Bicalho M L S, Machado V S, Higgins C H, Lima F S, Bicalho R C
Department of Population Medicine and Diagnostic Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853.
Department of Population Medicine and Diagnostic Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853.
J Dairy Sci. 2017 May;100(5):3850-3862. doi: 10.3168/jds.2016-12058. Epub 2017 Mar 2.
Metritis is a uterine disease that affects 10 to 30% of all lactating dairy cows and has detrimental effects on reproductive performance, milk production, and survival. Data regarding the identity and abundance of bacterial genes governing traits such as virulence, antibiotic resistance, and stress responses could enable identification of previously unknown agents that play a role in metritis pathogenesis. Moreover, such knowledge could lead to the development of improved treatments or preventive methods. Therefore, the objectives of this study were to characterize the uterine microbial population and to differentiate, for the first time, the microbial functional diversity in cows with metritis versus healthy cows. In addition, we aimed to identify relationships between microbial genes and postpartum uterine health. Uterine swabs were collected from 24 cows within 3 to 12 d in milk; 12 cows were diagnosed with metritis and the other 12 were healthy. Metritis was defined as a watery, reddish or brownish uterine discharge having a fetid smell, and rectal temperature greater than 39.5°C. Cows with a clear and viscous uterine discharge, not fetid or mucopurulent, were classified as healthy. Microbial metagenomic DNA from uterine swab samples was subjected to whole-genome shotgun sequencing on the Illumina MiSeq platform (Illumina Inc., San Diego, CA). The MG-RAST server (metagenomic rapid annotations using subsystems technology; http://metagenomics.anl.gov/) and STAMP software (http://kiwi.cs.dal.ca/Software/STAMP) were used to detect statistically significant differences in the abundance of taxonomic and functional features between the uterine microbial metagenomes of metritic and healthy cows. Our results showed an increased abundance of Fusobacteria and Bacteroidetes in metritic cows, confirming the potential role of those 2 taxa in the pathogenesis of metritis. The MG-RAST analysis revealed a significantly higher abundance of genes for protein transport across the cytoplasmic membrane and type VI bacterial secretion systems in the metritic microbiota. Additionally, genes coding for resistance to acid stress were exclusive to the metritis microbiota, suggesting that microbial resistance to acid stress is important for microbial survival in the infected uterus. On the other hand, genes coding for adhesion molecules, bacteriocins, and antibacterial peptides were significantly associated with the uterine microbiota of healthy cows, as was tolerance to colicin E2.
子宫炎是一种影响10%至30%泌乳奶牛的子宫疾病，对繁殖性能、产奶量和存活率有不利影响。关于控制毒力、抗生素抗性和应激反应等性状的细菌基因的身份和丰度的数据，能够帮助识别在子宫炎发病机制中起作用的先前未知的病原体。此外，这些知识可能会带来改进的治疗方法或预防措施。因此，本研究的目的是对子宫微生物群落进行表征，并首次区分子宫炎奶牛与健康奶牛的微生物功能多样性。此外，我们旨在确定微生物基因与产后子宫健康之间的关系。在产奶后3至12天内，从24头奶牛采集子宫拭子；12头奶牛被诊断为子宫炎，另外12头健康。子宫炎的定义为水样、微红或微褐色的子宫分泌物，有恶臭气味，直肠温度高于39.5°C。子宫分泌物清澈且粘稠、无恶臭或粘液脓性的奶牛被归类为健康。对子宫拭子样本的微生物宏基因组DNA在Illumina MiSeq平台（Illumina公司，加利福尼亚州圣地亚哥）上进行全基因组鸟枪法测序。使用MG-RAST服务器（使用子系统技术的宏基因组快速注释；http://metagenomics.anl.gov/）和STAMP软件（http://kiwi.cs.dal.ca/Software/STAMP）来检测子宫炎奶牛和健康奶牛的子宫微生物宏基因组在分类和功能特征丰度上的统计学显著差异。我们的结果显示，子宫炎奶牛中梭杆菌属和拟杆菌属的丰度增加，证实了这两个分类群在子宫炎发病机制中的潜在作用。MG-RAST分析显示，子宫炎微生物群中跨细胞质膜的蛋白质转运基因和VI型细菌分泌系统的丰度显著更高。此外，编码对酸应激抗性的基因是子宫炎微生物群所特有的，这表明微生物对酸应激的抗性对于受感染子宫中的微生物存活很重要。另一方面，编码粘附分子、细菌素和抗菌肽的基因与健康奶牛的子宫微生物群显著相关，对大肠杆菌素E2的耐受性也是如此。
