Genome-wide Identification of conditionally essential genes supportinggrowth in serum and cerebrospinal fluid.

Abstract

is a major cause of sepsis and meningitis in pigs, and zoonosis through the emergence of disease-associated lineages. The ability ofto adapt and survive in host environments, such as blood and cerebrospinal fluid (CSF), is important for pathogenesis. Here, we used Transposon Sequencing (Tn-seq) coupled with Nanopore sequencing to identify conditionally essential genes (CEGs) for the growth ofP1/7 in active porcine serum (APS) and CSF derived from choroid plexus organoids. To our knowledge, this is the first successful application of ONT to Tn-library screening, enabling rapid local runs and a publicly available analysis pipeline. Through comparative fitness analyses, we identified 33 CEGs that support growth in APS and 25 CEGs in CSF. These genes highlight the importance of pathways related to amino acid transport, nucleotide metabolism, and cell envelope integrity. Notably, the LiaFSR regulatory system and multiple ABC transporters were important for proliferation. We also identified several genes of unknown function as essential for growth, pointing to previously unrecognized genetic factors involved inadaptation during infection. These findings provide new insights into the genetic requirements forsurvival in host-like environments and a deeper understanding of its ability to adapt to distinct physiological niches.