Lactobacillus salivarius and application thereof in organic selenium rapeseed meal feed

Abstract

The application discloses a lactobacillus salivarius AKFX-005, a preservation number of which is CGMCC No.38802, and belongs to the technical field of microorganisms. The strain can be used for preparing a microbial agent. The strain or the agent is applied to a fermentation system containing rapeseed meal and sodium selenite to simultaneously degrade glucosinolates and transform organic selenium. 4-12 mg / kg of sodium selenite is added to the fermentation substrate containing rapeseed meal, and the strain or the agent is inoculated to carry out solid state fermentation for 48-72 hours, so that the fermentation product with greatly degraded glucosinolates and significantly increased organic selenium content can be obtained. The strain has good adaptability to the rapeseed meal substrate, and the glucosinolate degradation and the selenium transformation are synergistically enhanced. The experiment shows that the glucosinolate degradation rate in the fermentation product can be more than 55%, and the proportion of organic selenium can be more than 80%. The application provides an excellent strain and a simple and efficient method for the industrialized production of selenium-rich low-toxicity rapeseed meal feed.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, specifically relating to a strain of Lactobacillus salivarius and its application in organic selenium rapeseed meal feed. Background Technology

[0002] Selenium is an essential trace element for animal organisms, playing an irreplaceable role in maintaining life activities. Its physiological functions primarily depend on its participation in the synthesis of selenoproteins in the form of selenocysteine ​​(Sec), thereby regulating key biological processes such as redox homeostasis, immune response, thyroid hormone metabolism, and cell proliferation and differentiation. Among the 25 selenoproteins identified so far, family members such as glutathione peroxidase (GPx), thioredoxin reductase (TrxR), and deiodinase (DIO) have clearly defined functions in scavenging lipid peroxides, maintaining protein sulfhydryl states, and regulating energy metabolism. Numerous studies have shown that selenium deficiency can lead to stunted growth, decreased immunity, reproductive disorders, and muscle degeneration in animals, and exacerbate viral virulence expression and oxidative stress damage, especially in young animals and under stress conditions. Therefore, ensuring a balanced selenium diet for animals is not only fundamental to their healthy growth and efficient production but also an indispensable part of current precision nutrition strategies.

[0003] In animal husbandry practice, selenium supplementation in feed has become the most direct and effective nutritional regulation method for achieving selenium fortification in animals, improving production performance, and producing selenium-enriched livestock products. The core of the selenium supplementation strategy lies in the selection of the selenium source form. Based on chemical form, feed selenium sources can be divided into two main categories: inorganic selenium and organic selenium. Inorganic selenium mainly includes sodium selenite and sodium selenate. While it has the advantages of low cost and stable selenium content, its drawbacks are equally significant: low bioavailability, prominent effective absorption barriers, narrow safe dosage range, and excessive intake can easily induce oxidative stress and tissue damage, posing potential toxicity risks. In contrast, organic selenium is mainly in the form of selenomethionine (SeMet) and selenocysteine. It is efficiently absorbed through the intestinal active transport pathway and stored in the body as an "amino acid pool" in tissues such as muscle, mammary glands, and embryos, exhibiting a slow-release effect. In terms of bioavailability, tissue deposition efficiency, and immunomodulatory activity, organic selenium is significantly superior to inorganic forms. More importantly, the acute and subacute toxicity of organic selenium is far lower than that of inorganic selenium, ensuring its safety in high-dose fortification and long-term use scenarios. Therefore, replacing inorganic selenium with organic selenium is a clear direction for upgrading feed selenium fortification technology and an essential path to achieving standardized and functional production of selenium-enriched livestock products.

[0004] Rapeseed meal is a byproduct obtained from rapeseed through pre-pressing or direct leaching to extract oil. Its crude protein content typically reaches 35%–45%, with a relatively balanced amino acid composition, rich in sulfur-containing amino acids such as methionine and cysteine. It is an important plant-based protein feed resource in my country and globally. my country is a major producer and consumer of rapeseed meal, with annual production consistently at the tens of millions of tons level, widely used in feed for pigs, poultry, aquatic animals, and ruminants. However, the presence of various anti-nutritional factors and toxic substances in rapeseed meal severely limits its efficient application in feed. Among these, glucosinolates (thioglycosides) are the most prominent. While they themselves are not directly toxic, after ingestion by animals, they can be degraded by endogenous myrosinase or intestinal microorganisms, producing degradation products such as isothiocyanates, oxazolidinyl thiones, and nitrile compounds. These products not only have a pungent taste and reduce feed palatability and intake, but can also cause goiter, liver and kidney damage, growth inhibition, and decreased reproductive performance in animals. Rapeseed meal is also high in phytic acid, which strongly chelates minerals such as calcium, zinc, and iron, and inhibits the activity of endogenous proteases and amylases, reducing the utilization rate of feed minerals and energy. Furthermore, rapeseed meal has a high crude fiber content (up to 12% or more), which is poorly digestible in monogastric animals, affecting the effective value of the diet. The phenolic compounds it contains, such as sinapicin and tannins, not only affect palatability but may also produce trimethylamine in laying hens, leading to quality problems such as fishy-tasting eggs.

[0005] To address these shortcomings, various methods for detoxifying and improving rapeseed meal have been developed in existing technologies. Physical methods, such as high-temperature cooking and puffing, can inactivate myrosinase and degrade some heat-sensitive anti-nutritional factors to a certain extent. However, high temperatures easily lead to amino acid destruction and intensified Maillard reactions, resulting in decreased protein digestibility, and the degradation efficiency for glucosinolates is limited. Chemical methods, using acids, alkalis, or copper salts, can degrade glucosinolates or phytic acid, but they pose risks of chemical reagent residues, wastewater discharge, and nutrient loss, resulting in poor economic and environmental performance. Microbial fermentation, utilizing lactic acid bacteria, Bacillus, and yeast for solid-state fermentation of rapeseed meal, can simultaneously degrade glucosinolates, phytic acid, and crude fiber through enzyme systems produced by microbial metabolism, while accumulating microbial protein and beneficial metabolites. This is currently a hot research topic.

[0006] Regarding microbial fermentation, according to literature reports, strain ZUST49 can achieve a degradation rate of 61.3% for glucosinolates within 12 hours. It can secrete a novel glucosidase that converts inactive glucosinolates into highly active glucosinolates. In rapeseed meal fermentation, when lactic acid bacteria are mixed with other bacteria, the glucosinolate degradation rate can reach over 73.5% after 40 hours of fermentation, and it can significantly improve the protein digestibility of the feed.

[0007] While existing technologies include methods for detoxifying rapeseed meal through microorganisms, no microbial strains have been found that simultaneously degrade and detoxify glucosinolates in rapeseed meal and convert it into organic selenium. Furthermore, there are no related solutions for using microbial fermentation to directly incorporate inorganic selenium into the rapeseed meal protein matrix to prepare rapeseed meal feed that combines detoxification and organic selenium nutritional fortification. Summary of the Invention

[0008] In response to the lack of strains that are effective at both detoxifying and enriching rapeseed meal with selenium, this invention screened and obtained a strain of *Lactobacillus salivarius* AKFX-005, which has both high glucosinolate degradation capacity and high inorganic selenium conversion activity. This strain was applied to rapeseed meal fermentation to prepare organic selenium feed, which can simultaneously achieve glucosinolate detoxification and quality improvement of rapeseed meal and bioconversion of inorganic selenium to organic selenium. The glucosinolate content of the detoxified rapeseed meal is significantly reduced, the proportion of organic selenium is greatly increased, and both safety and nutritional value are effectively improved. It can be directly applied to the formulation of livestock and poultry feed.

[0009] This invention first provides a *Lactobacillus salivarius*. Lactobacillus salivarius This strain was deposited at the China General Microbiological Culture Collection Center on May 20, 2025. Lactobacillus salivarius The accession number is: CGMCC No.38802, Latin name: Lactobacillus salivarius AKFX-005, Address: Room H129, CGMCC, Institute of Microbiology, Chinese Academy of Sciences, No. 3, Beichen West Road, Chaoyang District, Beijing, 100101, China. Tel: 010-64807596.

[0010] This invention provides the application of the Lactobacillus salivarius AKFX-005 or the microbial agent in a fermentation system containing both rapeseed meal and sodium selenite, for degrading glucosinolates and converting sodium selenite into organic selenium during fermentation.

[0011] In this application, the preferred amount of sodium selenite added is 4–12 mg / kg based on the total weight of the fermentation substrate; the fermentation method is solid-state fermentation, and the fermentation time is 48–72 hours. More preferably, the amount of sodium selenite added is 9.5 mg / kg, and the fermentation time is 60 hours.

[0012] The present invention also provides a method for degrading glucosinolates in rapeseed meal, comprising the following steps: using rapeseed meal or a mixed feed containing rapeseed meal as a fermentation substrate, inoculating with the Lactobacillus salivarius AKFX-005 or the microbial agent, and adding sodium selenite at an addition rate of 4-12 mg / kg, and carrying out solid-state fermentation for 48-72 hours.

[0013] The present invention also provides a method for producing selenium-enriched low-sulfur glycoside feed, comprising the following steps: using rapeseed meal or a mixed feed containing rapeseed meal as a fermentation substrate, inoculating with the Lactobacillus salivarius AKFX-005 or the microbial agent, and adding sodium selenite at an addition rate of 4-12 mg / kg, and carrying out solid-state fermentation for 48-72 hours to obtain selenium-enriched low-sulfur glycoside feed. Beneficial effects

[0014] Compared with the prior art, the beneficial effects of the present invention are as follows: First, this invention provides a *Lactobacillus salivarius* strain, deposited at the China General Microbiological Culture Collection Center (CGMCC) under accession number CGMCC No. 38802. This strain was isolated from selenium-enriched rapeseed meal fermentation feed with a high organic selenium conversion rate. It exhibits natural adaptability to the rapeseed meal fermentation environment, can stably grow and exert metabolic activity in the rapeseed meal substrate, and can be directly applied without complex domestication or modification. Simultaneously, this strain possesses dual functions of highly efficient glucosinolate degradation and highly efficient inorganic selenium conversion. The screening process used both glucosinolate degradation rate and organic selenium conversion rate as dual screening indicators. The resulting strain has a clear genetic background and stable functional traits, solving the problem of the lack of dedicated strains capable of simultaneously performing both functions in existing technologies.

[0015] Secondly, the fermentation process provided by this invention can simultaneously complete the two core tasks of rapeseed meal detoxification and organic selenium bioconversion in the same fermentation system, without the need for step-by-step processing. The process steps are simple, adaptable to the processing requirements of existing feed solid-state fermentation production lines, and require no additional equipment investment or process complexity, thus possessing good prospects for industrial application. Compared with the traditional segmented processing scheme of first detoxifying and then supplementing selenium, this method shortens the production process, reduces processing energy consumption and production costs, and also avoids the risk of contamination by miscellaneous bacteria that may be introduced during secondary processing.

[0016] Finally, the selenium-enriched low-glucosinolate rapeseed meal feed prepared by the method of this invention has a glucosinolate degradation rate of over 55%, and the proportion of organic selenium in the total selenium can be stably maintained at over 80%. Compared with unfermented rapeseed meal, it not only reduces the anti-nutritional effects and toxicity risks of glucosinolates, but also improves the nutritional value and feeding safety of rapeseed meal. It also completes the organic conversion of selenium source, and the obtained organic selenium has high activity, low toxicity, and good bioavailability. It can be directly used to formulate selenium-enriched livestock and poultry feed and produce selenium-enriched livestock products, providing a new technical path for the high-value utilization of rapeseed meal, and has significant economic value and application prospects.

[0017] This invention provides key microbial resources and technical solutions for the low-cost, high-efficiency production of low-toxicity selenium-enriched rapeseed meal feed, and has good prospects for industrial application. Attached Figure Description

[0018] Figure 1 Microscopic images of the *Lactobacillus salivarius* selected in Example 1; Figure 2 Images of Lactobacillus salivarius colonies screened in Example 1; Figure 3 The DNA sequence of Lactobacillus salivarius screened in Example 1; Figure 4 The growth curve of Lactobacillus salivarius screened in Example 1 is shown. Detailed Implementation

[0019] Unless otherwise specified, the experimental methods used in the following examples are conventional methods. Unless otherwise specified, the experimental materials used in the following examples were all purchased from conventional biochemical reagent stores. All quantitative experiments in the following examples were performed in triplicate, and the results were averaged. In the following examples, the water used to prepare each culture medium was deionized water.

[0020] In the following examples, the OD600nm value was used to characterize the bacterial growth. Unless otherwise specified, all reagent kit materials used in the following examples are commercially available.

[0021] MRS liquid culture medium 20g glucose, 10g peptone, 10g beef extract, 5g yeast extract, 2g diammonium citrate, 5g sodium acetate, 0.58g magnesium sulfate, 0.25g manganese sulfate, 1mL Tween-80, 1000mL distilled water, pH adjusted to 6.2~6.4.

[0022] MRS solid culture medium 20g glucose, 10g peptone, 10g beef extract, 5g yeast extract, 2g diammonium citrate, 5g sodium acetate, 0.58g magnesium sulfate, 0.25g manganese sulfate, 1mL Tween-80, 15g agar, 1000mL distilled water, pH adjusted to 6.2-6.4, sterilized at 121℃ for 20min, shake well after sterilization, cool to about 50℃ and pour into plates.

[0023] The glucosinolate content was determined using the palladium chloride colorimetric method.

[0024] Glucoside degradation rate = [(Glucoside content in unfermented rapeseed meal - Glucoside content in fermented rapeseed meal) / Glucoside content in unfermented rapeseed meal] * 100%. Example 1

[0025] This embodiment provides a method for screening, purifying, and preserving Lactobacillus salivarius, as detailed below.

[0026] In the early stages of preparation of selenium-enriched rapeseed meal feed, the laboratory found that the organic selenium conversion rate of some rapeseed meal feeds was higher than that of other feeds. Therefore, the lactic acid bacteria in these rapeseed meal feeds were isolated and purified, and a strain of Lactobacillus salivarius was obtained. The specific steps are as follows.

[0027] (1) Sample collection and pretreatment Under aseptic conditions, 10 g of rapeseed meal feed sample with high organic selenium content was weighed and placed in an Erlenmeyer flask containing 90 mL of sterile physiological saline. The sample was shaken at 180 r / min for 30 min to disperse it fully, thus obtaining a sample suspension with a concentration of 0.1 g / mL.

[0028] (2) Enrichment culture of organic selenium-converting bacteria Based on MRS liquid medium, sodium selenite was added to a final concentration of 1.0 mmol / L to adjust the pH to 6.5. After aliquoting, the solution was sterilized at 121°C for 15 min. 5 mL of the above sample suspension was inoculated into 100 mL of enrichment medium and anaerobically cultured at 37°C for 48 h to obtain the enrichment culture.

[0029] (3) Isolation of lactic acid bacteria The enriched culture medium was serially diluted 10-fold with sterile physiological saline, and 10 were selected. -4 ~10 -6 Dilute each sample by taking 0.1 mL and spreading it onto an MRS solid-state screening plate.

[0030] The screening plates were prepared by adding 1.0 mmol / L sodium selenite and 2% CaCO3 to MRS solid medium, pouring the mixture onto plates, and then incubating it anaerobicly at 37°C for 48 h. Single colonies that were milky white, raised, with neat edges and a clear transparent calcium dissolution zone were selected and preliminarily identified as lactic acid bacteria strains.

[0031] (4) Initial screening of glucosinolate degradation ability The isolated strains were inoculated onto glucosinolate-based chromogenic plates. These plates were based on MRS agar, with 0.3 mmol / L potassium myrosinate and 0.5 mmol / L palladium chloride (PdCl2) added as chromogenic agents. The formation of a brown complex between glucosinolates and palladium chloride, and the appearance of a fading zone around the colony, indicated the presence of glucosinolate-degrading ability.

[0032] After cultivation, strains with a ratio of ≥3.0 between the diameter of the bleaching zone and the diameter of the colony were selected, transferred to MRS slant, and temporarily stored at 4℃ as excellent strains for initial screening.

[0033] (5) Secondary screening and strain identification The initially screened strains were inoculated into MRS liquid medium containing 0.3 mmol / L potassium glucosinolate and 5 mg / L sodium selenite, and cultured anaerobicly at 37°C for 48 h. After culture, the supernatant was collected by centrifugation, and the glucosinolate residue was determined by palladium chloride colorimetry to calculate the glucosinolate degradation rate. The organic selenium content in the culture was determined by atomic fluorescence spectrometry to calculate the organic selenium conversion rate. One isolate with a glucosinolate degradation rate ≥85% and a significantly better organic selenium conversion rate than other strains was retained and numbered as follows: AKFX-005 .

[0034] The 16S rDNA of the selected strains was amplified, and the purified PCR product was sequenced. The sequencing results are shown in Sequence 1 of the sequence listing. Figure 1 As shown in the figure.

[0035] Based on the combined results of morphological and molecular biological identification, the selected strain belongs to Lactobacillus salivarius.

[0036] (6) Purification Select Lactobacillus salivarius strains AKFX-005 The colonies were streaked three times on MRS plates, and a typical single colony was picked each time to transfer to the next generation. Colony morphology was observed, and colony photographs are shown below. Figure 2 See electron microscope images Figure 3 The selected strains exhibit round, transparent colonies with neat edges and smooth surfaces. Under an inverted microscope, the bacterial cells appear rod-shaped.

[0037] (7) Preservation Purified Lactobacillus salivarius AKFX-005 Inoculate the culture into MRS liquid medium and incubate at 37°C for 18 h until the late logarithmic growth phase. Take 600 μL of bacterial suspension and mix it with 400 μL of sterile 80% glycerol. Aliquot the mixture into 2 mL cryovials, label them, and store them in an ultra-low temperature freezer at -80°C. At the same time, prepare lyophilized bacterial powder using a vacuum freeze-drying method and store it at 4°C in the dark. This completes the screening, purification, and preservation of Lactobacillus salivarius.

[0038] The selected Lactobacillus salivarius AKFX-005 The strain has been deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 38802. The address is Room H129, CGMCC, Institute of Microbiology, Chinese Academy of Sciences, No. 3, Beichen West Road, Chaoyang District, Beijing, 100101, China. The telephone number is 010-64807596.

[0039] (8) Growth curve The preserved *Lactobacillus salivarius* was inoculated onto streak plates for culture, and single colonies from the plates were selected and incubated in 10 mL of MRS liquid medium at 37°C for 24 h to obtain *Lactobacillus salivarius* seed culture. The *Lactobacillus salivarius* seed culture was inoculated into 150 mL of MRS liquid medium at a 1% inoculation rate and incubated statically at 37°C for 48 h. Samples were taken every 3 h, and the optical density (OD) value was measured at 600 nm using a UV-Vis spectrophotometer. An OD600 versus time curve was plotted.

[0040] See results Figure 4 The *Lactobacillus salivarius* screened in Example 1 entered the logarithmic growth phase between 3 and 24 hours after a brief lag phase, reached its maximum growth concentration at 33 hours, and then entered the decline phase. Example 2

[0041] This embodiment provides a method for preparing Lactobacillus salivarius AKFX-005 into a lyophilized bacterial agent, the specific method of which is as follows.

[0042] 1. Streak preserved *Lactobacillus salivarius* AKFX-005 onto MRS solid plates and incubate anaerobically at 37°C for 24 h. Select typical single colonies and inoculate them into 10 mL of MRS liquid medium, then incubate statically at 37°C for 18 h to obtain the primary seed culture. Transfer the primary seed culture to 200 mL of MRS liquid medium at a 2% (v / v) inoculation rate and incubate statically at 37°C for 18 h to obtain the secondary seed culture.

[0043] 2. Inoculate the secondary seed culture at a rate of 5% (v / v) into a fermenter containing 5 L of MRS liquid medium. Incubate anaerobically at 37°C for 24 h, taking samples aseptically every 2 h to determine the OD600 value. When the OD600 value of the culture reaches above 3.0 and the pH drops to around 4.0, stop the culture and obtain the fermentation broth.

[0044] 3. Centrifuge the fermentation broth at 4℃ and 8000×g for 15 min, and discard the supernatant. Wash the bacterial precipitate twice with sterile physiological saline, and collect the wet bacterial cells by centrifugation again.

[0045] 4. Prepare a freeze-drying protectant by weight percentage: 10% skim milk powder, 5% sucrose, 3% glycerol, 1% monosodium glutamate, with the remainder being water. After thorough dissolution, sterilize at 115℃ for 20 min. Mix the collected wet bacterial cells with the protectant solution at a ratio of 1:2 (w / v) to obtain a bacterial suspension.

[0046] 5. Dispense the bacterial suspension into sterile vials, 2 mL per vial. Pre-freeze at -80℃ for 6 h, then place in a vacuum freeze dryer and freeze-dry at -50℃ and a vacuum of less than 10 Pa for 24 h to obtain Lactobacillus salivarius AKFX-005 freeze-dried bacterial powder. Example 3

[0047] This embodiment provides the use of preserved Lactobacillus salivarius. AKFX-005 The specific steps for fermenting rapeseed meal and reducing glucosinolates are as follows.

[0048] 1. After crushing the rapeseed meal, pass it through a 40-mesh sieve and sterilize it at 121℃ for 15 minutes. Accurately weigh 50g of rapeseed meal into a 250mL beaker to obtain the rapeseed meal fermentation substrate.

[0049] 2. The *Lactobacillus salivarius* prepared according to the method in Example 1 AKFX-005 The seed liquid and sterile water were mixed at a ratio of 1:10, and then mixed with rapeseed meal fermentation substrate at a ratio of 2:1. Finally, the mixture was sealed with a breathing membrane.

[0050] 3. Subsequently, the samples were placed under constant temperature for solid-state fermentation, with the samples turned over once every 12 hours to ensure uniform fermentation. The glucosinolate content of the fermented samples was measured at 24h, 36h, 48h, 60h, and 72h, and the glucosinolate degradation rate before and after fermentation was determined based on the glucosinolate content before fermentation. The results are shown in Table 1.

[0051] Table 1. Effect of fermentation time on glucosinolate content in rapeseed meal. Degradation rate of glucosinolates (%) 22.68 28.13 32.08 36.27 37.03 As can be seen from the data in Table 1, the degradation rate of glucosinolates gradually increases with the extension of fermentation time. When the fermentation time reaches 60h, the degradation rate slows down significantly. Further extending the fermentation time to 72h only increases the degradation rate of glucosinolates by 0.76 percentage points. Therefore, considering both production efficiency and degradation effect, the optimal time for this strain to degrade glucosinolates in rapeseed meal through solid-state fermentation can be set at 60h. Example 4

[0052] This embodiment provides the use of preserved Lactobacillus salivarius. AKFX-005 The specific steps of the method for fermenting rapeseed meal with added sodium selenite to reduce glucosinolates and convert it into organic selenium are as follows.

[0053] 1. After crushing the rapeseed meal, pass it through a 20-mesh sieve and sterilize it at 121℃ for 15 minutes. Accurately weigh 50g of rapeseed meal into a 250mL beaker to obtain the rapeseed meal fermentation substrate.

[0054] 2. The *Lactobacillus salivarius* prepared according to the method in Example 1 AKFX-005The seed liquid and sterile water were mixed at a ratio of 1:10, and then mixed with rapeseed meal fermentation substrate at a ratio of 2:1. Based on the weight of rapeseed meal, 0.2 mg, 0.3 mg, 0.4 mg, 0.5 mg, and 0.6 mg of sodium selenite were weighed out and added to the above mixture to prepare mixed fermentation substrates with sodium selenite content of 4, 6, 8, 10, and 12 mg / kg. Finally, the mixture was sealed with a breathing membrane.

[0055] 3. Subsequently, the samples were placed under constant temperature for solid-state fermentation, and turned over once every 12 hours to ensure uniform fermentation. The glucosinolate content of the fermented samples after 60 hours of fermentation was measured, and the glucosinolate degradation rate before and after fermentation was determined based on the glucosinolate content before fermentation. The results are shown in Table 2.

[0056] Table 2. Effects of sodium selenite addition on glucosinolate and selenium content in rapeseed meal. Degradation rate of glucosinolates (%) 36.75 42.03 55.27 56.09 56.18 L-Selenomethylselenocysteine ​​content (mg / kg) 1.12 2.10 3.84 4.82 4.96 Selenomethionine content (mg / kg) 0.76 0.98 1.95 2.26 2.41 Total selenium content (mg / kg) 3.72 5.65 7.03 8.59 10.25 Organic selenium percentage (%) 50.54 54.51 82.36 82.42 71.90 As can be seen from the data in Table 2, With increasing sodium selenite concentration, the degradation rate of glucosinolates gradually increased, reaching a peak of 56.09% at a concentration of 0.5 mg. Further increases in selenium concentration only slightly improved the degradation rate, with no further significant increase. The proportion of organic selenium also showed a trend of first increasing, then stabilizing, and then decreasing. When the sodium selenite concentration was 0.4–0.5 mg, the proportion of organic selenium remained stable above 82%, but decreased to 71.90% when the concentration increased to 0.6 mg. This indicates that excessively high sodium selenite concentrations inhibit the selenium conversion ability of *Lactobacillus salivarius* AKFX-005. Therefore, the addition of 0.8–1.2 mg / kg sodium selenite significantly improved the degradation activity of *Lactobacillus salivarius* AKFX-005 for glucosinolates. Simultaneously, the conversion rate of organic selenium also reached over 80%. Example 5

[0057] The inventors conducted an orthogonal experiment on the amount of sodium selenite added, and the result showed that the optimal amount of sodium selenite added was 9.5 mg / kg (based on the weight of the fermentation substrate). The applicant replaced the fermentation substrate with soybean meal, and the organic selenium content data after 60 hours of fermentation is shown in Table 3.

[0058] Table 3. Effects of different fermentation substrates on selenium content L-Selenomethylselenocysteine ​​content (mg / kg) 3.37 4.92 Selenomethionine content (mg / kg) 1.52 2.17 Total selenium content (mg / kg) 8.40 8.35 Organic selenium percentage (%) 58.21 84.91 As shown in Table 3, Lactobacillus salivarius AKFX-005 exhibits superior selenium conversion ability in rapeseed meal fermentation substrate. Its organic selenium ratio and target active selenium content are significantly higher than those of soybean meal fermentation substrate, indicating that this strain has better compatibility with rapeseed meal substrate and is more suitable for the production and processing of selenium-enriched rapeseed meal feed.

[0059] Based on these results, it is inferred that under conditions where the fermentation substrate contains both rapeseed meal and a specific concentration of sodium selenite, *Lactobacillus salivarius* AKFX-005 can simultaneously complete glucosinolate degradation and efficient selenium conversion. The specific mechanism requires further experimental investigation. Preliminary analysis suggests that rapeseed meal itself may contain a component that promotes the secretion of an enzyme by *Lactobacillus salivarius* AKFX-005 that degrades glucosinolates and converts organic selenium. Within a concentration range of 0.8–1.2 mg / kg, sodium selenite can be converted into organic selenium by the strain without causing oxidative damage that inhibits the strain's growth and metabolic activities, thus achieving a synergistic effect between glucosinolate degradation and selenium conversion. Example 6

[0060] This embodiment provides a method for fermenting a compound feed containing 20% ​​rapeseed meal and reducing glucosinolates using preserved Lactobacillus salivarius AKFX-005. The specific steps are as follows.

[0061] 1. Feed formulation (by weight): 50 parts corn, 15 parts soybean meal, 20 parts rapeseed meal, 10 parts wheat bran, 1 part vegetable oil, 3 parts DDGS, and 1 part salt. Grind all ingredients separately and pass through a 40-mesh sieve. Mix thoroughly and sterilize at 121℃ for 15 minutes. Accurately weigh 100 g of the mixed feed into a 500 mL beaker as the fermentation substrate.

[0062] 2. The Lactobacillus salivarius AKFX-005 seed culture prepared according to the method of Example 1 was mixed with sterile water at a volume ratio of 1:5. The mixture was then added to the above fermentation substrate at a material-to-liquid ratio of 1:1 (w / v), i.e., 100 mL of mixed bacterial culture was added. Then, 0.95 mg of sodium selenite was added at a ratio of 9.5 mg / kg. After thorough mixing, the mixture was placed in an aerobic and sterile environment.

[0063] 3. The sample was placed at a constant temperature of 37℃ for solid-state fermentation for 60 hours to obtain a standard-compliant detoxified selenium-enriched compound fermented feed. After fermentation, samples were taken to determine the glucosinolate content and selenium composition. The results showed that the glucosinolate degradation rate in the compound feed reached 86.37%, and the total organic selenium content reached 80.92%, meeting the feed hygiene standards and production requirements for selenium-enriched feed, and can be directly used for livestock and poultry feeding. This example verifies the feasibility of applying Lactobacillus salivarius AKFX-005 in compound feed production. This strain can normally exert its degradation activity in commercial compound feed systems containing rapeseed meal, and is suitable for the processing requirements of industrial feed production.

Claims

1. A strain of Lactobacillus salivarius AKFX-005 Its features are: The Lactobacillus salivarius AKFX-005 Its Latin name is: Lactobacillus salivarius It is deposited in the China General Microbiological Culture Collection Center, with accession number CGMCC No. 38802.

2. A microbial inoculant, characterized in that, It contains Lactobacillus salivarius AKFX-005 as described in claim 1.

3. The application of the *Lactobacillus salivarius* AKFX-005 according to claim 1 or the microbial agent according to claim 2, characterized in that: When the fermentation substrate contains both rapeseed meal and sodium selenite, it is used to degrade glucosinolates and convert sodium selenite into organic selenium.

4. The application of Lactobacillus salivarius AKFX-005 as described in claim 1 or the microbial agent as described in claim 2 in organic selenium-enriched rapeseed meal feed, characterized in that: The Lactobacillus salivarius AKFX-005 or microbial agent is used to ferment rapeseed meal feed containing sodium selenite, and to prepare organic selenium rapeseed meal feed.

5. The application of Lactobacillus salivarius AKFX-005 according to claim 3 or the microbial agent according to claim 2, characterized in that: The amount of sodium selenite added is 4–12 mg / kg based on the total weight of the fermentation substrate.

6. The application of Lactobacillus salivarius AKFX-005 according to claim 3 or the microbial agent according to claim 2, wherein the fermentation is solid-state fermentation and the fermentation time is 48-72 hours.

7. A method for degrading glucosinolates in rapeseed meal, characterized in that, The process includes the following steps: using rapeseed meal or a mixed feed containing rapeseed meal as the fermentation substrate, inoculating with Lactobacillus salivarius AKFX-005 as described in claim 1 or the microbial agent as described in claim 2, and adding sodium selenite at an addition rate of 4~12 mg / kg, and fermenting for 48~72 hours.

8. A method for producing selenium-enriched, low-sulfur glycoside feed, characterized in that, The process includes the following steps: using rapeseed meal or a mixed feed containing rapeseed meal as the fermentation substrate, inoculating with Lactobacillus salivarius AKFX-005 as described in claim 1 or the microbial agent as described in claim 2, and adding sodium selenite at an addition rate of 4~12 mg / kg, and fermenting for 48~72 hours.

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