柔嫩艾美耳球虫RON2L2ecto蛋白的原核表达及其免疫效果评价

doi:10.11843/j.issn.0366-6964.2025.11.024

Abstract

Abstract:

The aim of this study is to investigate the immune protection effect of recombinant Eimeria tenella RON2_L2ecto protein on chicks. The full-length sequence encoding EtRON2_L2 protein from Eimeria tenella cDNA was amplified and the extracellular domain of EtRON2_L2 was expressed through prokaryutic recombination in this study. A hundred and twenty 7-day-old healthy Roman Grey male chicks with similar body weight (raised in clean and coccidia-free environment, fed with diet without coccidiostats or artibiotic) were randomly allotted into 5 groups, including negative control group (without immune or coccidia infection), positive control group (without immune but infected with coccida), and 50 μg, 100 μg, 200 μg recombinant protein immune groups. Recombinant protein was mixed with Freund's adjuvant as a subunit vaccine to immunize chicks in three immune groups. The immune protection effect of recombinant protein was evaluated by measuring the body weight gain, cecal lesion score, number of oocysts per gram of feces (OPG), calculating the anticoccidial index, observing the histopathological changes in the cecum, detecting serum cytokines (IL-2, IL-10, IFN-γ) and serum IgG, and the secreting levels of cecal mucosal sIgA. The results showed that the extracellular domain of EtRON2_L2 was successfully expressed, with a size of 107 ku. The immune protection experiment indicated that compared with the non-immunized group, the average body weight gain of the 100 μg protein immunization group and the 200 μg protein immunization group significantly increased (P < 0.05), while the OPG significantly decreased (P < 0.05), the cecal lesion scores of each dose of immune group showed a significant decrease (P < 0.05), pathological histology observation shows that the degree of cecal lesions has been alleviated to some extent. The anti-coccidiosis index of the 100 μg protein immunization group and the 200 μg protein immunization group is greater than 160, indicating moderate anti-coccidiosis activity. The spleen index of three protein immune groups were also significantly increased (P < 0.05). At 28 days of age (the 7th day after infection), the serum cytokines IL-2, IFN-γ, and IL-10 in the protein immune groups were significantly higher (P < 0.01) than those in the two control groups, and the serum IgG levels were significantly higher than those in the negative control group (P < 0.05). The sIgA secretion levels in the 50 μg protein immune group were significantly higher (P < 0.05) than those in the other immune groups and the two control groups. The above results indicate that the extracellular domain of RON2_L2 has an immunoprotective effect on chicks infected with Eimeria tenella.

Key words: Eimeria tenella, moving junction, RON2_L2, immune protection effect

CLC Number:

S852.723

SHAN Yige, LIU Yongning, ZHOU Shuang, LIU Qianlin, LI Yilin, DUAN Sizhang, AN Jian, ZHANG Jianjun. Prokaryotic Expression and Immune Efficacy Evaluation of RON2_L2ecto Protein from Eimeria tenella[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(11): 5649-5659.

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References 25

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组别 Group	平均体增重/g Average weight gain	平均OPG/(×10⁵·g^-1) Average OPG
不免疫不攻虫组 Negative control group	65.29±1.32^a	0^a
不免疫攻虫组 Positive control group	42.58±1.11^b	7.04±0.26^b
50 μg蛋白免疫组 50 μg rEtRON2_L2ecto immune group	41.87±0.75^b	4.01±0.27^c
100 μg蛋白免疫组 100 μg rEtRON2_L2ecto immune group	47.02±0.86^c	0.40±0.19^d
200 μg蛋白免疫组 200 μg rEtRON2_L2ecto immune group	46.83±1.26^c	0.24±0.06^d

组别 Group	盲肠病变评分 Score of cecal lesions
不免疫不攻虫组 Negative control group	0^a
不免疫攻虫组 Positive control group	1.5±0.53^b
50 μg蛋白免疫组 50 μg rEtRON2_L2ecto immune group	0.62±0.74^ac
100 μg蛋白免疫组 100 μg rEtRON2_L2ecto immune group	0.25±0.46^ac
200 μg蛋白免疫组 200 μg rEtRON2_L2ecto immune group	0.25±0.46^ac

组别 Group	存活率/% Survival rate	相对增重率/% Relative weight gain rate	病变值 Lesion value	卵囊值 Oocyst value	抗球虫指数 ACI
不免疫不攻虫组 Negative control group	100	100.0	0	0	200
不免疫攻虫组 Positive control group	100	65.2	15	40	110.2
50 μg蛋白免疫组 50 μg rEtRON2_L2ecto immune group	100	64.1	6.2	20	137.9
100 μg蛋白免疫组 100 μg rEtRON2_L2ecto immune group	100	72.0	2.5	5	164.5
200 μg蛋白免疫组 200 μg rEtRON2_L2ecto immune group	100	71.7	2.5	5	164.2

组别 Group	脾脏指数 Spleen index	法氏囊指数 Bursa of Fabricius index
不免疫不攻虫组 Negative control group	21.8±1.0^a	52.6±2.2^a
不免疫攻虫组 Positive control group	24.6±2.6^a	44.5±1.1^b
50 μg蛋白免疫组 50 μg rEtRON2_L2ecto immune group	29.7±1.5^b	31.0±1.9^c
100 μg蛋白免疫组 100 μg rEtRON2_L2ecto immune group	31.3±4.5^b	24.7±4.0^d
200 μg蛋白免疫组 200 μg rEtRON2_L2ecto immune group	33.8.±2.2^b	30.5±1.3^c

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Prokaryotic Expression and Immune Efficacy Evaluation of RON2_L2ecto Protein from Eimeria tenella

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Prokaryotic Expression and Immune Efficacy Evaluation of RON2_L2ecto Protein from Eimeria tenella