Straw decomposing bacteria, application thereof and efficient straw returning method

Abstract

The invention discloses a straw decomposing bacterium, application thereof and an efficient straw returning method, and relates to the technical field of agriculture. The straw decomposing bacterium EQ-1 is preserved in the China Center for Type Culture Collection, the address is Wuhan University, Wuhan, China, 430072, the preservation number is CCTCC NO: M 20242458, the preservation date is November 7, 2024, and the straw decomposing bacterium EQ-1 has a 16SrDNA (Deoxyribose Nucleic Acid) sequence as shown in SEQ ID No: 1. The invention further discloses an efficient straw returning method. The straw decomposing bacterium (Enterobacter guaiaporachexii) EQ-1 provided by the invention has the capability of degrading cellulose, hemicellulose and lignin, and has a good decomposing effect on rice straws and wheat straws, so that the straw decomposing bacterium (Enterobacter guaiaporachexii) EQ-1 can be used for decomposing rice straws and wheat straws. The straw decomposing bacterium (Enterobacter guaihormacacei) EQ-1 is applied to efficient straw returning, the yield of rice can be remarkably increased, the content of humic acid and humin in soil is increased, the activity of alpha-glucosidase, beta-glucosidase and cellulase in the soil is improved, and the decomposing rate of straw returning to the field is remarkably increased.

Description

Technical Field

[0001] This invention relates to the field of agricultural technology, specifically to a straw decomposing bacteria and its application, and a method for efficient straw return to the field. Background Technology

[0002] Straw decomposition refers to the process of breaking down complex organic substances such as cellulose, hemicellulose, and lignin in crop straw into humus and small molecule compounds that can be absorbed and utilized by plants through physical, chemical, or biological means.

[0003] Currently, straw decomposition methods mainly include physical decomposition, chemical decomposition, and biological decomposition. Physical decomposition methods use mechanical crushing, heating, radiation, and other physical means to break down the straw's structure, accelerating the decomposition process. These methods are simple to operate, low in cost, and can be directly returned to the field to increase soil organic matter. However, incomplete crushing can lead to poor soil aeration, affecting seed germination, and requires the application of nitrogen fertilizer to adjust the carbon-nitrogen ratio. High-temperature crushing is energy-intensive and requires specialized equipment, making it suitable for large-scale processing. Chemical decomposition involves large investments in equipment and is currently mostly used in laboratory research, not yet widely applied. Chemical decomposition methods use chemical reagents (such as acids, alkalis, oxidants, etc.) to decompose lignin in straw, releasing cellulose and hemicellulose. This method has the advantage of high decomposition efficiency; however, it easily causes secondary pollution and damages the environment. Biological decomposition methods rely on the metabolic activities of microorganisms (such as bacteria, fungi, actinomycetes, etc.) to decompose straw. This method has the advantage of being green and environmentally friendly. However, when using existing biological decomposition methods, the stable structure of lignin in straw makes it difficult for microorganisms to directly decompose it, resulting in a long decomposition cycle.

[0004] Straw return technology is an agricultural technique that returns crop straw, either directly or after processing, to the field to achieve resource recycling and soil improvement. Generally, the straw is shredded and then plowed into the soil, with the depth of plowing used to improve crop growth. However, existing methods still have problems such as limited increases in crop yield.

[0005] Therefore, providing a green decomposition method for straw and an efficient straw return method for the field, so as to achieve efficient recycling of resources and improve crop yield, is an urgent problem to be solved by this invention. Summary of the Invention

[0006] The present invention provides a straw decomposing bacteria and its application, as well as a method for efficient straw return to the field, aiming to solve the problems existing in the above-mentioned background technology.

[0007] To achieve the above-mentioned technical objectives, the present invention mainly adopts the following technical solutions:

[0008] In a first aspect, the present invention discloses a straw-decomposing bacterium (Enterobacter guasihormacechei) EQ-1, deposited at the China Center for Type Culture Collection, address: Wuhan University, Wuhan, China, 430072, accession number: CCTCCNO: M20242458, deposit date: November 7, 2024, having a 16S rDNA sequence as shown in SEQ ID No:1.

[0009] In a second aspect, the present invention discloses a microbial agent containing Enterobacterguasihormacechei EQ-1 as described in the first aspect.

[0010] In a preferred embodiment of the present invention, the straw-decomposing bacteria (Enterobacter guasihormacechei) EQ-1 exists in the form of cultured live bacteria, fermentation broth, or bacterial suspension in the inoculum.

[0011] Thirdly, the present invention discloses the application of Enterobacterguasihormacechei EQ-1, as described in the first aspect, in the decomposition of crop straw.

[0012] Fourthly, this invention discloses the application of Enterobacter guasihormacechei EQ-1, as described in the first aspect, in the efficient return of straw to the field to improve rice yield.

[0013] Fifthly, this invention discloses the application of Enterobacter guasihormacechei EQ-1, as described in the first aspect, in the efficient return of straw to the field to improve soil organic matter content and the content of chemical indicators humic acid and humin.

[0014] In a sixth aspect, the present invention discloses the application of Enterobacter guasihormacechei EQ-1, as described in the first aspect, in improving the activity of soil biological indicators α-glucosidase, β-glucosidase, and cellulase through efficient straw return to the field.

[0015] In a seventh aspect, the present invention discloses the application of Enterobacter guasihormacechei EQ-1, as described in the first aspect, in improving the efficiency of straw decomposition through efficient straw return to the field.

[0016] Eighthly, this invention discloses a method for efficient straw return to the field, comprising the following steps:

[0017] Step 1: Crush the straw;

[0018] Step 2: Return the crushed straw from Step 1 to the field, and when returning it to the field, apply the straw decomposing microbial agent EQ-1 and activator product as described in claim 1 to the field;

[0019] Step 3: Apply base fertilizer according to conventional agricultural production practices. When the soil moisture content is ≤30%, carry out straw plowing operations.

[0020] Step 4: Flood the paddy field, level the surface, and transplant rice seedlings 1-3 days later.

[0021] In a preferred embodiment of the present invention

[0022] In step 1, the length of the crushed straw is less than 10cm, and the unevenness of the crushed straw distribution is ≤20%.

[0023] In step 2, the activator product is urea, the application rate of straw decomposing microbial agent EQ-1 is 15-30 kg / ha, and the application rate of the activator product is 25-35 kg / ha.

[0024] In step 3, when tilling the straw, the straw burial depth is controlled to be 13-17 cm and 18-22 cm respectively;

[0025] In step 4, the water depth in the paddy field is 4-6 cm, and the soaking time is 24-48 hours.

[0026] Compared with the prior art, the present invention has the following beneficial effects:

[0027] (1) The straw decomposing bacteria (Enterobacterguasihormacechei) EQ-1 provided by the present invention has the ability to degrade cellulose, hemicellulose and lignin, and has a good decomposition effect on rice and wheat straw;

[0028] (2) Applying the straw decomposing bacteria (Enterobacterguasihormacechei) EQ-1 of the present invention to the efficient return of straw to the field can significantly increase the yield of rice, increase the content of humic acid and humin in the soil, increase the activity of α-glucosidase, β-glucosidase and cellulase in the soil, and significantly increase the decomposition rate of straw returned to the field. Attached Figure Description

[0029] Figure 1 The colony structure and Gram staining results of Enterobacter guasihormacechei EQ-1 provided by the present invention;

[0030] Figure 2The growth curve of the straw-decomposing bacteria (Enterobacter guasihormacechei) EQ-1 provided by the present invention;

[0031] Figure 3 Figure showing the identification results of the straw decomposition ability of the straw-decomposing bacterium Enterobacter guasihormacechei EQ-1 provided by the present invention;

[0032] Figure 4 Figure showing the effect of different straw return methods on soil organic matter content;

[0033] Figure 5 This graph shows the change in straw decomposition amount with the time of burying bags under different straw return methods. Detailed Implementation

[0034] To better illustrate the purpose, technical solution, and advantages of the present invention, the present invention will be further described below in conjunction with the accompanying drawings and specific embodiments.

[0035] In the following embodiments, unless otherwise specified, the experimental methods used are conventional methods, and the materials and reagents used are commercially available unless otherwise specified.

[0036] The straw-decomposing bacterium Enterobacter guasihormacechei EQ-1 involved in this invention was deposited at the China Center for Type Culture Collection on November 7, 2024, with accession number CCTCC NO: M 20242458.

[0037] Example 1: Isolation and Identification of Straw-Decomposing Bacterium (Enterobacter guasihormacechei) EQ-1

[0038] (1) Separation method

[0039] A mixture of soil and decomposed straw from the rapid straw composting stage was selected and mixed with distilled water at a ratio of 1:9 (mass:volume), and then serially diluted to 10. -850 μL of the diluted solution was spread onto sodium carboxymethyl cellulose medium (2.5 g peptone, 0.5 g yeast extract, 2.5 g Na₂HPO₄, 1.5 g KH₂PO₄, 20 g agar, 1000 mL distilled water, pH 7.0), hemicellulose medium (20 g xylan, 2.5 g peptone, 0.5 g yeast extract, 0.5 g MgSO₄·7H₂O, 1.5 g KH₂PO₄, 20 g agar, 1000 mL distilled water, pH 7.0), and potato-guaiacol solid medium (200 g peeled potato, 20 g sucrose, 1000 mL tap water, 20 g agar, pH natural) plates for initial screening of cellulose, hemicellulose, and lignin-degrading microorganisms. The strains were then inoculated onto ordinary culture media for isolation and purification to obtain single strains. Subsequently, single strains were selected for secondary screening. Selected colonies were inoculated onto sodium carboxymethyl cellulose-Congo red medium, xylan medium, and aniline blue potato broth, respectively. After cultivation, the size of the clear degradation zone was measured to determine the bacteria's ability to decompose cellulose, hemicellulose, and lignin. Simultaneously, a filter paper disintegration test was used to further determine the decomposition ability of the screened bacteria. Bacterial genomic DNA extraction involved extracting DNA from the strains, amplifying fragments using 16S rDNA primers, and performing next-generation sequencing. The sequences were aligned using NCBI, and the highest-scoring sequence was selected. A phylogenetic tree was constructed using MEGA11 software, confirming the genus *Enterobacter guasihormacechei* to which the screened strains belonged.

[0040] Sodium carboxymethyl cellulose-Congo red medium: CMC-Na 20g, peptone 2.5g, yeast extract 0.5g, Na2HPO4 2.5g, KH2PO4 1.5g, agar 20g, Congo red 2g, distilled water 1000mL, pH 7.0.

[0041] Xylan medium: 20g xylan, 2.5g peptone, 0.5g yeast extract, 2.5g Na2HPO4, 1.5g KH2PO4, 20g agar, 1000mL distilled water, pH 7.0.

[0042] Aniline blue potato culture medium: 200g peeled potatoes, 20g sucrose, 1000mL tap water, 2g aniline blue, pH natural.

[0043] All the above culture media were sterilized at 121℃ for 30 min. After sterilization, the culture media were taken out and observed at room temperature for 2 days to confirm that there was no sterile growth before use.

[0044] (2) Morphological identification: such as Figure 1 As shown, the bacterial colonies are pale yellow, relatively large, and have smooth edges, indicating they are Gram-positive bacteria.

[0045] Molecular biological identification: Through 16S rDNA sequence analysis, the analysis results show that it has the 16S rDNA sequence shown in SEQ ID NO:1:

[0046] GGTAAGCGCCCTCCCGAAGGTTAAGCTACCTACTTCTTTTGCAACCCACTCCCATGGTGT

[0047] GACGGGCGGTGTGTACAAGGCCCGGGAACGTATTCACCGTGGCATTCTGATCCACGATTA

[0048] CTAGCGATTCCGACTTCATGGAGTCGAGTTGCAGACTCCAATCCGGACTACGACGCACTT

[0049] TATGAGGTCCGCTTGCTCTCGCGAGGTCGCTTCTCTTTGTATGCGCCATTGTAGCACGTGT

[0050] GTAGCCCTACTCGTAAGGGCCATGATGACTTGACGTCATCCCCACCTTCCTCCAGTTTATC

[0051] ACTGGCAGTCTCCTTTGAGTTCCCGGCCTAACCGCTGGCAACAAAGGATAAGGGTTGCG

[0052] CTCGTTGCGGGACTTAACCCAACATTTCACAACACGAGCTGACGACAGCCATGCAGCAC

[0053] CTGTCTCAGAGTTCCCGAAGGCACCAATCCATCTCTGGAAAGTTCTCTGGATGTCAAGAG

[0054] TAGGTAAGGTTCTTCGCGTTGCATCGAATTAAACCACATGCTCCACCGCTTGTGCGGGCC

[0055] CCCGTCAATTCATTTGAGTTTTAACCTTGCGGCCGTACTCCCCAGGCGGTCGACTTAACGC

[0056] GTTAGCTCCGGAAGCCACGCCTCAAGGGCACAACCTCCAAGTCGACATCGTTTACGGCG

[0057] TGGACTACCAGGGTATCTAATCCTGTTTGCTCCCCACGCTTTCGCACCTGAGCGTCAGTCT

[0058] TTGTCCAGGGGGCCGCCTTCGCCACCGGTATTCCTCCAGATCTCTACGCATTTCACCGCTA

[0059] CACCTGGAATTCTACCCCCCTCTACAAGACTCTAGCCTGCCAGTTTCGAATGCAGTTCCC

[0060] AGGTTGAGCCCGGGGATTTCACATCCGACTTGACAGACCGCCTGCGTGCGCTTTACGCCC

[0061] AGTAATTCCGATTAACGCTTGCACCCTCCGTATTACCGCGGCTGCTGGCACGGAGTTAGCC

[0062] GGTGCTTCTTCTGCGGGTAACGTCAATCGACAAGGTTATTAACCTTATCGCCTTCCTCCCC

[0063] GCTGAAAGTACTTTACAACCCGAAGGCCTTCTTCATACACGCGGCATGGCTGCATCAGGC

[0064] TTGCGCCCATTGTGCAATATTCCCCACTGCTGCCTCCCGTAGGAGTCTGGACCGTGTCTCA

[0065] GTTCCAGTGTGGCTGGTCATCCTCTCAGACCAGCTAGGGATCGTCGCCTAGGTGAGCCGT

[0066] TACCCCACCTACTAGCTAATCCCATCTGGGCACATCCGATGGCAAGAGGCCCGAAGGTCC

[0067] CCCTCTTTGGTCTTGCGACGTTATGCGGTATTAGCTACCGTTTCCAGTAGTTATCCCCCTCC

[0068] ATCAGGCAGTTTCCCAGACATTACTCACCCGTCCGCCACTCGTCAGCGAAGCAGCAAGCTGCTTCCTGTTACCGTTCGACTTGCATGTGTA, as shown in SEQ ID NO:1.

[0069] Example 2: Cultivation of Straw Decomposing Bacterium (Enterobacter guasihormacechei) EQ-1

[0070] The bacterial strain was inoculated into LB liquid medium and cultured with shaking at 28°C and 150 rpm. Samples were taken every hour to measure the OD and pH values ​​of the bacterial culture. All experimental treatments were performed in triplicate. Growth curves were plotted based on the results.

[0071] The results are as follows Figure 2 As shown, the bacteria entered a rapid growth phase after 20 hours of culture, and at the same time, the pH of the culture medium dropped rapidly from about pH 6.7 to about pH 5.2.

[0072] Example 3: Identification of the straw decomposition ability of straw-decomposing bacteria (Enterobacter guasihormacechei EQ-1)

[0073] The straw-degrading bacterium *Enterobacter guasihormacechei* EQ-1 was cultured in petri dishes on carboxymethyl cellulose sodium-Congo red medium (CMC-Na-Congo red medium), xylan medium, and aniline blue potato medium, respectively, for assessing its ability to degrade cellulose, hemicellulose, and lignin. Three replicates were recorded for each strain, with unclustered plates serving as controls. The dishes were incubated at 30°C, and the presence and size of a clear zone were observed daily. Results are as follows: Figure 3 As shown, the bacterium was found to produce clear zones in all three culture media, indicating that it has the ability to degrade cellulose, hemicellulose and lignin.

[0074] Add filter paper to the bacterial culture medium and incubate for 3 days. Observe the morphology of the filter paper. Figure 3 As shown, the filter paper was found to be intact in the culture medium without bacteria, but it disintegrated and was severely damaged in the culture medium with added bacteria, indicating that the bacteria have a strong ability to decompose cellulose.

[0075] Rice and wheat straw were added to liquid culture medium respectively, and cultured for 7 days. Figure 3As shown, the reduction in rice and wheat straw in the inoculated culture medium was significantly higher than that in the uninoculated control, further demonstrating that the bacteria have a good decomposition effect on rice and wheat straw.

[0076] Example 4: Efficient Straw Returning Method

[0077] The obtained straw-decomposing bacteria (Enterobacter guasihormacechei) EQ-1 was used in a field trial. A large-scale experimental design was adopted, with four treatments, each covering approximately 2 acres. The types and amounts of nitrogen, phosphorus, and potassium fertilizers were based on local practices. Base fertilizer and top dressing were mainly compound fertilizers, while tillering fertilizer was mainly urea. All straw was returned to the field, as shown in Table 2.

[0078] Efficient Straw Returning to the Field Steps: ① Straw Crushing and Returning to the Field: Harvest crops using a combine harvester equipped with a straw crushing and spreading device, or use a straw crushing and returning machine for straw crushing operations. The length of the crushed straw should be less than 10cm, and the unevenness of the scattered straw should be ≤20%. ② Application of Straw Decomposing Microbial Agent EQ-1 and Activator: When returning straw to the field, use a drone or a harvester equipped with a decomposing agent spraying device to apply straw decomposing microbial agent EQ-1 and activator products. The dosage of straw decomposing microbial agent EQ-1 is 15-30 kg / ha, and the dosage of activator such as urea is 30 kg / ha. ③ Straw Rotary Tillage and Returning to the Field: Apply base fertilizer according to conventional agricultural production methods. When the soil moisture content is ≤30%, use a rotary tiller or a combined plow and rotary tiller to till the straw. The straw burial depth is set at approximately 15 cm and 20 cm, respectively. ④ Soaking and preparing the field: The field is then flooded to a depth of about 5 cm for 24-48 hours. The surface is then leveled using a paddy harrow or a flat-bottomed threshing machine. Rice seedlings are transplanted 1-3 days after the field has settled.

[0079] Table 2. Experimental Treatments for Full Straw Return to Field

[0080]

[0081] (1) Investigate the effects of different straw return methods on rice yield and its components.

[0082] Compared with conventional straw return to the field (CK0), although the seed setting rate of rice decreased significantly after applying straw decomposing microbial agents EQ-1 (T1 and T2), the number of panicles per mu and the number of grains per panicle both significantly increased. This resulted in a significant increase in rice yield, from 650 kg / mu to 694 kg / mu and 678 kg / mu, respectively (Table 3).

[0083] Table 3. Effects of different treatments on rice yield and its components.

[0084]

[0085] (2) Investigate the effects of different straw return methods on soil chemical indicators

[0086] Compared to no straw return to the field (CK0), direct straw return to the field (CK1) significantly increased the organic matter content in the soil. The application of straw composting agents and the use of different tillage depths (T1 15cm and T2 20cm) further increased the organic matter content in the soil. Figure 4 As shown.

[0087] Compared with no straw return to the field (CK0), direct straw return to the field (CK1) can also increase the content of various humic substances in the soil, as shown in Table 3. Compared with direct straw return to the field (CK1), the application of straw decomposing microbial agent EQ-1 and different tillage depths (T1 and T2) increased the content of humic acid and humin in the soil, as shown in Table 4.

[0088] Table 4. Effects of different treatments on soil humus composition

[0089]

[0090] (3) Investigate the effects of different straw return methods on soil biological indicators.

[0091] Compared to no straw return to the field (CK0), direct straw return to the field (CK1) increased the activity of α-glucosidase in the soil. Compared to direct straw return to the field (CK1), the application of straw decomposing microbial agent EQ-1 and different tillage depths (T1 and T2) increased the activities of α-glucosidase, β-glucosidase, and cellulase in the soil, as shown in Table 5.

[0092] Table 5 Effects of different treatments on soil enzyme activity

[0093]

[0094] (4) Investigate the effects of different straw return methods on straw decomposition efficiency.

[0095] The application of straw decomposing microbial agent EQ-1, combined with different tillage depths (T1 and T2), significantly increased the decomposition rate of straw returned to the field. Figure 5 As shown, by Figure 5 It can be seen that, especially from 30 days after returning straw to the field, the straw decomposition rate increased by at least 17 percentage points.

[0096] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A straw-decomposing bacterium (Enterobacter guasihormacechei) EQ-1, characterized in that, It is deposited at the China Center for Type Culture Collection, Wuhan University, Wuhan, China 430072, China, accession number: CCTCC NO: M20242458, deposit date: November 7, 2024, and has a 16S rDNA sequence as shown in SEQ ID No:

1.

2. An inoculum containing the straw-decomposing bacteria (Enterobacterguasihormacechei) EQ-1 as described in claim 1.

3. The microbial agent according to claim 2, characterized in that, In the microbial agent, the straw-decomposing bacteria (Enterobacter guasihormacechei) EQ-1 exists in the form of cultured live bacteria, fermentation broth, or bacterial suspension.

4. The application of the straw-decomposing bacteria (Enterobacter guasihormacechei) EQ-1 as described in claim 1 in the decomposition of crop straw.

5. The application of the straw-decomposing bacteria (Enterobacter guasihormacechei) EQ-1 as described in claim 1 in the efficient return of straw to the field to improve rice yield.

6. The application of the straw-decomposing bacteria (Enterobacter guasihormacechei) EQ-1 as described in claim 1 in the efficient return of straw to the field to improve the content of soil organic matter and the content of chemical indicators humic acid and humin.

7. The application of the straw-decomposing bacteria (Enterobacter guasihormacechei) EQ-1 as described in claim 1 in improving the activity of soil biological indicators α-glucosidase, β-glucosidase and cellulase through efficient straw return to the field.

8. The application of the straw decomposing bacteria (Enterobacter guasihormacechei) EQ-1 as described in claim 1 in improving the efficiency of straw decomposition through efficient straw return to the field.

9. A method for efficient straw return to the field, characterized in that, Includes the following steps: Step 1: Crush the straw; Step 2: Return the crushed straw from Step 1 to the field, and when returning it to the field, apply the straw decomposing microbial agent EQ-1 and activator product as described in claim 1 to the field; Step 3: Apply base fertilizer according to conventional agricultural production practices. When the soil moisture content is ≤30%, carry out straw plowing operations. Step 4: Flood the paddy field, level the surface, and transplant rice seedlings 1-3 days later.

10. The method for efficient straw return to the field according to claim 9, characterized in that, In step 1, the length of the crushed straw is less than 10cm, and the unevenness of the crushed straw distribution is ≤20%. In step 2, the activator product is urea, the application rate of straw decomposing microbial agent EQ-1 is 15-30 kg / ha, and the application rate of the activator product is 25-35 kg / ha. In step 3, when tilling the straw, the straw burial depth is controlled to be 13-17 cm and 18-22 cm respectively; In step 4, the water depth in the paddy field is 4-6 cm, and the soaking time is 24-48 hours.

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