弓形虫PRU株速殖子感染小鼠产生包囊的试验研究

doi:10.11843/j.issn.0366-6964.2024.10.033

摘要/Abstract

摘要：

旨在获得弓形虫包囊，并对包囊传代保种，将其作为弓形虫慢性感染相关试验研究的感染性材料。本研究用四组不同数量的PRU株速殖子腹腔接种KM、ICR、BALB/c、C57BL/6四个品系小鼠，感染40 d后检测存活小鼠血清中抗体，取血清阳性小鼠各组织检查并分离包囊；以包囊灌胃感染不同品系小鼠，探究包囊传代剂量及最佳的传代小鼠品系。结果成功在感染低剂量的C57BL/6小鼠脑组织内分离得到包囊。包囊传代ICR小鼠死亡率低，且脑内成囊数显著多于其他品系小鼠(P < 0.05)。C57BL/6小鼠腹腔接种10~50个弓形虫速殖子可存活并在脑内形成包囊；以20~50个弓形虫包囊灌胃感染ICR小鼠，可扩增得到大量包囊。

关键词: PRU株, 速殖子, 包囊, 慢性感染, 包囊传代, 小鼠模型

Abstract:

This experiment was designed to obtain Toxoplasma cysts, which are often used as infectious materials in experimental studies related to chronic infection of Toxoplasma gondii, and to search for the most suitable mouse model for cysts passage. In this study, mice with KM, ICR, BALB/c and C57BL/6 strains were intraperitoneally inoculated with different numbers of tachyzoites of PRU strains. After 40 days of infection, antibodies in the serum of surviving mice were detected, and cysts were examined and isolated from the tissues of seropositive mice. Different strains of mice were infected by cyst intragastric administration to explore the cysts passage dose and the best passage mouse model. We successfully isolated cysts from the brains of C57BL/6 mice infected with low doses of Toxoplasma tachyzoites. When passing cysts, ICR mice had low mortality and significantly more cysts in the brain than other strains (P < 0.05). C57BL/6 mice can survive and form cysts in the brain when 10-50 tachyzoites are intraperitoneally inoculated; A larger number of toxoplasma cysts could be obtained by intragastric infecting ICR mice with 20-50 cysts.

Key words: PRU strain, tachyzoite, cysts, chronic infection, cyst passage, mouse model

中图分类号:

S852.72

李瑞芳, 张曼玉, 孙卿, 杜晶莹, 蒋蔚, 李增强, 夏炉明, 王权. 弓形虫PRU株速殖子感染小鼠产生包囊的试验研究[J]. 畜牧兽医学报, 2024, 55(10): 4620-4629.

Ruifang LI, Manyu ZHANG, Qing SUN, Jingying DU, Wei JIANG, Zengqiang LI, Luming XIA, Quan WANG. Experimental Study on the Production of Cysts in Mice Infected with Toxoplasma gondii Tachyzoites of PRU Strain[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(10): 4620-4629.

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目镜视场(FN) field of view	18	20	22	25	26.5
40×物镜下试剂视场面积/mm²	0.16	0.20	0.24	0.31	0.34
Reagent field area under 40×objective lens

分组 Group	小鼠品系 Mouse strain	接种速殖子数量/个 Number of tachyzoites	重复数/只 Number of replicates
K1	KM小鼠	500	6
K2	KM mice	200	6
K3		50	6
K4		10	6
I1	ICR小鼠	500	6
I2	ICR mice	200	6
I3		50	6
I4		10	6
B1	BALB/c小鼠	500	6
B2	BALB/c mice	200	6
B3		50	6
B4		10	6
C1	C57BL/6小鼠	500	6
C2	C57BL/6 mice	200	6
C3		50	6
C4		10	6

引物名称 Primer name	引物序列(5′→3′) Sequence(5′→3′)
外引物前	ACCTTTGAATCCCAAGCA
外引物后	AAGGTGCCATTTGCGTTC
内引物前	TTTGCATTCAAGAAGCGTG
内引物后	AAGGTGCCATTTGCGTTC

程序 Procedure	温度 Temperature	反应时间 Time	循环数 Cycle number
预变性	95℃	5 min	-
变性	95 ℃	50 s	$ \left. \begin{array}{l}\\\\\end{array} \right\}30 $
退火	56 ℃	40 s
延伸	72 ℃	45 s
最终延伸	72 ℃	10 min	-

小鼠编号 ID	450 nm吸光值 OD_{450 nm}	样本值/阴性值 S/N
1	3.138 0	29.600
2	2.978 0	28.094
3	1.806 0	17.037
4	1.041 0	9.821
5	1.179 8	11.130
6	1.461 2	13.785
7	1.661 7	15.676
8	2.162 9	20.405
9	2.261 9	21.339
10	0.889 4	8.391
11	1.183 7	11.117
12	1.134 7	10.705
阴性对照	0.106 0