Chinese Journal of Animal Nutrition >

2025 , Vol. 37 >Issue 1: 560 - 570

DOI: https://doi.org/10.12418/CJAN2025.048

RESEARCH PAPER

Effects of β-Glucan from Euglena gracilis on Growth Performance and Intestinal Microbiota and Tissue Protein Expression of Ira Meat Rabbits

  • WANG Guoyan ,
  • LI Yanping ,
  • REN Keliang ,
  • FAN Aifang ,
  • TANG Yaoping ,
  • DANG Wenqing ,
  • ZHAN Haijie ,
  • LI Jun ,
  • LIU Huadong ,
  • LI Tingting ,
  • TANG Juan ,
  • CAO Liang , *
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  • College of Animal Science, Shanxi Agricultural University, Taigu 030801, China
* associate professor, E-mail:

Received date: 2024-06-14

  Online published: 2025-01-10

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Abstract

This experiment was conducted to investigate the effects of dietary β-glucan from Euglena gracilis on growth performance and intestinal microbiota and tissue protein expression of Ira meat rabbits. A total of 120 35-day-old weaned Ila commercial meat rabbits (half male and half female) with similar body weight were randomly divided into 4 groups with 30 replicates per group and 1 rabbit per replicate. Rabbits in the control group were fed a basal diet, and those in the experimental groups were fed the basal diet supplemented with 300 (test group Ⅰ), 500 (test group Ⅱ) and 700 mg/kg (test group Ⅲ) β-glucan from Euglena gracilis, respectively. The pre-trial period lasted for 5 days and the experimental period lasted for 22 days. The results showed as follows: 1) there were no significant differences in the growth performance of meat rabbits among all groups (P>0.05). Compared with the control group, the mortality of meat rabbits in test groups Ⅱ and Ⅲ was significantly decreased (P<0.05). 2) Compared with the control group, the Simpson index of cecal microbiota of meat rabbits in test group Ⅱ was significantly increased (P<0.05). The results of analysis of similarities (ANOSIM) showed that there were significant differences in the composition of cecal microbiota between test group Ⅱ and other groups (P<0.05). Compared with the control group, at the phylum level, the relative abundances of Bacteroidetes and TM7 in cecum in test group Ⅱ were significantly increased (P<0.05); at the genus level, the Ruminococcus relative abundance in cecum in test group Ⅱ was significantly increased (P<0.05). 3) The heat shock protein 90 (HSP90) protein expression level in jejunum in test group Ⅱ was significantly lower than that in the other groups (P<0.05), and the Toll-like receptor 4 (TLR4) protein expression level in jejunum in test group Ⅱ was significantly lower than that in the control group and test group Ⅰ (P<0.05); the β-defensin-2 (BD-2) protein expression level in jejunum in test groups Ⅱ and Ⅲ was significantly lower than that in the control group and test group Ⅰ (P<0.05). In conclusion, under the conditions of this experiment, dietary supplementation of 500 mg/kg β-glucan from Euglena gracilis can reduce the mortality of Ila meat rabbits, increase the diversity of cecal microbiota, increase the relative abundances of beneficial bacteria in cecum, decrease the relative abundances of harmful bacteria in cecum, decrease the protein expression levels of HSP90, TLR4 and BD-2 in jejunum, enhance intestinal immunity and improve intestinal health.

Key words： β-glucan from Euglena gracilis; Ira rabbits; growth performance; intestinal microbiota; intestinal tissue protein

Cite this article

WANG Guoyan , LI Yanping , REN Keliang , FAN Aifang , TANG Yaoping , DANG Wenqing , ZHAN Haijie , LI Jun , LIU Huadong , LI Tingting , TANG Juan , CAO Liang . Effects of β-Glucan from Euglena gracilis on Growth Performance and Intestinal Microbiota and Tissue Protein Expression of Ira Meat Rabbits[J]. Chinese Journal of Animal Nutrition, 2025 , 37(1) : 560 -570 . DOI: 10.12418/CJAN2025.048

幼龄动物在断奶时,因其自身消化系统发育不完全,机体处于疾病易感期,当受到断奶、换料以及换窝等外界因素的刺激时,极易发生消化系统疾病。兔子的肠道系统十分庞大,其肠道总长度与体长比约为10:1,故肠道健康是其生产性能的重要影响因素。我国自2020年饲料全面禁抗以来[1],寻找能够提高动物生产性能且可提高动物疾病预防能力的绿色饲料添加物已成为研究热点。研究表明,β-葡聚糖不仅具有抗应激功能,而且可通过对肠道微生物区系的调节来增强肠道健康[2-3]。关于β-葡聚糖在饲粮中的应用研究已有报道,在幼龄反刍动物饲粮中添加酵母β-葡聚糖可提高犊牛平均日增重(ADG),改善腹泻程度[4];饲粮添加裸藻β-1,3-葡聚糖可以调节仔猪肠道微生物多样性[5]。而裸藻β-葡聚糖对兔肠道影响的研究报道相对较少。因此,本试验旨在探究裸藻β-葡聚糖对肉兔生长性能和肠道菌群及组织蛋白表达的影响,为在肉兔养殖过程中合理使用裸藻β-葡聚糖提供科学依据。

1 材料与方法

1.1 试验材料

本试验所用的裸藻β-葡聚糖为裸藻β-1,3-葡聚糖,为市售产品,纯度>95%。

1.2 试验设计和饲粮

本试验由山西农业大学实验动物伦理委员会批准进行,伦理批准编号为:SXAU-EAW-2023R.UG.004029177。
试验选用120只体重相近的35日龄断奶伊拉康大配套系商品肉兔(公母各占1/2),随机分为4组,每组30个重复,每个重复1只兔。对照组饲喂基础饲粮,试验组分别饲喂在基础饲粮的基础上添加300(试验Ⅰ组)、500(试验Ⅱ组)和700 mg/kg(试验Ⅲ组)裸藻β-葡聚糖的饲粮。预试期5 d,正试期22 d。
基础饲粮依据GB/T 14924.9—2001方法对豆粕、玉米、麸皮、玉米胚芽粕、葵花皮、棉籽粕、菜籽粕及苜蓿等原料进行营养成分测定后,参照《肉兔营养需要量》(NY/T 4049—2021)[6]配制,其组成及营养水平见表1。裸藻β-葡聚糖通过添加在预混料中后再添加到饲粮中。各组饲粮均制成直径为4 mm的颗粒料。原料和饲粮营养成分采用FOSS 2500近红外光谱仪测定。
表1 基础饲粮组成及营养水平(风干基础)

Table 1 Composition and nutrient levels of the basal diet (air-dry basis) %

项目Items 含量Content
原料Ingredients
玉米Corn 20.00
麸皮Wheat bran 15.00
豆粕Soybean meal 12.00
棉籽粕Cottonseed meal 2.00
菜籽粕Rapeseed meal 3.00
玉米胚芽粕Corn germ meal 7.00
苜蓿Alfalfa 5.00
葵花皮Sunflower skin 31.00
食盐NaCl 0.50
预混料Premix1) 4.50
合计Total 100.00
营养水平Nutrient levels2)
消化能DE/(MJ/kg) 9.67
粗蛋白质CP 15.09
粗纤维CF 18.15
粗灰分Ash 6.56
中性洗涤纤维NDF 35.68
酸性洗涤纤维ADF 24.35
酸性洗涤木质素ADL 5.04
赖氨酸Lys 0.89
蛋氨酸+胱氨酸Met+Cys 0.61
钙Ca 1.12
总磷TP 0.52

1)预混料为每千克基础饲粮提供 The premix provided the following per kg of the basal diet:VA 6 000 IU,VD3 950 IU,VE 40 mg,赖氨酸 Lys 2.2 g,蛋氨酸 Met 1.2 g,Cu (as copper sulfate) 10 mg,Fe (as ferrous sulfate) 50 mg,Mn (as manganese sulfate) 8.0 mg,Zn (as zinc sulfate) 50 mg,CaHPO4 2 500 mg。

2)消化能为计算值,其余为实测值。消化能为肉兔对各原料的消化能估值之和,计算公式为消化能=0.013×粗蛋白质+0.036×粗脂肪+0.017×非纤维碳水化合物+0.006×中性洗涤纤维[7]。DE was a calculate value, while the others were measured values; DE was the sum of the estimated digestible energy of all raw materials for meat rabbits, and the calculation formula was DE=0.013×CP+0.036×EE+0.017×NFC+0.006×NDF[7].

1.3 饲养管理

选取自然通风、采光好的兔舍作为试验兔舍。试验前,对兔舍及水线管道、饲养笼具等进行全面打扫和消毒,然后将试验兔称重、编号、分组,均采用单笼饲养;试验期间进行常规免疫,分别在每日08:30和17:30进行饲喂,自由饮水。

1.4 测定指标及方法

1.4.1 生长性能、腹泻率和死亡率

试验开始和结束时,于清晨对空腹后的试验兔进行称重,分别记为初重和末重,计算ADG;试验期间记录每只试验兔的采食量,计算平均日采食量(ADFI),并计算料重比(F/G);试验期间观察和记录试验兔腹泻和死亡情况,计算腹泻率和死亡率。计算公式如下:
腹泻率(%)=100×(各组腹泻试验兔只数×腹泻天数)/(各组试验兔总数×试验天数);
死亡率(%)=100×各组死亡兔只数/各组试验兔总数。

1.4.2 盲肠菌群

采集各组试验兔新鲜无污染的盲肠内容物,装入5 mL EP管中,迅速放入液氮中冻存,随后将盲肠内容物样品存放于-80 ℃冰箱保存备用。肠道菌群委托上海派森诺生物科技股份有限公司运用Illumina HiSeq平台对样本进行测序。

1.4.3 空肠组织蛋白表达量

试验结束时,禁食12 h,每组随机选取4只试验兔(2公2母),屠宰后用手术剪截取空肠段,并用0.9%氯化钠溶液冲洗,重复3次,将空肠内容物冲洗干净后,用滤纸吸干表面水分后,取1.0 g肠道组织,加入9.0 mL匀浆液[磷酸盐缓冲液(PBS),pH为7.2~7.4,浓度为0.01 mol/L),液氮研磨20 min,1 662×g离心15 min,取上清液。采用上海酶联生物科技有限公司的试剂盒检测空肠组织中热休克蛋白90(HSP90)、Toll样受体4(TLR4)和β-防御素-2(β-defensin-2,BD-2)蛋白表达量。

1.5 数据统计与分析

在Illumina HiSeq平台上,采用QIIME软件对各组样本中经过过滤、降噪后的有效序列进行多样性分析。采用SPSS 26.0统计软件对试验数据进行单因素方差分析(one-way ANOVA),并采用Duncan氏法进行多重比较,腹泻率和死亡率多重比较采用卡方检验;除腹泻率和死亡率外,试验结果数据以“平均值±标准误”形式表示,P<0.05表示差异显著。

2 结果与分析

2.1 裸藻β-葡聚糖对肉兔生长性能、腹泻率和死亡率的影响

由表2可知,各组间肉兔生长性能指标和腹泻率均无显著差异(P>0.05)。其中,试验Ⅱ组ADG最高,F/G和腹泻率最低。与对照组相比,试验Ⅱ组和试验Ⅲ组死亡率显著降低(P<0.05)。由此可见,本试验中试验Ⅱ组肉兔生长性能最佳。
表2 裸藻β-葡聚糖对肉兔生长性能、腹泻率和死亡率的影响

Table 2 Effects of β-glucan from Euglena gracilis on growth performance, diarrhea rate and mortality of meat rabbits

项目
Items		 对照组
Control group		 试验Ⅰ组
Test group Ⅰ		 试验Ⅱ组
Test group Ⅱ		 试验Ⅲ组
Test group Ⅲ		 P值
P-value
初重IBW/g 1 045.75±130.40 1 066.00±130.40 1 075.50±145.93 1 065.00±136.74 0.919
末重FBW/g 2 009.30±196.35 2 097.35±218.86 2 144.30±221.38 2 093.30±161.65 0.206
平均日采食量ADFI/g 116.06±11.37 121.81±10.09 123.47±8.60 119.21±7.55 0.083
平均日增重ADG/g 37.06±6.38 39.67±5.01 41.11±6.36 39.55±5.63 0.187
料重比F/G 3.19±0.41 3.10±0.35 3.05±0.34 3.07±0.46 0.701
腹泻率Diarrhea rate/% 6.06 4.09 2.27 4.85 0.705
死亡率Mortality/% 23.33a 13.33ab 3.33b 3.33b 0.035

同行数据肩标不同小写字母表示差异显著(P<0.05),相同小写字母或无字母表示差异不显著(P>0.05)。表3、表5、表6和表7同。

In the same row, values with different small letter superscripts mean significant difference (P<0.05), while with the same letter or no letter superscripts mean no significant difference (P>0.05). The same as Table 3, Table 5, Table 6 and Table 7.

2.2 裸藻β-葡聚糖对肉兔盲肠菌群的影响

2.2.1 裸藻β-葡聚糖对肉兔盲肠菌群α多样性的影响

由表3可知,与对照组相比,试验Ⅱ组肉兔盲肠菌群Simpson指数显著提高(P<0.05);同时,试验Ⅱ组Simpson指数显著高于试验Ⅲ组(P<0.05)。此外,随着饲粮裸藻β-葡聚糖添加水平的提高,各组盲肠菌群物种丰富度指数Observed_species指数和Chao1指数呈现先升高后降低的变化趋势,以试验Ⅱ组最高;物种多样性指数Shannon指数以及均匀度指数Pielou_e指数亦呈现先升高再降低的变化趋势,并同样以试验Ⅱ组最高,这表明饲粮添加裸藻β-葡聚糖可以提高肉兔盲肠菌群物种多样性、丰富度以及均匀度,但高水平添加会有负面影响。综上可知,试验Ⅱ组肉兔盲肠菌群物种丰富度、多样性以及均匀度最好。
表3 裸藻β-葡聚糖对肉兔盲肠菌群α多样性的影响

Table 3 Effects of β-glucan from Euglena gracilis on cecal microbiota α diversity of meat rabbits

项目
Items		 对照组
Control group		 试验Ⅰ组
Test group Ⅰ		 试验Ⅱ组
Test group Ⅱ		 试验Ⅲ组
Test group Ⅲ		 P值
P-value
Chao1指数Chao1 index 5 756.20±2 148.96 7 181.10±2 716.48 9 462.95±1 623.06 6 712.70±933.45 0.105
Observed_species指数
Observed_species index		 2 992.50±1 017.52 3 539.60±1 118.03 4 486.30±661.13 3 228.03±508.21 0.128
Simpson指数Simpson index 0.97±0.01b 0.99±0.01ab 0.99±0.00a 0.98±0.00b 0.011
Shannon指数Shannon index 8.35±1.04 8.94±0.74 9.62±0.42 8.46±0.39 0.092
Pielou_e指数Pielou_e index 0.73±0.06 0.76±0.04 0.79±0.02 0.73±0.02 0.066

2.2.2 裸藻β-葡聚糖对肉兔盲肠菌群β多样性的影响

如图1所示,肉兔盲肠菌群非度量多维尺度分析(NMDS)的应力值(stress)小于0.2(stress=0.098),说明NMDS较为可靠,图中点与点在坐标轴上的投影距离越近,说明样本间的微生物群落组成越相似。对照组与试验Ⅰ组有重叠区域,表明这2组间微生物群落组成差异不大。由表4可知,采用相似性分析(ANOSIM)检验方法对各组样本两两之间进行显著差异性分析,得到R值在0~1,说明各组间差异大于组内差异,R值越靠近1,表明各组间差异越大;经ANOSIM验证后,对照组与试验Ⅰ组间微生物群落组成差异不显著(P=0.140),而试验Ⅱ组与其他各组间差异均显著(0.029<P<0.034)。
图1 肉兔盲肠菌群非度量多维尺度分析

Fig.1 NMDS of cecal microbiota of meat rabbits

表4 肉兔盲肠菌群相似性分析

Table 4 ANOSIM of cecal microbiota of meat rabbits

项目
Items		 样本大小
Sample size/个		 置换次数
Permutations/次		 R值
R-value		 q值
q-value		 P值
P-value
对照组vs.试验Ⅰ组
Control group vs. test group Ⅰ		 8 999 0.281 250 0.140 0.140
对照组vs.试验Ⅱ组
Control group vs. test group Ⅱ		 8 999 0.572 917 0.034 0.034
对照组vs.试验Ⅲ组
Control group vs. test group Ⅲ		 8 999 0.604 167 0.031 0.031
试验Ⅰ组vs.试验Ⅱ组
Test group Ⅰ vs. test group Ⅱ		 8 999 0.343 750 0.031 0.031
试验Ⅰ组vs.试验Ⅲ组
Test group Ⅰ vs. test group Ⅲ		 8 999 0.510 417 0.034 0.034
试验Ⅱ组vs.试验Ⅲ组
Test group Ⅱ vs. test group Ⅲ		 8 999 0.697 917 0.029 0.029

2.2.3 裸藻β-葡聚糖对肉兔盲肠菌群组成的影响

如图2所示,各组肉兔盲肠菌群中的优势菌门为厚壁菌门(Firmicutes)、拟杆菌门(Bacteroidetes)、疣微菌门(Verrucomicrobia)和TM7等,相对丰度之和占总菌门的97%以上。由表5可知,与对照组相比,试验Ⅱ组盲肠拟杆菌门和TM7相对丰度显著提高(P<0.05);同时,试验Ⅱ组盲肠拟杆菌门和TM7相对丰度显著高于试验Ⅰ组和试验Ⅲ组(P<0.05)。随着饲粮裸藻β-葡聚糖添加水平的提高,各组盲肠厚壁菌门相对丰度呈现先降低再升高的变化趋势,其中以试验Ⅱ组最低。
图2 裸藻β-葡聚糖对肉兔盲肠菌群在门水平上组成的影响

Fig.2 Effects of β-glucan from Euglena gracilis on cecal microbiota composition of meat rabbits at phylum level

表5 裸藻β-葡聚糖对肉兔盲肠菌群在门水平上相对丰度的影响

Table 5 Effects of β-glucan from Euglena gracilis on relative abundance of cecal microbiota of meat rabbits at phylum level

项目
Items		 对照组
Control group		 试验Ⅰ组
Test group Ⅰ		 试验Ⅱ组
Test group Ⅱ		 试验Ⅲ组
Test group Ⅲ		 P值
P-value
厚壁菌门Firmicutes 93.31±3.49 90.86±9.26 85.31±3.01 94.57±1.88 0.115
拟杆菌门Bacteroidetes 2.06±0.98b 2.52±2.13b 6.54±0.33a 1.68±0.97b 0.001
疣微菌门Verrucomicrobia 1.59±0.93 3.83±5.79 3.65±3.65 0.80±0.49 0.539
TM7 0.79±0.38b 0.86±0.74b 1.79±0.52a 0.73±0.40b 0.045
变形菌门Proteobacteria 0.59±0.40 0.44±0.48 0.46±0.19 1.26±0.68 0.095
放线菌门Actinobacteria 0.56±0.41 0.54±0.59 1.04±0.11 0.41±0.12 0.136
软壁菌门Tenericutes 0.66±0.49 0.44±0.46 0.55±0.27 0.37±0.04 0.693
绿弯菌门Chloroflexi 0.28±0.13 0.37±0.30 0.31±0.10 0.13±0.05 0.292
蓝细菌门Cyanobacteria 0.07±0.07 0.08±0.08 0.11±0.02 0.06±0.03 0.673
由表6可知,各组肉兔盲肠菌群排名前8的优势菌属为阿克曼菌属(Akkermansia)、瘤胃球菌属(Ruminococcus)、粪球菌属(Coprococcus)、颤螺旋菌属(Oscillospira)、脱硫弧菌属(Desulfovibrio)、梭菌属(Clostridium)、布劳特氏菌属(Blautia)和多尔氏菌属(Dorea)。与对照组相比,试验Ⅱ组盲肠瘤胃球菌属相对丰度显著提高(P<0.05);同时,试验Ⅱ组盲肠瘤胃球菌属相对丰度显著高于试验Ⅰ组和试验Ⅲ组(P<0.05)。
表6 裸藻β-葡聚糖对肉兔盲肠菌群在属水平上相对丰度的影响

Table 6 Effects of β-glucan from Euglena gracilis on relative abundance of cecal microbiota of meat rabbits at genus level

项目
Items		 对照组
Control group		 试验Ⅰ组
Test group Ⅰ		 试验Ⅱ组
Test group Ⅱ		 试验Ⅲ组
Test group Ⅲ		 P值
P-value
阿克曼菌属Akkermansia 1.59±0.93 3.83±5.79 3.65±3.65 0.80±0.49 0.539
瘤胃球菌属Ruminococcus 2.18±0.99b 1.85±0.49b 4.30±2.27a 1.38±0.08b 0.031
粪球菌属Coprococcus 2.20±2.44 1.06±0.76 0.74±0.39 1.59±1.12 0.508
颤螺旋菌属Oscillospira 1.40±0.87 1.01±0.22 1.48±0.33 0.51±0.14 0.053
脱硫弧菌属Desulfovibrio 0.50±0.33 0.37±0.41 0.34±0.25 1.14±0.57 0.056
梭菌属Clostridium 0.27±0.12 0.39±0.08 0.62±0.31 0.36±0.15 0.103
布劳特氏菌属Blautia 0.35±0.07 0.48±0.33 0.25±0.05 0.41±0.13 0.363
多尔氏菌属Dorea 0.41±0.29 0.17±0.07 0.15±0.09 0.18±0.02 0.115
安德克氏菌属Adlercreutzia 0.20±0.12 0.16±0.21 0.17±0.10 0.20±0.13 0.702

2.2.4 肉兔盲肠菌群差异物种分析

采用线性判别分析(LDA)效应大小(LEfSe)方法对试验Ⅱ组肉兔盲肠菌群在不同分类水平上与其他组进行差异物种分析,筛选出相对丰度高且差异显著的物种,结果如图3所示。与其他组相比较,试验Ⅱ组肉兔盲肠菌群相对丰度较高的差异物种为拟杆菌门、拟杆菌纲(Bacteroidia)、拟杆菌目(Bacteroidales)中的理研菌科(Rikeneallaceae)和S24_7。
图3 试验Ⅱ组肉兔盲肠菌群差异物种进化分枝图

Fig 3 Cladogram of differential species in cecum microbiota of meat rabbits in test group Ⅱ

2.3 裸藻β-葡聚糖对肉兔空肠组织蛋白表达量的影响

由表7可知,随着饲粮裸藻β-葡聚糖添加水平的提高,各组肉兔空肠HSP90和TLR4蛋白表达量呈现先降低后升高的变化趋势,其中,试验Ⅱ组空肠HSP90蛋白表达量显著低于其他各组(P<0.05),空肠TLR4蛋白表达量显著低于对照组和试验Ⅰ组(P<0.05)。肉兔空肠BD-2蛋白表达量随饲粮裸藻β-葡聚糖添加水平的提高而逐渐降低,其中,试验Ⅱ组和试验Ⅲ组空肠BD-2蛋白表达量显著低于对照组和试验Ⅰ组(P<0.05),且试验Ⅱ组与试验Ⅲ组空肠之间BD-2蛋白表达量无显著差异(P>0.05)。
表7 裸藻β-葡聚糖对肉兔空肠组织蛋白表达量的影响

Table 7 Effects of β-glucan from Euglena gracilis on jejunal tissue protein expression level of meat rabbits

项目
Items		 对照组
Control group		 试验Ⅰ组
Test group Ⅰ		 试验Ⅱ组
Test group Ⅱ		 试验Ⅲ组
Test group Ⅲ		 P值
P-value
热休克蛋白90 HSP90 714.95±35.00a 637.03±37.45b 535.93±24.34c 658.53±24.19b <0.001
Toll样受体4 TLR4 16.98±0.86a 15.81±1.01b 14.62±0.43c 14.95±0.61bc 0.001
β-防御素-2 BD-2 212.30±13.14a 198.52±7.95a 183.38±12.73b 178.04±7.11b <0.001

3 讨论

3.1 裸藻β-葡聚糖对肉兔生长性能、腹泻率和死亡率的影响

在畜牧养殖过程中,动物的健康状态尤为重要。对肉兔养殖而言,腹泻率和死亡率是评价其健康状态的重要指标。研究表明,饲粮添加裸藻β-1,3-葡聚糖可降低仔猪腹泻率,尤其是在高水平添加时效果更加[5];裸藻β-1,3-葡聚糖可以增强小鼠免疫能力,改善其健康状态[8]。本试验中,饲粮添加裸藻β-1,3-葡聚糖可降低伊拉肉兔腹泻率和死亡率,改善肉兔健康状态,与上述报道结果一致。但高水平添加反而会增加肉兔的腹泻情况,这可能是由以下2种因素造成的:1)从生理结构上,兔子的消化系统比其他畜禽的肠道系统更脆弱、更敏感;2)裸藻β-葡聚糖是一种益生素,可以通过诱导免疫细胞激活和分化增殖,起到增强机体免疫能力的作用,但是免疫系统的过度反应也会对机体造成损伤。

3.2 裸藻β-葡聚糖对肉兔盲肠菌群的影响

本试验中,饲粮添加500 mg/kg裸藻β-葡聚糖显著提高肉兔盲肠菌群Simpson指数,同时Chao1指数、Shannon指数和Pielou_e指数等也以试验Ⅱ组最高,表明饲粮添加500 mg/kg裸藻β-葡聚糖可以提高盲肠菌群物种丰富度、多样性以及均匀度。同时,ANOSIM结果表明,试验Ⅱ组盲肠微生物群落组成也与其他组存在显著差异。
动物断奶后,机体的免疫能力及抗病性主要由肠道健康所决定[9],肠道微生物组成及多样性是影响肠道健康的主要因素之一[10]。拟杆菌门和厚壁菌门是家兔肠道菌群中的主要菌门[11],二者相辅相成,对稳定肠道菌群组成起到重要作用[12]。拟杆菌门和厚壁菌门可对木质纤维进行分解,并通过对多糖的降解为机体提供能量[13]。拟杆菌门相对丰度过低可能预示肠道炎症[14];厚壁菌门相对丰度与肥胖呈正相关,而肥胖易增加肠道通透性[15-16]。肠道中的TM7是一种寄生菌门,可以促进生物膜的形成,减少宿主细菌诱发的炎症损伤[17]。本研究中,饲粮添加500 mg/kg裸藻β-葡聚糖显著提高伊拉肉兔盲肠拟杆菌门和TM7相对丰度,并降低厚壁菌门相对丰度,这与王红明等[14]的研究中机体免疫力提高与拟杆菌门相对丰度升高的结果相吻合,提示饲粮添加500 mg/kg裸藻β-葡聚糖可能通过提高伊拉肉兔肠道中有益菌门的相对丰度使其免疫功能提高。
采用LEfSe方法进行差异物种分析发现,试验Ⅱ组显著富集理研菌科和S24_7(属于未分类拟杆菌科)。理研菌科既能促进植物生长,也能用来疾病治疗,属于一种益生菌[18-19];而未分类拟杆菌科可能通过与肠道致病菌竞争利用单糖,起到抑制致病菌的作用[20],也被视为益生菌的一种,本试验结果与赖碧玉等[21]的研究中肠道的病变伴随着未分类拟杆菌科相对丰度下降的结果相一致。
瘤胃球菌属可分解坚硬的植物物质,具有稳定肠道屏障的作用,是一种益生菌[22];颤螺旋菌属被列为下一代益生菌的候选者[23];大部分梭菌属如普拉梭菌、丁酸梭菌等,被列为益生菌,可抵抗病菌侵袭[24-25];多尔氏菌属和脱硫弧菌属的相对丰度与炎症性肠病正相关,均为有害菌[26-27]。本试验中,饲粮添加500 mg/kg裸藻β-葡聚糖可显著提高伊拉肉兔盲肠瘤胃球菌属相对丰度,这与杨馥嘉等[5]对裸藻β-1,3-葡聚糖在仔猪饲粮中的研究结果一致;同时,饲粮添加500 mg/kg裸藻β-葡聚糖可以降低有益菌属(颤螺旋菌属和梭菌属)相对丰度,降低有害菌属(多尔氏菌属和脱硫弧菌属)相对丰度,从而降低肠道潜在的炎症反应。
综上可知,饲粮添加500 mg/kg裸藻β-葡聚糖可通过提高有益菌相对丰度并降低有害菌相对丰度来调节肉兔肠道微生物群落结构,改善肠道微生态环境,降低肠道出现潜在炎性症状的可能性。

3.3 裸藻β-葡聚糖对肉兔空肠组织蛋白表达量的影响

当有害因子侵入机体或不良外界环境刺激机体时,机体的先天免疫系统会被激活,对有害因子及受损组织进行清除活动,在机体对自身健康的维护过程中起重要作用[28]。但是过度的炎症反应也会对机体造成损伤[29]。HSP90、TLR4和BD-2与炎症和免疫相关。Zhang等[30]对猪运输应激过程中热休克蛋白家族蛋白表达量变化的研究表明,在应激条件下,猪的心脏以及肝脏会发生急性病理性损伤,与此同时,热休克蛋白家族蛋白表达量会升高;李源等[31]对仔猪的断奶应激研究表明,低日龄断奶引起应激时,仔猪内脏中热休克蛋白家族蛋白量升高。HSP90表达水平的过度上调是癌细胞增殖和存活的关键因素之一[32],是机体存在潜在危害因子的表现。周琦璐等[33]研究发现,通过对TLR4信号通路表达的抑制,可缓解应激对仔猪造成的损伤状态。赵慧巧等[34]研究表明,在发生结肠病的肠道组织中BD-2的表达量会升高,通过给药对BD-2的分泌进行抑制,可以抑制Toll样受体/核因子-κB(NF-κB)通路中TLR4诱导的NF-κB被激活,从而限制炎性因子的分泌并促进抗炎因子的分泌,实现抗炎作用,提高机体免疫功能。本试验中,饲粮添加500 mg/kg裸藻β-葡聚糖可显著降低伊拉肉兔空肠组织中HSP90、TLR4和BD-2的蛋白表达量,减少潜在炎性反应,进而起到增强肠道免疫功能的作用。

4 结论

在本试验条件下,饲粮添加500 mg/kg裸藻β-葡聚糖可降低伊拉肉兔死亡率,提高盲肠菌群多样性,并提高盲肠有益菌相对丰度,降低有害菌相对丰度,同时降低空肠HSP90、TLR4和BD-2的蛋白表达量,增强肠道免疫能力,改善肠道健康。

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