更好地了解牛乳腺炎背后的分子机制是改善这种疾病管理的基础,这仍然是乳制品行业的主要关注点,尤其是亚临床形式。疾病的严重程度和进展取决于许多方面,比如家畜遗传学,以及致病因素之间的相互作用,主机,和环境。在这种情况下,表观遗传机制已被证明在控制动物对炎症的反应中起作用。因此,在这项研究中,我们旨在探索健康奶牛(n=15)和受链球菌自然发生的亚临床乳腺炎(Strep.)无乳(n=12)和无乳球菌属。(n=11),为了更好地了解体细胞甲基化在宿主对疾病的反应中的作用。比较评估差异甲基化区域(DMR):i)Strep。无乳感染与健康的;ii)原藻感染与健康,和iii)乳腺炎vs.健康和iv)链球菌。无乳感染与原藻感染。在2个水平下进行功能分析。首先,我们从启动子-DMR中提取差异甲基化基因(DMG),使用CytoscapeClueGO插件进行了分析。再加上这种DMG驱动的方法,所有与启动子甲基化区域相关的基因都被送入Pathifier算法。根据DMR分析,我们在Strep中鉴定出1,081个高甲基化和361个低甲基化启动子区.无乳感染的动物,虽然1,514个高甲基化和358个低甲基化的启动子区域在无绿藻感染的动物中被鉴定,与健康对照相比。当将受感染的动物视为一个整体(无论病原体如何)时,我们发现了1,576个高甲基化和460个低甲基化的启动子区域。两种病原体都与减数分裂相关通路基因的甲基化差异有关。繁殖和组织重塑。探索整个甲基化,在亚临床感染的奶牛中,我们观察到免疫相关途径的强烈失调,如核因子kB和toll样受体信号通路,以及与能量相关的途径,例如三羧酸循环和不饱和脂肪酸的生物合成。总之,本研究未检测到明显的病原体特异性SC甲基化组特征.总的来说,我们观察到亚临床乳腺炎时DNA甲基化对宿主免疫应答的明显调节.需要对更大的动物群体进行进一步的研究,以验证我们的结果,并可能确定一个独特的SC甲基化组,表明病原体特异性改变。
Better understanding of the molecular mechanisms behind bovine mastitis is fundamental for improving the management of this disease, which continues to be of major concern for the dairy industry, especially in its subclinical form. Disease severity and progression depend on numerous aspects, such as livestock genetics, and the interaction between the causative agent, the host, and the environment. In this context, epigenetic mechanisms have proven to have a role in controlling the response of the animal to inflammation. Therefore, in this study we aimed to explore genome-wide DNA methylation of milk somatic cells (SC) in healthy cows (n = 15) and cows affected by naturally occurring subclinical mastitis by Streptococcus agalactiae (n = 12) and
Prototheca spp. (n = 11), to better understand the role of SC methylome in the host response to disease. Differentially methylated regions (DMR) were evaluated comparing: (1) Strep. agalactiae-infected versus healthy; (2)
Prototheca-infected versus healthy, and (3) mastitis versus healthy and (4) Strep. agalactiae-infected versus
Prototheca-infected. The functional analysis was performed at 2 levels. To begin with, we extracted differentially methylated genes (DMG) from promoter DMR, which were analyzed using the Cytoscape ClueGO plug-in. Coupled with this DMG-driven approach, all the genes associated with promoter-methylated regions were fed to the Pathifier algorithm. From the DMR analysis, we identified 1,081 hypermethylated and 361 hypomethylated promoter regions in Strep. agalactiae-infected animals, while 1,514 hypermethylated and 358 hypomethylated promoter regions were identified in
Prototheca-infected animals, when compared with the healthy controls. When considering infected animals as a whole group (regardless of the pathogen), we found 1,576 hypermethylated and 460 hypomethylated promoter regions. Both pathogens were associated with methylation differences in genes involved in pathways related to meiosis, reproduction and tissue remodeling. Exploring the whole methylome, in subclinically infected cows we observed a strong deregulation of immune-related pathways, such as nuclear factor kB and toll-like receptors signaling pathways, and of energy-related pathways such as the tricarboxylic acid cycle and unsaturated fatty acid biosynthesis. In conclusion, no evident pathogen-specific SC methylome signature was detected in the present study. Overall, we observed a clear regulation of host immune response driven by DNA methylation upon subclinical mastitis. Further studies on a larger cohort of animals are needed to validate our results and to possibly identify a unique SC methylome that signifies pathogen-specific alterations.