Large-Scale CRISPRi and Transcriptomics of Staphylococcus epidermidis Identify Genetic Factors Implicated in Lifestyle Versatility

Staphylococcus epidermidis is a ubiquitous human commensal skin bacterium that is also one of the most prevalent nosocomial pathogens. The genetic factors underlying this remarkable lifestyle plasticity are incompletely understood, mainly due to the difficulties of genetic manipulation, precluding high-throughput functional profiling of this species. To probe the versatility of S. epidermidis to survive across a diversity of environmental conditions, we developed a large-scale CRISPR interference (CRISPRi) screen complemented by transcriptional profiling (RNA sequencing) across 24 diverse conditions and piloted a droplet-based CRISPRi approach to enhance throughput and sensitivity. We identified putative essential genes, importantly revealing amino acid metabolism as crucial to survival across diverse environments, and demonstrated the importance of trace metal uptake for survival under multiple stress conditions. We identified pathways significantly enriched and repressed across our range of stress and nutrient-limited conditions, demonstrating the considerable plasticity of S. epidermidis in responding to environmental stressors. Additionally, we postulate a mechanism by which nitrogen metabolism is linked to lifestyle versatility in response to hyperosmotic challenges, such as those encountered on human skin. Finally, we examined the survival of S. epidermidis under acid stress and hypothesize a role for cell wall modification as a vital component of the survival response under acidic conditions. Taken together, this study integrates large-scale CRISPRi and transcriptomics data across multiple environments to provide insights into a keystone member of the human skin microbiome. Our results additionally provide a valuable benchmarking analysis for CRISPRi screens and are a rich resource for other staphylococcal researchers. IMPORTANCE Staphylococcus epidermidis is a bacteria that broadly inhabits healthy human skin, yet it is also a common cause of skin infections and bloodstream infections associated with implanted medical devices. Because human skin has many different types of S. epidermidis, each containing different genes, our goal is to determine how these different genes allow S. epidermidis to switch from healthy growth in the skin to being an infectious pathogen. Understanding this switch is critical to developing new strategies to prevent and treat S. epidermidis infections.