Cinnamon streptomyces castanea and application of cinnamon streptomyces castanea in prevention and control of bemisia tabaci

Abstract

The invention belongs to the technical field of pest prevention and control, and particularly relates to streptomyces cinnamomi castanea and application thereof in prevention and control of bemisia tabaci. The invention discloses a cinnamon streptomyces castanea XJ33, which is preserved in the China Center for Type Culture Collection on March 10, 2025, and the preservation number of the cinnamon streptomyces castanea XJ33 is CCTCC (China Center for Type Culture Collection) M 2025414. The average death rate of the bemisia tabaci within 72 hours is 96.32%, 83.93%, 75.11% and 5.40% under the treatment of the stock solution of the cinnamon streptomyces castanea, the bacterium solution of the cinnamon streptomyces castanea diluted by 10 times, the bacterium solution of the cinnamon streptomyces castanea diluted by 100 times and clear water, which indicates that the cinnamon streptomyces castanea has a remarkable technical effect on prevention and treatment of the bemisia tabaci.

Description

Technical Field

[0001] The present invention belongs to the technical field of pest control, and particularly relates to a Streptomyces cinnamocastaneus and its application in controlling Bemisia tabaci. Background Art

[0002] Bemisia tabaci belongs to the Hemiptera, Aleyrodidae, and is an important pest on vegetables and cash crops. It is widely distributed globally and mainly damages crops and flowers such as Solanaceae, cucurbits, and legumes. Bemisia tabaci causes plant wilting and drying by sucking plant sap with nymphs and adults and secreting honeydew. Adults can also transmit plant viruses, resulting in crop yield reduction or even crop failure. At present, the application of chemical pesticides in production is the main measure for field control of Bemisia tabaci. Especially during the outbreak period of Bemisia tabaci, chemical control is the most effective emergency method. However, Bemisia tabaci has serious generation overlap, fast reproduction speed, and wide spread range. In particular, field populations have developed varying degrees of resistance to commonly used insecticides, increasing the difficulty of control. At present, Bemisia tabaci has developed resistance to various insecticides such as organophosphates, neonicotinoids, pyrethroids, and antibiotics. Some highly toxic, high-residue, and highly resistant insecticides no longer meet the requirements of current green agricultural development, and there is an urgent need to seek new insecticidal products with high efficiency, low toxicity, and low residue.

[0003] Microbial control is a method of using microorganisms and their metabolites to control crop pests and diseases. It includes directly infecting and killing pests with pathogenic microorganisms (such as bacteria, fungi, viruses, etc.), and using beneficial microorganisms (such as actinomycetes, yeasts, etc.) to produce antibiotics or other antibacterial substances to inhibit or kill harmful microorganisms. Microbial control has the characteristics of high efficiency, strong specificity, and environmental friendliness, and is an important part of the sustainable development of modern agriculture. Summary of the Invention

[0004] To solve the above technical problems, the present invention provides the following technical solutions:

[0005] The present invention discloses a Streptomyces cinnamocastaneus XJ33, which was deposited at the China Center for Type Culture Collection on March 10, 2025, with the deposit number CCTCC M2025414.

[0006] The present invention also discloses a microbial agent, which contains Streptomyces cinnamocastaneus XJ33.

[0007] The present invention also discloses a microbial fermentation broth, which is obtained by fermenting Streptomyces cinnamocastaneus XJ33.

[0008] Preferably, the preparation method of the microbial fermentation broth is as follows:

[0009] (1) Inoculate Streptomyces cinnamocastaneus XJ33 into TSBY medium and shake to activate it to obtain activated Streptomyces cinnamocastaneus XJ33;

[0010] (2) Inoculate the activated Streptomyces cinnamocastaneus XJ33 into YEME medium for fermentation to obtain the microbial fermentation broth.

[0011] Preferably, the preparation method of the TSBY medium in step (1) is: weigh 30 g / L of tryptic soy broth, 5 g / L of yeast extract, and sterilize at 121 °C with steam for 15 min.

[0012] Preferably, the conditions for shaking activation in step (1) are: adjust the shaker to 28 °C, 220 r / min, and shake for 48 h.

[0013] Preferably, the preparation method of the YEME medium in step (2) is: weigh 3 g / L of yeast extract, 3 g / L of malt extract, 5 g / L of protein, 340 g / L of sucrose, and 10 g / L of glucose, and sterilize at 113 - 114 °C at high temperature for 15 min; add MgCl 2 ·6H0 to make the final concentration of Mg 2+ 5 mM.

[0014] Preferably, the conditions for fermentation in step (2) are: adjust the shaker to 28 °C, 220 r / min, and shake for 72 h. The volume ratio of the activated Streptomyces cinnamocastaneus XJ33 to the YEME medium is 1:5.

[0015] Preferably, the concentration of Streptomyces cinnamocastaneus XJ33 in the microbial fermentation broth is 1.3 - 1.4 billion cfu / ml.

[0016] The application of Streptomyces cinnamocastaneus XJ33, a microbial inoculant containing the same, or a microbial fermentation broth fermented therefrom in the control of Bemisia tabaci.

[0017] The application of Streptomyces cinnamocastaneus XJ33, a microbial inoculant containing the same, or a microbial fermentation broth fermented therefrom in the preparation of a product for controlling Bemisia tabaci.

[0018] Advantages of the present invention:

[0019] The present invention discloses Streptomyces cinnamocastaneus XJ33, which was deposited at the China Center for Type Culture Collection on March 10, 2025, with the deposit number CCTCC M2025414. Under the treatments of the original solution of Streptomyces cinnamocastaneus, the bacterial solution of Streptomyces cinnamocastaneus diluted 10 times, the bacterial solution of Streptomyces cinnamocastaneus diluted 100 times, and water, the average mortality rates of Bemisia tabaci at 72 h were 96.32%, 83.93%, 75.11%, and 5.40%, respectively, indicating that Streptomyces cinnamocastaneus has significant technical effects in controlling Bemisia tabaci. Description of the drawings

[0020] Figure 1 Colony map of Streptomyces cinnamocastaneus XJ33;

[0021] Figure 2 Phylogenetic tree analysis map of Streptomyces cinnamocastaneus;

[0022] Figure 3 Effect of the original solution of Streptomyces cinnamocastaneus XJ33 on killing Bemisia tabaci;

[0023] Figure 4 Effect of the original solution of Streptomyces cinnamocastaneus XJ33 diluted 10 times on killing Bemisia tabaci;

[0024] Figure 5 Effect of the original solution of Streptomyces cinnamocastaneus XJ33 diluted 100 times on killing Bemisia tabaci;

[0025] Figure 6The figure shows the result of water treatment. Detailed implementation

[0026] To make the objectives, technical solutions and advantages of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are some, but not all, of the embodiments of the present invention. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0027] Example 1 Classification and identification of bacteria

[0028] The 16S rDNA identification technology was used to identify the species of strain XJ33. Universal primers for bacteria were used to amplify the 16S rRNA gene. After the PCR amplification was completed, the PCR products were detected by electrophoresis using 1% agarose gel; the gene products obtained by PCR were sequenced. The sequencing result of the 16S rRNA gene sequence of strain XJ33 is shown in SEQ ID NO.1:

[0029] SEQ ID NO.1

[0030]

[0031] The 16S rRNA gene sequence of strain XJ33 was input into the NCBI database and analyzed by BLAST software. Strains with higher homology to strain XJ33 were selected, and MEGA7.0 software was used for multiple sequence alignment and construction of a phylogenetic tree, as shown in Figure 2 Figure []. The 16S rRNA gene sequence of strain XJ33 was obtained by PCR amplification. The alignment results showed that strain XJ33 was most closely related to Streptomyces cinnamocastaneus. Therefore, strain XJ33 was identified as Streptomyces cinnamocastaneus. This strain was named Streptomyces cinnamocastaneus XJ33 and deposited in the China Center for Type Culture Collection. The deposit address is Wuhan, China (Wuhan University), and the strain deposit number is CCTCC M2025414. The deposit date was March 10, 2025.

[0032] Example 2 Preparation of microbial inoculum

[0033] Experimental method

[0034] (1) Take the Streptomyces strain XJ33 screened from the activated strain bank and purify it no more than 3 generations to ensure the activity of the bacteria.

[0035] (2) Use a 50 ml conical flask to fill it with 20 ml of TSBY medium. In a laminar flow hood, use a sterile inoculation stick to pick a single colony of Streptomyces strain XJ33 and inoculate it into a conical flask containing TSBY medium that has been pre-prepared and sterilized at 121 °C for 20 min and then cooled. Add 7 mm glass beads to it. Adjust the shaker to 28 °C and 220 r / min, and shake for 48 h to obtain TSBY bacterial liquid. Then transfer 20 ml of the TSBY bacterial liquid that has been shaken for 48 h to a 250 ml conical flask containing 100 ml of YEME medium and continue culturing. Adjust the shaker to 28 °C and 220 r / min, and shake for 72 h. Add 7 mm glass beads to it.

[0036] In the laminar flow hood, use sterile forceps and a pipette gun to take 10 μl of the bacterial liquid and drop it onto a hemocytometer, cover it with a cover slip, and count the number of viable bacteria under an electron microscope. The number of viable bacteria of XJ33 is 1.3 - 1.4 billion CFU / ml.

[0037] TSBY medium

[0038] Weigh 30 g of Tryptone Soya Broth and 5 g of yeast extract, make up the volume to 1 L with dd water, and sterilize it with steam at 121 °C for 15 min.

[0039] YEME medium

[0040] Weigh 3g of yeast extract, 3g of malt extract, 5g of protein, 340g of sucrose and 10g of glucose, dilute to 1L with dd water, sterilize at 113-114℃ for 15min; add MgCl before use. 2 ·6H0 makes Mg 2+ The final concentration is 5 mM.

[0041] Example 3 Application of microbial agent of Streptomyces cinnamomi XJ33

[0042] The bioassay method for Bemisia tabaci adults uses the leaf agar moisturizing method, and the specific steps are as follows:

[0043] (1) Prepare a flat-bottomed glass finger tube with a diameter of about 2.2 cm and a length of about 8 cm. Use a rubber-tipped dropper to add 2-3 mL of 2% agar solution to the bottom of the flat-bottomed glass finger tube. Avoid spilling the agar solution on the wall of the glass tube and avoid bubbles in the added agar solution. Seal the tube and cool it. After the agar solidifies and the water vapor on the wall of the flat-bottomed glass tube completely evaporates, the bioassay test can be carried out.

[0044] (2) Prepare the test insecticide concentration according to the recommended field concentration and the preliminary test results, dilute the test cinnamon marron Streptomyces XJ33 microbial agent into the test concentration according to the gradient, and use it as a control.

[0045] (3) Take the cultivated cotton leaves and use a puncher to punch out leaf discs with a diameter of 22 mm on the flat leaf part avoiding the large leaf veins. Immerse the leaf discs in the prepared microbial inoculum of Streptomyces cinnamomi XJ33 for 20 seconds and then take them out. Let the leaves dry naturally with the back facing up.

[0046] (4) Use blunt-tipped tweezers to pick up the dried cotton leaf disc and place it with the back facing up into a flat-bottomed glass finger-shaped tube where the agar has solidified. Fit the cotton leaf disc tightly against the agar and remove any bubbles. After inserting the cotton plug, mark the bottom of the flat-bottomed glass finger-shaped tube for later use.

[0047] (5) If whiteflies are densely distributed on leaves, they can be collected by tapping the leaves. Otherwise, use a sucker tool to drive the whiteflies into a flat-bottomed glass finger tube that has been treated and air-dried. Each tube contains a certain number of whiteflies. A cotton plug is inserted to half of the tube, allowing the whiteflies to crawl on the leaf disc in the flat-bottomed glass tube.

[0048] (6) Place the flat-bottomed glass finger tube containing whiteflies upside down in the prepared light incubator, set the temperature to 26°C ± 2°C, the humidity to 70% ± 10%, and the photoperiod to 16h:8h (L:D). After 48h, check the survival of the whiteflies in the flat-bottomed glass finger tube. If the whiteflies do not move or cannot move normally, they are considered dead.

[0049] The experimental results are as Figures 3 - 6 shown in Table 1:

[0050] Table 1 Experimental Results

[0051]

[0052] Note: The number before the slash in Table 1 is the number of dead whiteflies, and the number after the slash is the total number of whiteflies.

[0053] Result analysis: Under the treatment of the original solution of Streptomyces cinnamomeus, the bacterial solution of Streptomyces cinnamomeus diluted 10 times, the bacterial solution of Streptomyces cinnamomeus diluted 100 times, and water, the average mortality rates of whiteflies at 72 h were 96.32%, 83.93%, 75.11%, and 5.40%, respectively, indicating that Streptomyces cinnamomeus has significant technical effects in controlling whiteflies.

Claims

1. A strain of Streptomyces cinnamocastaneus XJ33, characterized in that: It was deposited in the China Center for Type Culture Collection on March 10, 2025, with the deposit number CCTCC M2025414.

2. A microbial agent, characterized in that: The microbial agent contains the Streptomyces cinnamocastaneus XJ33 described in claim 1.

3. A microbial fermentation broth, characterized in that: The microbial fermentation liquid is obtained by fermenting the Streptomyces cinnamocastaneus XJ33 described in claim 1.

4. The microbial fermentation liquid according to claim 3, characterized in that: The preparation method of the microbial fermentation liquid is as follows: (1) inoculating the Streptomyces cinnamocastaneus XJ33 described in claim 1 into TSBY medium for shaking and activation to obtain activated Streptomyces cinnamocastaneus XJ33; (2) The activated Streptomyces cinnamocastaneus XJ33 is inoculated into a YEME medium for fermentation to obtain the microbial fermentation liquid.

5. The microbial fermentation liquid according to claim 4, characterized in that: The preparation method of the TSBY medium in step (1) is as follows: weigh 30 g / L of tryptic soy broth and 5 g / L of yeast extract, and steam sterilize at 121° C. for 15 min.

6. The microbial fermentation liquid according to claim 5, characterized in that: The conditions for the shaking activation in step (1) are as follows: the shaking table is adjusted to 28° C., 220 r / min, and shaken for 48 hours.

7. The microbial fermentation liquid according to claim 4, characterized in that: The preparation method of the YEME medium in step (2) is as follows: weigh 3 g / L yeast extract, 3 g / L malt extract, 5 g / L protein, 340 g / L sucrose and 10 g / L glucose, sterilize at 113-114°C for 15 min; add MgCl2·6H0 to make MgCl2·6H0 2+ The final concentration is 5 mM.

8. The microbial fermentation liquid according to claim 7, characterized in that: In step (2), the fermentation conditions are as follows: the shaker is adjusted to 28° C., 220 r / min, and shaken for 72 h; the volume ratio of the activated Streptomyces cinnamocastaneus XJ33 to the YEME culture medium is 1:

5.

9. The microbial fermentation liquid according to claim 4, characterized in that: The concentration of the Streptomyces cinnamocastaneus XJ33 in the microbial fermentation broth is 1.3-1.4 billion cfu / ml.

10. Use of the Streptomyces cinnamocastaneus XJ33 according to claim 1, the microbial agent according to claim 2, or the microbial fermentation liquid according to any one of claims 3 to 9 in controlling Bemisia tabaci.

11. Use of the Streptomyces cinnamocastaneus XJ33 according to claim 1, the microbial agent according to claim 2, or the microbial fermentation liquid according to any one of claims 3 to 9 in preparing a product for controlling Bemisia tabaci.

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