In vitrophenotypic and transcriptomic variation inNeisseria musculimorphotypes correlate with colonization variability and persistencein vivo

Abstract

Asymptomatic colonization of the upper respiratory tract is a common trait of the two human restricted pathogens, Neisseria gonorrhoeae, and Neisseria meningitidis. In vivo models of pathogenic neisserial infections are heterologous systems that permit short-term colonization but do not fully recapitulate infections in humans. Studying Neisseria musculi (Nmus), an oral commensal, in laboratory mice allows investigation of Neisseria-host interactions that avoids host restriction barriers. Nmus produces smooth and rough morphotypes on solid media. We compared the in vitro phenotypes, biofilm transcriptomes, in vivo colonization patterns, and burdens of the two Nmus morphotypes. We observed that the two morphotypes differ in biofilm formation, pilin production, transformation frequency, and aggregation in vitro. These phenotypes strongly correlated with differential expression of a set of genes in the Nmus biofilms including those that encoded factors for bacterial attachment. In vivo, the smooth morphotype stably colonized the oral cavities of all inoculated A/J and C57BL/6J mice at higher burdens relative to the rough. Interestingly, both morphotypes colonized the oral cavities of A/Js at higher magnitudes than in C57BL/6Js. Gut colonization by the smooth morphotype was qualitatively higher than the rough. Nasal colonization in the A/Js was transient following nasal inoculations. Collectively, our results demonstrate that colonization by Nmus can be affected by various factors including Nmus morphotypes, inoculation routes, anatomical niches, and host backgrounds. The Nmus-mouse model can use variable morphotype-host combinations to study the dynamics of neisserial asymptomatic colonization and persistence in multiple extragenital niches.

Type IV pilus retraction is required for Neisseria musculi colonization and persistence in a natural mouse model of infection

“…The population size of Wt Nmus in C57Bl/6 mice was recently reported to fluctuate ( 54 ). The reason for this discrepancy in Nmus in vivo behavior in the two studies is unclear.…”

Unveiling the Immune Dynamics of Neisseria Persistent Oral Colonization: A Roadmap for Innovative Vaccine Strategies

“…Bacterial strains and growth conditions: A naturally occurring RifR smooth morphotype, strain NW742, of Neisseria musculi, was used for oral inoculations 41 . Neisseria musculi was cultivated on Gonococcal Base (GCB; Difco) agar plates containing rifampin (40 mg/L) and Kellogg's supplements routinely incubated at 37°C with 5% CO 2 for 48 hours.…”

“…The collected organs were each placed in a tube with organ transport medium (RPMI with 10% fetal bovine serum) containing penicillin and streptomycin and transported to the Ohio State University on wet ice. The tongues of mice were harvested as previously described 41 by making incisions made on both sides of the mouth of euthanized mice. The oral cavity was opened, and the tongue was held with sterile forceps from the tip and cut at the base with sterile scissors.…”

“…Weekly fecal and oral swabs were collected to quantify the colonization burden in the inoculated group and confirm the absence of Neisseria in the control group. Oral swabs were collected as described previously 41 and suspended in GCB + 20% glycerol, vortexed for one minute, and serially diluted in GC broth, for plating on GCB agar plates containing rifampin. Fecal pellets were collected in sterile microfuge tubes, weighed, and each pellet was mixed with 1 mL of GCB + 20% glycerol using a sterile plain wooden applicator.…”