Aravena Pamela, Pulgar Rodrigo, Ortiz-Severín Javiera, Maza Felipe, Gaete Alexis, Martínez Sebastián, Serón Ervin, González Mauricio, Cambiazo Verónica
Laboratorio de Bioinformática y Expresión Génica, Instituto de Nutrición y Tecnología de los Alimentos (INTA), Universidad de Chile, Santiago 7830490, Chile.
FONDAP Center for Genome Regulation, Santiago 8370415, Chile.
Pathogens. 2020 May 8;9(5):358. doi: 10.3390/pathogens9050358.
, the causative agent of piscirickettsiosis, is genetically divided into two genomic groups, named after the reference strains as LF-89-like or EM-90-like. Phenotypic differences have been detected between the genogroups, including antibiotic susceptibilities, host specificities and pathogenicity. In this study, we aimed to develop a rapid, sensitive and cost-effective assay for the differentiation of the genogroups. Using an in silico analysis of the 16S rDNA digestion patterns, we have designed a genogroup-specific assay based on PCR-restriction fragment length polymorphism (RFLP). An experimental validation was carried out by comparing the restriction patterns of 13 strains and 57 field samples obtained from the tissues of dead or moribund fish. When the bacterial composition of a set of field samples, for which we detected mixtures of bacterial DNA, was analyzed by a high-throughput sequencing of the 16S rRNA gene amplicons, a diversity of taxa could be identified, including pathogenic and commensal bacteria. Despite the presence of mixtures of bacterial DNA, the characteristic digestion pattern of the genogroups could be detected in the field samples without the need of a microbiological culture and bacterial isolation.
鱼类立克次氏体病的病原体，在基因上分为两个基因组群，以参考菌株命名为LF - 89样或EM - 90样。已在基因组群之间检测到表型差异，包括抗生素敏感性、宿主特异性和致病性。在本研究中，我们旨在开发一种快速、灵敏且经济高效的方法来区分基因组群。通过对16S rDNA消化模式的电子分析，我们设计了一种基于PCR - 限制性片段长度多态性（RFLP）的基因组群特异性检测方法。通过比较从死亡或濒死鱼类组织中获得的13株菌株和57份现场样本的限制性图谱进行了实验验证。当通过对16S rRNA基因扩增子进行高通量测序来分析一组现场样本的细菌组成时（我们检测到这些样本中存在细菌DNA混合物），可以鉴定出多种分类群，包括致病性和共生细菌。尽管存在细菌DNA混合物，但无需微生物培养和细菌分离即可在现场样本中检测到基因组群的特征性消化模式。
