不同底物对液体发酵饲料细菌群落结构的影响

陈倩倩; 刘欣; 蓝江林; 刘波; 刘国红; 史怀; 阮传清

doi:10.19303/j.issn.1008-0384.2023.09.005

不同底物对液体发酵饲料细菌群落结构的影响

doi: 10.19303/j.issn.1008-0384.2023.09.005

福建省农业学院农业生物资源研究所，福建　福州　350002

基金项目: 福建省科技计划公益类专项（2022R1034006）；福建省高质量发展超越“5511”协同创新工程项目（XTCXGC2021019-ZYS02）；福建省级农业种质资源保护项目（ZYBHDWZX202207）；福建省农业科学院科技创新平台专项（CXPT202104）

详细信息

作者简介:
陈倩倩（1985 — ），女，博士，主要从事微生物及次生代谢相关研究，E-mail：chenqqkib@163.com

通讯作者:
蓝江林（1972 —），女，博士，研究员，主要从事农业微生物及应用技术研究，529770269@qq.com

中图分类号: S816
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出版历程
- 收稿日期: 2022-12-13
- 修回日期: 2023-07-06
- 网络出版日期: 2023-10-25
- 刊出日期: 2023-09-28

Microbial Community in Fermented Forage Affected by Ingredients Used

Institute of Agricultural Biological Resource, Fujian Academy of Agriculture Sciences, Fuzhou, Fujian　350002, China

摘要

摘要: 目的液体发酵饲料有益于动物肠道健康，具有良好的应用前景。分析不同底物对液体发酵豆粕的理化特性和细菌群落的影响，为发酵饲料的开发利用提供参考。方法采用正交试验分析红糖、鱼粉和豆粕及其组合，对液体发酵饲料品质的影响；采用16S rDNA测序，分析不同底物对细菌群落结构的影响。结果液体发酵饲料外观与原料浓度相关，即原料浓度越大，发酵饲料颜色越深。添加豆粕和鱼粉提高了发酵液体中的有机质和氮含量。PCA分析显示发酵前后样本中具有完全不同的细菌群落结构。经发酵，液体饲料中的细菌种类和数量都有所增加。发酵初期，变形菌门细菌占主导，其中的优势属为黄单胞菌（Xanthomonas）；发酵结束后，厚壁菌门占主导，其中乳杆菌（Lactobacillus）和魏斯氏菌（Weissella）等乳酸菌是优势属。发酵底物是影响液体发酵饲料质量的重要因素，红糖与假单胞菌属（Pseudomonas）、黄单胞菌属（Xanthomonas）、魏斯氏菌属（Weissella）、片球菌属（Pediococcus）和乳杆菌（Lactobacillales）含量呈正相关。鱼粉与乳杆菌属含量呈负相关，与嗜冷杆菌属（Psychrobacter）、魏斯氏菌属和假单胞菌属呈正相关。豆粕含量与醋酸杆菌属（Acetobacter）、乳杆菌属、嗜冷杆菌属、魏斯氏菌属和片球菌属（Pediococcus）呈正相关。结论添加较高水平的红糖和豆粕、低水平的鱼粉能够提高液体发酵饲料中乳杆菌和魏斯氏菌等益生菌的丰度。
- 发酵底物 /
- 细菌群落 /
- 液体发酵饲料 /
- 高通量测序 /
- 相关性分析
Abstract: Objective Effects of major ingredients used in formulating fermented livestock forage on probiotics in animals fed with it were analyzed. Method Brown sugar, fish meal, soybean meal, and their combinations were used to formulate liquid culture media in an orthogonal experiment. Structure and diversity of the microbial community in the fermentation broths were determined using the high throughput 16S rDNA sequencing technique. Result The fermentation broth became darker as the concentration increased. The addition of soybean or fish meal raised the contents of organic matters and nitrogen, and the fermentation significantly altered the microbial community with a much-enriched diversity and abundance of the broth. At the beginning of the fermentation, Proteobacteria was the predominant phylum with Xanthomonas being the outstanding genus. In the end, Firmicutes with genera Lactobacillus and Weissella became dominant. The microbial community was significantly affected by the ingredients included in the fermentation medium. For instance, brown sugar enhanced the proliferation of Pseudomonas,Xanthomonas, Weissella, Pediococcus, and Lactobacillales, fish meal facilitated that of Psychrobacter, Weissella, and Pseudomonas but inhibited Lactobacillales, while soybean meal promoted the growth of Acetobacter, Lactobacillales, Psychrobacter, Weissella, and Pediococcus. Conclusion It appeared that a culture medium high on brown sugar and soybean meal but low on fish meal could significantly enhance the abundance of Lactobacillus and Weissella in the fermentation broth to be use as a forage for feeding the livestock.
- Forage ingredients /
- microbial community /
- fermented liquid forage /
- high throughput sequencing /
- correlation analysis

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图 1 液体发酵饲料外观

Figure 1. Appearance of fermentation broth

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图 2 液体发酵饲料pH（A）、有机质和总氮含量（B）变化

Figure 2. Changes on pH (A) and organic matter and nitrogen contents (B) in fermentation broth

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图 3 液体发酵前（A）后（B）物种venn图

Figure 3. Venn diagram denoting difference of OTUs before (A) and after (B) fermentation

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图 4 液体发酵饲料细菌组成

Figure 4. Relative abundance of predominant microbes in fermentation broth

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图 5 液体发酵过程细菌群落变化

Figure 5. Changes on predominant microbes in fermentation broth

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图 6 基于OTUs分析的液体发酵样本PCA图

Figure 6. PCA plot of different fermentation broths basing on OTUs

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图 7 发酵底物与优势菌属间的相关性热图

Figure 7. Heatmap of correlation between ingredients used and dominant microbes in fermentation broth

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表 1 红糖、鱼粉和豆粕添加量的正交设计

Table 1. Orthogonal experiment on forage formulation with brown sugar, fish meal, and soybean meal

因素 Elements	红糖 Brown sugar/%	鱼粉 Fish meal/%	豆粕 Soybean meal/%	质量分数 Mass fraction/%
处理1 a a a Treatment 1	1	2	2	5
处理2 a b b Treatment2	1	4	4	9
处理3 a c c Treatment 3	1	16	8	25
处理4 b a b Treatment 4	2	2	4	8
处理5 b b c Treatment 5	2	4	8	14
处理6 b c a Treatment 6	2	16	2	20
处理7 c a b Treatment 7	4	2	4	10
处理8 c b a Treatment 8	4	4	2	10
处理9 c c c Treatment 9	4	16	8	28

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表 2 液体发酵饲料 Aplha 多样性统计

Table 2. α-diversity of microbial community in fermentation broth

样本 Sampels	Chao指数 Chao index	Sobs指数 Sobs index	香农指数 Shannon index	辛普森指数 Simpson index	覆盖率 Coverage/%
S1	83.1	60	1.318	0.330	99.93
S1_7	96.2	69	2.049	0.295	99.96
S2	98.0	52	0.969	0.520	99.94
S2_7	123.8	118	2.522	0.152	99.97
S3	109.0	63	1.082	0.398	99.95
S3_7	141.9	105	2.457	0.128	99.90
S4	85.9	74	0.650	0.653	99.96
S4_7	163.9	127	2.447	0.182	99.92
S5	100.2	75	0.786	0.619	99.94
S5_7	113.0	78	1.872	0.268	99.95
S6	83.0	48	0.895	0.502	99.96
S6_7	89.4	80	1.791	0.347	99.98
S7	97.5	60	0.782	0.505	99.96
S7_7	84.1	78	1.169	0.519	99.97
S8	63.0	44	0.368	0.852	99.96
S8_7	69.2	62	1.411	0.441	99.98
S9	68.0	51	1.156	0.397	99.97
S09_7	107.0	77	2.023	0.231	99.97

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