CRISPRi-seq of Streptococcus pneumoniae in C+Y and in mouse pneumonia model

In this study, we applied CRISPRi-seq to study genetic factors important in C+Y mediuma and for causing pneumonia.Mice were infected intratracheally with our CRISPRi library and bacteria were collected at 24 hours (from lung) and 48 hours (from both lung and blood) after infection. Sequencing data showed a critical bottleneck after 48 hours of intratracheal infection, revealing that only a few bacteria can survive the brunt of the immune system and cause systemic infection.However, over short periods of time, at 24 hpi, CRISPRi-seq was able to identify fitness costs of the target genes in vivo. By comparison of the fitness cost between in vitro and in vivo conditions, many genes with significant differences were identified, including virulence factors, genes only essential in vivo, and genes only essential in vitro.