A method of shortening the growth period of tobacco

Abstract

The application provides a method for shortening the growth period of tobacco. The application finds that the high-altitude bacillus NM17 (collection number: CGMCC NO. 31273) has the following new uses: (1) shortening the harvesting period of tobacco and harvesting in advance; (2) improving the maturity and uniformity of tobacco leaves and promoting the stratified yellowing of tobacco leaves; and (3) promoting the decomposition of starch grains and improving the quality of tobacco leaves. The high-altitude bacillus NM17 has great application prospects in improving the maturity of tobacco leaves and promoting the early harvesting of tobacco leaves.

Description

Technical Field

[0001] This invention relates to the field of microbiology, and in particular to a method for shortening the growth period of tobacco. Background Technology

[0002] Tobacco is an important economic crop in my country, and the yield and quality of tobacco leaves are crucial to the development of the tobacco industry. However, with the increasing frequency of extreme weather events due to global climate change, coupled with unreasonable crop rotation patterns and a lack of scientific fertilization management, tobacco production faces severe challenges such as difficulty in late-stage yellowing, insufficient maturity, and declining quality, significantly impacting the sustainable development of the tobacco industry. Therefore, it is urgent to take effective measures to address the key issues during the tobacco leaf maturity stage, accelerate the accumulation and conversion of dry matter in tobacco plants, improve leaf maturity, and promote early harvesting. This is not only a core strategy for achieving high-quality flue-cured tobacco production but also a key path to promoting the high-quality development of the tobacco industry.

[0003] Currently, the main technical means to promote tobacco leaf ripening and shorten the growth period include chemical regulation, agronomic measures, and biotechnology applications. Chemical regulation primarily involves the exogenous application of plant growth regulators such as ethephon and abscisic acid to promote leaf ripening and yellowing. However, chemical regulation carries the risk of residue, affecting tobacco quality and safety, and has significant negative environmental impacts. Agronomic measures mainly involve reasonable planting density, optimized fertilization (such as nitrogen control and potassium increase), water management, and topping / removing side shoots, but their effectiveness is inconsistent and greatly limited by climate and soil conditions. Biotechnology applications have become a hot topic in recent years, but research mainly focuses on rhizosphere microorganisms with growth-promoting functions. These microorganisms regulate the physiological metabolism of tobacco plants, promoting dry matter accumulation and transformation, thereby accelerating tobacco leaf ripening. However, there is a lack of specific microbial agents targeting the later stages of tobacco leaf ripening.

[0004] While existing technologies can promote tobacco leaf maturation to some extent, they still suffer from unstable effects and high environmental risks. Therefore, it is necessary to develop a method to shorten the tobacco growth period. Summary of the Invention

[0005] The purpose of this invention is to provide a method for shortening the growth period of tobacco. This invention discovers that NM17 Pristella megaterium can shorten the growth period of tobacco, effectively regulate the ripening process of tobacco leaves, advance harvesting, and improve the uniformity of tobacco leaf harvesting, which has important production application value.

[0006] To achieve the above objectives, the present invention adopts the following technical solution:

[0007] In a first aspect of the present invention, a method for shortening the growth period of tobacco is provided, the method comprising:

[0008] The bacterial suspension of NM17 Primatex megaterium, with preservation number CGMCC NO.31273, was diluted and applied as a foliar spray.

[0009] Furthermore, the method for preparing the NM17 Pristinae bacterial culture includes: inoculating the Pristinae NM17 into a liquid culture medium for fermentation culture to obtain the NM17 fermentation broth, wherein the fermentation culture conditions include: a temperature of 26-30°C and a pH of 5-9.

[0010] Furthermore, the spraying method specifically includes:

[0011] The NM17 fermentation broth (OD600=0.3, 3×10⁻⁶) was used to ferment the NM17 fermentation broth. 8 After diluting the NM17 fermentation broth 100-300 times to obtain a diluted solution, spray with 30L of the diluted solution per acre. The density of NM17 *Primatex megaterium* in the fermentation broth is: OD600 0.2-0.4, 1×10⁻⁶. 8 -3×10 9 cfu / ml. Preferably, OD600 = 0.3, 3 × 10⁻⁶. 8 cfu / ml.

[0012] Furthermore, the foliar spraying is performed ≥ 2 times, with an interval of 7-10 days between sprayings.

[0013] Furthermore, the foliar spraying time includes: 7-10 days after topping or 20 days before harvest.

[0014] In a second aspect of the invention, the application of NM17 Precipitella megaterium in shortening the growth period of tobacco is provided, wherein the NM17 Precipitella megaterium has the accession number CGMCC NO.31273.

[0015] In a third aspect of the invention, a composition for shortening the growth period of tobacco is provided, the composition comprising:

[0016] NM17 Primate giantiformis (CGMCC NO.31273) 0.5-2wt%;

[0017] Brown algae oligosaccharides 1-3 wt%;

[0018] Carrier 95-98 wt%.

[0019] Furthermore, the molecular weight of the fucoidan is 80-300 Da.

[0020] Furthermore, the carrier is selected from chitosan oligosaccharide, amino acid, and glucose.

[0021] In a fourth aspect of the invention, the use of the composition in shortening the growth period of tobacco is provided.

[0022] One or more technical solutions in the embodiments of the present invention have at least the following technical effects or advantages:

[0023] This invention provides a method for shortening the tobacco growing season. The method involves foliar spraying with a diluted solution of *Plastrum megaterium* (NM17). This invention found that the NM17 strain can accelerate the senescence process of tobacco leaves, promote chlorophyll degradation, and increase leaf maturity, resulting in a maturity period 5-10 days earlier than the control group, which is beneficial for earlier harvesting. It also improves the uniformity of yellowing and leaf maturity in the tobacco field, reduces the field coefficient of variation, and creates favorable conditions for mechanized harvesting. The results show that the application of the NM17 strain can effectively regulate the tobacco leaf maturation process, advance harvesting, and improve the uniformity of tobacco leaf harvesting, demonstrating significant production application value.

[0024] The *Priestia megaterium* NM17 strain of this invention was deposited on July 11, 2024, with accession number CGMCC NO. 31273. Its taxonomic name is *Priestia megaterium*, and the depository is the China Center for Type Culture Collection, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, 100101, China (Institute of Microbiology, Chinese Academy of Sciences). *Priestia megaterium* NM17 was obtained by the inventors of this application through prior screening, and its patent number is CN119040221B. Attached Figure Description

[0025] To more clearly illustrate the technical solutions in the embodiments of the present invention, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the accompanying drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0026] Figure 1 Comparison of field growth after two applications of NM17 in conventional tobacco fields;

[0027] Figure 2 Comparison of field growth after spraying NM17 twice in late-maturing tobacco fields;

[0028] Figure 3 Comparison of field growth after spraying NM17 twice in black smoke fields;

[0029] Figure 4 Comparison of yellowing of tobacco leaves after two applications of NM17 spray;

[0030] Figure 5To observe the ultrastructure of tobacco leaves after spraying with NM17 bacterial solution; among them, AB are electron micrographs of the ultrastructure of tobacco leaves in the control group, with the arrows indicating chloroplasts; CD are electron micrographs of the ultrastructure of tobacco leaves in the treatment group sprayed with brown algae oligosaccharide. Detailed Implementation

[0031] The present invention will be described in detail below with reference to specific embodiments and examples, thereby making the advantages and various effects of the present invention more clearly apparent. Those skilled in the art should understand that these specific embodiments and examples are for illustrative purposes only and are not intended to limit the present invention.

[0032] Throughout this specification, unless otherwise specified, the terminology used herein should be understood as having the meaning commonly used in the art. Therefore, unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. In the event of any conflict, this specification shall prevail.

[0033] Unless otherwise specified, all raw materials, reagents, instruments and equipment used in this invention can be obtained by purchasing them from the market or by existing methods.

[0034] The following will describe in detail a method for shortening the growth period of tobacco according to this application, with reference to embodiments and experimental data. Example 1: Preparation of NM17 Pristella megaterium bacterial suspension.

[0035] The preserved Pristella giantiflora NM17 was activated and inoculated into NA liquid medium for fermentation culture to obtain NM17 fermentation broth. The fermentation culture conditions included: temperature of 26-30℃ and pH of 5-9.

[0036] The NM17 fermentation broth (OD600 = 0.3, 3 × 10⁻⁶) was used. 8 Dilute (cfu / ml) 100-300 times to obtain a diluted solution for subsequent spraying.

[0037] Example 2, Field Experiment

[0038] 1. Experimental Design

[0039] Representative tobacco fields in the Shandong tobacco-growing area were selected for foliar spraying treatment. The experimental design and site selection are shown in Table 1.

[0040] Table 1 Experimental Design and Site Selection

[0041]

[0042] The conventional treatments refer to those carried out according to local production practices or local tobacco production management technical standards, mainly including the following methods: 1. Foliar spraying of potassium dihydrogen phosphate; 2. Ring cutting at the base of the stem in black-leaved tobacco fields; 3. Ripening products from the agricultural input market (such as potassium dihydrogen phosphate and bud suppressant); 4. No treatment, allowing natural yellowing. In this study, the conventional treatment for normal tobacco fields was no treatment, allowing natural yellowing; the conventional treatment for late-maturing tobacco fields was spraying potassium dihydrogen phosphate (150g / mu); the conventional treatment for black-leaved tobacco fields was spraying potassium dihydrogen phosphate (150g / mu) + ring cutting.

[0043] 2. Indicator Measurement

[0044] 2.1 Harvesting period records

[0045] Based on the maturity of tobacco leaves in the field, the harvest time of tobacco leaves treated with different methods was recorded.

[0046] 2.2 Field photos of plant growth

[0047] After two applications, drones were used to photograph the field growth of different treatments and compare the changes in leaf color.

[0048] 2.3 Observation of blade ultrastructure

[0049] Several small pieces, approximately 2mm × 2mm, were cut from the lower right side of the midrib of fresh tobacco leaves. These pieces were then placed in 2.5% glutaraldehyde phosphate buffer (pH 7.2) and aerated at room temperature until the material settled. The samples were then fixed in a 4°C refrigerator for at least 72 hours. The samples were then washed, dehydrated, dried, mounted, and observed and photographed under a Hitachi S-3000 scanning electron microscope.

[0050] Test results

[0051] 3.1 Comparison of harvest time for different treatments

[0052] Table 2. Records of harvest time in tobacco fields under different treatments

[0053]

[0054] The table shows that NM17 strain treatment promotes tobacco leaf maturity and advances harvest. In normal tobacco fields, the third harvest is advanced by 2 days, and the final harvest by 5 days. In late-maturing tobacco fields, the second harvest is advanced by 3 days, the third by 5 days, and the final harvest by 7 days. In fields affected by severe weather, the second harvest is advanced by 4 days, the third by 6 days, and the final harvest by 10 days. The more difficult the tobacco harvesting and curing process, the more advanced the harvest becomes with foliar spraying of NM17 strain.

[0055] 3.2 Comparison of yellowing of tobacco leaves in the field under different treatments

[0056] The results of field growth comparison in conventional tobacco fields after conventional treatment and after two applications of NM17 are as follows: Figure 1 As shown;

[0057] Comparison of field growth in late-maturing tobacco fields after conventional treatment and after two applications of NM17: Figure 2 As shown;

[0058] Comparison of tobacco growth in black-spotted fields after conventional treatment and after two applications of NM17: Figure 3 As shown;

[0059] Comparison of yellowing tobacco leaves after two applications of NM17 spray: Figure 4 As shown;

[0060] By observing and comparing the growth of tobacco in different fields, the treatment group sprayed with NM17 strain twice showed significant physiological effects. Specifically, the effects were: (1) the senescence process of tobacco leaves was significantly accelerated, the chlorophyll degradation rate was increased, and the tobacco leaves matured earlier; (2) the uniformity of yellowing was significantly improved, the field coefficient of variation was reduced, and the maturity was consistent.

[0061] 3.3 Observation and Analysis of Blade Ultrastructure

[0062] Observation of the ultrastructure of tobacco leaves (e.g.) Figure 5 As shown in the image, the cell membranes and cell walls of the control group leaves were intact and clearly defined. Organelles were compressed to the cell edges by vacuoles and arranged in an orderly manner. The thylakoid structures of the chloroplasts were clearly visible, and a small amount of starch grains were present. This indicates that the control group leaves were actively synthesizing starch and other substances, and were still in the developmental stage. In contrast, electron microscopy results of the NM17 strain treatment group showed that chloroplasts and other organelles were gradually decomposed, plastid membranes ruptured, and starch grains accumulated in large quantities and began to degrade. This indicates that the leaves of the treatment group were already in the mature stage.

[0063] 4. Experimental Conclusions

[0064] Analysis of data from different tobacco fields and comparison of growth patterns revealed that the treatment group sprayed with NM17 strain twice exhibited significant physiological effects. NM17 strain accelerated tobacco leaf senescence, promoted chlorophyll degradation, and increased leaf maturity, resulting in a maturity period 5-10 days earlier than the control group, which is beneficial for earlier harvesting. It also improved the uniformity of yellowing and leaf maturity in the tobacco field, reduced the field coefficient of variation, and created favorable conditions for mechanized harvesting. The results indicate that the application of NM17 strain can effectively regulate the tobacco leaf maturity process, advance harvesting, and improve the uniformity of tobacco leaf harvesting, demonstrating significant value for production applications.

[0065] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

[0066] Finally, it should be noted that 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 process, method, article, or apparatus.

[0067] Although embodiments of the invention have been described, those skilled in the art, upon learning the basic inventive concept, can make further changes and modifications to these embodiments. Therefore, the appended claims are intended to be interpreted as including the embodiments as well as all changes and modifications falling within the scope of the invention.

[0068] Obviously, those skilled in the art can make various modifications and variations to this invention without departing from its spirit and scope. Therefore, if these modifications and variations fall within the scope of the claims of this invention and their equivalents, this invention also intends to include these modifications and variations.

Claims

1. A method for shortening the growth period of tobacco, characterized in that, The method includes: Foliar spraying is performed using diluted NM17 *Primatex megaloides* bacterial suspension, the preservation number of which is CGMCC NO.31273. The preparation method of the NM17 bacterial suspension includes: inoculating *Primatex megaloides* NM17 into a liquid culture medium for fermentation to obtain NM17 fermentation broth. The fermentation conditions include: temperature 26–30℃, pH 5–9. The foliar spraying method specifically includes: diluting the NM17 fermentation broth 100–300 times, absorbing the diluted solution, and spraying 30 L of the diluted solution per acre. The foliar spraying is performed at least twice, with an interval of 7–10 days. The foliar spraying time includes: 7–10 days after topping or 20 days before harvest.

2. The application of NM17 *Primatex megaterium* in shortening the growth period of tobacco, characterized in that... The NM17 Precipitella megaterium has the accession number CGMCC NO.31273.

3. A composition for shortening the growth period of tobacco, characterized in that, The composition contains 0.5-2 wt% of NM17 Primate macrocephala, which has the accession number CGMCC NO.31273.

4. The use of the composition according to claim 3 in shortening the growth period of tobacco.

Images