The Broad Institute of MIT and Harvard, Cambridge, MA, USA; Department of Molecular Biology, Massachusetts General Hospital, Boston, MA, USA; Center for Computational and Integrative Biology, Massachusetts General Hospital, Boston, MA, USA; Department of Genetics, Harvard Medical School, Boston, MA, USA.
The Broad Institute of MIT and Harvard, Cambridge, MA, USA.
Cell. 2023 Feb 16;186(4):877-891.e14. doi: 10.1016/j.cell.2023.01.002. Epub 2023 Jan 27.
We introduce BacDrop, a highly scalable technology for bacterial single-cell RNA sequencing that has overcome many challenges hindering the development of scRNA-seq in bacteria. BacDrop can be applied to thousands to millions of cells from both gram-negative and gram-positive species. It features universal ribosomal RNA depletion and combinatorial barcodes that enable multiplexing and massively parallel sequencing. We applied BacDrop to study Klebsiella pneumoniae clinical isolates and to elucidate their heterogeneous responses to antibiotic stress. In an unperturbed population presumed to be homogeneous, we found within-population heterogeneity largely driven by the expression of mobile genetic elements that promote the evolution of antibiotic resistance. Under antibiotic perturbation, BacDrop revealed transcriptionally distinct subpopulations associated with different phenotypic outcomes including antibiotic persistence. BacDrop thus can capture cellular states that cannot be detected by bulk RNA-seq, which will unlock new microbiological insights into bacterial responses to perturbations and larger bacterial communities such as the microbiome.
我们介绍了 BacDrop，这是一种用于细菌单细胞 RNA 测序的高可扩展技术，它克服了许多阻碍细菌 scRNA-seq 发展的挑战。BacDrop 可应用于来自革兰氏阴性和革兰氏阳性物种的数千到数百万个细胞。它具有通用的核糖体 RNA 耗竭和组合条形码，可实现多路复用和大规模并行测序。我们将 BacDrop 应用于研究肺炎克雷伯菌临床分离株，并阐明它们对抗生素应激的异质反应。在假定为同质的未受干扰的群体中，我们发现群体内异质性主要由促进抗生素耐药性进化的可移动遗传元件的表达驱动。在抗生素胁迫下，BacDrop 揭示了与不同表型结果（包括抗生素持续存在）相关的转录上不同的亚群。因此，BacDrop 可以捕获通过批量 RNA-seq 无法检测到的细胞状态，这将为细菌对扰动和更大的细菌群落（如微生物组）的反应提供新的微生物学见解。
