Neisseria genes required for persistence identified via in vivo screening of a transposon mutant library

Species of the Neisseria genus harbor close relationships with their mamallian hosts, including humans. The outcome of these interactions can result in commensalism, asymptomatic carriage, or acute inflammatory responses. Little is known of the mechanisms Neisseria use to maintain and shape their host niche in the context of asymptomatic infection or long term commensal colonization and persistence. To identify novel host interaction factors contributing to host persistence, a Tn5 transposon library was created in Neisseria musculi, and inoculated into a permissive strain of laboratory mice (CAST/EiJ). At various time points post inoculation, transposon mutants were recovered from fecal and oral samples, and analyzed to identifiy mutants incapable of maintaining a host niche at either site. A Tn5 transposon library was generated in Neisseria musculi (Nmus), and inoculated into the oral cavity of mice. Two library replicates were tested in 5 mice per replicate. For each library replicate, DNA was isolated from the initial stock passage and the inoculum. In host screen samples, bacteria were recovered from oral and fecal samples on selective media, and CFUs were pooled by timepoint, sample site and dose prior to DNA extraction. Transposon junction sites were then amplified by PCR and sequenced.