Finally, we examined virulence in vivo using a murine acute infection model of listeriosis (Fig. 2C). Most mutants were significantly attenuated for virulence in both the spleen and the liver. Interestingly, nrdD::Tn mutants, which should be unable to grow anaerobically (21), were attenuated only moderately in the spleen and were not attenuated in the liver, suggesting that these particular niches for L. monocytogenes are not completely devoid of oxygen. The most attenuated strains were pdhC::Tn, menD::Tn, menF::Tn, and yvcJ::Tn mutants. Together, these data show that, independently of cell type-specific survival defects, genes required for cytosolic survival are essential for virulence in vivo.