Impact of Glutamine Transporters on Pneumococcal Fitness under Infection-Related Conditions

The genomic analysis of Streptococcus pneumoniae predicted six putative glutamine uptake systems, which are expressed under in vitro conditions, as shown here by reverse transcription-PCR. Four of these operons consist of glnHPQ, while two lack glnH, which encodes a soluble glutamine-binding protein. Here, we studied the impact of two of these glutamine ATP-binding cassette transporters on S. pneumoniae D39 virulence and phagocytosis, which consist of GlnQ and a translationally fused protein of GlnH and GlnP. Mice infected intranasally with D39 Delta gln0411/0412 showed significantly increased survival times and a significant delay in the development of pneumococcal pneumonia compared to those infected with D39, as observed in real time using bioluminescent pneumococci. In a mouse sepsis model, the mutant D39 Delta gln0411/0412 showed only moderate but significant attenuation. In contrast, the D39 Delta gln1098/1099 knockout strain was massively attenuated in the pneumonia and septicemia mouse infection model. To cause pneumonia or sepsis with D39 Delta gln1098/1099, infection doses 100- to 10,000-fold higher than those used for wild-type strain D39 were required. In an experimental mouse meningitis model, D39 Delta gln1098/1099 produced decreased levels of white blood cells in cerebrospinal fluid and showed decreased numbers of bacteria in the bloodstream compared to D39 and D39 Delta gln0411/0412. Phagocytosis experiments revealed significantly decreased intracellular survival rates of mutants D39 Delta gln1098/1099 and D39 Delta gln0411/0412 compared to wild-type D39, suggesting that the deficiency of Gln uptake systems impairs resistance to oxidative stress. Taken together, our results demonstrate that both glutamine uptake systems are required for full virulence of pneumococci but exhibit different impacts on the pathogenesis of pneumococci under in vivo conditions.