Effects of Probiotics on Growth Performance, Plasma Immune and Antioxidant Indexes, and Intestinal Health of Lactating Calves

KONG Xiaoli; LIU Xiaonnu; REN Wenyi; YANG Shuangming; LIU Miao; KONG Weihao; LIU Min; ZHANG Lili; XU Xiaofeng

doi:10.12418/CJAN2026.220

2026 , Vol. 38 >Issue 4: 2730 - 2740

DOI: https://doi.org/10.12418/CJAN2026.220

RESEARCH PAPER

Effects of Probiotics on Growth Performance, Plasma Immune and Antioxidant Indexes, and Intestinal Health of Lactating Calves

KONG Xiaoli ^,¹ ,
LIU Xiaonnu ¹ ,
REN Wenyi ¹ ,
YANG Shuangming ¹ ,
LIU Miao ¹ ,
KONG Weihao ¹ ,
LIU Min ² ,
ZHANG Lili ¹ ,
XU Xiaofeng ^,¹^,^*

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¹ College of Animal Science and Technology, Ningxia University, Yinchuan 750021, China
² Ningxia Xingken Animal Husbandry Co., Ltd., Yinchuan 750021, China

*professor, E-mail: xuxiaofengnd@126.com

Received date: 2025-09-12

Online published: 2026-04-14

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Abstract

This experiment was conducted to investigate the effects of Bacillus licheniformis, Bacillus subtilis, and Lactobacillus plantarum on growth performance, plasma immune and antioxidant indices, and intestinal health of lactating Holstein calves. Forty-eight healthy newborn Holstein calves with similar body weight [(36.73±1.57) kg] and parity [(1.53±0.51) parities]were randomly divided into four groups, with 12 calves per group. Control group (CON) was fed milk + starter feed; the Bacillus licheniformis group (BL group) was fed milk + 1 g/d Bacillus licheniformis + starter feed; the Bacillus subtilis group (BS group) was fed milk + 1 g/d Bacillus subtilis + starter feed; and the Lactobacillus plantarum group (LP group) was fed milk + 1 g/d Lactobacillus plantarum + starter feed. The trial lasted for 70 days. The results showed as followed: 1) compared with the CON group, the final body weight (FBW), average daily gain (ADG), dry matter intake (DMI) and feed-to-gain ratio (F/G) did not significantly change in the BL, BS, and LP groups (P>0.05). 2) Compared with the CON group, plasma immunoglobulin A (IgA) content was significantly increased in the BS and LP groups at 7 and 21 days of age (P<0.05), and in the LP group at 70 days of age (P<0.05). Plasma immunoglobulin G (IgG) content was significantly increased in the BL group at 7 days of age (P<0.05). Plasma interleukin-1β (IL-1β) content was significantly decreased in the BL and LP groups at 7 days of age (P<0.05), and in the LP group at 21 days of age (P<0.05). 3) Compared with the CON group, plasma total antioxidant capacity (T-AOC) was significantly increased in the BS and LP groups at 21 days of age (P<0.05). Plasma total superoxide dismutase (T-SOD) activity was significantly increased in the BS group at 7 days of age (P<0.05). Plasma glutathione peroxidase (GSH-Px) activity was significantly increased in the BS group at 21 days of age (P<0.05), while plasma malondialdehyde (MDA) content was significantly decreased in the BS group at 21 days of age (P<0.05). 4) Compared with the CON group, the diarrhea rate was significantly reduced in the LP group from 1 to 35 days of age (P<0.05). Fecal scores and diarrhea rates were significantly reduced in the BL, BS, and LP groups from 36 to 70 days of age (P<0.05). The fecal score was significantly reduced in the LP group from 1 to 70 days of age (P<0.05), and diarrhea rates were significantly reduced in the BS and LP groups from 1 to 70 days of age (P<0.05). 5) Compared with the CON group, plasma diamine oxidase (DAO) activity at 21 days of age and plasma D-lactic acid (D-LA) content at 70 days of age were significantly decreased in the BL, BS, and LP groups (P<0.05). In conclusion, under the conditions of this trial, supplementing milk with Bacillus licheniformis, Bacillus subtilis, or Lactobacillus plantarum do not significantly improve the growth performance of lactating calves, but it significantly enhances immune function and antioxidant capacity, reduces diarrhea incidence, and improves intestinal barrier function, thereby promoting intestinal health.

Key words： probiotics; calves; growth performance; immune function; antioxidant capacity; intestinal health

Cite this article

KONG Xiaoli , LIU Xiaonnu , REN Wenyi , YANG Shuangming , LIU Miao , KONG Weihao , LIU Min , ZHANG Lili , XU Xiaofeng . Effects of Probiotics on Growth Performance, Plasma Immune and Antioxidant Indexes, and Intestinal Health of Lactating Calves[J]. Chinese Journal of Animal Nutrition, 2026 , 38(4) : 2730 -2740 . DOI: 10.12418/CJAN2026.220

犊牛阶段是生长发育的关键时期,新生犊牛的免疫和抗氧化系统不成熟,导致其抗病能力低。因此,这生理特点使得新生犊牛容易感染呼吸系统和肠道疾病,从而影响其后续生长和整体健康^[1]。由于抗生素存在动物产品残留的风险且可能诱导抗生素耐药菌株的产生,我国已禁止使用抗生素作为饲料添加剂。因此,开展益生菌和天然饲料添加剂等相关技术和方法的研究与应用,成为确保犊牛达到最佳生长发育状态和保障健康的重要途径^[2]。益生菌作为抗生素生长促进剂的替代品,已被广泛用于维护肠道健康领域,其可通过抑制有害病原体、增强肠道屏障功能和调节机体免疫力等方式发挥作用^[3]。研究表明,在犊牛断奶前,饲喂益生菌进行早期干预,可以通过促进胃肠道健康和维持断奶前犊牛的肠道屏障完整性,达到预防腹泻、降低死亡率的目的,进而改善犊牛健康状况^[4];在羔羊中,益生菌能促进羔羊肠道健康,减少腹泻的发生率^[5]。在各类益生菌中,芽孢杆菌因具备产生内孢子的能力,能够耐受极端环境条件,并在胃肠道内的存储和运输过程中仍能保持活性^[6]。地衣芽孢杆菌(Bacillus licheniformis)属于芽孢杆菌属,在自然界中广泛分布,具有抑制病原肠道细菌的能力及非致病性特点,与枯草芽孢杆菌(Bacillus subtilis)等其他芽孢杆菌物种相同^[7]。其中,枯草芽孢杆菌是畜牧业公认的强效益生菌,有研究表明,饲喂枯草芽孢杆菌可改善犊牛生长性能和健康状况^[8]。另有研究证明,植物乳杆菌(Lactobacillus plantarum)在促进营养物质消化率、增强机体免疫功能及减少病原体定植方面均具有有益作用^[9-10]。

因此,本试验通过在哺乳期荷斯坦犊牛奶中添加地衣芽孢杆菌、枯草芽孢杆菌和植物乳杆菌,探究益生菌对犊牛生长性能、血浆免疫和抗氧化指标及肠道健康的影响,为进一步优化犊牛饲喂管理策略、改善犊牛健康状况提供理论依据和实践指导。

1 材料与方法

1.1 试验材料

试验所用益生菌包括地衣芽孢杆菌(活菌数为1×10¹² CFU/g)、枯草芽孢杆菌(活菌数为1×10¹² CFU/g)和植物乳杆菌(活菌数为1×10¹¹ CFU/g),均由山东某生物工程有限公司提供。

1.2 动物试验伦理声明

动物试验由宁夏大学科技伦理委员会批准(批准编号:NXU-2024-145)。

1.3 试验设计

选择48头健康的新生荷斯坦犊牛,按初始体重[(36.73±1.57) kg]和胎次[(1.53±0.51)胎]相近原则随机分为4组,每组12头犊牛。对照组(CON组)饲喂牛奶+开食料,地衣芽孢杆菌组(BL组)饲喂牛奶+1 g/d地衣芽孢杆菌+开食料,枯草芽孢杆菌组(BS组)饲喂牛奶+1 g/d枯草芽孢杆菌+开食料,植物乳杆菌组(LP组)饲喂牛奶+1 g/d植物乳杆菌+开食料。试验期70 d。各组益生菌均通过牛奶加入,每日饲喂1次,在犊牛70日龄断奶并结束试验。

1.4 饲养管理

饲养试验于2024年7月至2024年10月在宁夏中宁县宁夏某牧业有限公司犊牛场进行。犊牛出生以后,从产房转至犊牛岛且在1 h之内灌服初乳4 L,以获得足够的被动免疫,24 h以后开始饲喂常乳。整个饲喂期间,采用奶量先增后减的牛奶饲喂程序:2~6日龄每天2.5 L牛奶,7~11日龄每天3.0 L牛奶,12~16日龄每天3.5 L牛奶,17~21日龄每天4.0 L牛奶,22~26日龄每天4.5 L牛奶,27~31日龄每天5.0 L牛奶,32~36日龄每天5.5 L牛奶,37~41日龄每天6.0 L牛奶,42~46日龄每天6.5 L牛奶,47~66日龄每天6.0 L牛奶,67日龄5.0 L牛奶,68日龄4.0 L牛奶,69日龄2.0 L牛奶(每天06:30和16:00各饲喂1次)。在犊牛4日龄时开始补饲开食料(购自宁夏某科技有限公司),每天记录投料量和剩余料。每隔2周收集1次开食料和牛奶样本,于-20 ℃保存待测,牛奶的营养水平和开食料组成及营养水平分别见表1和表2,其中牛奶的乳成分分析通过Ekomilk Bond牛奶分析仪(保加利亚Bulteh 2000公司)进行测定。所有犊牛单栏饲养,自由采食和饮水,并定期对犊牛岛进行清理消毒。

表1 牛奶的营养水平(鲜样)

Table 1 Nutrient levels of cow’s milk (fresh sample) %

项目Items	含量Content
乳脂Milk fat	3.82
非脂固体Non-fat solids	8.48
乳蛋白Milk protein	3.21
乳糖Lactose	4.67

表2 开食料组成及营养水平(风干基础)

Table 2 Composition and nutrient levels of the starter (air-dry basis) %

项目Items	含量Content
原料Ingredients
玉米Corn	27.50
豆粕Soybean meal	26.00
小麦麸Wheat bran	18.00
玉米干酒糟及其可溶物Corn DDGS	8.00
大豆皮Soybean hulls	15.50
预混料Premix¹⁾	5.00
合计Total	100.00
营养水平Nutrient levels²⁾
干物质DM	88.50
粗蛋白质CP	22.25
粗脂肪EE	3.50
中性洗涤纤维NDF	24.60
酸性洗涤纤维ADF	12.60
粗灰分Ash	5.36
钙Ca	0.98
磷P	0.52

1)预混料为每千克开食料提供 Premix provided the following per kg of the starter:VA 12 000 IU,VD 5 000 IU,VE 40 IU,Fe 64 mg,Cu 11 mg,Mn 42 mg,Zn 60 mg,Se 0.3 mg,I 0.8 mg,Co 0.2 mg。
2)营养水平为实测值。Nutrient levels were measured values.

1.5 样品采集及指标测定

1.5.1 开食料营养水平

开食料中的干物质含量参照GB/T 6435—2014进行测定,粗蛋白质含量参照GB/T 6432—2018进行测定,粗脂肪含量参照GB/T 6433—2006进行测定,中性洗涤纤维含量参照GB/T 20806—2022进行测定,酸性洗涤纤维含量参照NY/T 1459—2022进行测定,粗灰分含量参照 GB/T 6438—2007进行测定,钙含量参照GB/T 6436—2018进行测定,磷含量参照GB/T 6437—2018进行测定。

1.5.2 生长性能

在犊牛1、71日龄晨饲前测定犊牛的体重,每天记录每头试验犊牛开食料的投料量及剩料量,计算试验期间犊牛的平均日增重(ADG)、干物质采食量(DMI)和料重比(F/G)。其中料重比计算公式如下:

料重比=干物质采食量/平均日增重。

1.5.3 血浆指标

在犊牛7、21、70日龄晨饲前分别在颈静脉处采集血液样本20 mL置于肝素钠管,1 000×g离心15 min,取上层抗凝血浆于冻存管,然后置于液氮暂冻后在-80 ℃冰箱保存待测血浆免疫指标、抗氧化指标和肠道屏障指标。

免疫指标:采用酶联免疫吸附法测定免疫球蛋白A(IgA)、免疫球蛋白G(IgG)、白细胞介素-1β(IL-1β)、白细胞介素-2(IL-2)和白细胞介素-10(IL-10)含量。

抗氧化指标:用微板法测定总抗氧化能力(T-AOC),用羟胺法测定总超氧化物歧化酶(T-SOD)活性,用比色法测定谷胱甘肽过氧化物酶(GSH-Px)活性,用钼酸铵法测定过氧化氢酶(CAT)活性,用TBA法测定丙二醛(MDA)含量。

肠道屏障指标:采用紫外微板法测定二胺氧化酶(DAO)活性,用比色法测定D-乳酸(D-LA)含量。

1.5.4 粪便评分

在犊牛1~70日龄,由1名固定评分员根据评分标准对犊牛粪便外形进行评分,具体粪便评分标准见表3。若连续3 d评分≥3,则记为腹泻,并在评分完成后计算腹泻率。计算公式如下:

腹泻率(%)=[(腹泻头数×腹泻天数)/(总头数×试验天数)]×100^[11]。

表3 粪便评分标准

Table 3 Fecal scoring criteria

外形Shape	流动性Fluidity	评分Score
正常Normal	成型良好、稳定,但不硬	1
松软Soft	虽稍有柔软但成堆,布丁状	2
软膏Pasty	煎饼或软膏状,易扩散	3
水状Watery	固液分离,飞溅	4

1.6 数据统计与分析

试验数据中,腹泻率通过SPSS 26.0软件进行卡方检验,结果用平均值和卡方值(χ²)表示;其他试验数据通过SPSS 26.0软件进行单因素方差分析,并采用Duncan氏法进行多重比较,结果用平均值和均值标准误(SEM)表示。P<0.05表示差异显著,P>0.05表示差异不显著,P<0.01表示差异极显著,0.05≤P<0.10表示存在变化趋势。

2 结果

2.1 益生菌对哺乳犊牛生长性能的影响

由表4可知,试验犊牛初始体重组间无显著差异(P>0.05),符合试验设计要求。与对照组相比,在犊牛牛奶中添加益生菌对哺乳犊牛终末体重、平均日增重、干物质采食量和料重比均无显著影响(P>0.05)。

表4 益生菌对哺乳犊牛生长性能的影响

Table 4 Effects of probiotics on growth performance of lactating calves

项目 Items	组别Groups				均值标准误 SEM	P值 P-value
项目 Items	CON	BL	BS	LP	均值标准误 SEM	P值 P-value
初始体重IBW/kg	36.92	36.40	36.46	36.58	0.232	0.868
终末体重FBW/kg	99.13	100.86	102.63	100.13	1.078	0.720
平均日增重ADG/(kg/d)	0.89	0.92	0.95	0.90	0.015	0.641
干物质采食量DMI/(kg/d)	1.34	1.37	1.36	1.37	0.009	0.448
料重比F/G	1.58	1.61	1.53	1.60	0.031	0.788

	同行数据肩标无字母或相同字母表示差异不显著(P>0.05),不同小写字母表示差异显著(P<0.05)。下表同。
	In the same row, values with no letter or the same letter superscripts mean no significant difference (P>0.05), while with different small letter superscripts mean significant difference (P<0.05). The same as below.

2.2 益生菌对哺乳犊牛血浆免疫和抗氧化指标的影响

2.2.1 益生菌对哺乳犊牛血浆免疫指标的影响

由表5可知,与对照组相比,益生菌显著或者极显著提高7(P<0.01)、21(P<0.05)和70日龄(P<0.01)血浆IgA含量,其中,BS组和LP组7和21日龄血浆IgA含量显著提高(P<0.05),LP组70日龄血浆IgA含量显著提高(P<0.05);益生菌显著提高7日龄血浆IgG含量(P<0.05),其中,BL组7日龄血浆IgG含量显著提高(P<0.05),LP组21日龄血浆IgG含量有升高的趋势(0.05≤P<0.10);益生菌者极显著降低7和21日龄血浆IL-1β含量(P<0.01),其中,BL组和LP组7日龄血浆IL-1β含量显著降低(P<0.05),LP组21日龄血浆IL-1β含量显著降低(P<0.05),BS组70日龄血浆IL-1β含量有降低的趋势(0.05≤P<0.10);益生菌对7、21和70日龄血浆IL-2和IL-10含量没有显著影响(P>0.05),其中,BS组7日龄血浆IL-10含量有升高的趋势(0.05≤P<0.10)。

表5 益生菌对哺乳犊牛血浆免疫指标的影响

Table 5 Effects of probiotics on plasma immunity indexes of lactating calves

项目 Items	组别Groups				均值标准误 SEM	P值 P-value
项目 Items	CON	BL	BS	LP	均值标准误 SEM	P值 P-value
免疫球蛋白A IgA/(μg/mL)
7日龄7 days of age	1 111.68^c	1 253.17^bc	1 328.23^a	1 200.62^ab	23.315	0.003
21日龄21 days of age	1 243.00^b	1 274.58^ab	1 341.00^a	1 339.93^a	14.360	0.022
70日龄70 days of age	1 133.58^b	1 108.78^b	1 130.76^b	1 246.27^a	19.694	0.019
免疫球蛋白G IgG/(μg/mL)
7日龄7 days of age	37.15^b	43.47^a	39.87^ab	40.66^ab	0.779	0.019
21日龄21 days of age	37.39	41.20	39.23	43.85	0.965	0.078
70日龄70 days of age	43.50	44.43	42.82	43.73	0.775	0.769
白细胞介素-1β IL-1β/(pg/mL)
7日龄7 days of age	707.85^a	567.66^b	623.39^ab	556.72^b	18.539	0.006
21日龄21 days of age	675.63^a	587.74^ab	638.64^a	520.31^b	18.711	0.008
70日龄70 days of age	471.90	454.96	390.86	481.83	13.115	0.062
白细胞介素-2 IL-2/(pg/mL)
7日龄7 days of age	233.83	232.80	211.75	214.60	6.328	0.408
21日龄21 days of age	235.23	254.42	241.09	256.50	5.620	0.502
70日龄70 days of age	290.60	263.96	268.96	255.73	6.128	0.208
白细胞介素-10 IL-10/(pg/mL)
7日龄7 days of age	84.27	83.48	99.48	87.26	2.466	0.056
21日龄21 days of age	101.75	107.01	102.95	104.95	2.256	0.868
70日龄70 days of age	128.52	130.67	135.31	129.40	3.389	0.909

2.2.2 益生菌对哺乳犊牛血浆抗氧化指标的影响

由表6可知,与对照组相比,益生菌显著或者极显著影响7日龄血浆T-SOD活性(P<0.05)及21日龄血浆T-AOC(P<0.01)、GSH-Px活性(P<0.01)和MDA含量(P<0.05),其中,BS组和LP组21日龄血浆T-AOC显著提高(P<0.05);BS组7日龄血浆T-SOD活性显著提高(P<0.05),BS组21日龄血浆GSH-Px活性显著提高(P<0.05),BS组21日龄血浆MDA含量显著降低(P<0.05),BS组70日龄血浆MDA含量有降低的趋势(0.05≤P<0.10);益生菌对7、21和70日龄血浆CAT活性没有显著影响(P>0.05)。

表6 益生菌对哺乳犊牛血浆抗氧化指标的影响

Table 6 Effects of probiotics on plasma antioxidant indexes of lactating calves

项目 Items	组别Groups				均值标准误 SEM	P值 P-value
项目 Items	CON	BL	BS	LP	均值标准误 SEM	P值 P-value
总抗氧化能力T-AOC/(mmol/L)
7日龄7 days of age	0.42	0.47	0.44	0.46	0.035	0.966
21日龄21 days of age	0.41^c	0.53^bc	0.62^b	0.75^a	0.032	<0.001
70日龄70 days of age	0.50	0.53	0.51	0.65	0.028	0.188
总超氧化物歧化酶T-SOD/(U/mL)
7日龄7 days of age	65.92^b	67.89^b	74.34^a	71.13^ab	1.124	0.025
21日龄21 days of age	73.12	72.07	73.63	71.26	0.644	0.622
70日龄70 days of age	67.22	69.86	69.97	71.40	0.804	0.314
谷胱甘肽过氧化物酶GSH-Px/(U/mL)
7日龄7 days of age	209.01	219.14	262.48	210.47	9.140	0.144
21日龄21 days of age	114.96^b	147.98^ab	175.61^a	121.00^b	8.251	0.005
70日龄70 days of age	141.33	134.50	159.11	160.44	14.482	0.921
丙二醛MDA/(nmol/mL)
7日龄7 days of age	7.19	7.11	7.11	7.48	0.244	0.953
21日龄21 days of age	5.04^a	4.83^a	3.53^b	5.12^a	0.257	0.045
70日龄70 days of age	6.78	6.25	5.10	5.75	0.242	0.055
过氧化氢酶CAT/(U/mL)
7日龄7 days of age	38.51	24.28	36.99	37.24	2.981	0.294
21日龄21 days of age	41.44	63.03	48.63	46.11	5.045	0.502
70日龄70 days of age	19.53	23.81	20.59	23.29	1.954	0.879

2.3 益生菌对哺乳犊牛肠道健康的影响

2.3.1 益生菌对哺乳犊牛粪便评分和腹泻率的影响

由表7可知,与对照组相比,益生菌对36~70日龄、1~70日龄粪便评分及1~35日龄、36~70日龄、1~70日龄腹泻率均有极显著影响(P<0.01),其中,LP组1~35日龄腹泻率显著降低(P<0.05),且粪便评分有降低的趋势(0.05≤P<0.10);BL组、BS组和LP组36~70日龄粪便评分和腹泻率均显著降低(P<0.05),LP组1~70日龄粪便评分显著降低(P<0.05),BS组和LP组1~70日龄腹泻率显著降低(P<0.05)。

表7 益生菌对哺乳犊牛粪便评分和腹泻率的影响

Table 7 Effects of probiotics on fecal score and diarrhea rate of lactating calves

项目 Items	组别Groups				均值标准误/ 卡方值 SEM/χ²	P值 P-value
项目 Items	CON	BL	BS	LP	均值标准误/ 卡方值 SEM/χ²	P值 P-value
粪便评分Fecal score¹⁾
1~35日龄1 to 35 days of age	1.64	1.67	1.60	1.38	0.042	0.054
36~70日龄36 to 70 days of age	1.53^a	1.32^b	1.23^b	1.15^b	0.037	<0.001
1~70日龄1 to 70 days of age	1.58^a	1.49^a	1.41^ab	1.26^b	0.033	0.002
腹泻率Diarrhea rate/%²⁾
1~35日龄1 to 35 days of age	6.90^a	7.14^a	6.49^a	1.43^b	17.821	<0.001
36~70日龄36 to 70 days of age	4.05^a	0.86^b	0.00^b	0.86^b	27.753	<0.001
1~70日龄1 to 70 days of age	5.95^a	4.00^ab	3.25^b	1.07^c	29.806	<0.001

1)对应统计量为均值标准误。The corresponding statistic was SEM.
2)对应统计量为卡方值。The corresponding statistic was χ².

2.3.2 益生菌对哺乳犊牛血浆肠道屏障指标的影响

由表8可知,与对照组相比,益生菌显著降低21日龄血浆DAO活性和70日龄血浆D-LA含量(P<0.05),对7、70日龄血浆DAO活性以及7、21日龄血浆D-LA含量没有显著影响(P>0.05),其中,BL组、BS组和LP组21日龄血浆DAO活性和70日龄血浆D-LA含量均显著降低(P<0.05)。

表8 益生菌对哺乳犊牛血浆肠道屏障指标的影响

Table 8 Effects of probiotics on plasma intestinal barrier indexes of lactating calves

项目 Items	组别Groups				均值标准误 SEM	P值 P-value
项目 Items	CON组	BL组	BS组	LP组	均值标准误 SEM	P值 P-value
二胺氧化酶DAO/(ng/mL)
7日龄7 days of age	5.80	5.80	6.13	5.55	0.139	0.584
21日龄21 days of age	6.82^a	5.76^b	6.06^b	6.03^b	0.138	0.021
70日龄70 days of age	11.10	10.16	10.83	10.05	0.226	0.307
D-乳酸D-LA/(mol/L)
7日龄7 days of age	45.07	44.03	46.05	45.33	0.767	0.891
21日龄21 days of age	47.32	47.71	49.64	48.68	0.572	0.375
70日龄70 days of age	50.75^a	42.63^b	42.61^b	45.40^b	1.046	0.010

3 讨论

3.1 益生菌对哺乳犊牛生长性能的影响

哺乳阶段是犊牛生长发育的关键时期,影响犊牛未来的健康和生产力。该阶段犊牛肠道功能尚未发育完善,特别容易受到肠道疾病的影响,若在出生后最初几周感染肠道,可能会导致犊牛生长受阻、死亡风险增加,且会长期抑制其生产性能^[12]。在本试验中,各组犊牛的终末体重、平均日增重、干物质采食量及料重比均未表现出显著差异。与本试验结果一致,Zhang等^[13]研究发现,饲喂植物乳杆菌和枯草芽孢杆菌不会影响荷斯坦犊牛的干物质采食量和体重。Górka等^[14]报道,在代乳品中补充地衣芽孢杆菌不会影响犊牛的采食量、体重和平均日增重;针对羔羊的研究也显示,饲粮中添加地衣芽孢杆菌对羔羊的干物质采食量和平均日增重没有影响^[15]。此外,与单一益生菌相比,补充益生菌混合物对断奶前犊牛的生长和采食量具有更显著的促进作用^[16]。在本试验中,与对照组相比,益生菌对犊牛的增重和采食量未表现出显著影响,可能原因在于本试验各试验组仅使用了单一种益生菌进行饲喂。在本试验中,从数值来看,饲喂枯草芽孢杆菌犊牛的终末体重和平均日增重均高于其他组,表明饲喂枯草芽孢杆菌具有促进犊牛增重和提高平均日增重的潜力。这与Sun等^[17]研究发现饲喂枯草芽孢杆菌可以提高犊牛平均日增重的结论相符。此外,有研究表明,在新生犊牛的代乳品中添加4 g/d植物乳杆菌能够使犊牛体重提高11.8%,而添加8 g/d植物乳杆菌能够使犊牛平均日增重提高14.2%^[18]。本试验中,益生菌未显著改善犊牛生长性能,这可能与试验期内哺乳犊牛的基础营养水平充足、饲养环境应激较小、饲喂剂量偏低以及试验周期较短等因素有关,此推论尚需后续试验进一步证实。

3.2 益生菌对哺乳犊牛血浆免疫和抗氧化指标的影响

3.2.1 益生菌对哺乳犊牛血浆免疫指标的影响

血液免疫球蛋白和炎症因子含量是评估机体免疫功能的重要指标。在犊牛出生后最初几周,犊牛的免疫系统功能较弱,易受到病原体侵袭而引发呼吸道和肠道疾病,导致群体疾病率及死亡率升高^[19]。其中,血浆IgG和IgA含量可反映机体体液免疫功能状态^[20],而IgA是黏膜表面外分泌液中的主要抗体,在免疫保护方面中发挥关键作用^[21]。在本试验中,与对照组相比,饲喂枯草芽孢杆菌或植物乳杆菌显著提高了7、21日龄犊牛血浆IgA含量,饲喂地衣芽孢杆菌显著提高了7日龄犊牛血浆IgG含量,饲喂植物乳杆菌有提高21日龄犊牛血浆IgG含量的趋势,这些结果表明益生菌可以改善犊牛的免疫功能。相关研究显示,芽孢杆菌菌株在促进犊牛免疫力方面具有积极作用^[22]。与本次试验结果一致,在全脂牛奶中添加植物乳杆菌,可通过提高犊牛血清IgG含量改善犊牛健康状态^[16]。据报道,添加枯草芽孢杆菌的乳制品可显著提高兔子血清IgG和IgA含量^[23],说明枯草芽孢杆菌具有潜在的免疫调节作用。另外,在禽类的一项研究中发现,在补充枯草芽孢杆菌或地衣芽孢杆菌42 d后,肉鸡血清IgA和IgG[其禽类对应物免疫球蛋白Y(IgY)]含量显著提高,表明肉鸡的免疫功能得到了改善^[24]。

细胞因子包括促炎因子(如IL-1β和IL-2)和抗炎因子(如IL-10),其在启动、调节和终止炎症反应方面发挥着重要作用,一方面保护机体免受感染和损伤,另一方面防止过度炎症引起的组织损伤^[25]。作为炎症介质,这类细胞因子能激活中性粒细胞和巨噬细胞,并上调内皮细胞表面细胞黏附分子的表达^[26]。其中,IL-1β可诱导自身表达,并能促进IL-6、IL-8和TNF-α等促炎细胞因子、黏附分子和趋化因子的表达,进而引发炎症反应与中性粒细胞和巨噬细胞等抗菌细胞的募集^[27]。在本试验中,与对照组相比,饲喂地衣芽胞杆菌或植物乳杆菌显著降低了7日龄犊牛血浆IL-1β含量,饲喂植物乳杆菌显著降低了21日龄犊牛血浆IL-1β含量,饲喂枯草芽孢杆菌有提高7日龄犊牛血浆IL-10含量的趋势,这些结果表明益生菌可能减轻犊牛炎症反应,防止体内组织损伤。本试验结果与前人报道一致,包括多种芽孢杆菌的益生菌能够促进奶犊牛体内IL-10的产生^[17,28]。Xu等^[24]研究发现,补充枯草芽孢杆菌或地衣芽孢杆菌能降低肉鸡血清IL-1β含量并提高IL-10含量。

3.2.2 益生菌对哺乳犊牛血浆抗氧化指标的影响

新生犊牛抗氧化系统在出生时尚未完全发育,这使其易出现严重氧化应激。根据定义,氧化应激是氧化剂和抗氧化剂之间平衡失调的结果,该失调会造成氧化还原信号传导及调控功能紊乱,或引发分子损伤^[29]。其中,氧化剂主要为活性氧(ROS),包括过氧化氢(H₂O₂)、羟基自由基(·OH)和超氧阴离子(

${O}_{2}^{-}$

·),这类物质主要通过细胞的有氧代谢等活动产生^[30]。ROS过度积累会导致氧化应激,从而破坏氧化还原稳态,氧化还原稳态失衡不仅对细胞产生毒性作用,还会破坏生物分子并生成氧化产物,如MDA和8-羟基-2’-脱氧鸟苷(8-OHDG)^[31]。MDA是脂质氧化过程的特异性产物,其含量反映机体脂质过氧化程度和细胞损伤程度^[32]。T-AOC反映机体的抗氧化防御潜力,其由许多抗氧化化合物及酶类共同构成。SOD、GSH-Px、CAT是体内广泛分布的酶促抗氧化剂,可通过清除自由基发挥保护作用。SOD和GSH-Px可以消除超氧化物和脂质氢过氧化物,减少有机氢过氧化物对机体的损害^[20]。CAT则可催化H₂O₂分解为分子氧和水,实现体内的H₂O₂的有效清除。在本试验中,与对照组相比,饲喂枯草芽孢杆菌或植物乳杆菌显著提高了21日龄犊牛血浆T-AOC;饲喂枯草芽孢杆菌显著提高了7日龄犊牛血浆T-SOD活性和21日龄犊牛血浆GSH-Px活性,降低21日龄犊牛血浆MDA含量,并有降低70日龄犊牛血浆MDA含量的趋势,这些结果表明,枯草芽孢杆菌和植物乳杆菌可提高犊牛抗氧化能力,且枯草芽孢杆菌有降低犊牛血浆中MDA含量的潜力,有助于保护细胞免受氧化损伤,从而保证犊牛的健康。与本试验结果一致,有研究报道,饲粮中添加枯草芽孢杆菌或地衣芽孢杆菌的肉鸡具有更强的抗氧化能力^[24];还有研究表明,在牛奶中添加植物乳杆菌犊牛能提高犊牛抗氧化能力^[33],原因可能是乳酸菌通过释放氢离子或氢电子,能够中断自由基的链式反应,从而发挥抗氧化作用^[34]。

3.3 益生菌对哺乳犊牛肠道健康的影响

3.3.1 益生菌对哺乳犊牛粪便评分和腹泻率的影响

粪便评分系统通常用于预测幼年动物的腹泻研究^[35]。腹泻是造成犊牛死亡的主要原因之一,会给乳品和养牛业带来高昂的治疗费用和巨额经济损失。此外,腹泻通常发生在犊牛出生后2周左右。在本试验中,与对照组相比,益生菌对36~70日龄、1~70日龄粪便评分及1~35日龄、36~70日龄、1~70日龄腹泻率均有极显著影响,饲喂植物乳杆菌显著降低了1~35日龄犊牛腹泻率,饲喂地衣芽孢杆菌、枯草芽孢杆菌或植物乳杆菌均显著降低了36~70日龄犊牛粪便评分和腹泻率,饲喂枯草芽孢杆菌或植物乳杆菌显著降低了1~70日龄犊牛腹泻率,这些结果表明地衣芽孢杆菌、枯草芽孢杆菌和植物乳杆菌能改善犊牛肠道健康。与本试验结果一致,已有相关研究表明,饲喂植物乳杆菌有助于改善犊牛的肠道健康状况^[18];在牛奶中添加植物乳杆菌可降低断奶前犊牛的腹泻率^[33];断奶前仔猪补充地衣芽孢杆菌和枯草芽孢杆菌可降低仔猪腹泻率^[36]。

3.3.2 益生菌对哺乳犊牛血浆肠道屏障指标的影响

肠道屏障作为复杂的防御系统,是机体抵御病原体入侵的第1道防线,包括机械屏障、化学屏障、微生物屏障及免疫屏障^[37]。肠道黏膜环境的稳定性在很大程度上取决于肠道屏障的完整性。在炎症性肠病中,肠道通透性是判断胃肠道处于正常或病理状态的关键指标^[38]。D-LA和DAO可通过受损的肠黏膜进入血液循环,二者已成为评价肠黏膜屏障功能的重要标志物^[39]。血浆D-LA含量升高可反映肠黏膜通透性增加^[40],因此其可能作为揭示肠黏膜的损伤程度的指标。DAO活性是用于评估肠道机械屏障完整性和黏膜绒毛损伤程度^[41],故血浆DAO活性可反映肠腔的损伤和恢复情况。血浆D-LA含量与DAO活性的升高可反映肠道屏障功能的异常或损伤^[42]。在本试验中,与对照组相比,饲喂地衣芽孢杆菌、枯草芽孢杆菌或植物乳杆菌的21日龄犊牛血浆DAO活性和70日龄血浆D-LA含量均显著降低,与本试验结果一致,Yun等^[43]发现,饲粮中添加枯草芽孢杆菌可降低哺乳仔猪的血浆DAO活性。由此可见,地衣芽孢杆菌、枯草芽孢杆菌和植物乳杆菌可能通过维持犊牛肠道屏障完整性,降低肠道黏膜绒毛损伤程度,从而达到缓解犊牛腹泻的效果。

4 结论

在本试验条件下,通过在牛奶中添加地衣芽孢杆菌、枯草芽孢杆菌和植物乳杆菌虽未改善哺乳犊牛生长性能,但可增强其免疫功能与抗氧化能力,同时降低腹泻率并改善肠道屏障功能,从而促进肠道健康。

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