The aim of our study was to assess the sequencing of unique nucH gene fragment based on performed bioinformatics analysis as a novel diagnostic method for the identification of difficult to identify staphylococcal human pathogenic strains. Initially, PCR-RFLP-rrn analysis specific to the spacers between 16SrDNA and 23SrDNA followed by HhaI restriction analysis was performed. Further, sequencing of nucH and 16S rDNA genes fragments was carried out. Blast analysis from the NCBI showed 99% similarity of nucH gene fragment with reference genomic DNA for S. succinus with the accession no. CP018199. This result was also confirmed by MALDI-TOF analysis. Sequencing analysis of 16S rDNA gene fragment allowed for 100% identification of two strains isolated from human samples as Staphylococus succinus subsp. casei. Sequencing of identified unique nucH gene fragment seems to be a promising diagnostic assay for the identification of Staphylococcus species. Based on our results, we can assume that probably other Staphylococcus species originated from different clinical samples could be identified using nucH gene sequencing method we developed. However, an extension of the genetic databases with a substantially bigger number of reference staphylococcal species for nucH gene is needed to make this method better than widely used standard 16S rDNA sequencing assay. To the best of our knowledge, it is the second published isolation of S. succinus subsp. casei from human clinical specimens. Moreover, possibility of decreasing the number of dimensions from multi-PCR-bands results using ribotyping analysis is also described.

中文翻译：

---

用于鉴定人类相关葡萄球菌琥珀亚种的新型葡萄球菌 nucH 分类标记。干酪

我们研究的目的是评估基于生物信息学分析的独特nucH基因片段的测序，作为一种新的诊断方法，用于鉴定难以鉴定的人类葡萄球菌致病菌株。最初，进行了对 16SrDNA 和 23SrDNA 之间的间隔物特异性的PCR-RFLP- rrn分析，然后进行了Hha I 限制性分析。此外，进行了nucH和16S rDNA基因片段的测序。来自 NCBI 的 Blast 分析显示nucH基因片段与S. succinus的参考基因组 DNA 有 99% 的相似性入藏号 CP018199。MALDI-TOF分析也证实了这一结果。16S rDNA 基因片段的测序分析允许 100% 鉴定从人类样品中分离的两种菌株为葡萄球菌琥珀亚种。干酪_ 对鉴定的独特nucH基因片段进行测序似乎是鉴定葡萄球菌属的一种有前途的诊断方法。根据我们的结果，我们可以假设可能来自不同临床样本的其他葡萄球菌物种可以使用nucH进行鉴定我们开发的基因测序方法。然而，为了使这种方法优于广泛使用的标准 16S rDNA 测序分析，需要扩展具有大量参考葡萄球菌种类的nucH基因的遗传数据库。据我们所知，这是第二次发表的S. succinus subsp 的分离。来自人类临床标本的干酪。此外，还描述了使用核糖分型分析从多 PCR 条带结果中减少维数的可能性。