Abstract
Riemerella anatipestifer is a gram-negative bacterium that is the first naturally competent bacterium identified in the family Flavobacteriaceae. However, the determinants that influence the natural transformation and the underlying mechanism remain unknown. In this study, we evaluated the effects of various nutritional factors of the GCB medium [glucose, L-glutamine, vitamin B1, Fe (NO3)3, NaCl, phosphate, and peptone], on the natural transformation of R. anatipestifer ATCC 11845. Among the assayed nutrients, peptone and phosphate affected the natural transformation of R. anatipestifer ATCC 11845, and the transformation frequency was significantly decreased when phosphate or peptone was removed from the GCB medium. When the iron chelator 2,2′-dipyridyl (Dip) was added, the transformation frequency was decreased by approximately 100-fold and restored gradually when Fe (NO3)3 was added, suggesting that the natural transformation of R. anatipestifer ATCC 11845 requires iron. Given the importance of TonB in nutrient transportation, we further identified whether TonB is involved in the natural transformation of R. anatipestifer ATCC 11845. Mutation of tonBA or tonBB, but not tbfA, was shown to inhibit the natural transformation of R. anatipestifer ATCC 11845 in the GCB medium. In parallel, it was shown that the tonBB mutant, but not the tonBA mutant, decreased iron acquisition in the GCB medium. This result suggested that the tonBB mutant affects the natural transformation frequency due to the deficiency of iron utilization.
Highlights
Natural transformation is the process by which bacteria take up and integrate exogenous DNA into their genomes (Seitz and Blokesch, 2013)
Since R. anatipestifer is naturally competent in all growth phases, and the transformation frequency is highest in the exponential growth phase (Liu et al, 2017b) and cell density do not play a significant role in competence development, we suspected that the nutritional environment affects bacterial natural transformation
We chose to culture R. anatipestifer ATCC 11845 in the GC broth (GCB) medium containing 60 μM Dip and found that the growth of bacteria was significantly inhibited but gradually restored when Fe (NO3)3 was added (Figure 1F). These results indicated that phosphate, peptone, and iron affected the natural transformation of R. anatipestifer ATCC 11845 in the GCB medium
Summary
Natural transformation is the process by which bacteria take up and integrate exogenous DNA into their genomes (Seitz and Blokesch, 2013). Most gram-negative bacteria use the type IV pilus family (T4P) to retract DNA into the periplasm, where one strand is degraded and the other is internalized into the cytoplasm through the ComEC transmembrane channel. Environmental determinants are important factors affecting natural transformation. Most bacteria possess competence genes, numerous conditions or signals trigger competence and are often species specific (Seitz and Blokesch, 2013). The expression of competence genes is influenced by the growth phase, cell density, metabolic activity, nutritional stress, and DNA damage (Johnsborg et al, 2007). In S. pneumoniae and Helicobacter pylori, competence is induced by antibiotic stress or DNA damage (Prudhomme et al, 2006; Dorer et al, 2010)
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