通过定量聚合酶链反应评估亚种在牛子宫内膜细胞中的细胞内存活情况。
Evaluation of intracellular survival of subsp. in bovine endometrial cells by qPCR.
作者信息
Campos Muzquiz L G, Martínez Gómez D, Reyes Cruz T, Méndez Olvera E T
机构信息
Department of Genetics and Virology, Faculty of Chemical Sciences, Autonomous University of Coahuila, Saltillo, Mexico.
Laboratory of Agricultural Microbiology, Department of Agricultural and Animal Production, Metropolitan Autonomous University-Xochimilco, Mexico City, Mexico.
出版信息
Iran J Vet Res. 2021 Spring;22(2):94-99. doi: 10.22099/ijvr.2021.38693.5632.
BACKGROUND
subsp. is the causal agent of sporadic abortion and infertility in bovines that produces economic losses in livestock
AIMS
This study evaluates the capability of subsp. to invade and survive in bovine endometrial epithelial cells and attempts to describe a pathogenic mechanism of this microorganism.
METHODS
Primary culture of bovine endometrial epithelial cells was challenged with subsp. . Intracellular bacteria, represented by the number of genomic copies (g.c.) were quantified at 0, 2, 4, 10, and 24 hours post-infection (h.p.i.), by quantitative polymerase chain reaction (qPCR). The presence of intracellular bacteria was evaluated by immunofluorescence and immunohistochemistry.
RESULTS
The results showed that only viable subsp. could invade endometrial cells. The g.c. number in assays with viable subsp. reached an average value of 656 g.c., remained constant until 4 h.p.i., then decreased to 100 g.c, at 24 h.p.i. In assays with non-viable microorganisms, the average value of g.c. was less than 1 g.c. and never changed. The intracellular presence of this bacteria was confirmed at 2 h.p.i. by immunofluorescence and immunohistochemistry.
CONCLUSION
The results suggest that only subsp. viable can invade bovine endometrial epithelial cells but will not replicate in them, indicating that the endometrial cells do not represent a replication niche for this pathogen. Nonetheless, this invasion capability suggests that this type of cell could be employed by the pathogen to spread to other tissues.
背景
亚种是牛散发性流产和不育的病原体，会给畜牧业造成经济损失。
目的
本研究评估亚种侵入牛子宫内膜上皮细胞并在其中存活的能力，并试图描述这种微生物的致病机制。
方法
用亚种对牛子宫内膜上皮细胞原代培养物进行攻击。在感染后0、2、4、10和24小时，通过定量聚合酶链反应（qPCR）对以基因组拷贝数（g.c.）表示的细胞内细菌进行定量。通过免疫荧光和免疫组织化学评估细胞内细菌的存在情况。
结果
结果表明，只有活的亚种能够侵入子宫内膜细胞。在使用活的亚种进行的试验中，g.c.数达到平均656 g.c.，在4 h.p.i.之前保持恒定，然后在24 h.p.i.时降至100 g.c.。在使用非活微生物进行的试验中，g.c.的平均值小于1 g.c.且从未改变。通过免疫荧光和免疫组织化学在2 h.p.i.时证实了这种细菌在细胞内的存在。
结论
结果表明，只有活的亚种能够侵入牛子宫内膜上皮细胞，但不会在其中复制，这表明子宫内膜细胞不是这种病原体的复制位点。尽管如此，这种侵入能力表明病原体可能利用这种类型的细胞扩散到其他组织。
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