Lactobacillus amyloliquefaciens for improving lean meat percentage of Bama pigs and application of lactobacillus amyloliquefaciens
By screening and applying Lactobacillus amyloliquefaciens WANRU1108 from the intestines of Bama pigs as a feed additive, the problem of poor colonization of probiotic strains in the intestines of Bama pigs in existing technologies has been solved, achieving the effects of reducing fat deposition and increasing lean meat percentage.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-12-29
- Publication Date
- 2026-03-13
AI Technical Summary
Existing technologies have failed to effectively combine the physiological characteristics and metabolic patterns of Bama pigs, resulting in poor colonization of the selected probiotic strains in their intestines, making it difficult to effectively regulate fat deposition and affecting meat quality and economic benefits.
A strain of Lactobacillus amylovorus WANRU1108 was screened from the intestines of Bama pigs and prepared as a feed additive. By adding it to the feed of Bama pigs, it can reduce the expression of genes that absorb nutrients in the intestines, reduce fat deposition, and increase lean meat percentage.
It stably colonizes in the intestines of Bama pigs, significantly reduces fat deposition, increases lean meat percentage by 9.8%, improves meat quality, reduces subcutaneous fat by 15.3%, and backfat thickness by 12.7%, without affecting growth performance.
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Abstract
Description
Technical Field
[0001] This invention relates to the field of microbial technology, specifically to a strain of Lactobacillus amyloliquefaciens that improves the lean meat percentage of Bama pigs and its application. Background Technology
[0002] Pork is one of the most consumed meats in the world. However, with the improvement of people's living standards, the demand for pork quality is also increasing. Bama miniature pigs are a famous local breed from Guangxi, China, and one of the smallest pig breeds among the world's original breeds. Compared with imported lean-type pigs, they have a better taste, higher fat content, and lower feed conversion ratio, which greatly affects the production and promotion of Bama miniature pigs. Animal fat includes subcutaneous fat, abdominal fat, visceral fat, intramuscular fat, and intramuscular fat. Their deposition has temporal and spatial characteristics, with subcutaneous fat depositing first, followed by abdominal and visceral fat. These fats primarily protect the animal and are therefore sometimes referred to as "waste fat" in production. Finally, fat is deposited in the muscle, and its content affects the taste of pork and consumer preference, making it one of the most important traits.
[0003] Importantly, the deposition of nutrients in muscle plays a crucial role in pork quality, including amino acids, fatty acids, and volatile substances. Studies have found that organic iron and zinc improve the quality of raw pork (color, pH, shear force, marbling score, intramuscular fat content, intramuscular fat content, and zinc content) and the sensory characteristics of cooked meat (odor, flavor, tenderness, juiciness, and freshness of juices) by increasing flavor compounds, altering fatty acid composition, and changing the metabolic profile. Research on the quality of Bama miniature pigs is relatively limited, with most previous studies focusing on the medical field. Therefore, this study investigated a strain of *Lactobacillus amyloliquefaciens* isolated from Bama pig feces and studied its ability to reduce fat deposition in Bama miniature pigs by decreasing the expression of genes involved in intestinal nutrient absorption.
[0004] Bama pigs are a national geographical indication product of Guangxi Zhuang Autonomous Region. Their superior meat quality has won consumer favor. However, excessive fat accumulation during their growth process affects both the taste and texture of the meat, increasing feeding costs and reducing economic efficiency. Currently, there are no nutritional strategies or technical solutions specifically designed to improve the meat quality of Bama pigs. In Bama pig farming, antibiotics have been widely used to prevent disease and indirectly improve growth performance. However, the overuse of antibiotics not only leads to the proliferation of drug-resistant strains in pigs, posing public health and safety risks by transmitting them to humans through the food chain, but also completely disrupts the intestinal flora structure of Bama pigs, inhibiting the growth of beneficial bacteria and further exacerbating lipid metabolism disorders. Studies have shown that Bama pigs treated with tetracycline antibiotics for a long period experienced a reduction of more than 40% in the number of short-chain fatty acid-producing bacteria in their intestines, while the number of endotoxin-producing harmful bacteria significantly increased, resulting in decreased fat decomposition efficiency.
[0005] With the increasing application of probiotic regulation technology in pig farming, its use in regulating fat metabolism in Bama pigs has been explored. However, existing strain screening techniques do not fully consider the physiological characteristics and metabolic patterns of Bama pigs, resulting in limited effectiveness and poor applicability of the selected strains. Currently, most probiotic strains used for fat regulation in pigs originate from the intestines of ordinary commercial pigs, fermented foods (such as kimchi and yogurt), or the soil environment. Only a few studies have isolated strains from the intestines of Bama pigs themselves. As a local breed, Bama pigs exhibit significant differences in intestinal pH (average 6.2), digestive enzyme activity, and gut microbiota composition compared to imported commercial pigs, making it difficult for imported strains to colonize their intestines. Furthermore, existing screening methods primarily focus on "improving growth performance" and "improving gut health," with evaluation indicators concentrated on characteristics such as acid resistance, bile salt tolerance, and antibacterial activity, without considering "regulating fat metabolism" as a core screening objective. Therefore, this study screened a probiotic strain—Lactobacillus amyloliquefaciens—from the intestines of Bama pigs to investigate its effect on fat deposition in Bama pigs, which can be applied to Bama pig farming to increase the lean meat ratio.
[0006] It should be noted that the information disclosed in the background section above is only used to enhance the understanding of the background of this disclosure, and therefore may include information that does not constitute prior art known to those skilled in the art. Summary of the Invention
[0007] The purpose of this invention is to provide a strain of Lactobacillus amyloliquefaciens that can improve the lean meat percentage of Bama pigs, thereby addressing the problems of high fat content and low feed conversion rate in Bama pigs. At the same time, it provides a safe and effective microbial technology solution for targeted improvement of the body composition and lean meat percentage of Bama pigs.
[0008] To achieve the above objectives, this invention provides a strain of *Lactobacillus amyloliquefaciens* that improves the lean meat percentage of Bama pigs. Lactobacillus amylovorus WANRU1108, this strain was deposited on December 16, 2024 at the China Center for Type Culture Collection (CCTCC), Wuhan University, Wuhan, China, with accession number CCTCC NO: M20242823.
[0009] The Lactobacillus amyloliquefaciens WANRU1108 provided by this invention can be used to prepare feed additives that improve the lean meat percentage of Bama pigs.
[0010] The present invention also provides a feed additive for improving the lean meat percentage of Bama pigs, the additive comprising the aforementioned Lactobacillus amyloliquefaciens WANRU1108.
[0011] Preferably, the live bacterial concentration of *Lactobacillus amyloliquefaciens* WANRU1108 in the feed additive provided by the present invention is 4.4 × 10⁻⁶. 9 CFU / mL.
[0012] The Lactobacillus amyloliquefaciens WANRU1108 or feed additive provided by this invention can be applied in the field of Bama pig farming, including reducing fat deposition in Bama pigs, increasing lean meat percentage in Bama pigs, and improving the quality of Bama pork.
[0013] The present invention also provides a method for improving the lean meat percentage of Bama pigs by adding the above-mentioned Lactobacillus amyloliquefaciens WANRU1108 to the basal diet and feeding Bama pigs.
[0014] Preferably, the feeding method is as follows: feed once daily for the first week, and then feed three times a week thereafter; each feeding amount is 4.4 × 10⁻⁶ live bacteria concentration. 9 50 mL of bacterial solution per head with a CFU / mL concentration should be fed for at least 10 weeks.
[0015] The present invention has the following advantages: This invention screened a novel *Lactobacillus amyloliquefaciens* strain, WANRU1108, which can be prepared as a feed additive. It can stably colonize in the intestinal environment at pH 6.2. Animal experiments verified that it can be successfully colonized at a growth rate of 4.4 × 10⁻⁶. 9 Adding CFU / mL to the feed and feeding Bama pigs for 10 weeks significantly reduced the expression of nutrient absorption genes such as SGLT1, GLUT2, and FATP4 in the jejunum and ileum, reduced fat deposition, decreased subcutaneous fat weight percentage by 15.3%, backfat thickness by 12.7%, while increasing lean meat percentage by 9.8% and improving pork shear force by 22.4%. This invention provides a green nutritional regulation scheme for Bama pig farming that can replace antibiotics. Attached Figure Description
[0016] Figure 1 Phylogenetic tree constructed based on the sRNA sequence of strain WANRU110816.
[0017] Figure 2 This presents the results of the effects of Lactobacillus amyloliquefaciens on the growth and slaughter performance of Bama pigs in this invention.
[0018] Figure 3 This invention describes the effect of Lactobacillus amyloliquefaciens on the quality of Bama pork.
[0019] Figure 4 The results show the effects of *Lactobacillus amyloliquefaciens* on serum biochemical indicators in Bama pigs in this invention.
[0020] Figure 5 This invention illustrates the effect of *Lactobacillus amyloliquefaciens* on the expression of lipid metabolism genes in the jejunum of Bama pigs.
[0021] Figure 6 This invention illustrates the effect of *Lactobacillus amyloliquefaciens* on the expression of lipid metabolism genes in the ileum of Bama pigs. Detailed Implementation
[0022] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.
[0023] Note: Unless otherwise specified, the experimental methods in the following examples are conventional methods, performed according to the techniques or conditions described in the literature in this field or according to the product instructions. Unless otherwise specified, the materials and reagents used in the following examples are commercially available.
[0024] Example 1: Isolation and Identification of Strains Feces from Bama pigs were collected, and a bacterial strain was screened from the fecal samples. Its 16SRNA was amplified, and its sequence is shown in SEQ ID NO. 17. A phylogenetic tree was constructed, as shown below. Figure 1 The strain was identified as *Lactobacillus amyloliquefaciens* (Lactobacillus). Lactobacillus amylovorus The strain was named WANRU1108 and deposited for patent protection. The deposit information is as follows: deposited on December 16, 2024 at the China Center for Type Culture Collection (CCTCC), Wuhan University, Wuhan, China, with accession number CCTCC NO: M20242823.
[0025] The 16SRNA sequence of strain WANRU1108 (SEQ ID NO.17):
[0026] Example 2: Application Validation of the Strains The screened strains were cultured on MRS liquid medium (MRSB, Qingdao Haibo Biotechnology Co., Ltd.) or MRS solid medium (MRSA) plates at 37°C and pH 6.2. After culture, the *Lactobacillus amyloliquefaciens* strains were counted using the plate count method. Finally, the concentration was adjusted to 4.4 × 10⁻⁶. 9 Prepare a bacterial culture at CFU / mL for later use.
[0027] Bama pigs were purchased from the Rural Revitalization Research Institute of Bama Yao Autonomous County, Guangxi. Eighteen Bama pigs of similar weight (25±0.5 kg) and health status were divided into two groups: a control group and a Lactobacillus amyloliquefaciens group, with nine pigs in each group. The control group was fed a basal diet; the composition of the basal diet and the nutritional levels after measurement are shown in Table 1. The Lactobacillus amyloliquefaciens group (LA group) was fed a basal diet plus Lactobacillus amyloliquefaciens: the prepared live bacteria concentration was 4.4×10⁻⁶. 9 The bacterial culture at CFU / mL was added to a small amount of food at a rate of 50 mL / head and stirred before being fed to the pigs within 30 minutes. For the first week, the pigs were fed Lactobacillus amyloliquefaciens daily at 9:00 AM. For the following nine weeks, the pigs were fed three times a week, with the experimental period lasting 10 weeks.
[0028] Table 1. Composition and nutrient levels of basal diet (air-dried basal, %) Each kilogram of the premix diet contains the following: Vitamin A 1300 IU, Vitamin D 3150 IU, Vitamin E 11 IU, Vitamin K 30.5 mg, Vitamin B1 2.5 mg, Vitamin B2 1.0 mg, Vitamin B6 1.0 mg, and Vitamin B12.5 mg. 12 0.01mg, niacin 30.0mg, folic acid 0.30 mg, pantothenic acid 10.0 mg, biotin 0.05 mg, Fe 60mg, Zn 60 mg, Mn 2.0 mg, Cu 4.0 mg, I 0.14mg, Se 0.20 mg.
[0029] The growth and physiological indicators of Bama pigs after 10 weeks of feeding were measured, as follows: 1. Assessment of slaughter performance and pork quality After the experiment, carcasses of Bama pigs from each group were collected, and lean meat, back fat, and subcutaneous fat were separated. Backfat thickness (located between the third and fourth ribs from the bottom) was measured. Additionally, samples of the longissimus dorsi muscle were collected between the 10th and 12th ribs. Meat color (L*, a*, b*), shear force, and pH value were measured using instruments (Nanjing Mingao Instrument Equipment Co., Ltd.), and meat color and marbling pattern were scored using a pork color chart.
[0030] The effects of Lactobacillus amyloliquefaciens on the growth and slaughter performance of Bama pigs are shown in [the table below]. Figure 2 As shown, where, Figure 2 A, B, C, D, E, F, G, H, I, and J represent body weight, average daily weight gain, average daily feed intake, feed conversion ratio, ketone percentage, lean meat percentage, back fat percentage, subcutaneous fat percentage, body fat percentage, and backfat thickness, respectively. *Lactobacillus amyloliquefaciens* did not affect its growth performance. Figure 2 (AD in the middle), but significantly improved the lean meat weight / body weight ratio ( Figure 2 G in the middle), and significantly reduced the subcutaneous fat weight / body weight ratio ( Figure 1 (F in the text), subcutaneous fat weight / body weight ratio ( Figure 2 H), total fat weight / body weight ratio, and back fat thickness (in the text). Figure 2 (I and J in the text).
[0031] Meat quality determines consumers' choice of pork. Therefore, the impact of *Lactobacillus amyloliquefaciens* on the quality of Bama pork was further investigated and evaluated. The results are shown in [Figure 1]. Figure 3 As shown, where, Figure 3 A, B, C, D, and E in the table represent meat color, meat pH, meat shear force, meat color score, and meat marbling score, respectively. The results showed that *Lactobacillus amyloliquefaciens* significantly reduced L* and b* values after 48 hours. Figure 3 (A in the sample), and the pH value decreased at 45 minutes, but the situation reversed after 48 hours. Figure 3 (B in the text). *Lactobacillus amyloliquefaciens* significantly reduced the shear force in pork and improved the meat's color score (…). Figure 3 (CD in the data). These data indicate that Lactobacillus amyloliquefaciens improves the meat quality of Bama pigs.
[0032] 2. Serum biochemical index analysis Blood samples were collected from each group of Bama pigs. After standing for 40 minutes, the blood was centrifuged at 1500 rpm and 4°C for 10 minutes, and the supernatant was collected. The levels of blood urea nitrogen (BUN), creatinine (CRE), total protein (TP), albumin (ALB), globulin (GLB), alanine aminotransferase (ALT), aspartate aminotransferase (AST), interleukin-10 (IL-10), and interleukin-22 (IL-22) were measured. The kits used were purchased from Nanjing Jiancheng Biotechnology Institute.
[0033] Serum biochemical parameters reflect the metabolic state of animals and the absorption and deposition of nutrients. Serum biochemical parameters of each group of samples were measured, and the results are shown below. Figure 4 As shown, where, Figure 4 A, B, C, D, E, F, G, H, I, and J represent the serum urea nitrogen, creatinine, total protein, albumin, globulin, albumin / globulin ratio, alanine aminotransferase, aspartate aminotransferase, interleukin-10, and interleukin-22 levels, respectively. It can be seen that *Lactobacillus amyloliquefaciens* significantly increased serum BUN, CRE, ALB, and the A / G ratio. Figure 4 (A, B, D, F), while GLB, AST, IL-10 and IL-22 were significantly reduced ( Figure 4 (E, H, I, J in the text).
[0034] 3. Gene expression assay Gene expression quantification was performed using real-time PCR. Jejunal and ileal mucosal samples were frozen and ground in liquid nitrogen, and total RNA was extracted using TRIzol reagent (Genesta GmbH, Germany), followed by treatment with DNase I (Genesta GmbH, Germany). Reverse transcription was performed at 37°C for 15 minutes, followed by 95°C for 5 seconds. Primers used in this study were designed based on porcine sequences (see Table 2) for PCR amplification.
[0035] Table 2 Primers used for RT-PCR gene expression analysis in, SGLT1 = Sodium / Glucose Cotransporter 1; GLUT2 = Glucose Transporter2; GLUT5 = Glucose Transporter 5; B 0 AT1= Sodium-dependent Neutral Amino AcidTransporter 1; PEPT1=Peptide Transporter 1; SNAT2= Sodium-coupled Neutral AminoAcid Transporter 2; FATP4= Fatty Acid Transport Protein 4.
[0036] The small intestine is the primary site of nutrient absorption. The absorption of carbohydrates, proteins, and fats by the intestines affects the body's metabolism and the deposition of nutrients in meat. The absorption of nutrients by the intestines depends on the expression of carrier proteins, such as SGLT1, GLUT2, GLUT5, and B. 0 AT1, PEPT1, SNAT2, and FATP4. The results of the determination of each gene index in the jejunum are shown below. Figure 5 As shown, the results of the determination of various gene indicators in the ileum are as follows: Figure 6 As shown, where, Figure 5 , 6 A, B, C, D, E, F, and G represent the expression levels of different genes. It can be seen that *Lactobacillus amyloliquefaciens* significantly reduced the expression of SGLT1, GLUT2, GLUT5, and B genes in the jejunum. 0 Gene expression of AT1, SNAT2, and FATP4, as well as SGLT1, GLUT2, GLUT5, and B in the ileum. 0 Gene expression of AT1, PEPT1, and FATP4 was observed. However, *Lactobacillus amyloliquefaciens* increased PEPT1 gene expression in the jejunum, suggesting that *Lactobacillus amyloliquefaciens* reduces fat deposition by decreasing the expression of genes related to nutrient absorption in the gut.
[0037] In summary, the *Lactobacillus amyloliquefaciens* strain screened in this invention, as a feed additive, can effectively reduce the problem of excessive fat accumulation in Bama pig farming, reduce body fat percentage, and increase lean meat percentage, showing potential application prospects in the field of Bama pig breeding.
[0038] Although the present invention has been described in detail through the preferred embodiments above, it should be understood that the above description should not be considered as a limitation of the present invention. Various modifications and substitutions to the present invention will be apparent to those skilled in the art after reading the above description. Therefore, the scope of protection of the present invention should be defined by the appended claims.
Claims
1. A strain of Lactobacillus amyloliquefaciens that improves the lean meat percentage of Bama pigs ( Lactobacillus amylovorus WANRU1108, characterized in that, This strain was deposited on December 16, 2024, at the China Center for Type Culture Collection (CCTCC), Wuhan University, Wuhan, China, with accession number CCTCC NO: M20242823.
2. The application of Lactobacillus amyloliquefaciens WANRU1108 as described in claim 1 in the preparation of feed additives to improve the lean meat percentage of Bama pigs.
3. A feed additive for improving the lean meat percentage of Bama pigs, characterized in that, It contains Lactobacillus amyloliquefaciens WANRU1108 as described in claim 1.
4. The feed additive according to claim 3, characterized in that, The viable bacterial concentration of the *Lactobacillus amyloliquefaciens* WANRU1108 was 4.4 × 10⁻⁶. 9 CFU / mL.
5. The application of Lactobacillus amyloliquefaciens WANRU1108 as described in claim 1 or the feed additive as described in any one of claims 3-4 in the field of Bama pig farming.
6. The application according to claim 5, characterized in that, The application includes: Reduce fat deposition in Bama pigs; Increase the lean meat percentage of Bama pigs; Improve the quality of Bama pork.
7. A method for increasing the lean meat percentage of Bama pigs, characterized in that, The Lactobacillus amyloliquefaciens WANRU1108 described in claim 1 was added to the basal diet and fed to Bama pigs.
8. The method according to claim 7, characterized in that, The feeding method is as follows: feed once daily for the first week, then three times a week thereafter; each feeding amount is 4.4 × 10⁻⁶ live bacteria concentration. 9 50 mL of bacterial culture per head with a concentration of CFU / mL.
9. The method of obtaining according to any one of claims 7 or 8, characterized in that, The feeding cycle is no less than 10 weeks.