Lactobacillus plantarum LSR-22 and application thereof in preparation of silage

Abstract

The invention discloses lactobacillus plantarum LSR-22 and application thereof in preparation of silage, the biological preservation number of the lactobacillus plantarum LSR-22 is CCTCC M 20242927, the lactobacillus plantarum LSR-22 is used for preparing ruminant silage, the lactobacillus plantarum LSR-22 is inoculated and added into a mixed silage raw material, the water content of the silage raw material is adjusted to a preset value, and the silage raw material is sealed and packaged. And fermenting in a dark place for 45-70 days to prepare the silage. When the strain is used for fermenting the silage, the contents of dry matters and crude proteins can be remarkably increased, water-soluble carbohydrates are increased, and the fiber content is reduced, so that the nutritional value and digestibility of the silage are improved, the content of ammoniacal nitrogen can be reduced, the content of lactic acid can be increased, and the fermentation quality of the silage is further improved.

Description

Technical Field

[0001] The present invention belongs to the fields of microbial fermentation technology and agricultural biotechnology, and relates to a Lactiplantibacillus plantarum, the application of this strain in the preparation of silage, and the silage fermented using this strain. Background Art

[0002] Silage is a kind of soft, juicy and nutrient-rich roughage made by chopping fresh green forage and then making it in a vacuum and sealed environment through a microbial fermentation process. In animal husbandry, especially in alpine mountainous areas where forage is scarce in winter and spring, silage can effectively preserve the fresh state of forage, ensure that livestock can still obtain high-quality feed when there is a lack of green forage, and thus maintain a high production performance.

[0003] During the ensiling process, lactic acid bacteria play a crucial role. They convert carbohydrates in the silage into organic acids such as lactic acid and propionic acid through fermentation, reduce the pH value, inhibit the growth of harmful microorganisms, and ensure the quality and safety of the feed. Existing studies have shown that lactic acid bacteria can not only promote good acidification during fermentation, prevent the growth of harmful bacteria (such as Crossiella, Salmonella, etc.), but also reduce the generation of toxic by-products, improve the fermentation effect and quality of silage, optimize the composition of beneficial microbial communities in the feed, and further improve the nutritional value of the feed.

[0004] Lactiplantibacillus plantarum is an important strain among lactic acid bacteria and is widely present in various ecological environments such as various fermented foods (such as yogurt, pickles, sauerkraut, etc.), plant surfaces, animal intestines, and soil. Lactiplantibacillus plantarum has characteristics such as a large quantity, diversity, and strain specificity, which make it show different performances in different application environments.

[0005] Large quantity: Lactiplantibacillus plantarum is widely distributed in the natural environment, especially on fermented foods and plant surfaces, and its quantity is very large. For example, during the fermentation of pickles and sauerkraut, Lactiplantibacillus plantarum can account for a large proportion of the total flora.

[0006] Diversity: Different strains of Lactiplantibacillus plantarum have significant differences in metabolic ability, tolerance, functional characteristics, etc.

[0007] Specificity: Although Lactiplantibacillus plantarum has diversity, some strains show unique specificity in specific fermentation environments. For example, some strains can produce specific metabolites such as lactic acid, propionic acid, and acetic acid during fermentation. These metabolites can not only reduce the pH value, but also inhibit the growth of harmful microorganisms, improve the nutritional components and fermentation quality of silage.

[0008] At present, most silage feeds rely on natural fermentation methods, which are inefficient and difficult to fully improve the fermentation quality of the feeds. During the natural fermentation process, the composition and activity of the microbial community are unstable, resulting in uneven fermentation effects, large losses of nutritional components, and being easily affected by environmental factors. In addition, not all Lactobacillus plantarum can improve the fermentation quality of silage feeds. Different strains show different performances during fermentation. Some strains may show excellent fermentation effects under specific conditions, while some strains may have poor effects or even have negative impacts. Summary of the Invention

[0009] In view of this, the present invention aims to provide a specific Lactobacillus plantarum strain and its application method. Through scientific screening and cultivation, it can improve the fermentation efficiency and quality of silage feeds, ensure the safety and nutritional value of the feeds, and provide a higher-quality silage feed additive for the livestock industry. Specifically, the objectives of the present invention include:

[0010] Provide a Lactobacillus plantarum strain with excellent fermentation characteristics, which can effectively reduce the pH value during fermentation, inhibit the growth of harmful microorganisms, and improve the nutritional components and fermentation quality of silage feeds.

[0011] Provide an efficient lactic acid bacteria additive: Prepare an efficient lactic acid bacteria additive, which can significantly improve the nutritional components and fermentation quality of silage feeds, and improve the palatability and digestibility of the feeds.

[0012] Optimize the fermentation conditions: Through experimental research, determine the optimal fermentation conditions, including temperature, pH value, inoculation amount, etc., to improve the fermentation efficiency and reduce the production cost.

[0013] Provide a safe and efficient method for preparing silage feeds: Provide a safe and efficient method for preparing silage feeds for the livestock industry, ensuring that livestock can obtain high-quality feeds in winter and spring seasons and improving the production performance of livestock.

[0014] Through these measures, the present invention will provide important technical support for the development of the livestock industry, improve the production performance of livestock, and promote the sustainable development of the livestock industry.

[0015] Through long-term exploration and attempts, as well as multiple experiments and efforts, and continuous reform and innovation, the inventors provide a technical solution to solve the above technical problems. The present invention provides a Lactobacillus plantarum LSR-22, and the biological deposit number of the Lactobacillus plantarum LSR-22 is: CCTCC M20242927, the deposit date: December 27, 2024, the deposit unit: China Center for Type Culture Collection, the deposit address: Wuhan University.

[0016] Compared with the prior art, the beneficial effects of the present invention are:

[0017] This strain has efficient fermentation performance, can produce acid rapidly within a short time, quickly reduce the pH value of silage, effectively inhibit the growth of harmful microorganisms, and ensure the quality and safety of the feed. The LSR-22 strain can grow within the temperature range of 25°C to 40°C, with the optimal growth temperature being 37°C. It reaches the highest growth value at pH = 7.0, showing good acid tolerance and salt tolerance, and can even grow well in a lower concentration of bile salts. Using this strain to ferment silage can not only significantly increase the dry matter and crude protein content, increase water-soluble carbohydrates, and reduce the fiber content, thereby enhancing the nutritional value and digestibility of the feed, but also reduce the ammonia nitrogen content and increase the lactic acid content, further improving the fermentation quality of silage. In addition, the application method of the LSR-22 strain is simple and the cost is low. It is applicable to a variety of silage raw materials, such as buckwheat straw, silage corn, and alfalfa. By reasonable proportioning and simple fermentation conditions, high-quality silage can be prepared, which has broad application prospects and important economic and environmental benefits.

[0018] The present invention also provides an application of Lactobacillus plantarum LSR-22 as described in claim 1 in the preparation of silage, which is used for preparing silage for ruminants.

[0019] Preferably, the ruminants include cattle and sheep.

[0020] The present invention also provides a lactic acid bacteria additive containing the Lactobacillus plantarum LSR-22, and this additive is used to improve the fermentation quality and nutritional components of silage.

[0021] The present invention also provides a method for preparing silage, which inoculates and adds the Lactobacillus plantarum LSR-22 to the mixed silage raw materials, adjusts the moisture content of the silage raw materials to a preset value, seals and packages, and ferments in the dark for 45 to 70 days.

[0022] Preferably, the fermentation time is 60 days.

[0023] Based on the above technical solutions, the present invention can also be improved as follows:

[0024] Further: The silage raw materials include one or more of buckwheat straw, silage corn, and alfalfa.

[0025] Preferably, the buckwheat straw, silage corn, and alfalfa are all dried.

[0026] Further: The silage raw materials are buckwheat straw, silage corn, and alfalfa, and the mass ratio is 4:6:3.

[0027] Further, the inoculation amount of the Lactobacillus plantarum LSR-22 is 1.0×10 5~1.0×10 7 cfu / g.

[0028] Preferably, the inoculation amount of Lactobacillus plantarum LSR-22 is 1.0×10 6 cfu / g.

[0029] Furthermore: the water content of the silage raw material is 70% - 80%.

[0030] Furthermore: after sealed packaging, vacuum treatment is carried out, and the fermentation temperature is 25 - 40°C.

[0031] Preferably, the fermentation temperature is 37°C.

[0032] The present invention also provides a silage, which is prepared by the aforementioned preparation method, and its nutritional components include:

[0033] Dry matter DM: 23.00% ± 0.54%;

[0034] Crude ash Ash: 8.06% ± 0.72% DM;

[0035] Crude protein CP: 9.99% ± 1.39% DM;

[0036] Water-soluble carbohydrates WSC: 22.34% ± 3.22% DM;

[0037] Neutral detergent fiber NDF: 64.00% ± 0.05% DM;

[0038] Acid detergent fiber ADF: 50.09% ± 0.03% DM.

[0039] Compared with the prior art, the beneficial effects of the present invention are:

[0040] The nutritional components of silage have a decisive impact on its quality. The appropriate level of dry matter content ensures the stability of the feed during storage and reduces nutrient loss, while increasing the energy value of the feed. The reasonable range of crude ash content means that the feed can provide an appropriate amount of minerals to meet the needs of animal growth and development. The high content of crude protein directly supports animal protein synthesis and promotes its healthy growth. The sufficient amount of water-soluble carbohydrates provides energy for lactic acid bacteria fermentation, helps to reduce the pH value, inhibits harmful microorganisms, and ensures the quality of the feed. The lower content of neutral detergent fiber and acid detergent fiber improves the digestibility of the feed, enabling animals to absorb nutrients more effectively. Overall, these nutritional components of the silage of the present invention not only improve the nutritional value of the silage, but also enhance its palatability and digestibility, thus providing high-quality feed resources for the livestock industry and helping to improve animal production performance and economic benefits. Brief Description of the Drawings

[0041] To more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present invention, and thus should not be regarded as limiting the scope. For those of ordinary skill in the art, other related drawings can be obtained based on these drawings without creative efforts.

[0042] Figure 1 is the OD value and pH value after culturing 10 strains for 36 h during the strain screening process. 600 value and pH value.

[0043] Figure 2 is the morphological and microscopic observation results of strain LSR-22; Figure 2 In it, Figure a is the morphological diagram of the strain, and Figure b is the Gram staining result under a 400-fold microscope.

[0044] Figure 3 is the 16S rDNA sequence similarity alignment result of strain LSR-22.

[0045] Figure 4 is the phylogenetic tree of strain LSR-22.

[0046] Figure 5 is the growth rate curve of strain LSR-22.

[0047] Figure 6 is the acid production rate curve of strain LSR-22.

[0048] Figure 7 is the OD value of strain LSR-22 after culturing for 24 h at different temperatures. 600 value,

[0049] Figure 8 is the OD value of strain LSR-22 after culturing for 24 h at different pH values. 600 value.

[0050] Figure 9 is the OD value of strain LSR-22 after culturing for 24 h in media with different NaCl concentrations. 600 value. Detailed Embodiments

[0051] The following is described in conjunction with specific embodiments.

[0052] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the following will, in conjunction with the embodiments of the present invention, clearly and completely describe the technical solutions in 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 belong to the scope of protection of the present invention. Therefore, the following detailed description of the embodiments of the present invention is not intended to limit the scope of the claimed present invention, but merely represents selected embodiments of the present invention.

[0053] In the present invention, unless otherwise specified, all devices and raw materials can be purchased from the market or are commonly used in this industry. The methods in the following embodiments, unless otherwise specified, are conventional methods in this field.

[0054] In this embodiment, the culture media and their components are as follows:

[0055] MRS liquid medium:

[0056] Tryptone 10 g / L, beef powder 10 g / L, yeast powder 4 g / L, ammonium citrate 2 g / L, sodium acetate 5 g / L, magnesium sulfate 0.2 g / L, manganese sulfate 0.05 g / L, dipotassium hydrogen phosphate 2 g / L, glucose 20 g / L, Tween 80 1 mL / L. Adjust the pH value to 5.7 and sterilize at 115 °C for 30 min.

[0057] MRS-CaCO3 solid medium:

[0058] Add 2% agar to the MRS liquid medium formula, adjust the pH value to 5.7, sterilize at 115 °C for 30 min, and add 10 g / L of dry-heat sterilized calcium carbonate.

[0059] Example 1

[0060] This example describes the process of isolating, screening, identifying, and determining the characteristics of Lactobacillus plantarum LSR-22 and its strains.

[0061] The dilution plate method was used to screen lactic acid bacteria.

[0062] The silage samples included buckwheat straw, silage corn, and alfalfa. They were chopped into fragments less than about 1 cm in size, sealed, and naturally fermented. Take 20 g of the fermented silage sample, put it into 180 mL of sterilized MRS liquid medium, shake for 1 h, and then perform 10 -5 、10 -6 、10 -7 gradient dilutions.

[0063] Subsequently, the above three kinds of gradient culture media were respectively spread on MRS-CaCO3 solid medium, and three parallel replicates were made for each dilution. Anaerobic culture was carried out at 37°C for 48 h.

[0064] Subsequently, according to the color, size and state of the colonies, appropriate plates were selected, and typical colonies with calcium dissolution zones were picked and streaked for single colony isolation on MRS plates. Anaerobic culture was continued at 37°C for 48 h.

[0065] Preliminarily screen lactic acid bacteria strains.

[0066] According to the method of "Classification and Identification of Lactic Acid Bacteria and Experimental Methods", single colonies were picked from MRS plates for Gram staining and catalase test, and the strains with positive Gram staining and negative catalase test were numbered.

[0067] The results showed that 10 lactic acid bacteria strains were isolated. The cells had no endospores and were non-motile. The colonies were circular protrusions of different sizes on the medium, with a moist and smooth surface, and mostly milky white in color. Single colony isolation was carried out again, and the confirmed purified single strains were transferred to MRS slant medium and stored at 4°C in a refrigerator for later use.

[0068] The 10 strains preliminarily screened were inoculated into MRS liquid medium and cultured at 37°C for 36 hours, and the OD 600 value and pH value were measured. Using the OD 600 value as the abscissa and the pH value as the ordinate, a scatter plot was drawn, as shown in Figure 1 .

[0069] The results showed that one of the strains had the highest cell density (OD 600 =1.97) and the lowest pH value (pH = 3.49), which was the best strain screened in the present invention and was named LSR-22.

[0070] The morphological characteristics and microscopic examination results of this strain are shown in Figure 2 .

[0071] The DNA of the activated strain was extracted using a bacterial total DNA kit. 1 ml of the activated bacterial liquid was aspirated and added to a 1.5 ml EP tube, centrifuged at 12,000 r / min for 1 minute, and the supernatant was removed as much as possible. DNA was extracted using the alkaline lysis method, rinsed and centrifuged repeatedly to obtain high-quality bacterial genomic DNA, which was the DNA template for identification. Universal primers were used for amplification, and the amplified products were sent to the company for sequencing after electrophoresis detection.

[0072] The universal primers are:

[0073] Upstream primer 27F: 5’-AGAGTTTGATCCTGGCTCAG-3’,

[0074] Downstream primer 1492R: 5’-GGCTACCTTGTTACGACTT-3’.

[0075] The 16S rRNA of LSR-22 strain was sequenced. The full length of the 16S rRNA sequence of LSR-22 was 1462 bp, and the sequence was as shown in SEQ ID NO.1.

[0076] The sequencing sequence was submitted to the BLAST module in the NCBI database for sequence alignment (see Figure 3 ), and the Neighbor Joining Method in the MAGA10.0 software was used to construct the strain phylogenetic tree (see Figure 4 ).

[0077] Sequence alignment showed that it had the highest similarity (99.79%) with Lactiplantibacillus plantarun. Combining its biochemical characteristics, 16S rRNA sequence, and phylogenetic tree results, this bacterium was determined to be L. plantarun.

[0078] L. plantarun LSR-22, biological deposit number: CCTCC M 20242927, deposit date: December 27, 2024, deposit unit: China Center for Type Culture Collection, deposit address: Wuhan University.

[0079] Subsequently, the biochemical characteristics, acid production rate, temperature growth, acid and alkali tolerance, salt tolerance, and bile salt tolerance of L. plantarun LSR-22 were determined. Excel and SPSS24.0 software were used to analyze the data. The SPSS24.0 software was used for one-way ANOVA and LSD test (P<0.05) to statistically analyze the research data. GraphPad Prism 8.0 and Excel software were used for image processing.

[0080] Detection of biochemical characteristics. According to the methods in the "Manual for Systematic Identification of Common Bacteria" and the instructions for the new bacterial biochemical identification tubes produced by Guangdong Huankai Microbial Science and Technology Co., Ltd., the effects of LSR-22 on glucose (acid production and gas production), lactose, maltose, mannitol, sucrose, indole, MR (methyl red), VP, Simmons citrate, urea, hydrogen sulfide, indole, motility, etc. were measured respectively. The results of biochemical characteristics detection are shown in Table 1. The LSR-22 strain produced acid but not gas in glucose, was negative in peptone water, and the results of MR, VP, and Gram staining tests were positive. In addition, its results for Simmons citrate, hydrogen sulfide, and urea (lyophilized) were all negative and positive in semi-solid agar and catalase test.

[0081] Table 1 Detection Results of Biochemical Characteristics

[0082]

[0083] Note: “+”: more than 90% positive; “(+)”: 75 - 89% positive; “v”: 25 - 74% positive; “(-)”: 10 - 24% positive; “-”: 0 - 9% positive.

[0084] Growth and Acid Production Rate Determination Test.

[0085] Inoculate the strain into MRS liquid medium at an inoculation amount of 1%, set 3 replicates for each group, culture in an incubator at 37°C for 36 h, sample every 2 h to measure the OD 600 value and pH value. Use the culture time as the abscissa, and the OD 600 value and pH value as the ordinates to plot the growth curve and acid production rate curve, and infer the growth status and acid production ability of the strain according to the changing trend of the curve. The growth curve and acid production rate curve are as shown in Figure 5 and Figure 6 . The LSR-22 strain grows slowly before 6 h, then enters the logarithmic growth phase, and enters the stationary phase after 18 h. The pH value of the medium decreases slowly within 8 h, the acid production rate is relatively high from 8 to 16 h, the acid production rate gradually decreases from 16 to 24 h, and the pH value remains basically stable within 24 to 36 h, indicating that this strain may have good acid production efficiency.

[0086] Growth Test at Different Temperatures.

[0087] Inoculate the strain into MRS liquid medium at an inoculation amount of 1%, culture at 25°C, 30°C, 37°C, and 40°C for 24 h respectively, set 3 replicates for each group, measure the OD 600 value of each bacterial liquid. Use the culture temperature as the abscissa and the OD 600 value as the ordinate to plot a bar chart. The results of the temperature growth experiment are as shown in Figure 7 . The LSR-22 strain can grow within the temperature range of 25 - 40°C, reaches the highest value (OD 600 = 1.38) at 37°C, and its optimal temperature is 37°C.

[0088] Acid and Alkali Tolerance Test.

[0089] Inoculate the strain into MRS liquid medium with different pH values at a rate of 1% respectively. The pH values are 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5. Set 3 replicates for each group. After culturing at 37°C for 24 h, measure the OD 600 value of each bacterial liquid. Use the pH value of the medium as the abscissa and the OD600 The values were plotted as a bar graph with the vertical axis. The acid and alkali resistance results are as Figure 8 shown. The LSR-22 strain reached the highest value (OD 600 = 1.74) at pH = 7.0, and its optimal pH was 7.0.

[0090] Salt tolerance test.

[0091] The strains were inoculated into MRS liquid media with 6 different NaCl concentrations at a rate of 1% respectively. The NaCl concentrations were 0%, 2%, 4%, 6%, 8%, and 10%. Three replicates were set for each group. After culturing at 37 °C for 24 h, the OD 600 value of each sample bacterial solution was measured. With the NaCl concentration of the medium as the horizontal axis and the OD 600 value as the vertical axis, a bar graph was plotted. The salt tolerance results are as Figure 9 shown. The LSR-22 strain still had good growth effect in 6% NaCl, indicating that it had good salt tolerance ability.

[0092] Bile salt tolerance test.

[0093] The pH of the MRS medium was adjusted to 2.0 and 3.0 respectively with a mixed acid, and 0.2 wt%, 0.3 wt%, 0.4 wt%, and 0.5 wt% of bile salts were added respectively. The MRS medium was used as a blank control. In different media, inoculate at an inoculation amount of 1%, and incubate in a 37 °C incubator for 1 h, 2 h, 3 h, 4 h, 5 h, and 6 h respectively. Using 0 h as the control, measure the OD value at a wavelength of 600 nm. The results of bile salt tolerance are shown in Table 2 and Table 3. Under the conditions of pH 3.0 and 0.2 wt% bile salt, the growth of the LSR-22 strain showed an upward trend within 0 - 5 hours and reached the highest value at 5 hours (OD 600 = 0.52), indicating that the strain grew well in lower concentration bile salts. Under the condition of 0.3 wt% bile salt, the growth of the strain was relatively slow, and the growth density reached the peak at 6 hours (OD 600 = 0.27), but still showed good tolerance. In 0.4 wt% and 0.5 wt% bile salts, the growth of the strain was significantly inhibited, indicating that high concentration bile salts had a strong inhibitory effect on the growth of the strain. Under the condition of pH 2.0, the growth of the LSR-22 strain was inhibited to a certain extent compared with that at pH 3.0, and the maximum OD 600 value was 0.08, showing a certain bile salt tolerance.

[0094] Table 2 Growth of LSR-22 in different bile salts at pH 3.0

[0095]

[0096] Table 3 Growth of LSR-22 in different bile salts at pH 2.0

[0097]

[0098] Example 2

[0099] This example synchronously describes the application of L. plantarun LSR-22, the method of fermentation using L. plantarun LSR-22, and the silage obtained by fermentation.

[0100] The raw materials are dried materials of buckwheat straw, silage corn and alfalfa. Appropriate amount of water is added according to the proportion for preparation, and the moisture content of the fermented materials is 70 - 80%.

[0101] Mix buckwheat straw and silage corn in a weight ratio of 4:6, and then add 30% of alfalfa to form a mixture for silage simulation experiment.

[0102] The experimental group was added with L. plantarun LSR-22 at an inoculation amount of 1.0×106 cfu / g, and the control group was added with the same proportion of sterile water. After mixing evenly, it was vacuum packaged and fermented for 60 days at room temperature in the dark to determine the effects of the screened strain on the nutritional quality and fermentation quality of the silage.

[0103] The results of the nutritional components of the treated silage are shown in Table 4.

[0104] Table 4 Effects of adding L. plantarun LSR-22 on the nutritional components in silage

[0105] Treatment group CK group LSR-22 group Dry matter (DM) / % 21.71±0.22 23.00±0.54 Crude ash (Ash) / % DM 7.97±0.81 8.06±0.72 Crude protein (CP) / % DM 6.10±0.78 9.99±1.39 Water-soluble carbohydrates (WSC) / % DM 7.82±2.51 22.34±3.22 Neutral detergent fiber (NDF) / % DM 70.03±0.01 64.00±0.05 Acid detergent fiber (ADF) / % DM 64.00±0.05 50.09±0.03

[0106] Dry matter content is one of the important indicators to measure the quality of silage, and its level directly affects the nutritional value and storage characteristics of the feed. Crude protein is an important nutritional component in silage, which directly affects the growth and production performance of animals. After 60 days of fermentation, the dry matter and crude protein of the silage increased by 5.90% and 63.80% respectively compared with the control group. Therefore, this feed can significantly improve the growth performance and economic benefits of animals. Increasing the crude protein content in the feed not only helps to improve the production performance of animals, but also can reduce environmental pollution by reducing nitrogen emissions, thus achieving a double improvement in economic and environmental benefits.

[0107] The water-soluble carbohydrate (WSC) content of the LSR-22 group increased by 185.4% compared with the control group, indicating that this strain can effectively inhibit the growth of harmful microorganisms and significantly reduce the loss of water-soluble carbohydrates. A significant increase in the water-soluble carbohydrate (WSC) content in silage usually means that during the ensiling process, there is an adequate fermentation substrate for beneficial microorganisms, which can effectively promote the fermentation activity of lactic acid bacteria, improve the fermentation quality and nutritional value of the feed, and enhance the utilization efficiency of the feed.

[0108] The contents of neutral detergent fiber and acid detergent fiber decreased by 8.61% and 1.63% respectively compared with the control group, indicating that the addition of this bacterium can reduce the fiber content in silage, thereby improving the digestibility of silage.

[0109] The fermentation quality results of the silage after treatment are shown in Table 5.

[0110] Table 5 Effects of adding L. plantarun LSR-22 on the fermentation quality of silage

[0111] Treatment group CK group LSR-22 group Ammoniacal nitrogen content (g / kg) 1.08±0.10 0.96±0.07 pH value 4.18±0.01 4.08±0.02 Lactic acid content (g / kg) 1.46±0.19 3.01±0.58 Acetic acid content (g / kg) 0.06±0.02 0.13±0.05 Propionic acid content (g / kg) 0.21±0.17 0.03±0.04 Butyric acid content (g / kg) - -

[0112] The ammonia nitrogen content in silage is an important indicator to measure the fermentation quality of feed. The ammonia nitrogen content of the silage in this example decreased by 11.1% compared with the control group, indicating that the protein decomposition in the feed is reduced, the nutritional components are relatively more stable, and it is not easy to deteriorate and spoil.

[0113] A significant increase in the lactic acid content in silage has an important impact on the quality preservation and nutritional value of the feed. The lactic acid content of the silage in this example increased by 106.2%, further inhibiting the growth of other harmful microorganisms, and no butyric acid was detected in both the control group and the inoculation group, proving that adding L. plantarun LSR-22 can effectively improve the fermentation quality of silage.

[0114] As an efficient lactic acid bacteria additive, L. plantarun LSR-22 has broad application prospects. In fermentation practice, adding this strain can significantly improve the quality of silage, reduce nutrient loss, and increase the nutritional value of the feed. This has important economic and environmental benefits for livestock production, especially for the feeding of ruminants such as cattle and sheep.

[0115] As a further implementation method, those skilled in the art can further develop lactic acid bacteria additives based on this example.

[0116] As a further implementation, those skilled in the art can further optimize the cultivation conditions of L. plantarun LSR-22 and study its application effects under different raw materials and different environments, with a view to providing higher-quality silage feed additives for the livestock industry.

[0117] In the description of the present invention, it should be understood that "-" and "~" represent the range between two numerical values, and this range includes the endpoints. For example: "A-B" represents a range greater than or equal to A and less than or equal to B. "A~B" represents a range greater than or equal to A and less than or equal to B.

[0118] In the description of the present invention, the term "and / or" herein is merely an associative relationship describing associated objects, indicating that there can be three relationships. For example, A and / or B can represent: A exists alone, A and B exist simultaneously, and B exists alone.

[0119] In the description of the invention, numerical values such as time, temperature, ratio, and mass can be based on actual measurements, equipment standard parameters, simplified rounding results, or within an acceptable error range, ensuring the practicability and reproducibility of the invention.

[0120] In the description of the present invention, the term "about" or "approximately" is used to express an approximate value of a numerical value or interval, allowing for a certain error to ensure the flexibility and practicability of the description, while remaining within an acceptable error range, and the maximum error range does not exceed 10% of the corresponding numerical value or numerical range.

[0121] The above are only the preferred implementation manners of the present invention. It should be noted that the above preferred implementation manners should not be regarded as a limitation of the present invention. The protection scope of the present invention should be subject to the scope defined by the claims. For those of ordinary skill in the art in this technical field, several improvements and refinements can be made without departing from the spirit and scope of the present invention, and these improvements and refinements should also be regarded as within the protection scope of the present invention.

Claims

1. A Lactobacillus plantarum LSR-22, characterized in that The biological preservation number of the Lactobacillus plantarum LSR-22 is: CCTCC M 20242927.

2. An application of Lactobacillus plantarum LSR-22 according to claim 1 in preparing silage, characterized in that: Used to prepare silage for ruminants.

3. A lactic acid bacteria additive, characterized in that The additive comprises the Lactobacillus plantarum LSR-22 as described in claim 1, and is used for improving the fermentation quality and nutritional content of silage.

4. A method for preparing silage, characterized in that: The Lactobacillus plantarum LSR-22 described in claim 1 is inoculated and added to the mixed silage raw materials, the moisture content of the silage raw materials is adjusted to a preset value, the silage raw materials are sealed and packaged, and fermented in the dark for 45 to 70 days.

5. The method according to claim 4, characterized in that The silage raw materials include: one or more of buckwheat straw, silage corn, and alfalfa.

6. The method according to claim 5, characterized in that The silage raw materials are buckwheat straw, silage corn and alfalfa, with a mass ratio of 4:6:

3.

7. The method according to claim 4, characterized in that The inoculation amount of the Lactobacillus plantarum LSR-22 was 1.0×10 5 ~1.0×10 7 cfu / g.

8. The method according to claim 4, characterized in that The moisture content of the silage raw material is 70% to 80%.

9. The method according to claim 4, characterized in that The sealed packaging is followed by vacuum treatment, and the fermentation temperature is 25-40°C.

10. A silage, characterized in that: The silage is prepared by the preparation method according to any one of claims 5 to 9, and its nutritional components include: Dry matter DM: 23.00% ± 0.54%; Ash: 8.06% ± 0.72% DM; Crude protein CP: 9.99% ± 1.39% DM; Water-soluble carbohydrates WSC: 22.34% ± 3.22% DM; Neutral detergent fiber NDF: 64.00% ± 0.05% DM; Acid detergent fiber ADF: 50.09% ± 0.03% DM.

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