Signature-tagged transposon mutagenesis studies demonstrate the dynamic nature of cecal colonization of 2-week-old chickens by Campylobacter jejuni.

We have constructed plasmids to be used for in vitro signature-tagged mutagenesis (STM) of Campylobacter jejuni and used these to generate STM libraries in three different strains. Statistical analysis of the transposon insertion sites in the C. jejuni NCTC 11168 chromosome and the plasmids of strain 81-176 indicated that their distribution was not uniform. Visual inspection of the distribution suggested that deviation from uniformity was not due to preferential integration of the transposon into a limited number of hot spots but rather that there was a bias towards insertions around the origin. We screened pools of mutants from the STM libraries for their ability to colonize the ceca of 2-week-old chickens harboring a standardized gut flora. We observed high-frequency random loss of colonization proficient mutants. When cohoused birds were individually inoculated with different tagged mutants, random loss of colonization-proficient mutants was similarly observed, as was extensive bird-to-bird transmission of mutants. This indicates that the nature of campylobacter colonization in chickens is complex and dynamic, and we hypothesize that bottlenecks in the colonization process and between-bird transmission account for these observations.

Lymph Node Colonization Dynamics after Oral Salmonella Typhimurium Infection in Mice

An understanding of how pathogens colonize their hosts is crucial for the rational design of vaccines or therapy. While the molecular factors facilitating the invasion and systemic infection by pathogens are a central focus of research in microbiology, the population biological aspects of colonization are still poorly understood. Here, we investigated the early colonization dynamics of Salmonella enterica subspecies 1 serovar Typhimurium (S. Tm) in the streptomycin mouse model for diarrhea. We focused on the first step on the way to systemic infection - the colonization of the cecal lymph node (cLN) from the gut - and studied roles of inflammation, dendritic cells and innate immune effectors in the colonization process. To this end, we inoculated mice with mixtures of seven wild type isogenic tagged strains (WITS) of S. Tm. The experimental data were analyzed with a newly developed mathematical model describing the stochastic immigration, replication and clearance of bacteria in the cLN. We estimated that in the beginning of infection only 300 bacterial cells arrive in the cLN per day. We further found that inflammation decreases the net replication rate in the cLN by 23%. In ccr7-/- mice, in which dendritic cell movement is impaired, the bacterial migration rate was reduced 10-fold. In contrast, cybb-/- mice that cannot generate toxic reactive oxygen species displayed a 4-fold higher migration rate from gut to cLN than wild type mice. Thus, combining infections with mixed inocula of barcoded strains and mathematical analysis represents a powerful method for disentangling immigration into the cLN from replication in this compartment. The estimated parameters provide an important baseline to assess and predict the efficacy of interventions. © 2013 Kaiser et al.