Grain storage pest sex identification method and application

Abstract

The invention provides a stored grain pest sex identification method and application, and relates to the field of grain storage technology and agricultural insect and pest control. The method for identifying the sex of the stored grain pests comprises the following steps: (1) measuring the length and width of each section of tentacles of female and male adult stored grain pests; (2) analyzing the relationship between the total tentacle length, the length and / or width of each section and the gender of the adult stored grain pests; and (3) measuring the full-length antenna length and the length and / or width of each section of the adult stored grain pests to be measured, and identifying the gender of the adult stored grain pests according to the relationship in the step (2). The method for identifying the sex of the stored grain pests is simple, convenient, accurate and efficient, the stored grain pests still survive after identification is finished, and subsequent scientific research is facilitated.

Description

Technical Field

[0001] The invention relates to the fields of grain storage technology, agricultural insects and pest control, and in particular to a method for sex identification of stored-grain pests and its application. Background Art

[0002] Stored grain pests are one of the important biological factors that cause the loss of grain quantity and quality. The stable environment and sufficient food of grain storage provide an ideal breeding ground for their growth and reproduction. The control methods of stored grain pests include physical control, chemical control, biological control and ecological control methods. Identifying the sex of stored grain pests when they are alive is of great significance for pest control research, because stored grain pests of different sexes differ in behavioral habits and reproductive capacity. For example, female pests are usually responsible for laying eggs, while male pests are mainly involved in mating. By identifying sex, targeted control measures can be formulated. For example, during the peak period of female pests laying eggs, strengthen the protection of grain to reduce the chance of pest reproduction; or set up specific trapping devices based on the activity characteristics of male pests to reduce their mating success rate, thereby effectively controlling the pest population. In addition, identifying the sex of stored grain pests helps to gain a deeper understanding of the biological characteristics, reproductive laws, behavioral habits, etc. of pests, enrich the basic theoretical research on stored grain pests, and provide a more solid theoretical support for the integrated control of pests. . However, in the prior art, the sex of stored-grain pests is identified by observing the reproductive organs of stored-grain pests through dissection. Since the pests are dead after dissection, subsequent research cannot be carried out. Therefore, it is urgent to establish a method for sex identification of stored-grain pests while ensuring the survival of the pests. Summary of the invention

[0003] The purpose of the present invention is to provide a method for sex identification of stored-grain pests. After the identification, the stored-grain pests are still alive, and the method is simple, accurate and efficient.

[0004] The purpose of the present invention is achieved by the following technical solutions:

[0005] A method for identifying sex of stored-grain pests comprises the following steps:

[0006] (1) Measure the length and width of each segment of the antennae of female and male stored-grain pest adults;

[0007] (2) Analyze the relationship between the total length, segment length and / or width of antennae of adult stored-grain pests and their sex;

[0008] (3) measuring the total length of the antennae, the length of each segment and / or the width of the adult stored-grain pest to be tested, and identifying the sex of the adult stored-grain pest according to the relationship described in step (2).

[0009] In the present invention, the stored-grain pest is the grain borer.

[0010] In the present invention, the relationship between the total length of antennae of adult grain borers and gender is as follows: the total length of antennae of male adult grain borers is greater than 620 μm, and the total length of antennae of female adult grain borers is less than 620 μm.

[0011] In the present invention, the relationship between the width of each segment of the antennae of the adult grain borer and the sex is as follows: the width of the third flagellum of the male adult grain borer is less than 36 μm, and the width of the third flagellum of the female adult grain borer is greater than 36 μm.

[0012] In the present invention, the relationship between the length of each segment of the antennae of the adult grain borer and the sex is as follows: the length of the sixth flagellum of the male adult grain borer is greater than 100 μm, and the length of the sixth flagellum of the female adult grain borer is less than 100 μm.

[0013] In the present invention, in step (1), after taking photos with an electron microscope or a camera, the total length of the antennae, the length and width of each segment of the adult stored-grain pests are measured using a built-in ruler or image processing software. The image processing software may be CameraMeasure, SigmaScan Pro, Digimizer, Zonson2D image measurement software, Metus, Image-Pro Plus, etc.

[0014] Beneficial effects of the present invention: The method for sex identification of stored-grain pests provided by the present invention is simple, accurate and efficient. After the identification, the stored-grain pests are still alive, which is beneficial to subsequent scientific research. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 : Photo of adult grain borer.

[0016] Figure 2 The antennae of the grain borer and the morphology, width and length of each segment are shown. Sc: peduncle; Pe: pedunculate; F: flagellum. F1 is the first flagellum, and the others are similar. DETAILED DESCRIPTION

[0017] The following examples are used to further illustrate the present invention, but should not be construed as limiting the present invention. Without violating the essence and spirit of the present invention, any optimization and replacement of the present invention shall fall within the scope of the present invention.

[0018] Unless otherwise stated, the technical means involved in the following embodiments are all conventional techniques commonly used by technicians in this scientific field.

[0019] Example 1

[0020] 1. Collection and identification of test insects

[0021] (1) Collection of test insects

[0022] More than 3,000 adult insects were collected from 139 locations including farmers, processing plants, and granaries in various provinces and cities in China as test insects. Due to the large number of sources, only some of the test insect sources are listed in Table 1.

[0023] (1) Species identification

[0024] The test insects from the above 139 sources were morphologically identified according to GB / T 37719.4-2024 "Auxiliary Atlas for Inspection of Pests in Grain and Oil Storage Part 4: Boring Pests (Curculionidae, Leguminosae, Mylostomidae, Mylostomidae, Lycopodiidae)" and were determined to be Grain Borer ( Figure 1 ).

[0025] Table 1 Sources and collection locations of adults of Cephalopoda spp.

[0026] Serial number source Collection location Serial number source Collection location 1 Processing Plant Qiqihar, Heilongjiang 23 Grain depot Xuancheng, Anhui 2 Farmers Daqing, Heilongjiang 24 Farmers Shijiazhuang, Hebei 3 Grain depot Handan, Hebei 25 Processing Plant Tangshan, Hebei 4 Processing Plant Zhangjiakou, Hebei 26 Processing Plant Qinhuangdao, Hebei 5 Grain depot Cangzhou, Hebei 27 Grain depot Hunan Hengyang 6 Grain depot Zhengzhou, Henan 28 Grain depot Yichang, Hubei 7 Processing Plant Suihua, Heilongjiang 29 Grain depot Hubei Huanggang 8 Farmers Heze, Shandong 30 Grain depot Changsha, Hunan 9 Grain depot Anyang, Henan 31 Grain depot Zhangjiajie, Hunan 10 Farmers Liaocheng, Shandong 32 Grain depot Leshan, Sichuan 11 Grain depot Beijing 33 Grain depot Guang'an, Sichuan 12 Processing Plant Tianjin 34 Grain depot Loudi, Hunan 13 Grain depot Shanghai 35 Grain depot Nanchang, Jiangxi 14 Processing Plant Taiyuan, Shanxi 36 Grain depot Ziyang, Sichuan 15 Grain depot Jiaozuo, Henan 37 Grain depot Nanjing, Jiangsu 16 Grain depot Nanyang, Henan 38 Grain depot Xuzhou, Jiangsu 17 Grain depot Hefei, Anhui 39 Grain depot Yichun, Jiangxi 18 Grain depot Anhui Bozhou 40 Grain depot Huai'an, Jiangsu 19 Grain depot Xinyang, Henan 41 Grain depot Zhenjiang, Jiangsu 20 Grain depot Huaibei, Anhui 42 Grain depot Yingtan, Jiangxi 21 Grain depot Anhui Tongling 43 Grain depot Chengdu, Sichuan 22 Grain depot Wuhan, Hubei 44 Grain depot Suqian, Jiangsu

[0027] 2. Anatomical identification of sex and electron microscopy observation and determination

[0028] (1) Anatomical identification of sex

[0029] The collected test insects were cleaned by nitrogen purging, and placed under a stereo microscope for dissection and observation of their reproductive organs to identify their sex. Those with ovaries were females, and those with testes were males.

[0030] (2) Scanning electron microscope observation

[0031] After the test insects were identified by dissection, their heads and antennae were removed, placed in 0.1mol / L phosphate buffer, and treated with ultrasound for 30s, and then rinsed with ultrapure water three times. They were fixed in 2.5% glutaraldehyde for 2h; then washed with 0.1mol / L phosphate buffer and ultrapure water, three times each. The washed antennae were dehydrated with 20% ethanol aqueous solution, 25% ethanol aqueous solution, 30% ethanol aqueous solution, 35% ethanol aqueous solution, 40% ethanol aqueous solution, 45% ethanol aqueous solution, 50% ethanol aqueous solution, 55% ethanol aqueous solution, 60% ethanol aqueous solution, 65% ethanol aqueous solution, 70% ethanol aqueous solution, 75% ethanol aqueous solution, 80% ethanol aqueous solution, 85% ethanol aqueous solution, 90% ethanol aqueous solution, and 95% ethanol aqueous solution in sequence, with each dehydration lasting 5min. Finally, gradient drying is performed in sequence using 30% tert-butanol aqueous solution, 50% tert-butanol aqueous solution, 70% tert-butanol aqueous solution, 90% tert-butanol aqueous solution, 100% tert-butanol aqueous solution, tert-butanol-acetonitrile (3:1), tert-butanol-acetonitrile (2:1), tert-butanol-acetonitrile (1:1), tert-butanol-acetonitrile (1:2), and 100% acetonitrile. Among them, tert-butanol-acetonitrile (3:1) refers to a mixed solvent of tert-butanol and acetonitrile with a volume ratio of 3:1, and the others are analogous.

[0032] The dehydrated and dried insect sample (antenna) was glued to the scanning electron microscope sample stage by the ventral surface with conductive double-sided tape, gold was sprayed by ion sputtering, and the sample was observed and photographed by TM3000 scanning electron microscope at an accelerating voltage of 15 kV.

[0033] The antennae of the adult grain borer are composed of the stalk segment (Sc), the pedicel segment (Pe) and the flagellum segment (F) connected in sequence. Figure 2 ), wherein the flagellum is composed of the 1st to 8th flagellum. The microscopic image analysis and measurement software Digimizer 3.2 was used to measure the length and width of the stalk segment (Sc), pedicel segment (Pe), and the 1st to 8th flagellum segments in the antennae of each test insect. The length of each segment from the pedicel segment to the 7th flagellum refers to: the line connecting the midpoint of the line connecting the segment with the previous segment and the midpoint of the line connecting the segment with the next segment on the photo of the grain borer. For example, the length of F3 (the 3rd flagellum) refers to the line CD connecting the midpoint D of the line connecting F3 and F2 and the midpoint C of the line connecting F3 and F4. The length of F8 refers to: the maximum value of the line connecting the midpoint H of the line connecting F8 and F7 and the end of F8 on the photo of the grain borer. The length of the stalk segment refers to: the line AB connecting the midpoint B of the line connecting the stalk segment to the head of the grain borer and the midpoint A of the line connecting the stalk segment to the stalk on the photo of the grain borer. The width of each segment from the peduncle to the 8th whip segment refers to the largest dimension of each segment in the direction perpendicular to its length in the photo of the grain borer, for example, the width of F3 is MN, and the width of F6 is JK. The total antenna length is calculated as follows: total antenna length = peduncle length + peduncle length + 1st whip segment length + 2nd whip segment length + 3rd whip segment length + 4th whip segment length + 5th whip segment length + 6th whip segment length + 7th whip segment length + 8th whip segment length. Due to the large amount of data on more than 3,000 grain borer adults, only some of the data can be listed, see Tables 2 and 3 for details.

[0034] The total length of antennae, the length of each segment and the width of each segment of antennae of more than 3,000 male and female adults of P. pygmaea were analyzed, and it was found that the total length of antennae of male adults of P. pygmaea was greater than 620 μm, the width of the third flagellum was less than 36 μm, and the length of the sixth flagellum was greater than 100 μm; the total length of antennae of female adults of P. pygmaea was less than 620 μm, the width of the third flagellum was greater than 36 μm, and the length of the sixth flagellum was less than 100 μm.

[0035] Therefore, the sex of the adult rice borer can be identified by the following method (referred to as the method of the present invention): the adult rice borer is photographed by an electron microscope or an optical microscope or a camera (such as a professional camera, a high-resolution camera of a mobile phone, etc.), the length of each segment of the antennae, the width of the third flagellum and the length of the sixth flagellum are measured, and the total length of the antennae = the length of the pedicel + the length of the pedicel + the length of the first flagellum + the length of the second flagellum + the length of the third flagellum + the length of the fourth flagellum + the length of the fifth flagellum The total length of the antennae is calculated by adding the length of the 3rd flagellum + the length of the 6th flagellum + the length of the 7th flagellum + the length of the 8th flagellum; if the total length of the antennae of the adult grain borer is greater than 620μm, the width of the 3rd flagellum is less than 36μm, and the length of the 6th flagellum is greater than 100μm, the corresponding test insect is identified as a male grain borer; if the total length of the antennae of the adult grain borer is less than 620μm, the width of the 3rd flagellum is greater than 36μm, and the length of the 6th flagellum is less than 100μm, the corresponding test insect is identified as a female grain borer.

[0036] Table 2 Total length of antennae, width of the third flagellum and length of the sixth flagellum of male adults (partial data)

[0037]

[0038]

[0039]

[0040]

[0041]

[0042]

[0043]

[0044] Note: In Table 2, F3 is the third flagellum of antennae, and F6 is the sixth flagellum of antennae, the same below.

[0045] Table 3 Total length of antennae, width of the third flagellum and length of the sixth flagellum of female adults (partial data)

[0046]

[0047]

[0048]

[0049]

[0050]

[0051]

[0052] Example 2 Verification

[0053] From each source in Table 1 (Table 1), 826 adult grain borers were collected as test insects, and N2 was blown on their surface to reduce surface impurities. The sex was identified by the method of the present invention: the cleaned test insect (the sample stage with the ventral surface facing the microscope) was placed under an optical microscope for shooting, and the length of each section of its antenna, the width of the third whip section and the length of the sixth whip section were measured by the microscopic image analysis and measurement software Digimizer3.2. According to the total length of antennae=stalk length+stalk length+1st whip section length+2nd whip section length+3rd whip section length+4th whip section length+5th whip section length+6th whip section length+7th whip section length+8th whip section length, the total length of antennae was calculated. Wherein, the length of each section, the width of the third whip section and the length of the sixth whip section are defined as in Example 1. After the measurement, each test insect was dissected under an optical microscope, the genitalia were checked, and its sex was identified to verify the accuracy of the identification method of the present invention. Due to the large amount of data, only some of the results are listed in Table 4. Results: The identification results of the above 826 grain borers using the dissection method were consistent with the identification results of the method of the present invention.

[0054] In addition, the adult grain borer whose sex is to be identified can also be photographed with a camera (such as a professional camera, a high-resolution camera on a mobile phone, etc.), and the above dimensions can be measured using the ruler provided in the image processing software to identify its sex.

[0055] Table 4: Consistency test table of identification results of the method of the present invention and the dissection method (partial data)

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[0057]

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[0059]

[0060]

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[0062]

[0063]

[0064]

[0065]

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[0068]

[0069]

[0070]

Claims

1. A method for sex identification of stored-grain pests, characterized in that: The steps include: (1) Measure the length and width of each segment of the antennae of female and male stored-grain pest adults; (2) Analyze the relationship between the total length, segment length and / or width of antennae of adult stored-grain pests and their sex; (3) measuring the total length of the antennae, the length of each segment and / or the width of the adult stored-grain pest to be tested, and identifying its sex based on the relationship described in step (2).

2. The identification method according to claim 1, characterized in that The stored-grain pest is the grain borer.

3. The identification method according to claim 1 or 2, characterized in that The relationship between the total length of antennae of adult grain borers and their gender is as follows: the total length of antennae of male adult grain borers is greater than 620 μm, while the total length of antennae of female adult grain borers is less than 620 μm.

4. The identification method according to claim 3, characterized in that The relationship between the width of each segment of the antennae of adult grain borers and their sex is as follows: the width of the third flagellum of male adult grain borers is less than 36μm, while the width of the third flagellum of female adult grain borers is greater than 36μm.

5. The identification method according to claim 4, characterized in that The relationship between the length of each segment of the antennae of adult grain borers and their sex is as follows: the length of the sixth flagellum of male adult grain borers is greater than 100 μm, while the length of the sixth flagellum of female adult grain borers is less than 100 μm.

6. The identification method according to claim 5, characterized in that In step (1), after taking pictures with an electron microscope or a camera, the total length, length and width of the antennae of the adult stored-grain pests are measured using a built-in ruler of the instrument or image processing software.

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