Method for efficiently breeding tomato leaf miners based on tobacco tomato double-host mode

Abstract

The invention discloses a method for efficiently breeding tomato leaf miners based on a tobacco tomato double-host mode, and belongs to the field of agricultural pest biological control. The method comprises the following steps: taking a tobacco plant as a primary host, and providing a spawning site for the tomato leaf miner; after the tomato leaf miners lay eggs, the tobacco plants with the eggs are separated, and after the eggs are hatched into larvae, the tomato plants serve as secondary hosts to replace the tobacco plants so that the larvae can grow; when the damage rate of the tomato plants reaches 80%, replacing part of the tomato plants with new tomato plants; part of the replaced tomato plants are used for continuous development of the larvae, and finally the emerged tomato leaf miners are obtained. Compared with the quantity of the tomato leaf miner larvae bred by a single tomato plant, the quantity of the tomato leaf miner larvae bred by the method is larger, the larva development stage is shorter, the adult eclosion rate is higher, and the egg laying amount is larger. And a foundation is laid for further treatment and research on the tomato leaf miners under laboratory conditions.

Description

Technical Field

[0001] The invention relates to the field of biological control of agricultural pests, and in particular to a method for efficiently breeding tomato leafminers based on a tobacco-tomato dual-host model. Background Art

[0002] Tuta absoluta, a tomato leafminer, belongs to the order Lepidoptera and the family Gelechiidae. It is commonly known as the tomato leafminer. It causes damage with its larvae. After hatching, the larvae will burrow into the host plant tissues to feed on the mesophyll and form tiny tunnels on the leaves. They can also bore into the fruit to form holes, or feed at the junction of the calyx and the fruit, eventually causing a serious reduction in the yield of the host plant, or even destroying the plant for re-seeding. This pest is most fond of eating tomatoes, which can cause a 80% to 100% reduction in yield in severe cases. It can also harm other Solanaceae plants such as potatoes, eggplants, and tobacco. It poses a serious threat to the yield of important Solanaceae crops, mainly tomatoes and potatoes.

[0003] The research on agricultural pest control is of great significance to the research on natural enemy resources. First, it can not only provide an in-depth understanding of the life history of pests, but also find the critical period of control by mastering the habits of pests. At the same time, the occurrence of pests in the field can be predicted based on their response to different environmental conditions; second, the interaction between host insects and natural enemies can be studied experimentally to evaluate the effectiveness of natural enemies in reducing the number of agricultural pests. Apply effective research results to practice to achieve efficient control of harmful organisms; third, provide a scientific basis for the rational use and protection of natural enemy resources and the development of more efficient and environmentally friendly biological control methods. Therefore, in order to solve the problem of Solanaceae crops being harmed by tomato leafminers, it is first necessary to effectively solve the breeding problem of tomato leafminers, and providing a sufficient number of experimental insects required by the laboratory is the primary basis.

[0004] Due to the factors such as the binge eating period and large appetite of the larvae of tomato leafminers, it is difficult to raise adults in large numbers under laboratory conditions. At present, the research on the breeding of this kind of pest mainly focuses on feed development and single-head breeding. Such as patents ZL202310145504.1, ZL202310556990.6 and ZL202411302823.X. Although a lot of preliminary preparation work is reduced under feed breeding conditions, there are many problems. Since this type of pest belongs to leafmining pests, feed feeding will affect its growth and development to a certain extent, such as the increase in the hatching time of tomato leafminer eggs, the decrease in the hatching rate of tomato leafminer larvae eggs, and the decrease in the number of eggs laid by tomato leafminers compared to the single-host tomato plant breeding conditions; and although the single-head breeding of tomato leafminers is convenient for studying the various stages of growth of a single larva, it is not conducive to a large number of host egg experiments. Therefore, how to solve the breeding problem of tomato leafminers under laboratory conditions is an urgent problem to be solved. Summary of the invention

[0005] The larval stage of the tomato leafminer has characteristics such as a long transition time and voracious appetite. In a laboratory environment, a large number of tomato plants are required to maintain the normal survival of its larvae. However, due to factors such as temperature and humidity that cannot be at the optimal value at a specific time and the sudden occurrence of pests and diseases, it is often difficult to ensure the number of required plant hosts.

[0006] The purpose of the present invention is to provide a method for efficiently breeding tomato leafminers based on a tobacco-tomato dual host model to solve the problems existing in the above-mentioned prior art. The present invention uses tobacco as a transitional plant because it has the advantages of large leaf area, higher tolerance, and shorter growth cycle. The dual host breeding system of tomatoes and tobacco can provide a larger living space, maximum number of utilization, etc., which appropriately reduces the competitiveness between tomato leafminers and improves the survival rate of larvae, thereby obtaining tomato leafminers with better quantity and quality.

[0007] To achieve the above object, the present invention provides the following solutions:

[0008] The present invention provides a method for efficiently breeding tomato leafminers based on a tobacco-tomato dual-host model, comprising the following steps:

[0009] Tobacco plants are used as primary hosts, providing a place for the tomato leafminer to lay eggs;

[0010] After the tomato leafminer lays eggs, the tobacco plants with the eggs are separated, and after the eggs hatch into larvae, the tomato plants are used as secondary hosts to replace the tobacco plants for the larvae to grow;

[0011] When the damage rate of the tomato plants reaches 80%, replacing some of the tomato plants with new tomato plants;

[0012] The replaced tomato plants are used for the larvae to continue to develop, and finally the tomato leaf miners after emergence are obtained.

[0013] Optionally, when the tobacco plant is used as a primary host to provide an egg-laying site for the tomato leafminer, the release amount of the tomato leafminer is 20 pairs of tomato leafminers per tobacco plant.

[0014] The leaf area ratio of tomato plants and tobacco plants is about 1:2. The present invention uses tobacco plants as primary hosts, and tobacco leaves provide more choices of egg-laying locations for tomato leaf miners than tomato leaves.

[0015] Optionally, the tobacco plants carrying eggs are separated 72 hours after the tomato leafminer lays eggs.

[0016] Optionally, the eggs hatching into larvae are hatching into 1st instar larvae.

[0017] Optionally, replacing some of the tomato plants is replacing 1 / 3 of the tomato plants.

[0018] Optionally, the larvae continue to develop into mature larvae of the fourth instar.

[0019] Optionally, after the larvae continue to develop into fourth-instar mature larvae, the fourth-instar mature larvae are moved to a new container, and after cocooning and pupation, the pupae are moved to another new container again, and the tomato leafminer is obtained after rain.

[0020] Optionally, the breeding conditions of the tomato leafminer are a temperature of 24-26° C., a humidity of 55-65%, and a photoperiod of 16L:8D.

[0021] Optionally, the tobacco plant is a tobacco plant with 6 to 8 true leaves.

[0022] Optionally, the tomato plant is a tomato plant with 8 to 10 true leaves.

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

[0024] The present invention uses tobacco as a transitional plant because it has the advantages of large leaf area, higher tolerance, shorter growth cycle, etc. Therefore, the tomato and tobacco dual-host breeding system can provide a larger living space, maximum number of utilization, etc., appropriately reduce the competitiveness between tomato leafminers, increase the survival rate of larvae, shorten the larval development period, and ensure the quality and quantity of adults.

[0025] The number of tomato leafminer larvae bred by the present invention is greater than that bred on a single tomato plant, the larval development stage is shorter, the adult emergence rate is higher, and the egg-laying amount is larger, laying a foundation for further control and research of tomato leafminers under laboratory conditions. DETAILED DESCRIPTION

[0026] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0027] It should be understood that the terms described in the present invention are only for describing a particular embodiment and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0028] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may 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 associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0029] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to those skilled in the art. The present invention description and examples are exemplary only.

[0030] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0031] All the embodiments of the present invention were carried out in the greenhouse of the Institute of Biological Control of Jilin Agricultural University and the Natural Enemy Insect Industry R&D Base, and the implementation time was March 2024.

[0032] Example 1: Mass breeding of tomato leafminer adults in a tobacco and tomato dual host model

[0033] Test insects: Tomato leafminer Tuta absoluta (collected from the vegetable base in Nong'an County, Changchun City, Jilin Province in 2024), bred for 20 generations in the breeding room of the Institute of Biological Control, Jilin Agricultural University.

[0034] Host plants for testing: Tobacco (Beijing tobacco) plants, when the tobacco plants grow to 6 to 8 true leaves; tomato (Rich No. 1) plants, when the tomato plants grow to 8 to 10 true leaves.

[0035] (1) Cultivation of tobacco host plants

[0036] Tobacco seeds were placed in seedling trays for seedling cultivation (single seed sowing, 72 holes), and the seedlings were observed daily. When the tobacco seedlings grew to 4 true leaves, they were transplanted to 1.5L pots (one plant per pot). The nutrient soil used for the above seedling cultivation and transplanting was humus black soil, collected from Jilin Shenzhibei Fertilizer Co., Ltd. When the tobacco seedlings grew to 6-8 true leaves, they were ready for testing.

[0037] (2) Cultivation of tomato host plants

[0038] Tomato seeds were placed in seedling trays (105 holes) for seedling cultivation. The seedlings were observed daily and transplanted into 1.5L pots (one plant per pot) when the tomato seedlings grew to 4 true leaves. The nutrient soil used for the above seedling cultivation and transplanting was humus black soil, collected from Jilin Shenzhibei Fertilizer Co., Ltd. When the tomato plants grew to 8-10 true leaves, they were ready for testing.

[0039] In steps (1) and (2), the host plants are cultivated in a greenhouse of the Institute of Biological Control of Jilin Agricultural University from March to November each year. The daily light intensity in the greenhouse is higher than 12 hours through artificial lighting, the temperature is controlled between 20 and 35° C. through artificial temperature control, and the relative humidity is between 50 and 60%.

[0040] (3) Preparation of adult tomato leafminers

[0041] A tobacco plant cultivated in step (1) is placed in an insect cage, and test insects are released at a ratio of 20 pairs (one male and one female) of tomato leafminer adults per tobacco plant. After the tomato leafminer adults lay eggs for 72 hours, the tobacco plant cultivated in step (1) is used to replace the tobacco plant in the insect cage for standby use, and the replaced tobacco plant with tomato leafminer eggs is continued to be used for hatching eggs.

[0042] When the eggs are hatched into first-instar larvae, the tobacco plants are replaced with the tomato plants cultivated in step (2), and the plants are transferred to a new insect cage. When the damage rate of the tomato plants in the insect cage reaches 80%, 1 / 3 of the tomato plants in the insect cage are promptly replaced with the tomato plants cultivated in step (2), and the replaced tomato plants are placed in a new insect cage. When the larvae of the tomato leafminer develop into the fourth-instar mature larvae, the larvae are moved to a plastic box covered with a plastic film, and after they are cocooned and pupated, all the pupae are picked out and placed in a test tube. After the adult insects emerge, they are experimental insects.

[0043] The breeding climate conditions are 24-26℃, 55-65%RH, and 16:8L:D.

[0044] Example 2 Comparison of development time of tomato leafminer larvae

[0045] Test insects: Tomato leafminer Tuta absoluta (collected from the vegetable base in Nong'an County, Changchun City, Jilin Province in 2024), bred for 20 generations in the breeding room of the Institute of Biological Control, Jilin Agricultural University.

[0046] Host plants for testing: Tobacco (Beijing tobacco) plants, when the tobacco plants grow to 6 to 8 true leaves; tomato (Rich No. 1) plants, when the tomato plants grow to 8 to 10 true leaves.

[0047] (1) Comparison of the hatching rate of tomato leafminer eggs: The tomato leafminer adults (initial eclosion < 6h) bred on single-host tomato plants and the leafminer adults (initial eclosion < 6h) obtained by the invasive pest biological control research group of Jilin Agricultural University were obtained according to the tomato and tobacco dual-host breeding method in Example 1. The tomato leafminers obtained by the two different breeding methods were fully mated to provide the required amount of eggs for the experiment. After 72 hours of egg laying, the tobacco and control group plants were removed. Eggs were randomly selected from the two plants, 30 eggs per group, and repeated 3 groups. The number of eggs laid by the tomato leafminer on the plants was counted, and the number of larvae hatched was observed regularly every day, and the number of unhatched eggs was counted until all eggs were successfully hatched. The hatching rate of larval eggs under the two feeding methods was compared. Hatching rate (%) = number of hatched insects / total egg laying × 100%.

[0048] (2) Comparison of the hatching time of tomato leafminers: The invasive pest biological control research group of Jilin Agricultural University obtained tomato leafminers adults (initial eclosion <6h) bred on single-host tomato plants and leafminers adults (initial eclosion <6h) obtained according to the tomato and tobacco dual-host breeding method in Example 1. The tomato leafminers obtained by the two different breeding methods each provided the required amount of eggs for the experiment after sufficient mating. Randomly select eggs, 30 eggs per group, and repeat 3 groups. Place them in test tubes respectively, observe and record them regularly every day until all eggs hatch into first-instar larvae. Compare the time it takes for larval eggs to hatch into first-instar larvae under the two feeding systems.

[0049] (3) Comparison of the duration of tomato leafminers: The invasive pest biological control research group of Jilin Agricultural University obtained tomato leafminers adults (initial eclosion <6h) bred on single-host tomato plants and leafminers obtained according to the tomato and tobacco dual-host breeding method in Example 1. The tomato leafminers obtained by the two different breeding methods each provided eggs required for the experiment after sufficient mating. After hatching and developing into the larval stage, they were the larvae required for the experiment. Use a brush to move the eggs on the plant to a new tobacco plant, leaving only 30 eggs, 30 eggs in a group, and repeat 3 groups. Check regularly every day, record when the eggs hatch into first-instar larvae, until the larvae develop to the pupal stage, and compare the duration of the larvae under the two feeding systems.

[0050] (4) Comparison of the survival rate of tomato leafminers from larvae to pupae: Tomato leafminers (<6 hours after initial emergence) bred on single-host tomato plants and leafminers (<6 hours after initial emergence) obtained by the Biological Control of Invasive Pests Research Group of Jilin Agricultural University were obtained according to the dual-host breeding method of tomato and tobacco in Example 1. The tomato leafminers obtained by the two different breeding methods provided the eggs required for the experiment after sufficient mating, and the larvae required for the experiment were hatched and developed into larvae after hatching. Mature larvae were randomly selected with a brush, and 30 larvae were grouped together, and repeated for 3 groups. Check regularly every day, record the number of mature larvae that pupated every day, and compare the larval life under the two feeding systems.

[0051] Example 3 Comparison of the reproductive capacity of tomato leafminers

[0052] Test insects: Tomato leafminer Tuta absoluta (collected from the vegetable base in Nong'an County, Changchun City, Jilin Province in 2024), bred for 20 generations in the breeding room of the Institute of Biological Control, Jilin Agricultural University.

[0053] Host plants for testing: Tobacco (Beijing tobacco) plants, when the tobacco plants grow to 6 to 8 true leaves; tomato (Rich No. 1) plants, when the tomato plants grow to 8 to 10 true leaves.

[0054] (1) Determination of the individual size of tomato leafminers: Tomato leafminers adults (<6 hours after initial emergence) bred on single-host tomato plants and leafminers adults (<6 hours after initial emergence) obtained by the Biological Control of Invasive Pests Research Group of Jilin Agricultural University were obtained according to the dual-host breeding method of tomato and tobacco in Example 1. The body lengths of female and male and female adults under the two rearing methods were measured using a micrometer.

[0055] (2) Determination of the emergence rate of tomato leafminer: Tomato leafminer pupae were obtained from Implementation Case 1, and 30 pupae were grouped together for three times. After the pupae were fully emerged, the number of moths that emerged was investigated. Similarly, tomato leafminer pupae bred on a single host tomato plant were obtained from the Invasive Pest Biological Control Research Group of Jilin Agricultural University. 30 pupae were grouped together for three times. After the pupae were fully emerged, the number of moths that emerged was investigated. The above research was carried out in an artificial climate room with the climatic conditions of (25±1)℃, humidity RH (60±5)%, and photoperiod 14L:10D. Emergence rate (%) = number of moths that emerged / 30×100%.

[0056] (3) Determination of the number of eggs laid by female tomato leafminers: Adult tomato leafminers (dual hosts) were obtained from Case 1, and adults tomato leafminers (single hosts) bred on single-host tomato plants were obtained from the research group on biological control of invasive pests at Jilin Agricultural University. The number of adults of tomato leafminers raised on single hosts and those raised on dual hosts were sucked out using an insect sucker. A pot of cultivated tobacco plants was placed in an insect cage, and 20 pairs of adult tomato leafminers were released in the cage. After 72 hours of egg laying, the tobacco plants were taken out and placed in an insect cage containing tomato plants. They were observed regularly every day. After 72 hours of egg laying, the total number of eggs laid was recorded. The above experiment was repeated three times.

[0057] (4) Comparison of the lifespan of tomato leafminers: Tomato leafminers (dual hosts) (initial eclosion <6h) were obtained from Case Study 1, and tomato leafminers (single host) (initial eclosion <6h) were obtained from the research group on biological control of invasive pests at Jilin Agricultural University and bred on single-host tomato plants. The lifespans of male and female adults under the two breeding systems were compared.

[0058] Results and Analysis

[0059] (1) Comparison of hatching rates of tomato leafminer larvae eggs

[0060] By comparing the hatching rates of larvae and eggs, it was found that the hatching rate of larvae and eggs of tomato leafminer bred on dual hosts was as high as 85.7%, which was significantly higher than the hatching rate of 81.4% of larvae and eggs of tomato leafminer bred on single-host tomato plants (Table 1).

[0061] Table 1 Comparison of hatching rates of tomato leafminer larvae eggs bred by two methods

[0062]

[0063] (2) Comparison of hatching time of tomato leafminer larvae eggs

[0064] By comparing the hatching time of larval eggs, it was found that the hatching time of the larval eggs of tomato leafminer bred on dual-host plants was 4.83 days, and the hatching time of the larval eggs of tomato leafminer bred on single-host tomato plants was 4.87 days, and there was no significant difference between the two (Table 2).

[0065] Table 2 Comparison of hatching time of tomato leafminer larvae eggs bred by two methods

[0066]

[0067] (3) Comparison of development time of tomato leafminer larvae

[0068] By comparing the larval development time, it was found that the larval development time of tomato leafminer bred on dual hosts was 13.8 days, which was shorter than the larval development time of tomato leafminer bred on single-host tomato plants (15.7 days) (Table 3).

[0069] Table 3 Comparison of development time of tomato leafminer larvae bred by two methods

[0070]

[0071] (4) Comparison of survival rates of tomato leafminers from larvae to pupae

[0072] By comparing the larval development time, it was found that the survival rate of tomato leafminer larvae to the pupal stage in dual-host breeding was 88.6%, which was higher than the survival rate of tomato leafminer larvae to the pupal stage in single-host tomato plants (85.2%) (Table 4).

[0073] Table 4 Comparison of the survival rate of tomato leafminer larvae to pupae bred by two methods

[0074]

[0075] (5) Comparison of the individual sizes of tomato leafminers

[0076] By comparing the sizes of tomato leafminers bred in two different plant host models, it was found that the tomato leafminers raised in the dual-host system were the largest, with females averaging 7.1 mm in length and males averaging 6.4 mm in length; the tomato leafminers bred in the single-host system had an average female length of 6.7 mm and males averaging 5.5 mm (Table 5). Overall, the tomato leafminers bred in the dual-host model of tobacco and tomatoes were significantly taller.

[0077] Table 5 Comparison of body length of tomato leafminer bred by two methods

[0078]

[0079] (6) Comparison of emergence rate

[0080] By comparing the emergence rates, it was found that the emergence rate of tomato leafminers bred on dual hosts was as high as 91%, which was higher than the 86% emergence rate of tomato leafminers bred on single-host tomato plants (Table 6).

[0081] Table 6 Comparison of emergence rates of tomato leafminers bred by two methods

[0082]

[0083] (7) Comparison of the average number of eggs laid by each female tomato leafminer

[0084] By comparing the number of eggs laid by adult tomato leafminers raised in two different ways, it was found that the number of eggs laid by female tomato leafminers bred on two hosts was as high as 78.4, which was significantly higher than the 52.7 eggs laid by tomato leafminers bred on single-host tomato plants (Table 7).

[0085] Table 7 Comparison of egg laying of female tomato leafminers bred by two methods

[0086]

[0087] (8) Comparison of the lifespan of tomato leafminers

[0088] By comparing the lifespan of adult tomato leafminers raised in two different ways, it was found that the lifespan of female tomato leafminers bred on two hosts was as high as 25.52 days, which was higher than the lifespan of female tomato leafminers bred on a single host tomato plant (21.23 days) (Table 8).

[0089] Table 8 Comparison of the life span of adults of the tomato leafminer under two different breeding methods

[0090]

[0091] Based on the results of (1), (2), (3), (4), (5), (6), (7), and (8), the dual-host system for rearing tomato leafminer larvae has a higher hatching rate, a shorter larval life span, larger adult individuals, a higher emergence rate, and a larger egg-laying amount, which can more efficiently breed tomato leafminer.

[0092] The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. A method for efficiently breeding tomato leafminers based on a tobacco-tomato dual-host model, characterized in that: The following steps are involved: Tobacco plants are used as primary hosts, providing a place for the tomato leafminer to lay eggs; After the tomato leafminer lays eggs, the tobacco plants with the eggs are separated, and after the eggs hatch into larvae, the tomato plants are used as secondary hosts to replace the tobacco plants for the larvae to grow; When the damage rate of the tomato plants reaches 80%, replacing some of the tomato plants with new tomato plants; The replaced tomato plants are used for the larvae to continue to develop, and finally the tomato leaf miners after emergence are obtained.

2. The method according to claim 1, characterized in that When the tobacco plant is used as a primary host to provide an egg-laying site for the tomato leafminer, the release amount of the tomato leafminer is 20 pairs of tomato leafminers per tobacco plant.

3. The method according to claim 1, characterized in that The tobacco plants with eggs are separated 72 hours after the tomato leafminer lays eggs.

4. The method according to claim 1, characterized in that: The eggs hatching into larvae means hatching into 1st instar larvae.

5. The method according to claim 1, characterized in that The replacing of some tomato plants means replacing 1 / 3 of the tomato plants.

6. The method according to claim 1, characterized in that The larvae continue to develop into mature larvae up to the fourth instar.

7. The method according to claim 6, characterized in that After the larvae continue to develop into fourth-instar mature larvae, the fourth-instar mature larvae are moved to a new container, and after cocooning and pupation, the pupae are moved to another new container again, and the tomato leaf miner is obtained after rain.

8. The method according to claim 1, characterized in that The breeding conditions of the tomato leafminer are a temperature of 24-26° C., a humidity of 55-65%, and a photoperiod of 16L:8D.

9. The method according to claim 1, characterized in that: The tobacco plant is a tobacco plant with 6 to 8 true leaves.

10. The method according to claim 1, characterized in that The tomato plant is a tomato plant with 8 to 10 true leaves.

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