基于转录组学探究日粮添加酵母β-葡聚糖对新城疫疫苗免疫鸡肠道免疫的影响

doi:10.11843/j.issn.0366-6964.2025.03.042

畜牧兽医学报 ›› 2025, Vol. 56 ›› Issue (3): 1441-1452.doi: 10.11843/j.issn.0366-6964.2025.03.042

基于转录组学探究日粮添加酵母β-葡聚糖对新城疫疫苗免疫鸡肠道免疫的影响

李常营¹(), 李俊²^,³, 李锡锋³, 毕师诚³^,⁴, 曹立亭³^,⁴^,*()

1. 西南大学动物科学技术学院，重庆 402460
2. 宜宾市翠屏区动物防疫检疫中心，宜宾 644000
3. 西南大学动物医学院，重庆 402460
4. 西南大学中兽医药研究所，重庆 402460

收稿日期:2024-06-25 出版日期:2025-03-23 发布日期:2025-04-02
通讯作者: 曹立亭 E-mail:licy1983@163.com;caoliting@swu.edu.cn
作者简介:李常营(1983-)，男，山东滕州人，讲师，博士，主要从事动物营养与疾病防控研究，E-mail: licy1983@163.com，Tel：023-46751588
基金资助:
重庆市技术创新与应用发展专项资助项目(cstc2019jsccx-lyjsBX0003；cstc2021jscx-lyjsAX0008)

Effect of Dietary Yeast β-glucan Supplementation on Intestinal Immune Function in Chickens Immunized against Newcastle Disease Vaccine based on Transcriptomic

LI Changying¹(), LI Jun²^,³, LI Xifeng³, BI Shicheng³^,⁴, CAO Liting³^,⁴^,*()

1. College of Animal Science and Technology, Southwest University, Chongqing 402460, China
2. Cuiping Animal Epidemic Prevention and Quarantine Center of Yibin, Yibin 644000, China
3. College of Veterinary Medicine, Southwest University, Chongqing 402460, China
4. Institute of Traditional Chinese Veterinary Medicine, Southwest University, Chongqing 402460, China

Received:2024-06-25 Online:2025-03-23 Published:2025-04-02
Contact: CAO Liting E-mail:licy1983@163.com;caoliting@swu.edu.cn

摘要/Abstract

摘要：

旨在探讨基础日粮中添加酵母β-葡聚糖(G70)对新城疫疫苗免疫鸡肠道免疫功能的影响。试验选取16只1日龄健康乌骨鸡随机分为2个组，对照组(Vaccine)和G70组(G70+Vaccine)，G70组在饲喂基础日粮的基础上添加1 g·kg^-1的G70，分别于14、28日龄进行新城疫疫苗首次免疫和加强免疫。35日龄时，采集鸡的空肠组织，测定空肠黏膜IgA⁺细胞数量，空肠CD4⁺CD8⁺双阳性T细胞占比与空肠免疫相关基因mRNA表达水平，随后进行转录组学测序，并对差异表达基因进行GO功能和KEGG通路富集分析。结果表明，与对照组相比，饲料添加G70显著提高空肠黏膜内IgA⁺细胞的数量(P < 0.05)，空肠CD4⁺CD8⁺双阳性T细胞占比显著上升(P < 0.05)，GATA-3、MHC-Ⅰ、MHC-Ⅱ、CCR7和IFN-γ mRNA表达水平显著上调(P < 0.05)；转录组学研究表明，添加G70后空肠组织差异表达基因有559个，其中54个基因上调和505个基因下调，基因本体(GO)富集分析揭示差异表达基因多数被注释到胺代谢和分解、蛋白质合成、刺激生长因子(TGF-β)和调节酶活性等相关的GO条目上；KEGG信号通路分析显示差异表达基因主要富集在与核糖体、MAPK、细胞黏附和局部黏附等与细胞合成和代谢相关的信号通路。上述结果提示，饲粮中添加酵母β-葡聚糖(G70)可增加新城疫免疫鸡空肠中IgA⁺细胞的数量，提高空肠CD4⁺CD8⁺双阳性T细胞占比，上调空肠中免疫相关基因的表达，其机制可能是通过调节IL-17RD、TGF-β和TMEM158等免疫相关基因的表达，影响IL-17受体、TGF-β和MAPK信号通路提高鸡的肠道免疫功能。

关键词: 酵母β-葡聚糖, 新城疫疫苗, 肠道免疫, 转录组学

Abstract:

The aim of this study is to investigate the effect of adding yeast β-glucan (G70) to the diet on intestinal immune function in chickens immunized Newcastle disease (ND) vaccine. Sixteen 1-day-old healthy black-bone chickens were randomly divided into 2 groups, the Vaccine group and the G70+Vaccine group, and in the G70+Vaccine group, yeast β-glucan was added at 1 g·kg^-1 in the basal diet. At 14 and 28 days of age, ND vaccine was administered for the 1^st immunization and booster immunization. At 35 days of age, jejunum tissue was collected to determine the number of IgA⁺cells, the proportions of CD4⁺CD8⁺double-positive T cells, and the mRNA expression levels of immune-related genes in jejunum, then transcriptome sequencing was performed, and the differentially expressed genes were analyzed by GO function and KEGG pathway. The results showed that, compared with the Vaccine group, dietary G70 significantly increased the number of IgA⁺cells (P < 0.05) and the proportions of CD4⁺CD8⁺double-positive T cells in the jejunum (P < 0.05), and the mRNA expression levels of GATA-3, MHC-Ⅰ, MHC-Ⅱ, CCR7 and IFN-γ were significantly up-regulated (P < 0.05). RNA-seq showed that there were 559 differentially expressed genes in G70+Vaccine group, of which 54 were up-regulated and 505 were down-regulated, the GO analysis revealed that most of the differentially expressed genes were annotated to the GO entries related to amine metabolism and catabolism, protein synthesis, stimulation of TGF-β, and regulation of enzyme activities. KEGG signaling pathway analysis revealed that the differentially expressed genes were mainly enriched in signaling pathways related to cell synthesis and metabolism, such as ribosomes, MAPK, cell adhesion and local adhesion. In summary, the addition of yeast β-glucan (G70) to the diet increased the number of IgA⁺cells, promoted the proportions of CD4⁺CD8⁺double-positive T cells, and up-regulated the expression of immune-related genes in jejunum. The possible mechanism of improving chicken's intestinal immune function might be affecting the IL-17 receptor, TGF-β and MAPK signaling pathways by regulating the expression of immune-related genes such as IL-17RD, TGF-β and TMEM158.

Key words: yeast β-glucan, Newcastle disease vaccine, intestinal immune function, transcriptomics

中图分类号:

S831.7

李常营, 李俊, 李锡锋, 毕师诚, 曹立亭. 基于转录组学探究日粮添加酵母β-葡聚糖对新城疫疫苗免疫鸡肠道免疫的影响[J]. 畜牧兽医学报, 2025, 56(3): 1441-1452.

LI Changying, LI Jun, LI Xifeng, BI Shicheng, CAO Liting. Effect of Dietary Yeast β-glucan Supplementation on Intestinal Immune Function in Chickens Immunized against Newcastle Disease Vaccine based on Transcriptomic[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(3): 1441-1452.

图/表 12

表 1

表 2

RT-qPCR引物序列"

基因 Genes	引物序列(5′→3′) Primer sequences(5′→3′)	扩增长度/bp Length
β-actin	5′-GAGAAATTGTGCGTGACATCA-3′ 5′-CCTGAACCTCTCATTGCCA-3′	134
TGF-β	F: 5′-TTCCAACACCAGGTCCTACTCCAG-3′ R: 5′-AAGCAGACAGGTCCAGCAATAACAG-3′	88
IL-6	F: 5′-GAAATCCCTCCTCGCCAATCTGAAG-3′ R: 5′-GCCCTCACGGTCTTCTCCATAAAC-3′	108
IFN-γ	F: 5′-ACGACACCATCCTGGACACC-3′ R: 5′-TTTGGCGTTGGCTGTCGTTC-3′	129
NF-κB	F: 5′-CTTGCCAAGCGTCACTGCAA-3′ R: 5′-GCAGTGAGATGGCGCTGAAC-3′	89
CD40	F: 5′-GGGCTCGTGGTGAAGGTGAAAG-3′ R: 5′-GGATCAGCACTGACAGCGATGAG-3′	85
CD80	F: 5′-TCGTTCAGAGTCTCCAGTCTTCACC-3′ R: 5′-CAGCGGTAACAAAGAGGGTCACAG-3′	84
CCR7	F: 5′-CCGACGACTATGACGCCAACAC-3′ R: 5′-GGAGGAAGGCAGCACGGAAATC-3′	96
CCR-9	F: 5′-AGTGCTCCTCCTTGGGTGACAG-3′ R: 5′-GGCATGAGGCAGGAACAACAGAG-3′	132
TLR-3	F: 5′-GCAAGCTATTGAGCAAAGTCGAGAC-3′ R: 5′-GCCAGTTCAAGATGCAGCAAGATC-3′	128
TLR-4	F: 5′-CATCCCAACCCAACCACAGTAGC-3′ R: 5′-CCACTGAGCAGCACCAATGAGTAG-3′	119
TLR-5	F: 5′-ACTCCCTTCCTTCCCACATCTGAC-3′ R: 5′-TGTGTTGCTACTATTGCCGTGTGAG-3′	87
GATA-3	F: 5′-CTACTTGTGTAACGCCTGTGGACTC-3′ R: 5′-GTGGTGGTGGTCTGACAGTTAGC-3′	132
J-CHAIN	F: 5′-GGTTCGTCCTTGTGGCAGGTTATC-3′ R: 5′-GAGGTCACCGTTACGCACTTACAC-3′	88
pIgR	F: 5′-AAGGGCTCTCCAACAGGGTCAG-3′ R: 5′-AGAACTTCCTCGTGCTGGCATAATC-3′	144
MHC-Ⅰ	F: 5′-GCACAGCCCCATCCTCT-3′ R: 5′-TGGCCCATCATTTTATTTCA-3′	101
MHC-Ⅱ	F: 5′-CAGCGTTCTTCTTCTGCGGT-3′ R: 5′-GGTTGTAGATTTCCCGTTCCAG-3′	90
AN04	5′-CAGCCCTTCCAAAGATGATGACTCC-3′ 5′-TTCCTCTCCCACCACTTCCAACC-3′	94
ASPA	5′-AATCTTGGACAGACAGTGGTGGAAG-3′ 5′-TGTTAGCAGTGGTGTTGTGAAGGTC-3′	136
IL-17RD	5′-GCCTTCCTCCTCGCTCTCCTC-3′ 5′-ATTGCTCTTGGTGACAGATGACAGG-3′	135
MYH	5′-AGCCGAAGCACAAGCCAATCTG-3′ 5′-GCCAATATCCCTTGACTTGCTCCTC-3′	133
CLDN3	5′-CTTCATCGGCAACAACATCGTGAC-3′ 5′-CCAGCATGGAGTCGTACACCTTG-3′	113
TMEM158	5′-GCTCAACTTCTGCTGCTTGGATTTC-3′ 5′-CCACGCTCCACACGATGATGAC-3′	134
RRS1	5′-GGCAAGAAACGGCGCTTCCAG-3′ 5′-CTCGCGGAGCTGCTTGTTGAC-3′	138
MT3	5′-ACCTGCACGTGTGGAGACAA-3′ 5′-GCACACTTGGCACATCCTGC-3′	101

表 2

图 1

图 2

图 3

图 4

图 5

表 3

部分下调和上调的差异表达基因"

基因ID Gene ID	基因名称 Gene name	差异倍数 log₂FC	基因描述 Gene characterization
下调基因 Down-regulated genes
ENSGALG00000053991	SERF2	-3.070914732	small EDRK-rich factor 2
ENSGALG00000039489	5-8S rRNA	-3.027282516	5_8S ribosomal RNA
ENSGALG00000030890	RRS1	-2.198902274	ribosome biogenesis regulator homolog
ENSGALG00000034326	TMEM158	-2.198603339	transmembrane protein 158
ENSGALG00000041752	CD164L2	-1.965298427	CD164 molecule like 2
ENSGALG00000049538	RPS28	-1.921395818	ribosomal protein S28
ENSGALG00000002517	TMEM35B	-1.741653236	transmembrane protein 35B
ENSGALG00000043641	JUND	-1.618482633	JunD proto-oncogene
ENSGALG00000037713	FNDC10	-1.458563877	fibronectin type Ⅲ
ENSGALG00000049153	PDF	-1.400669956	peptide deformylase
ENSGALG00000029768	PTGDR	-1.375135892	prostaglandin D2 receptor-like
ENSGALG00000037716	UBA52	-1.330577817	ubiquitin A-52
ENSGALG00000031067	TMEM132A	-1.305910743	transmembrane protein 132A
ENSGALG00000030407	IL-6R	-1.268521422	interleukin 6 receptor
ENSGALG00000041232	TMEM223	-1.142077014	transmembrane protein 223
上调基因 Up-regulated genes
ENSGALG00000011614	ANO4	2.392598397	anoctamin 4
ENSGALG00000015358	MYH15	1.956091863	myosin, heavy chain 15
ENSGALG00000051773	JHY	1.822528177	chromosome 11 open reading frame 63
ENSGALG00000005499	IL17-RD	1.615012462	interleukin 17 receptor
ENSGALG00000003876	TIMD4	1.524616906	T-cell immunoglobulin and mucin domain containing 4
ENSGALG00000043027	HEPHL1	1.417362402	hephaestin like 1
ENSGALG00000005360	CA4	1.377146878	carbonic anhydrase 4
ENSGALG00000049288	CD86	1.278387892	CD86 molecule
ENSGALG00000028790	DNASE2B	1.267707784	deoxyribonuclease 2 beta
ENSGALG00000019651	AGR3	1.241116654	anterior gradient 3
ENSGALG00000030703	CYBRD1	1.197753577	cytochrome b reductase 1
ENSGALG00000002390	CA6	1.123467369	carbonic anhydrase 6
ENSGALG00000037077	CCR5	1.038338392	C-C motif chemokine receptor 5
ENSGALG00000013748	CD226	1.02970962	CD226 molecule

表 3

图 6

图 7

表 4

图 8

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日粮组成 Feed ingredient	1~21日龄 1~21 day-old	22-35日龄 22~35 day-old
玉米Corn	55.34	60.53
麸皮Wheat bran	2.70	2.70
植物油Vegetable oil	3.17	3.83
鱼粉Fish meal	5.01	4.90
豆粕Soybean meal	19.00	12.00
胡麻饼Hemp cake	4.50	5.20
棉籽粕Cottonseed meal	3.20	4.10
菜籽粕Rapeseed meal	2.70	3.06
DL-蛋氨酸DL-Methionine	0.12	0.02
磷酸氢钙CaHPO₄	3.13	2.46
石粉Limestone	0.15	0.33
食盐NaCl	0.27	0.26
氯化胆碱Choline chloride	0.12	0.02
预混料^a Premix^a	0.50	0.50
合计Total	100	100
营养水平^b Nutrient content^b
粗蛋白质Crude protein	20.00	18.90
代谢能/(MJ·kg^－1)ME	12.11	12.54
蛋氨酸Methionine	0.48	0.38
赖氨酸Lysine	1.09	0.94
钙Calcium	1.00	0.90
有效磷Effective phosphorus	0.45	0.41

通路 Pathway	数量 Count	q值 q-value	基因 Gene
核糖体 Ribosome	11	0.001	RPS28/MRPL9、MRPS14/MRPL17、MRPL4/UBA52、MRPL20/RPS19、MRPS12/RPL35A、MRPS21
MAPK信号通路 MAPK signaling pathway	9	0.23	JUND/ECSIT NFKB2/TGFA/GADD45G/PGF
细胞黏附分子 Cell adhesion molecules	7	0.032	CLDN3/BLB1、CD40/ITGB7、CD86/CD226
黏着斑 Focal adhesion	7	0.26	PPP1R12C/ITGB7/PGF

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基于转录组学探究日粮添加酵母β-葡聚糖对新城疫疫苗免疫鸡肠道免疫的影响

Effect of Dietary Yeast β-glucan Supplementation on Intestinal Immune Function in Chickens Immunized against Newcastle Disease Vaccine based on Transcriptomic

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