Due to its lifestyle as a commensal and occasional pathogen in the upper and lower respiratory tracts of humans, Haemophilus influenzae needs to protect itself from endogenously and exogenously generated reactive oxygen species. To better understand the oxygen radical resistance and to investigate a correlation with virulence, randomly generated paraquat-sensitive H. influenzae transposon mutants were analyzed in an infant rat model of infection. Among 25 different paraquat-sensitive mutants only one mutant harbouring a Tn-insertion within the tRNA-Ser1 gene specific for the rare serine codon UCC, was highly attenuated for intraperitoneal infectivity. Compared to the wild-type strain, the tRNA-Ser1 mutant was also more sensitive to neutrophil-mediated killing, deficient for DNA transformation but showed similar growth rates under laboratory conditions. However, by comparative analysis using an oxyR mutant strain, we could show that neutrophil-mediated killing might not be relevant for intraperitoneal infectivity. Therefore, the increased ROS sensitivity observed for tRNA-Ser1 mutant may not be directly responsible for the observed virulence deficiency in the intraperitoneal infection. We speculate that a reduced translation efficiency of several UCC containing mRNAs results in a delay of protein synthesis and consequently in the loss of cellular mechanisms which are necessary for ROS resistance and virulence.