Sugarcane tip ensiling method and application thereof

Abstract

The invention discloses a method for ensiling sugarcane tips and application thereof, and belongs to the technical field of preparation of silage. The method comprises the following steps: (1) treating sugarcane tips: removing impurities from fresh sugarcane tips, cleaning, airing, and cutting the sugarcane tips into 2-4cm long for later use; and (2) mixing the treated sugarcane tips with compound lactic acid bacteria and an enzyme preparation, and ensiling for 40-50 days to obtain the ensiled sugarcane tips. The silage sugarcane tips prepared by the method are applied to preparation of buffalo weight increasing feed. According to the scheme provided by the invention, the nutrient components of the silage sugarcane tips can be improved, the abundance of microorganisms in the intestinal tracts of the buffalo can be promoted, and the diversity of the microorganisms in the intestinal tracts of the fed buffalo can be effectively improved. The research finds that a feeding application popularization experiment is further designed, and finds that the growth and weight gain of the buffalo can be promoted and the death rate of the buffalo can be effectively reduced when the prepared silage sugarcane tip compound auxiliary material is used for feeding the buffalo, so that the economic benefit of buffalo breeding is improved.

Description

Technical Field

[0001] The invention belongs to the technical field of preparing silage, and relates to a method for silaging sugarcane tips and application thereof. Background Art

[0002] my country is rich in buffalo (Bubalus bubalis) resources. According to the 2020 statistics of the Food and Agriculture Organization of the United Nations (FAO), the number of buffaloes in the world is about 205.205 million, and China has 27.431 million buffaloes, accounting for about 13.37% of the total number of buffaloes in the world. Buffaloes are known for their tolerance to roughage, strong adaptability, resistance to high temperature and humidity, strong disease resistance, tolerance to roughage, and easy breeding. Buffalo milk is thick and of excellent quality, and is the best of milk, known as the "king of milk". Guangxi Zhuang Autonomous Region is the main production area of ​​buffaloes. With the development of land transfer and agricultural mechanization, buffalo breeding is changing from the past use of draught to dairy and milk and meat, becoming an effective way to alleviate the contradiction between supply and demand in the beef and milk markets in southern my country.

[0003] Sugarcane is a tropical or subtropical crop, and its growth conditions have special requirements for temperature, light, and water. Guangxi Zhuang Autonomous Region has a subtropical climate and is the main sugarcane producing area. In the past 10 years, the sugarcane planting area in Guangxi Zhuang Autonomous Region has been around 1 million hm2. 2 The sugarcane output is more than 70 million tons, accounting for 60% of the country (Wu Duoguang et al., 2017). Sugarcane tips are the 2 to 3 tender nodes on the top of the sugarcane and the entire leaves attached to them after the sugarcane is harvested, accounting for about 20% of the total weight of sugarcane (Zhou Bo et al., 2019). Guangxi Zhuang Autonomous Region is a major sugarcane planting province in the country. The output of sugarcane tail leaves is about 15 million tons, and the feed utilization rate is about 20% (Chen Siye, 2020). Local superior feed resources are seriously wasted. Sugarcane tips have a relatively comprehensive nutritional value and can be directly fed to ruminants, but fresh sugarcane tips are prone to dehydration and mildew during storage. Silage can extend its storage time, improve the utilization efficiency of sugarcane tail leaves, improve feed palatability, and improve animal production performance (Zhou Xiong et al., 2015). At present, the utilization of sugarcane tops in my country is mainly through silage by adding microbial additives after appropriate treatment. This is one of the most effective ways to solve the waste of sugarcane top resources in my country and ensure the long-term supply of livestock forage (Mu Shenglong et al., 2017).

[0004] Lactic acid bacteria are commonly used microbial additives for silage. The lactic acid they produce can effectively lower the pH of silage and inhibit the growth of harmful bacteria.

[0005] Enzyme preparations are a very important type of biocatalyst, which are proteins or RNAs produced by living cells and have high specificity and high catalytic efficiency for their substrates. Adding enzyme preparations to silage can improve the nutritional value and fermentation quality of silage. Adding enzyme preparations helps to decompose plant cell walls, increase the content of soluble carbohydrates, provide sufficient substrates for the reproduction and fermentation of lactic acid bacteria, and quickly reduce pH, thereby improving the fermentation quality of silage.

[0006] There are many reports on the research of lactic acid bacteria and enzyme preparations on some other silages, and the silage quality has been improved to a certain extent, but there are few reports on sugarcane top silage. Sugarcane tops are a very rich feed resource in southern my country. Guangxi Zhuang Autonomous Region is the main production area of ​​sugarcane and buffalo. The feed consumption is large, and the requirements for the quantity and quality of forage are also increasing. If we can make full use of local agricultural and sideline product resources while studying high-quality sugarcane top silage processing methods, it will be an effective way to improve the economic benefits of buffalo breeding. Summary of the invention

[0007] The purpose of the present invention is to provide a method and application of sugarcane top silage in view of the above problems. The present invention finds that the use of a specific composite lactic acid bacteria and enzyme preparation combination to ensilage fresh sugarcane tops can increase the content of acetic acid and total volatile fatty acids in in vitro fermentation of ensilage, and reduce the content of ammonia nitrogen; increase the abundance of Ruminobacter microorganisms that are significantly positively correlated with the acetic acid content, and reduce the abundance of Rikenellaceae_Rc9_gut_group microorganisms that are positively correlated with NH3-N. Through the scheme experiment of this application, the nutrient composition of ensiled sugarcane tops can be improved, the abundance of microorganisms in the intestinal tract of buffaloes can be promoted, and the diversity of microorganisms in the intestinal tract of fed buffaloes can be effectively increased. Through this research discovery, a feeding application promotion experiment was further designed, and it was found that feeding buffaloes with the prepared ensiled sugarcane top compound auxiliary material of the present application can promote the growth and weight gain of buffaloes and effectively reduce the mortality rate of buffaloes, thereby improving the economic benefits of buffalo breeding.

[0008] In order to achieve the above object, the technical solution adopted by the present invention is:

[0009] One of the purposes is to provide a method for silage of sugarcane tips, comprising the following steps:

[0010] (1) Sugarcane tip processing: remove impurities from fresh sugarcane tips, wash, dry, cut into 2-4 cm lengths, and set aside;

[0011] (2) mixing the treated sugarcane tops with the composite lactic acid bacteria and the enzyme preparation, and silaging for 40-50 days to obtain silage sugarcane tops;

[0012] The composite lactic acid bacteria consists of Lactobacillus plantarum and Pediococcus pentosaceus;

[0013] The enzyme preparation consists of cellulase, xylanase and glucose oxidase.

[0014] It is further described that the enzyme activity of the cellulase is 50,000 U / g, the enzyme activity of the xylanase is 50,000 U / g; and the enzyme activity of the glucose oxidase is 10,000 U / g.

[0015] Further description, the content of the plant lactobacillus is 1×10 5 -1×10 7 cfu / g; the content of Pediococcus pentosaceus is 1×10 5 -1×10 7 cfu / g; the amount of the cellulase is 1×10 4 -3×10 4 u / g; the dosage of the xylanase is 1×10 4 -3×10 4 u / g; the concentration of glucose oxidase added is 0.1-0.3% per kilogram of fresh weight of sugarcane shoots.

[0016] Further description, the content of the plant lactobacillus is 1×10 6 cfu / g; the content of Pediococcus pentosaceus is 1×10 6 cfu / g; the amount of the cellulase is 1×10 6 cfu / g; the dosage of the xylanase is 3×10 4 u / g; the concentration of glucose oxidase added is 0.3% per kilogram of fresh weight of sugarcane shoots.

[0017] It is further explained that the silage condition is light-proof anaerobic silage.

[0018] The present invention also provides an application of the silage sugarcane tops prepared by the method described above in preparing feed for weight gain of buffaloes.

[0019] It is further described that, in terms of weight percentage, the feed includes 15%-25% of silage sugarcane tips prepared by the above method, 40%-50% of corn stalk silage and 35% of auxiliary materials.

[0020] It is further specified that the auxiliary materials include, by weight percentage, 42 parts of corn flour, 25 parts of soybean meal, 15 parts of bran, 10 parts of rice bran, 5 parts of mineral premix, and 3 parts of vitamin premix.

[0021] Due to the adoption of the above technical solution, the present invention has the following beneficial effects:

[0022] The results obtained by the inventor through research are:

[0023] After the experimental study designed in this application, it was found that the concentration of plant lactobacillus added per kilogram of fresh weight of sugarcane tops in group T6 was 1×10 6 cfu / g, the added concentration of Pediococcus pentosaceus was 1×10 6 cfu / g, the added concentration of cellulase was 1×10 4 u / g, the added concentration of xylanase was 3×10 4 u / g, when the addition concentration of glucose oxidase is 0.3%), a higher crude protein content and lower neutral detergent fiber and acid detergent fiber can be obtained. Silage in vitro fermentation has a significant effect on total gas production, hydrogen production, pH, acetic acid content, total volatile fatty acids, and ammonia nitrogen content. The T6 group can increase the acetic acid and total volatile fatty acid content of silage in vitro fermentation and reduce the ammonia nitrogen content. The T6 group can increase the abundance of Ruminobacter microorganisms that are significantly positively correlated with acetic acid content, and reduce the abundance of Rikenellaceae_Rc9_gut_group microorganisms that are positively correlated with NH3-N. Based on the above results, a buffalo feeding experiment was designed, and it was found that feeding buffalo with the silage sugarcane tops composite auxiliary material prepared in this application can effectively improve the immunity of the buffalo, promote the growth and weight gain of the buffalo, and effectively reduce the incidence of buffalo, thereby improving the economic benefits of buffalo breeding. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 This is the OTU-venn diagram of microorganisms added to the in vitro fermentation of sugarcane top silage with different enzymes and bacteria mixed in the present invention.

[0025] Figure 2 This is the distribution diagram of rumen microbial species at the phylum level of in vitro fermentation of sugarcane top silage with different enzyme and bacteria mixtures added according to the present invention.

[0026] Figure 3 This is a distribution diagram of rumen microbial species at the genus level in the in vitro fermentation of sugarcane top silage with different enzyme and bacteria mixtures added according to the present invention.

[0027] Figure 4 Heatmap of the correlation between relative abundance of bacteria at the genus level and rumen in vitro fermentation parameters. DETAILED DESCRIPTION

[0028] The specific implementation of the invention is further described below with reference to the accompanying drawings.

[0029] The specific verification experiments are as follows:

[0030] 1. Effects of different levels of mixed addition of lactic acid bacteria and enzyme preparations on the fermentation quality of sugarcane top silage

[0031] 1.1 Test materials.

[0032] 1.1.1 Test materials

[0033] The experimental raw material sugarcane tops were taken from Fusui County, Chongzuo City, Guangxi Zhuang Autonomous Region, and the sugarcane tops were cut into lengths of about 1 to 2 cm using a grass cutter. The nutritional components of sugarcane tops are shown in Table 1.

[0034] Table 1 Nutritional composition of sugarcane shoots (absolute dry basis) %

[0035] raw material DM CP OM NDF ADF Sugarcane shoots 24.06±0.32 8.45±0.18 92.57±0.03 74.30±0.35 40.17±0.65

[0036] 1.1.2 Test additives

[0037] Lactobacillus plantarum (10 billion iu / g) was purchased from Shaanxi Baichuan Biotechnology Co., Ltd.; Pediococcus pentosaceus (10 billion cfu / g) was purchased from Xi'an Qiancao Biotechnology Co., Ltd.; cellulase (50,000 U / g) and xylanase (50,000 U / g) were both purchased from Beijing Solebow Technology Co., Ltd.; glucose oxidase (10,000 U / g) was purchased from Nanning Pangbo Bioengineering Co., Ltd.

[0038] 1.2 Silage preparation

[0039] The experiment used fresh sugarcane tops as raw materials and used L18 (3 5 ) Orthogonal experimental design was used to select the best results for the mixed addition of different levels of Lactobacillus plantarum, Pediococcus pentosaceus, cellulase, xylanase and glucose oxidase. The addition levels are shown in Table 2 and the experimental groups are shown in Table 3. The experiment used a small-scale fermentation method. The prepared lactic acid bacteria and enzyme preparation solution were evenly sprayed on the surface of the cut sugarcane tips and mixed evenly. Then, they were packed into polyethylene bags for vacuum packaging. Each bag was 500g and sealed using a vacuum packaging machine (DZ500-ZD, Guangzhou Rifu Packaging Machinery Co., Ltd.). Each group was repeated 5 times and stored at room temperature in the dark for 45 days. The silage nutrients and fermentation parameters were determined to analyze their effects on the silage quality of sugarcane tips.

[0040] Table 2 Sugarcane tip silage bacteria and enzyme addition concentration (kgFW)

[0041]

[0042] Table 3 Grouping of sugarcane shoot silage experiment

[0043]

[0044] 1.3 Silage sample collection and determination of nutritional composition and fermentation parameters

[0045] On the 45th day of the experiment, the silage was opened to determine the silage fermentation parameters, and the fermentation products were determined according to Cai's method. Take 20g of the sample from the middle of the packaging bag, put it in a 250mL wide-mouth bottle, add 180mL of distilled water, seal it and place it in a refrigerator at 4℃ for 24h, then filter it through two layers of gauze, collect the filtrate and test the pH, organic acids (lactic acid (LA), acetic acid (AA), propionic acid (PA) and butyric acid (BA)) and ammonia nitrogen (NH 3 -N) content. The pH was measured using a portable pH meter (PH8180-0-00, Dongguan Wanchuang Electronic Products Co., Ltd.). The organic acid content was measured using an Agilent 1260 high performance liquid chromatograph according to the method of Xie. The analysis conditions were as follows: chromatographic column ShodexRSpakKC-811, DAD detector SPD-M10AVP, eluent 3mmol / l perchloric acid, column temperature 50℃, flow rate 1mL / min, detection wavelength 210nm, injection volume 5μL. NH 3 -N was determined by phenol-sodium hypochlorite colorimetry. Take 200 grams of sample and dry it at 65℃, grind it through a 40-mesh sieve, and test dry matter (DM), crude ash (Ash), crude protein (CP), neutral detergent fiber (NDF), and acid detergent fiber (ADF) according to Zhang Liying's method.

[0046] 1.4 Data Statistical Analysis

[0047] The data was initially sorted by Excel, and the range analysis method and comprehensive balance method were used to perform orthogonal analysis on the total score of silage fermentation quality. Ki is the average value of the level of factor i (i = 1, 2, 3). The larger the Ki, the better the level. The range (R) is the difference between the maximum and minimum values ​​of the factor Ki. The larger the range, the greater the impact of the factor on the silage quality. The comprehensive balance analysis method was used to analyze the results and select the optimal combination. The fuzzy mathematics membership function method was used to calculate the membership function value of each treatment single indicator, and then the average membership function value of each treatment was calculated to comprehensively evaluate the silage quality, and the 6 better combinations were selected according to the size.

[0048] 1.5 Results and Analysis

[0049] 1.5.1 Effects of different bacterial enzyme addition levels on the nutritional components of sugarcane top silage

[0050] It can be seen from Table 4 that the factors affecting the DM content of silage are A>C>D>B>E, and the optimal combination of each factor is A3C1D1B2E2. It can be seen from Table 5 that the factors affecting the CP content of silage are D>E>A>C>B, and the optimal combination of each factor is D3E3A2C1B2. It can be seen from Table 6 that the factors affecting the OM content of silage are C>B>E>D>A, and the optimal combination of each factor is C3B3E3D1A1. It can be seen from Table 7 that the factors affecting the NDF content of silage are D>A>E>C>B, and the optimal combination of each factor is D3A2E2C1B2. It can be seen from Table 8 that the factors affecting the ADF content of silage are D>E>A>B>C, and the optimal combination of each factor is D3E3A2B2C1.

[0051] Table 4 Effects of different bacterial enzyme addition levels on DM content in sugarcane shoot silage

[0052]

[0053]

[0054] Table 5 Effects of different bacterial enzyme addition levels on CP content of sugarcane tip silage

[0055]

[0056]

[0057] Table 6 Effects of different bacterial enzyme addition levels on OM content in sugarcane shoot silage

[0058]

[0059]

[0060] Table 7 Effects of different bacterial enzyme addition levels on NDF content of sugarcane shoot silage

[0061]

[0062] Table 8 Effects of different bacterial enzyme addition levels on ADF content of sugarcane tip silage

[0063]

[0064] 1.5.2 Effects of different bacterial enzyme addition levels on the fermentation quality of sugarcane top silage

[0065] As shown in Table 9, the factors affecting pH content are B>A>D>C>E, and the optimal combination of each factor is B1A1D3C3E3. As shown in Table 10, the factors affecting acetic acid content in silage are A>E>C>D>B, and the optimal combination of each factor is A2E2C3D3B1. As shown in Table 11, the factors affecting ammonia nitrogen content are C>B>E>A>D, and the optimal combination of each factor is C2B3E1A2D2. As shown in Table 12, the factors affecting lactic acid content are B>C>D>A>E, and the optimal combination of each factor is B3C2D1A2E1.

[0066] Table 9 Effects of different bacterial enzyme addition levels on pH content of sugarcane tip silage

[0067]

[0068] Table 10 Effects of different bacterial enzyme addition levels on AA content in sugarcane tip silage

[0069]

[0070]

[0071] Table 11 Effects of different bacterial enzyme addition levels on NH3-N content in sugarcane shoot silage

[0072]

[0073]

[0074] Table 12 Effects of different bacterial enzyme addition levels on LA content in sugarcane tip silage

[0075]

[0076]

[0077] 1.5.3 Comprehensive scoring analysis of silage quality

[0078] The results of the comprehensive balance analysis in Table 13 show that the optimal combination is A2B2C1D3E3, that is, the optimal concentration of plant lactobacillus is 1×10 6 cfu / g, Pediococcus pentosaceus is 1×10 6 cfu / g, cellulase is 1×10 4 u / g, xylanase is 3×10 4 u / g, glucose oxidase is 0.3%.

[0079] Table 13 Analysis results of orthogonal test comprehensive balance method

[0080]

[0081] 1.5.4 Comprehensive scoring of silage quality analysis using fuzzy mathematical average membership function method

[0082] Table 14 Membership function analysis and comprehensive value ranking of sugarcane shoot silage at different bacterial enzyme addition levels

[0083]

[0084]

[0085] The fuzzy mathematical average membership function method was used to take DM, CP, OM, acetic acid, and lactic acid as positive correlation indicators, and NDF, ADF, pH, and NH3-N as negative correlation indicators. As shown in Table 14, the six groups with the highest to lowest comprehensive scores are groups 17, 16, 12, 2, and 3, and the silage quality of these five groups is better.

[0086] 1.6 Summary

[0087] When the concentration of Lactobacillus plantarum added per kg of fresh sugarcane shoots was 1×10 6 cfu / g, the added concentration of Pediococcus pentosaceus was 1×10 6 cfu / g, the added concentration of cellulase was 1×10 4 u / g, the added concentration of xylanase was 3×10 4 u / g, when the addition concentration of glucose oxidase was 0.3%, the silage quality was the best. The five better groups in this test were group 17 (A3B2C1D3E1), group 16 (A3B1C3D2E3), group 12 (A1B3C2D2E1), group 2 (A1B2C2D2E2), and group 3 (A1B3C3D3E3).

[0088] 2 Effects of mixed addition of lactic acid bacteria and enzyme preparations on in vitro fermentation of sugarcane top silage

[0089] 2.1 Test materials

[0090] 2.1.1 In vitro fermentation substrates

[0091] The first step of orthogonal test screened out 6 better sugarcane top silages, which were group 17, group 16, group 12, group 2, group 3 and the best group corresponding to the following substrates of group T1, group T2, group T3, group T4, group T5 and group T6.

[0092] 2.1.2 Rumen fluid is supplied to animals.

[0093] Buffalo with permanent rumen fistula.

[0094] 3.2.2 Test methods

[0095] 2.2.1 The preparation method of in vitro fermented artificial saliva refers to the method of Gan Jie (2007).

[0096] 2.2.2 Preparation of mixed culture medium and pretreatment before culture

[0097] According to the method of Menke et al. (1979), the rumen fluid collected before morning feeding was filtered through 100 mesh four-layer gauze and mixed with anaerobic buffer at a ratio of 1:2. In this study, 10 ml of rumen fluid was mixed with 20 ml of buffer to form the culture medium. The mixed culture medium was quickly purged with CO before fermentation. 2 After at least 2-3 minutes, place it in a shaker at 39°C in a water bath and wait for incubation.

[0098] 2.2.3 Substrate preparation and recording of gas production and methane production

[0099] One day before in vitro culture, 1 g of feed sample was accurately weighed into each fermentation bottle in advance, and 6 replicates were set for each sample, and 3 blanks (i.e., only fermentation medium was injected) were set for each batch of culture. After the start of the experiment, the cumulative gas production was recorded at 3, 6, 9, 12, 24, 36, 48, and 72 hours respectively. The gas production sampling syringe was used to directly measure the gas production, and then the gas chromatograph was used to detect the hydrogen and methane production.

[0100] 2.2.4 Sample processing and analysis methods

[0101] After the fermentation, the fermentation liquid was filtered through 100 mesh nylon cloth, and the pH, VFA, NH3-N, MCP, and microbial flora of the culture liquid were tested. After filtration, the residue was transferred to a glass pot without damage and repeatedly rinsed with distilled water, and dried at 105℃ to determine the dry matter disappearance rate.

[0102] 2.3 Data processing The data were preliminarily sorted using Excel and SPSS23.0 was used for one-way ANOVA analysis of variance. After significant differences were found, Duncan's method was used for multiple comparisons. The results were expressed as "mean ± standard deviation" and P < 0.05 indicated a significant difference.

[0103] 2.4 Results and Analysis

[0104] 2.4.1 Effects of different enzyme and bacteria mixtures on the nutritional components of sugarcane tip silage

[0105] As shown in Table 15, the DM content of group IV was significantly higher than that of other groups (P<0.05), the CP content of groups Ⅰ and Ⅱ was significantly higher than that of other groups (P<0.05), the Ash content of groups Ⅰ, Ⅱ and Ⅳ was significantly higher than that of other groups (P<0.05), and there was no significant difference in NDF and ADF contents among the groups (P>0.05).

[0106] Table 15 Effects of different enzyme and bacteria mixed addition on the nutritional components of sugarcane tip silage (absolute dry basis) (%)

[0107]

[0108] Note: Data in the same row with different letters in the shoulder indicate significant differences (P>0.05), while data with the same letters or no shoulder indicate no significant differences (P>0.05).

[0109] 2.4.2 Effects of different enzyme and bacteria mixtures on sugarcane top silage fermentation parameters

[0110] As shown in Table 16, the pH of T4, T5 and T6 groups was significantly lower than that of other groups (P<0.05), the pH of T6 group was the lowest, the lactic acid content of T6 and T3 groups was significantly higher than that of other groups (P<0.05), the acetic acid content was not significantly different among the groups (P>0.05), the ammonia nitrogen content of T6 group was the lowest, and the MCP content was not significantly different among the groups (P>0.05).

[0111] Table 16 Effects of adding cellulase and xylanase on fermentation parameters of sugarcane top silage 2.4.3 Effects of adding different enzymes and bacteria on gas production and fermentation parameters of sugarcane top silage in vitro

[0112] As shown in Table 17, there was no significant difference in dry matter digestibility among the groups (P>0.05). The total gas production in group T3 was the highest, which was significantly higher than other groups (P<0.05) except that there was no significant difference with group T4 (P>0.05). The total hydrogen production in group T3 was the highest, which was significantly higher than other groups (P<0.05) except that there was no significant difference with group T4 (P>0.05). There was no significant difference in total methane production among the groups (P>0.05). The pH value was the highest in group T1 and the lowest in group T4. The pH of all groups was at normal levels. The total VFA and acetic acid contents in groups T5 and T6 were significantly higher than those in other groups (P<0.05). There was no significant difference in propionic acid and butyric acid contents among the groups (P>0.05). The ammonia nitrogen content in group T6 was the lowest, which was significantly lower than other groups (P<0.05) except that there was no significant difference with groups T3 and T4 (P>0.05).

[0113] Table 17 Effects of different enzyme-bacteria mixtures on gas production and fermentation parameters of sugarcane tip silage in vitro 2.4.4 Changes in microbial structure of sugarcane tip silage in vitro fermentation with different enzyme-bacteria mixtures

[0114]

[0115] 2.4.4.1 Analysis of α-diversity index of in vitro fermentation bacteria

[0116] As shown in Table 18, the coverage index of this sequencing analysis is greater than 98%, indicating that the sequencing results can reflect the true situation of the sample microorganisms. The Sobs index, Chao1 index, Ace index, Shannon index and Simpson index of each group were not significantly different (P>0.05).

[0117] Table 18 Rumen fluid bacterial alpha diversity index

[0118]

[0119] 2.4.4.2 OTU-Venn diagram of in vitro fermentation samples

[0120] Depend on Figure 1 It can be seen that there are 2177 OTUs in the fermentation broth of the six groups. The number of OTUs in groups T1, T2, T3, T4, T5, and T6 are 2908, 2744, 3036, 2632, 2952, and 3151, respectively. The three groups share 2177 OTUs, accounting for 33.29% of the total number of OTUs. The number of unique OTUs in each group is 731, 567, 859, 455, 775, and 975, respectively, accounting for 11.17%, 8.67%, 13.13%, 6.96%, 11.85%, and 14.91% of the total number of OTUs, respectively.

[0121] 2.4.4.3 Species distribution of rumen fluid bacteria in each group at the phylum level

[0122] Depend on Figure 2 As shown in Table 19, at the phylum level, the rumen fluid flora in this experiment mainly consisted of the following five phyla: Firmicutes, Bacteroidota, Verrucomicrobiota, Spirochaetota, and Proteobacteria. The dominant flora in each group was Firmicutes and Bacteroidota. There were certain differences in the main microbial colonies at the phylum level. The Firmicutes in T4 group was significantly higher than those in T1, T2, T3 and T6 groups (P<0.05), except that there was no significant difference between T5 group and T4 group (P>0.05). The Bacteroidota in T1, T2 and T3 groups were significantly higher than those in T4, T5 and T6 groups (P<0.05). The Verrucomicrobiota in T5 group was significantly higher than those in T1, T2, T3 and T6 groups (P<0.05), except that there was no significant difference between T4 and T4 group (P>0.05). The Spirochaetota in T4 and T6 groups were significantly higher than those in T1, T2, T3 and T5 groups (P<0.05). The Proteobacteria in T4 group was significantly higher than that in other groups (P<0.05).

[0123] 2.4.4.4 Species distribution of rumen fluid bacteria in each group at the genus level

[0124] Depend on Figure 3 As shown in Table 19, at the genus level, the rumen microorganisms consisted of 17 genera, including Succiniclasticum, Rikenellaceae_Rc9_gut_group, Unclassified_f_F082, and Prevotella. The dominant genera in each group were Succiniclasticum and Rikenellaceae. At the genus level, there were certain differences in the microbial colonies except NK4A214_group. The relative abundance of norank_f_vadinBE97 in group T1 increased significantly (P<0.05), the relative abundance of Rikenellaceae_RC9_gut_group in group T2 increased significantly (P<0.05), the relative abundance of Prevotella in group T3 increased significantly (P<0.05), the relative abundance of Succiniclasticum in group T4 increased significantly (P<0.05), the relative abundance of norank_f_norank_o_WCHB1-41 in group T5 increased significantly (P<0.05), and the relative abundance of Ruminobacter in group T6 increased significantly (P<0.05).

[0125] Table 19 Effects of different proportions of sugarcane top silage on the relative abundance of rumen flora 2.4.4.5 Correlation analysis between relative abundance of bacteria at genus level and rumen fermentation parameters

[0126]

[0127] Figure 4The correlation between the relative abundance of rumen bacteria at the genus level (the top 15 genera in content) and fermentation parameters in buffalo was shown. Among them, dry matter digestibility, methane, and propionic acid were not significantly correlated with the content of each bacteria. Gas production was significantly negatively correlated with Rikenellaceae_Rc9_gut_group (P<0.05). Hydrogen was significantly positively correlated with norank_f_norank_o_Clostridia_UCG-014 and Ruminobacter (P<0.05), and significantly negatively correlated with Rikenellaceae_Rc9_gut_group (P<0.05). pH was significantly positively correlated with Rikenellaceae_Rc9_gut_group (P<0.05), and significantly negatively correlated with norank_f_norank_o_Clostridia_UCG-014, Candidatus_Saccharimonas, and Ruminobacter (P<0.05). Acetic acid was significantly positively correlated with Ruminobacter (P<0.05). Butyric acid was significantly positively correlated with Saccharofermentans, NK4A214_group and Rikenellaceae_Rc9_gut_group (P<0.05). SaccharochaetaTVFA was significantly positively correlated with Candidatus_Saccharimonas and Ruminobacter (P<0.05). NH3-N was significantly positively correlated with Rikenellaceae_Rc9_gut_group (P<0.05), and significantly negatively correlated with Candidatus_Saccharimonas, Sphaerochaeta and prevotella (P<0.05).

[0128] Summary: Under the experimental conditions, the mixed addition of different concentrations of Lactobacillus plantarum, Pediococcus pentosaceus, cellulase, xylanase and glucose oxidase can have a significant effect on the dry matter, crude protein, ash content, neutral detergent fiber, pH, lactic acid and ammonia nitrogen of silage. The T6 group (A2B2C1D3E3) can obtain higher crude protein content and lower neutral detergent fiber and acid detergent fiber. The in vitro fermentation of silage has a significant effect on the total gas production, hydrogen production, pH, acetic acid content, total volatile fatty acids and ammonia nitrogen content. The T6 group (A2B2C1D3E3) can increase the acetic acid and total volatile fatty acid content of silage in vitro fermentation and reduce the ammonia nitrogen content. The T6 group (A2B2C1D3E3) can increase the abundance of Ruminobacter microorganisms that are significantly positively correlated with the acetic acid content and reduce the abundance of Rikenellaceae_Rc9_gut_group microorganisms that are positively correlated with NH3-N.

[0129] Application experiment:

[0130] In summary, the silage quality of group T6 (A2B2C1D3E3) was the best, that is, when the concentration of Lactobacillus plantarum added per kilogram of fresh weight of sugarcane tops was 1×10 6 cfu / g, the added concentration of Pediococcus pentosaceus was 1×10 6 cfu / g, the added concentration of cellulase was 1×10 4 u / g, the added concentration of xylanase was 3×10 4 u / g, and the addition concentration of glucose oxidase was 0.3%, the silage quality was the best. And through in vitro fermentation treatment, it was found that the T6 group (A2B2C1D3E3) could increase the content of acetic acid and total volatile fatty acids in in vitro fermentation of silage, reduce the content of ammonia nitrogen, increase the abundance of Ruminobacter microorganisms that were significantly positively correlated with the content of acetic acid, and reduce the abundance of Rikenellaceae_Rc9_gut_group microorganisms that were positively correlated with NH3-N.

[0131] Through this study, the applicant conducted a feeding experiment: according to the applicant's experience, the following feeding feed experimental group was designed:

[0132] Group 1: 10% sugarcane tops + 55% corn stalk silage + 35% auxiliary materials silaged according to the T6 group silage plan.

[0133] Group 2: 15% sugarcane tops + 50% corn stalk silage + 35% auxiliary materials silaged according to the silage plan of Group T6.

[0134] Group 3: 20% sugarcane tops + 45% corn stalk silage + 35% auxiliary materials silaged according to the silage plan of Group T6.

[0135] Group 4: 25% sugarcane tops + 40% corn stalk silage + 35% auxiliary materials silaged according to the silage plan of Group T6.

[0136] Group 5: 30% sugarcane tops + 35% corn stalk silage + 35% auxiliary materials silaged according to the silage plan of Group T6.

[0137] Group 6: corn stalk silage 65% + auxiliary materials 35%.

[0138] The auxiliary materials of this experiment are composed of 42 parts of corn flour, 25 parts of soybean meal, 15 parts of bran, 10 parts of rice bran, 5 parts of mineral premix and 3 parts of vitamin premix according to the weight ratio.

[0139] The corn stalk silage used in this experiment was purchased from the original product sold by Hengxian Sitong Agriculture and Animal Husbandry Co., Ltd. in Guangxi Zhuang Autonomous Region.

[0140] 60 16-month-old buffaloes of similar weight were selected and randomly divided into 6 groups, with 10 buffaloes in each group. Each group was kept in a single-pen buffalo pen of the same size and environment, and the routine feeding and management procedures of the buffalo farm were followed. Free access to water and food, and conventional management and breeding were carried out for 70 days.

[0141] Record ① Average daily weight gain: weigh the buffaloes in each pen on an empty stomach at 9:00 a.m. on the first day of the experiment and the last day of the experiment, and calculate the average daily weight gain based on the initial and final weights and the number of days of the experiment.

[0142] ② The incidence of intestinal diseases in buffaloes was observed throughout the experimental period. The period from the first onset to recovery was recorded as 1 time. If the disease recurred after recovery, it was continued to be recorded as 2 times. Regardless of whether the disease occurred in the same buffalo, the final incidence rate was calculated as follows: The incidence rate calculation formula is: Incidence rate (%) = number of onsets × 100 / total number of buffaloes at the beginning of the period.

[0143] The specific record results are shown in Table 20.

[0144] Table 20 Buffalo records

[0145]

[0146] From the data in the above table, it can be seen that the weight gain effect is ranked as 4 groups>3 groups>2 groups>5 groups>1 groups>6 groups. From the perspective of morbidity, feeding buffaloes with the silage sugarcane top compound auxiliary material prepared in the present application can effectively reduce the morbidity. It can be seen that the feeding scheme of the silage sugarcane top compound auxiliary material prepared in the present application can effectively improve the immunity of cattle, promote the growth and weight gain of buffaloes, and effectively reduce the morbidity of buffaloes, thereby improving the economic benefits of buffalo breeding.

[0147] Although the present invention has been disclosed as above with preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art may make some modifications and improvements without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention shall be based on the definition of the claims.

Claims

1. A method for silage of sugarcane tips, characterized in that: The following steps are involved: (1) Sugarcane tip processing: remove impurities from fresh sugarcane tips, wash, dry, cut into 2-4 cm lengths, and set aside; (2) mixing the treated sugarcane tops with the composite lactic acid bacteria and the enzyme preparation, and silaging for 40-50 days to obtain silage sugarcane tops; The composite lactic acid bacteria consists of Lactobacillus plantarum and Pediococcus pentosaceus; The enzyme preparation consists of cellulase, xylanase and glucose oxidase.

2. The method for silaging sugarcane tips according to claim 1, characterized in that: The enzyme activity of the cellulase is 50,000 U / g, the enzyme activity of the xylanase is 50,000 U / g; and the enzyme activity of the glucose oxidase is 10,000 U / g.

3. The method for silaging sugarcane tips according to claim 1, characterized in that: The content of the plant lactobacillus is 1×10 5 -1×10 7 cfu / g; the content of Pediococcus pentosaceus is 1×10 5 -1×10 7 cfu / g; the amount of the cellulase is 1×10 4 -3×10 4 u / g; the dosage of the xylanase is 1×10 4 -3×10 4 u / g; the concentration of glucose oxidase added is 0.1-0.3% per kilogram of fresh weight of sugarcane shoots.

4. The method for silaging sugarcane tips according to claim 1, characterized in that: The content of the plant lactobacillus is 1×10 6 cfu / g; the content of Pediococcus pentosaceus is 1×10 6 cfu / g; the amount of the cellulase is 1×10 6 cfu / g; the dosage of the xylanase is 3×10 4 u / g; the concentration of glucose oxidase added is 0.3% per kilogram of fresh weight of sugarcane shoots.

5. The method for silaging sugarcane tips according to claim 1, characterized in that: The silage condition is light-proof anaerobic silage.

6. Use of the silage sugarcane tops prepared according to any one of claims 1 to 5 in preparing feed for weight gain of buffaloes.

7. The use according to claim 6, characterized in that: Calculated by weight percentage, the feed comprises 15%-25% of silage sugarcane tops prepared according to any one of claims 1 to 5, 40%-50% of corn stalk silage and 35% of auxiliary materials.

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