由于缺乏已建立的培养方法,口腔微生物组中的细菌鉴定不佳。因此,这项研究旨在使用无文化分析技术,包括细菌单细胞基因组测序,鉴定细菌种类并研究唾液中的基因分布。来自同一个体的唾液样品被分类为灭活的或活的,然后使用16SrRNA测序进行分析。宏基因组鸟枪测序,和细菌单细胞测序。16SrRNA测序的结果揭示了两个样本中相似的微生物群结构,链球菌是主要的属。宏基因组鸟枪测序显示,样本中大约80%的DNA是非细菌来源的,而单细胞测序显示每个基因组的平均污染率为10.4%。单细胞测序还产生了用于灭活样品的48个孔中的43个和用于活样品的48个孔中的45个的基因组序列。关于抗性基因,88个分离物中有4个携带cfxA,它编码β-内酰胺酶,四个分离株携带红霉素抗性基因。在9种细菌中发现了四环素抗性基因。宏基因组鸟枪测序提供了cfxA的完整序列,ermF,和ermX,而其他抗性基因,例如tetQ和tetM,被检测为碎片。此外,肺炎链球菌的毒力因子是最常见的,检测到13个基因。我们的平均核苷酸同一性分析还表明五种单细胞分离的细菌是潜在的新物种。这些数据将有助于扩大口腔微生物组数据资源。
Bacteria in the oral microbiome are poorly identified owing to the lack of established culture methods for them. Thus, this study aimed to use culture-free analysis techniques, including bacterial single-cell genome sequencing, to identify bacterial species and investigate gene distribution in saliva. Saliva samples from the same individual were classified as inactivated or viable and then analyzed using 16S rRNA sequencing, metagenomic shotgun sequencing, and bacterial single-cell sequencing. The results of 16S rRNA sequencing revealed similar microbiota structures in both samples, with Streptococcus being the predominant genus. Metagenomic shotgun sequencing showed that approximately 80 % of the DNA in the samples was of non-bacterial origin, whereas single-cell sequencing showed an average contamination rate of 10.4 % per genome. Single-cell sequencing also yielded genome sequences for 43 out of 48 wells for the inactivated samples and 45 out of 48 wells for the viable samples. With respect to resistance genes, four out of 88 isolates carried cfxA, which encodes a β-lactamase, and four isolates carried erythromycin resistance genes. Tetracycline resistance genes were found in nine bacteria. Metagenomic shotgun sequencing provided complete sequences of cfxA, ermF, and ermX, whereas other resistance genes, such as tetQ and tetM, were detected as fragments. In addition, virulence factors from Streptococcus pneumoniae were the most common, with 13 genes detected. Our average nucleotide identity analysis also suggested five single-cell-isolated bacteria as potential novel species. These data would contribute to expanding the oral microbiome data resource.