Rapid identification method for aphid resistance of broad beans

Abstract

The invention relates to the technical field of pest resistance identification, and discloses a rapid identification method for resistance of broad beans to aphids. Insect inoculation and observation are carried out under the manual control condition, the problems of uneven population density and the like in the natural environment are avoided, the identification period is shortened, only several days to several weeks are needed from insect inoculation to obtaining of a reliable identification result, compared with several months in the prior art, the efficiency is greatly improved, and the method is suitable for large-scale popularization and application. Manual control conditions can ensure that environmental conditions of each experiment are consistent, interference of environmental factors on experiment results is reduced, meanwhile, strict experiment operation and a data processing method also improve the accuracy and repeatability of the results, the identification results are more reliable, operation is relatively simple and convenient, complex equipment and a large amount of labor force are not needed, and the method is suitable for large-scale popularization and application. And the identification process can be completed only by carrying out simple insect inoculation and observation recording work in a manually controlled environment.

Description

Technical Field

[0001] This invention relates to the field of pest resistance identification technology, specifically a rapid method for identifying broad bean resistance to aphids. Background Technology

[0002] Aphids are one of the major pests affecting broad bean production. They feed on the sap of broad bean plants with their piercing-sucking mouthparts, leading to poor plant growth, curled and deformed leaves, and severely impacting the yield and quality of broad beans. Therefore, accurately identifying the resistance level of broad bean varieties to aphids is of great significance for the breeding and promotion of broad bean varieties.

[0003] In existing technologies, two methods are commonly used to identify broad bean resistance to aphids: one method involves planting the variety to be tested in the field, relying on natural insect infestations; however, this method is greatly affected by environmental conditions, with significant differences in insect population density across different regions and seasons, leading to inaccurate identification results. Furthermore, the identification process can take several months, requiring substantial time, manpower, and resources from planting to obtaining reliable results. In addition, due to the uncertainty of environmental factors, the results have poor repeatability, making it difficult to guarantee the stability and reliability of the identification results.

[0004] Another method involves inoculating aphids onto single leaves or plants during the seedling stage and assessing their severity by observing changes in aphid populations or the degree of damage to the plants. While this method reduces the impact of environmental factors to some extent, it still requires a long period of rearing and observation. The reproduction and growth rate of aphids are influenced by a combination of factors. Under natural or simple artificial conditions, accurately counting changes in aphid populations and assessing the degree of damage to plants requires continuous and prolonged tracking and observation, which increases the time and cost of identification and is not conducive to quickly screening out broad bean varieties with excellent resistance. Summary of the Invention

[0005] The purpose of this invention is to provide a rapid method for identifying the resistance of broad beans to aphids, so as to solve the problems mentioned in the background art.

[0006] To achieve the above objectives, the present invention provides the following technical solution: a rapid identification method for broad bean resistance to aphids, comprising the following steps: Step S1: Seed selection: Select seeds from the broad bean variety to be tested and the control broad bean variety with known resistance levels; Step S2, Seedling raising: Sow the seeds of the broad bean variety to be tested and the control broad bean variety in the same and suitable growing environment for seedling raising; Step S3, Seedling Selection: When the broad bean seedlings grow to a specific growth stage, select broad bean seedlings to be tested and control seedlings with consistent growth status. Step S4, Inoculation: Under artificial control conditions, inoculate the same number of adult aphids onto the leaves of the selected broad bean seedlings to be tested and the control broad bean seedlings. Step S5, Data Recording: After inoculation, at preset time intervals, regularly record the number of surviving aphids, the number of aphids that reproduce on the leaves of the broad bean seedlings to be tested and the control broad bean seedlings, as well as the symptoms of damage to the plants. Step S6, Index Calculation: Based on the recorded data, calculate the aphid survival rate inhibition rate, reproduction rate inhibition rate, and plant damage degree difference indicators of the tested broad bean variety compared with the control broad bean variety. Step S7, Resistance Identification: Based on the calculated aphid survival rate inhibition rate, reproduction rate inhibition rate, and plant damage severity difference indicators, combined with the preset resistance rating standards, the resistance level of the broad bean variety to aphids is identified.

[0007] Preferably, the specific growth period in step S3 is when the broad bean seedlings have grown 3-5 true leaves.

[0008] Preferably, the artificial control conditions in step S4 are: temperature maintained at 15-25℃, relative humidity maintained at 60%-70%, light intensity at 1200-1500 lx, and light exposure time at 12-14 hours per day.

[0009] Preferably, in step S4, the same number of adult aphids are inoculated, specifically 5-10 adult aphids are inoculated on each leaf.

[0010] Preferably, the preset time interval in step S5 is 12-24 hours.

[0011] Preferably, the formula for calculating the aphid survival rate inhibition rate in step S6 is: Survival inhibition rate = (Survival rate of aphids in control variety / Survival rate of aphids in control variety − Survival rate of aphids in test variety) × 100%; Among them, the aphid survival rate of the control variety is the ratio of the number of surviving aphids on the leaves of the control broad bean variety after inoculation to the number of inoculated aphids; the aphid survival rate of the test variety is the ratio of the number of surviving aphids on the leaves of the test broad bean variety after inoculation to the number of inoculated aphids.

[0012] Preferably, the formula for calculating the reproduction rate inhibition rate in step S6 is: Reproduction rate inhibition rate = (Reproduction rate of aphids in control variety / Reproduction rate of aphids in control variety − Reproduction rate of aphids in test variety) × 100%; Among them, the aphid reproduction rate of the control variety is the ratio of the number of aphids reproduced on the leaves of the control broad bean variety after inoculation to the number of inoculated aphids; the aphid reproduction rate of the test variety is the ratio of the number of aphids reproduced on the leaves of the test broad bean variety after inoculation to the number of inoculated aphids.

[0013] Preferably, the plant damage severity index in step S6 is obtained by comprehensively evaluating the degree of leaf curling, discoloration, and growth inhibition.

[0014] Preferably, the preset resistance rating standard in step S7 is: When the aphid survival rate inhibition rate is ≥70%, the reproduction rate inhibition rate is ≥70%, and the plant damage level is 1 or 2, it is identified as a highly resistant variety.

[0015] Preferably, when the aphid survival rate inhibition rate is less than 70% and the reproduction rate inhibition rate is less than 70% and the plant damage level is rated as level 3, it is identified as a moderately resistant variety; when the aphid survival rate inhibition rate is less than 30% and the reproduction rate inhibition rate is less than 30% and the plant damage level is rated as level 4, it is identified as a susceptible variety.

[0016] Compared with the prior art, the beneficial effects of the present invention are as follows: 1. By conducting insect inoculation and observation under artificially controlled conditions, problems such as uneven insect population density in the natural environment are avoided, shortening the identification cycle. From insect inoculation to obtaining reliable identification results, only a few days to a few weeks are needed, which is much more efficient than the months required by existing technologies. Furthermore, artificially controlled conditions can ensure that the environmental conditions are consistent for each experiment, reducing the interference of environmental factors on the experimental results. At the same time, strict experimental operations and data processing methods also improve the accuracy and repeatability of the results, making the identification results more reliable.

[0017] 2. The identification method of this invention has clear steps and is relatively simple to operate. It does not require complex equipment or a large amount of labor. The identification process can be completed simply by carrying out insect inoculation and observation and recording in an artificially controlled environment. This makes it easy to promote and apply in agricultural production. Moreover, the rapid and accurate resistance identification method can promptly screen out broad bean varieties with excellent aphid resistance characteristics, providing an important basis for the breeding of broad bean varieties, helping to improve the yield and quality of broad beans, and promoting the sustainable development of the broad bean industry. Attached Figure Description

[0018] Figure 1 A flowchart of a preferred embodiment of the rapid identification method for broad bean resistance to aphids provided by the present invention; Detailed Implementation

[0019] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0020] Please see Figure 1 As shown, a rapid method for identifying broad bean resistance to aphids includes the following steps: Step S1: Seed selection: Select seeds from the broad bean variety to be tested and the control broad bean variety with known resistance levels; Step S2, Seedling raising: Sow the seeds of the broad bean variety to be tested and the control broad bean variety in the same and suitable growing environment for seedling raising; Step S3, Seedling Selection: When the broad bean seedlings grow to a specific growth stage, select broad bean seedlings to be tested and control seedlings with consistent growth status. Step S4, Inoculation: Under artificial control conditions, inoculate the same number of adult aphids onto the leaves of the selected broad bean seedlings to be tested and the control broad bean seedlings. Step S5, Data Recording: After inoculation, at preset time intervals, regularly record the number of surviving aphids, the number of aphids that reproduce on the leaves of the broad bean seedlings to be tested and the control broad bean seedlings, as well as the symptoms of damage to the plants. Step S6, Index Calculation: Based on the recorded data, calculate the aphid survival rate inhibition rate, reproduction rate inhibition rate, and plant damage degree difference indicators of the tested broad bean variety compared with the control broad bean variety. Step S7, Resistance Identification: Based on the calculated aphid survival rate inhibition rate, reproduction rate inhibition rate, and plant damage severity difference indicators, combined with the preset resistance rating standards, the resistance level of the broad bean variety to aphids is identified.

[0021] It should be noted that using a control broad bean variety with known resistance levels as a reference standard helps to more accurately assess the resistance status of the broad bean variety being tested. Using the same growing environment can minimize the impact of environmental factors on the growth of broad bean seedlings and ensure the reliability of subsequent identification results. Suitable growing environment includes appropriate soil, temperature, humidity and other conditions, such as fertile, loose and well-aerated soil, and temperature maintained within the appropriate range for broad bean seed germination and seedling growth.

[0022] The optimal growth period is when the broad bean seedlings have grown 3-5 true leaves. At this time, the growth of the broad bean seedlings is relatively stable, their resistance to aphids is also more obvious, and it is easier to carry out subsequent insect inoculation operations. The consistent growth status is mainly reflected in the plant height, leaf size and color, ensuring that the starting conditions of the experiment are the same.

[0023] Artificially controlled conditions include a temperature maintained at 15-25℃, relative humidity maintained at 60%-70%, light intensity at 1200-1500 lx, and light duration at 12-14 hours per day. These environmental conditions can simulate the suitable living environment for aphids while ensuring the reproducibility of the experiment. Each leaf is inoculated with 5-10 adult aphids. This number ensures that there are enough aphids for observation and statistics, but also avoids excessive competition among aphids that could affect the experimental results.

[0024] The preset time interval is 12-24 hours. This time interval can track the growth and reproduction dynamics of aphids and the changes in the degree of damage to plants in a timely manner, providing detailed and accurate information for subsequent data analysis.

[0025] The formula for calculating the aphid survival rate inhibition rate is: Survival inhibition rate = (Survival rate of aphids in control variety / Survival rate of aphids in control variety − Survival rate of aphids in test variety) × 100%; Among them, the aphid survival rate of the control variety is the ratio of the number of surviving aphids on the leaves of the control broad bean variety after inoculation to the number of inoculated aphids; the aphid survival rate of the test variety is the ratio of the number of surviving aphids on the leaves of the test broad bean variety after inoculation to the number of inoculated aphids.

[0026] The formula for calculating the reproduction rate inhibition rate is: Reproduction rate inhibition rate = (Reproduction rate of aphids in control varieties / Reproduction rate of aphids in control varieties − Reproduction rate of aphids in the tested varieties) × 100%; Among them, the aphid reproduction rate of the control variety is the ratio of the number of aphids reproduced on the leaves of the control broad bean variety after inoculation to the number of inoculated aphids; the aphid reproduction rate of the test variety is the ratio of the number of aphids reproduced on the leaves of the test broad bean variety after inoculation to the number of inoculated aphids; the difference index of plant damage degree is obtained by comprehensively evaluating the degree of leaf curling, discoloration and growth inhibition of the plant.

[0027] The severity of plant damage is assessed by comprehensively evaluating the degree of leaf curling, discoloration, and growth inhibition. For example, detailed evaluation criteria can be established: Level 1 (minor damage, no obvious symptoms) is defined as leaves without curling or discoloration and with normal growth; Level 2 (mild damage, visible symptoms) is defined as leaves with slight curling and discoloration and slightly inhibited growth; Level 3 (moderate damage, obvious symptoms) is defined as leaves with severe curling and obvious discoloration and significantly inhibited growth; and Level 4 (severe damage, serious injury) is defined as leaves withered and plant growth stagnant.

[0028] The preset resistance rating criteria are: When the aphid survival rate inhibition rate is ≥70%, the reproduction rate inhibition rate is ≥70%, and the plant damage severity difference index is low, it is identified as a highly resistant variety. When the aphid survival rate inhibition rate is less than 70% and the reproduction rate inhibition rate is less than 70% and the plant damage severity difference index is moderate, it is identified as a moderately resistant variety. When the aphid survival rate inhibition rate is <30%, the reproduction rate inhibition rate is <30%, and the plant damage severity difference index is high, the plant is identified as a susceptible variety. For example: Select seeds of broad bean variety A to be tested and broad bean variety B, which is known to be highly susceptible to aphids.

[0029] Seedling raising: Sow seeds of variety A and variety B separately in flowerpots filled with fertile, loose and breathable soil, and place them in an artificial climate chamber with a temperature of 22℃, relative humidity of 65%, light intensity of 1300lx and light duration of 13 hours per day for seedling raising.

[0030] Seedling selection: When the broad bean seedlings have grown 4 true leaves, select 10 seedlings each of variety A and variety B with the same growth status (similar plant height, leaf size and color).

[0031] Inoculation: In the artificial climate chamber described above, 8 adult aphids were inoculated onto each leaf of the selected seedlings of variety A and variety B.

[0032] Data recording: After inoculation, the number of surviving aphids on the leaves, the number of aphids that reproduced, and the symptoms of damage to the plants were recorded every 18 hours for 5 consecutive days.

[0033] Indicator Calculation: Calculate the aphid survival rate inhibition rate: The average number of aphids surviving after inoculation on the leaves of variety B was 7, while the number of aphids surviving after inoculation was 8. Therefore, the survival rate of aphids in the control variety was: 7 ÷ 8 × 100% = 87.5%.

[0034] If the average number of aphids surviving after inoculation on the leaves of variety A is 2, then the aphid survival rate of the tested variety is (2÷8)×100%=25%; Aphid survival rate inhibition rate = [(87.5%−25%) / 87.5%]×100%≈71.4%.

[0035] Calculate the reproduction rate inhibition rate: The average number of aphids on the leaves of variety B after inoculation was 15. Therefore, the reproduction rate of aphids in the control variety was (15÷8)×100%=187.5%.

[0036] If the average number of aphids on the leaves of variety A after inoculation is 4, then the aphid reproduction rate of the tested variety is (4÷8)×100%=50%.

[0037] Reproduction suppression rate = [(187.5%−50%) / 187.5%]×100%≈73.3%.

[0038] Calculate the index of differences in plant damage severity: Variety B plants have severely curled and discolored leaves, and their growth is significantly inhibited, classified as level 3.

[0039] Variety A plants have slightly curled and discolored leaves, and their growth is slightly inhibited, so they are rated as level 2.

[0040] Resistance identification: Based on the criteria of aphid survival rate inhibition rate ≥70%, reproduction rate inhibition rate ≥70%, and low evaluation of plant damage severity, the tested broad bean variety A was identified as a highly resistant variety.

[0041] As can be seen from the above embodiments, the rapid identification method for broad bean resistance to aphids of the present invention can accurately and quickly identify the resistance level of broad bean varieties to aphids, and has high practicality and reliability.

[0042] It should be noted that, in this document, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitations, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0043] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A rapid method for identifying broad bean resistance to aphids, characterized in that, Includes the following steps: Step S1: Seed selection: Select seeds from the broad bean variety to be tested and the control broad bean variety with known resistance levels; Step S2, Seedling raising: Sow the seeds of the broad bean variety to be tested and the control broad bean variety in the same and suitable growing environment for seedling raising; Step S3, Seedling Selection: When the broad bean seedlings grow to a specific growth stage, select broad bean seedlings to be tested and control seedlings with consistent growth status. Step S4, Inoculation: Under artificial control conditions, inoculate the same number of adult aphids onto the leaves of the selected broad bean seedlings to be tested and the control broad bean seedlings. Step S5, Data Recording: After inoculation, at preset time intervals, regularly record the number of surviving aphids, the number of aphids that reproduce on the leaves of the broad bean seedlings to be tested and the control broad bean seedlings, as well as the symptoms of damage to the plants. Step S6, Index Calculation: Based on the recorded data, calculate the aphid survival rate inhibition rate, reproduction rate inhibition rate, and plant damage degree difference indicators of the tested broad bean variety compared with the control broad bean variety. Step S7, Resistance Identification: Based on the calculated aphid survival rate inhibition rate, reproduction rate inhibition rate, and plant damage severity difference indicators, combined with the preset resistance rating standards, the resistance level of the broad bean variety to aphids is identified.

2. The rapid identification method for broad bean resistance to aphids according to claim 1, characterized in that: The specific growth period in step S3 is when the broad bean seedlings have grown 3-5 true leaves.

3. The rapid identification method for broad bean resistance to aphids according to claim 1, characterized in that: The artificial control conditions in step S4 are: temperature maintained at 15-25℃, relative humidity maintained at 60%-70%, light intensity at 1200-1500 lx, and light exposure time at 12-14 hours per day.

4. The rapid identification method for broad bean resistance to aphids according to claim 1, characterized in that: In step S4, the same number of adult aphids are inoculated, specifically 5-10 adult aphids are inoculated on each leaf.

5. The rapid identification method for broad bean resistance to aphids according to claim 1, characterized in that: The preset time interval in step S5 is 12-24 hours.

6. The rapid identification method for broad bean resistance to aphids according to claim 1, characterized in that: The formula for calculating the aphid survival rate inhibition rate in step S6 is as follows: Survival inhibition rate = (Survival rate of aphids in control variety / Survival rate of aphids in control variety − Survival rate of aphids in test variety) × 100%; Among them, the aphid survival rate of the control variety is the ratio of the number of surviving aphids on the leaves of the control broad bean variety after inoculation to the number of inoculated aphids; the aphid survival rate of the test variety is the ratio of the number of surviving aphids on the leaves of the test broad bean variety after inoculation to the number of inoculated aphids.

7. The rapid identification method for broad bean resistance to aphids according to claim 1, characterized in that: The formula for calculating the reproduction rate inhibition rate in step S6 is: Reproduction rate inhibition rate = (Reproduction rate of aphids in control variety / Reproduction rate of aphids in control variety − Reproduction rate of aphids in test variety) × 100%; Among them, the aphid reproduction rate of the control variety is the ratio of the number of aphids reproduced on the leaves of the control broad bean variety after inoculation to the number of inoculated aphids; the aphid reproduction rate of the test variety is the ratio of the number of aphids reproduced on the leaves of the test broad bean variety after inoculation to the number of inoculated aphids.

8. The rapid identification method for broad bean resistance to aphids according to claim 1, characterized in that: The plant damage severity index in step S6 is obtained by comprehensively evaluating the degree of leaf curling, discoloration, and growth inhibition.

9. A rapid identification method for broad bean resistance to aphids according to claim 1, characterized in that: The preset resistance rating standard in step S7 is as follows: A highly resistant variety is identified when the aphid survival rate inhibition rate is ≥70%, the reproduction rate inhibition rate is ≥70%, and the plant damage severity difference index is low.

10. A rapid identification method for broad bean resistance to aphids according to claim 9, characterized in that: When the aphid survival rate inhibition rate is less than 70% and the reproduction rate inhibition rate is less than 70% with a moderate evaluation of the plant damage severity index, it is identified as a moderately resistant variety; when the aphid survival rate inhibition rate is less than 30% and the reproduction rate inhibition rate is less than 30% with a high evaluation of the plant damage severity index, it is identified as a susceptible variety.

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