Application of a Dot Blot Hybridization Platform to Assess Streptococcus uberis Population Structure in Dairy Herds.
Application of a Dot Blot Hybridization Platform to Assess Streptococcus uberis Population Structure in Dairy Herds.
复制标题
DOT印迹杂交平台的应用来评估Uberis链球菌种群结构。
DOI：
10.3389/fmicb.2017.00054
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发表时间：
2017
影响因子：
5.2
通讯作者：
Tavares F
中科院分区：
文献类型：
--
作者：
Albuquerque P;Ribeiro N;Almeida A;Panschin I;Porfirio A;Vales M;Diniz F;Madeira H;Tavares F
Uberis链球菌被认为是与牛乳房炎相关的最重要的病原体之一。虽然传统上被认为是一种环境病原体，但Uberis已经被证明在几个奶牛群中采用了传染性流行病学模式。由于不同的传播方式采取了不同的控制策略，因此对Uberis种群的深入研究对于确定控制这种病原体的最佳做法是必不可少的。在这项工作中，我们优化和验证了一个斑点杂交平台，结合自动图像分析，快速评估感染性Uberis的种群结构，并评估其与多位点序列分析(MLSA)基因分型的效率。对两个感染Uberis的奶牛群进行了为期6个月的跟踪调查，以收集和鉴定持续感染奶牛的分离株。这些位于葡萄牙(巴塞罗那和马亚地区)的牛群有类似的管理做法，来自巴塞罗那的牛群规模较小，挤奶厅管理更好，因为感染的奶牛是立即隔离的。从两个牛群的24头不同奶牛身上共分离到54株Uberis菌株。为了克服对斑点杂交分析的操作者依赖性，提高技术的一致性和可靠性，杂交信号被转换为概率值，平均概率大于0.5被认为是阳性结果。这些数据允许使用分类群特定的标记来确认分离株作为S.uberis的身份，并确定与毒力和抗生素耐药性相关的基因的存在。此外，MLSA允许披露两个牛群中最普遍的Uberis克隆谱系。确定了七个不同的集群，其中巴塞罗那表现出高度的克隆多样性，而Maia是感染大多数奶牛的优势谱系，这表明了不同的流行病学模式，而S.uberis根据牛群表现出一种环境或传染性的传播模式。总体而言，这项工作显示了斑点杂交和MLSA在表征引起乳房炎的Uberis的种群结构和流行病学模式方面的有效性。这种方法能够揭示Uberis在两个不同群体中的流行毒力模式和克隆谱系，并获得关于群体管理做法对病原菌种群结构的影响的见解。
Streptococcus uberis is considered one of the most important pathogens associated with bovine mastitis. While traditionally acknowledged as an environmental pathogen, S. uberis has been shown to adopt a contagious epidemiological pattern in several dairy herds. Since different control strategies are employed depending on the mode of transmission, in-depth studies of S. uberis populations are essential to determine the best practices to control this pathogen. In this work, we optimized and validated a dot blot platform, combined with automatic image analysis, to rapidly assess the population structure of infective S. uberis, and evaluated its efficiency when compared to multilocus sequence analysis (MLSA) genotyping. Two dairy herds with prevalent S. uberis infections were followed in a 6 month period, in order to collect and characterize isolates from cows with persistent infections. These herds, located in Portugal (Barcelos and Maia regions), had similar management practices, with the herd from Barcelos being smaller and having a better milking parlor management, since infected cow segregation was immediate. A total of 54 S. uberis isolates were obtained from 24 different cows from the two herds. To overcome operator-dependent analysis of the dot blots and increase the technique's consistency and reliability, the hybridization signals were converted into probability values, with average probabilities higher than 0.5 being considered positive results. These data allowed to confirm the isolates' identity as S. uberis using taxa-specific markers and to determine the presence of virulence- and antibiotic resistance-related genes. In addition, MLSA allowed to disclose the most prevalent S. uberis clonal lineages in both herds. Seven different clusters were identified, with Barcelos showing a high clonal diversity and Maia a dominant lineage infecting most cows, suggesting distinct epidemiological patterns, with S. uberis displaying an environmental or contagious transmission pattern depending on the herd. Overall, this work showed the utility of dot blot and MLSA to characterize population structure and epidemiological patterns of mastitis-causing S. uberis. This approach allowed to disclose prevalent virulence patterns and clonal lineages of S. uberis in two distinct herds, and gain insights on the impact of herd management practices on pathogen population structure.