = (Hatching rate of nematodes in the control group - Hatching rate of nematodes in the treatment group) / Hatching rate of nematodes in the control group × 100%

[0045] 4. Experimental Results

[0046] The results showed that the inhibition rate of hatching of southern root-knot nematode eggs increased with increasing nicotinamide concentration. Figure 1 Nicotinamide solutions at concentrations of 25 mg / mL, 20 mg / mL, 15 mg / mL, 10 mg / mL, and 5 mg / mL showed inhibition rates of 25.94%, 37.40%, 91.44%, 94.61%, and 95.90% on the hatching of Southern Root-knot Nematodes eggs, respectively. Furthermore, the hatching inhibition rate was above 90% when the concentration range was 15-25 mg / mL.

[0047] In summary, when the concentration of nicotinamide is between 15 and 25 mg / mL, the lethality and egg hatching inhibition rate against southern root-knot nematodes are both above 90%, demonstrating significant nematicidal activity.

[0048] Although the present invention has been described in detail above with general descriptions and specific embodiments, modifications or improvements can be made to it, which will be obvious to those skilled in the art. Therefore, all such modifications or improvements made without departing from the spirit of the present invention fall within the scope of protection claimed by the present invention.

Claims

1. Application of nicotinamide in the control of southern root-knot nematode (Meloidogyne incognita).

2. Application of nicotinamide in the preparation of insecticides for southern root-knot nematodes.

3. The application according to claim 2, characterized in that, The active ingredient of the insecticide is nicotinamide, and the concentration of nicotinamide is 15-25 mg / mL when the insecticide is applied.

4. The application according to claim 2, characterized in that, The insecticide exists in solution form.

5. The application according to claim 4, characterized in that, The insecticide is prepared with water.

6. The application according to any one of claims 1-5, characterized in that, The applications include: (1) Nicotinamide has an inhibitory and killing effect on the second instar larvae of the southern root-knot nematode. (2) Nicotinamide inhibits the hatching of Southern Root-knot Nematode eggs.

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