Application of bacillus velezensis TCS001 to improvement of strawberry growth traits

Abstract

According to the application of the bacillus velezensis TCS001 serving as the medicine for improving the growth traits of the strawberries, after the strawberries are treated by the bacillus velezensis TCS001, plant growth can be remarkably promoted, and strawberry leaves can be induced to generate faster and stronger defensive reaction for protecting an enzyme system.

Description

Technical Field

[0001] The present invention relates to the field of plant growth regulation, in particular to the field of strawberry growth trait regulation, and in particular to the use of Bacillus velezensis TCS001 for improving strawberry growth traits. Background Art

[0002] Strawberry (Fragaria ananassa Duch) is an important economic crop with a wide cultivation area and is grown in many countries. It is popular for its soft and juicy fruits, delicious taste, rich nutrition, short growing period, early fruiting and high yield, and has an important economic position in the international market.

[0003] Bacillus velezensis is a novel biocontrol bacterium. Spanish researchers Ruiz-García et al. first isolated two strains, CR-14b and CR-502T, in 2005. These strains are capable of producing large quantities of lipopeptides and exhibit strong antibacterial activity. Bacillus velezensis is widely distributed and can survive adverse conditions such as drought and salinity. However, few related products exist, suggesting promising development prospects.

[0004] Bacillus velezensis TCS001 was isolated from Bohai Sea silt. The original strain was Bacillus velezensis CT2628. After mutagenesis and stabilization, it was identified and named Bacillus velezensis TCS001. It has been deposited with the General Microbiology Center of the China General Culture Collection (CGMCC) under the accession number CGMCC No. 8921 and was first published in Patent ZL201410168402.2. When grown on NA medium, TCS001 colonies are nearly round, light yellow, and opaque. Initially, the surface is smooth with neat edges, but later, the surface becomes wrinkled, with slightly irregular edges and a central bulge, which diffuses in a cloud-like pattern. Gram staining showed that the Bacillus Velezii TCS001 strain is Gram-positive and rod-shaped; it has a certain inhibitory effect on cucumber gray mold, cucumber vine blight, cucumber sclerotinia, cucumber brown spot, cotton wilt and banana leaf spot, among which the inhibition rate against cucumber gray mold is the highest, reaching 87.66%.

[0005] When the inventors of the present application used Bacillus Velez TCS001 to prevent and control gray mold of strawberry, they unexpectedly discovered that Bacillus Velez TCS001 had a significant improvement effect on the growth traits of strawberry, and therefore proposed the present invention. Summary of the Invention

[0006] The main purpose of the present invention is to provide the use of Bacillus velezensis TCS001 as a drug for improving the growth traits of strawberries.

[0007] Another main object of the present invention is to provide a method for improving the growth traits of strawberries using Bacillus Velez TCS001.

[0008] Another main object of the present invention is to provide a composition for improving the growth traits of strawberries, characterized in that it includes a fermentation broth of Bacillus Velez TCS001.

[0009] The strawberry growth traits include leaf chlorophyll concentration, single plant leaves, stem and leaf fresh weight, root fresh weight, leaf area, plant height, leaf IAA content, ABA content, SOD activity, leaf GA content, CAT activity, POD activity, etc.

[0010] Preferably, the Bacillus Velez subtilis TCS001 is a fermentation broth of Bacillus Velez subtilis TCS001.

[0011] Preferably, the Bacillus Velez subtilis TCS001 fermentation broth is prepared as a suspension concentrate.

[0012] Preferably, the formula of the suspension concentrate is: 40% of Bacillus velez TCS001 fermentation broth, 3% of dispersant SXC, 3% of dispersant UNA, 0.15% of thickener, 0.2% of preservative kason, 0.2% of preservative sodium benzoate, and 53.45% of water.

[0013] Preferably, the application concentration of the suspension is 200-500 times diluted.

[0014] Preferably, the application concentration of the suspension is 300 times diluted.

[0015] Preferably, the Bacillus Velez subtilis TCS001 is applied during the seedling stage.

[0016] Preferably, the Bacillus Velez subtilis TCS001 is applied by root irrigation and / or spraying.

[0017] Beneficial technical effects of the present invention

[0018] The present invention discovers for the first time that Bacillus Velez TCS001 can improve the growth traits of strawberries. After treatment with a 300-fold spray concentration of Bacillus Velez TCS001 suspension concentrate, the chlorophyll concentrations of strawberry leaves at 14 days and 21 days are the highest, increasing by 11% and 14.89% respectively compared with CK. The fresh weight of leaves, stems and leaves, and roots of individual strawberries at 28 days increase by 26.5%, 79.4%, and 120% respectively compared with CK.

[0019] After root irrigation with 300 times the concentration of Bacillus Velez TCS001 suspension concentrate, the leaf area and plant height of strawberries increased by 68.41% and 29.68% respectively compared with CK on day 28.

[0020] After root irrigation with 300 times the concentration of Bacillus Velez TCS001 suspension concentrate, the IAA content, ABA content and SOD activity of strawberry leaves increased by 38.47%, 36.04% and 21.41% respectively compared with CK on day 28.

[0021] After treatment with 300 times the concentration of Bacillus Velez TCS001 suspension concentrate, the GA content, CAT activity and POD activity of strawberry leaves increased by 34.04%, 21.23% and 23.21% respectively compared with CK on day 28.

[0022] Treatment with Bacillus Velez TCS001 can significantly promote the growth of strawberry plants and induce strawberry leaves to produce faster and stronger protective enzyme defense responses. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 For each treated strawberry plant of the present invention;

[0024] Figure 2 The changes in strawberry chlorophyll content in each treatment group of the present invention at different time periods;

[0025] Figure 3 The changes in the number of leaves per strawberry plant in each treatment group of the present invention at different time periods;

[0026] Figure 4 The height changes of strawberries in different time periods in each treatment group of the present invention;

[0027] Figure 5 The fresh weight changes of strawberry stems and leaves in different time periods in each treatment group of the present invention;

[0028] Figure 6 The fresh weight changes of strawberry roots in each treatment group of the present invention at different time periods;

[0029] Figure 7 The strawberry leaf area of ​​each treatment group of the present invention after 28 days;

[0030] Figure 8 The changes in the content of abscisic acid in strawberry leaves of each group treated with different days in the present invention;

[0031] Figure 9 The changes in auxin content in strawberry leaves of each group treated with different days in the present invention;

[0032] Figure 10 The changes in gibberellin content in strawberry leaves of each group at different treatment days of the present invention;

[0033] Figure 11 The changes in catalase activity in strawberry leaves of each group treated with different days in the present invention;

[0034] Figure 12 The changes in superoxide dismutase activity in strawberry leaves of each group under different treatment days of the present invention;

[0035] Figure 13 The figure shows the changes in peroxidase activity in strawberry leaves of each group treated with different days in the present invention. Specific embodiments

[0036] Materials and Methods

[0037] Material:

[0038] The test bacterial agent of the present invention is the suspension concentrate of Bacillus velezensis TCS001 (3.9×10 9 CFU / mL), the specific formula is: 40% of Bacillus Velez TCS001 fermentation broth, 3% of dispersant SXC, 3% of dispersant UNA, 0.15% of thickener, 0.2% of preservative Kathon, 0.2% of preservative sodium benzoate, and 53.45% of water.

[0039] Test plants:

[0040] Experimental variety: Four-leaf red strawberry.

[0041] Main instruments

[0042] ME104E electronic balance (Sartorius Scientific Instruments Co., Ltd.), chlorophyll tester (Zhejiang Top Yunnong Technology Co., Ltd.), electric constant temperature blast drying oven (DHG-9023A)

[0043] method

[0044] Treatment

[0045] Table 1 Treatment methods of strawberry pot test with Bacillus velezensis TCS001 suspension concentrate

[0046]

[0047] The suspension concentrate of Bacillus Velez TCS001 was diluted with sterile water to the corresponding concentration and then used for root irrigation and spraying treatments respectively.

[0048] The potting soil was sterilized by dry heat sterilization at 160°C for 2 h in an oven, and each pot contained 960 g of sterile soil.

[0049] Root irrigation: After the strawberry seedlings were transplanted, each plant was irrigated with 100 mL of 300-fold diluted Bacillus Velez subtilis TCS001 suspension concentrate, and each treatment was repeated in 5 pots.

[0050] Spraying: The first spraying treatment was performed one week after transplanting the strawberry seedlings. The spraying was repeated every 7 days with 5 mL of solution per plant (1, 2, and 3 times). Each treatment was repeated for 5 pots at a spray pressure of 0.1 MPa. Growth index measurements were performed.

[0051] Determination of strawberry morphological indicators

[0052] The basic morphological indexes (plant height) of strawberries after treatment with Bacillus Velez TCS001 suspension concentrate (14d, 21d, 28d) were measured using a ruler (graduation value 1mm). Five plants with consistent growth were selected for measurement in each treatment.

[0053] Determination of chlorophyll content in strawberry leaves

[0054] The relative chlorophyll content (SPAD) of the second expanded leaf outside the heart leaf of strawberry after treatment with TCS001 suspension concentrate (14d, 21d, and 28d) was determined using a chlorophyll meter (Zhejiang Top Yunnong Technology Co., Ltd.). Five plants with consistent growth were selected for measurement in each treatment.

[0055] Determination of fresh weight of strawberry roots and leaves

[0056] After treatment with TCS001 suspension concentrate (14d, 21d, 28d), the strawberry plants were dried with paper towels and weighed with an electronic balance to determine the fresh weight of their roots and leaves. The fresh weight of the roots and leaves of a single strawberry plant was considered one replicate, and each treatment had 5 replicates.

[0057] Determination of leaf number per strawberry plant

[0058] The number of leaves per strawberry plant was determined after treatment with TCS001 suspension concentrate (14d, 21d, and 28d). The number of leaves per strawberry plant was considered one replicate, and each treatment had 5 replicates.

[0059] Strawberry leaf area measurement

[0060] The leaf area of ​​strawberries was measured 28 days after treatment with TCS001 suspension concentrate using a leaf area meter (Zhejiang Top Yunnong Technology Co., Ltd.). The third expanded leaf outside the heart leaf of the strawberry was taken, and each treatment was replicated five times.

[0061] Enzyme activity and endogenous hormones

[0062] After 7 days of incubation following spray treatment (1, 2, or 3 times), root length, plant height, stem and leaf fresh and dry weight, and chlorophyll were measured. Leaf peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT) activities were determined using corresponding enzyme-linked immunosorbent assay kits. The abscisic acid (ABA), indoleacetic acid (IAA), and gibberellin (GA) contents in strawberry leaves were also measured using corresponding enzyme-linked immunosorbent assay kits.

[0063] Experimental results

[0064] The plants after spraying and root irrigation with 300 times of Bacillus Velez TCS001 suspension were as follows: Figure 1 shown.

[0065] Growth index determination results such as Figure 2-7 shown.

[0066] according to Figure 2-7 It can be seen that after the treatment with 300 times of suspension concentrate of Bacillus Velez TCS001, the chlorophyll concentration of strawberry leaves ( Figure 2 ) were the highest, and compared with CK, they increased by 11% and 14.89% respectively. On the 28th day, the leaves of single strawberry plants ( Figure 3 ), stem and leaf fresh weight ( Figure 5 ), fresh root weight ( Figure 6 ), which were increased by 26.5%, 79.4% and 120% respectively compared with CK.

[0067] After irrigating the roots with 300 times the concentration of Bacillus Velez TCS001 suspension concentrate, the leaf area of ​​strawberry ( Figure 7 ), plant height( Figure 4 ), which are increased by 68.41% and 29.68% respectively compared with CK.

[0068] Endogenous hormone and enzyme activity assay results

[0069] according to Figure 8-13 It can be seen that after the root irrigation treatment with 300 times of Bacillus Velez TCS001 suspension concentrate, the IAA content of strawberry leaves ( Figure 9 ), ABA content ( Figure 8 )、SOD activity ( Figure 12 ), compared with CK, it increased by 38.47%, 36.04% and 21.41%.

[0070] After treatment with 300 times of suspension concentrate of Bacillus Velez TCS001, the GA content of strawberry leaves ( Figure 10 ), CAT activity ( Figure 11 )、POD activity ( Figure 13 ), compared with CK, it increased by 34.04%, 21.23% and 23.21%.

[0071] The results showed that treatment with Bacillus Velez TCS001 could significantly promote the growth of strawberry plants and induce faster and stronger protective enzyme defense responses in strawberry leaves.

[0072] The specific implementation methods described above further illustrate the objectives, technical solutions and beneficial effects of the present invention in detail. It should be understood that the above description is only a specific implementation method of the present invention and is not intended to limit the scope of protection of the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. Application of Bacillus velez TCS001 as a drug for improving the growth traits of strawberry.

2. The use according to claim 1, characterized in that: The strawberry growth traits include one or more of leaf chlorophyll concentration, single plant leaves, stem and leaf fresh weight, root fresh weight, leaf area, plant height, leaf IAA content, ABA content, SOD activity, leaf GA content, CAT activity, POD activity, etc.

3. A method for improving strawberry growth traits using Bacillus Velez TCS001, characterized in that: The strawberry growth traits include one or more of leaf chlorophyll concentration, single plant leaves, stem and leaf fresh weight, root fresh weight, leaf area, plant height, leaf IAA content, ABA content, SOD activity, leaf GA content, CAT activity, POD activity, etc.

4. The use according to any one of claims 1 to 2 or the method according to claim 3, characterized in that: The Bacillus Velez TCS001 is the fermentation liquid of Bacillus Velez TCS001.

5. The use according to any one of claim 4 or the method according to claim 4, characterized in that: The Bacillus Velez TCS001 fermentation liquid is prepared into a suspension.

6. The use according to claim 5 or the method according to claim 5, characterized in that: The suspending agent comprises 40% of Bacillus Velez TCS001 fermentation liquid, 3% of dispersant SXC, 3% of dispersant UNA, 0.15% of thickener, 0.2% of preservative Kathon, 0.2% of preservative sodium benzoate and 53.45% of water.

7. The use according to claim 6 or the method according to claim 6, characterized in that: The application concentration of the suspension is 200-500 times diluted, preferably, the application concentration is 300 times diluted.

8. The use according to any one of claims 1 to 2 or the method according to claim 3, characterized in that: The application time of the Bacillus Velez TCS001 is the seedling stage.

9. The use according to any one of claims 1 to 2 or the method according to claim 3, characterized in that: The application method of the Bacillus Velez TCS001 is root irrigation and / or spraying treatment.

10. A composition for improving the growth characteristics of strawberries, characterized in that: Including Bacillus velez TCS001 fermentation broth.

Images