FIG 3.
Virulence screen of clinical L. monocytogenes isolates in murine spleen. CD1 mice (nonpregnant) were infected i.v. with bacterial pools containing differentially tagged L. monocytogenes strains at equal ratios (total of 10 pools). Pools A to I contained 9 clinical strains and 2 10403S strains per pool; the 10403S pool contained 11 laboratory reference strains. Statistically significant differences in splenic bacterial burden from those in the control group were determined using one-way ANOVA with Dunnett's multiple comparisons posttest. ***, P < 0.0001; **, P < 0.01; *, P < 0.05. (A) Bacterial burden in murine spleen at 48 hpi with 2 × 105 CFU per pool. For pools A to I there were 10 mice per pool; the 10403S pool contained 15 mice. Each circle represents the bacterial burden in one spleen, and each pool is represented by a different color. Red lines represent medians. (B) The average relative abundance of each strain in mouse spleen was quantified by qPCR. To accurately compare values across pools, the average relative abundance for each isolate was then normalized to the average for the reference strain in each pool. Significance Z-scores were calculated for the deviation from the range expected on the basis of the results for the 10403S pool (black circles). Blue circles indicate isolates with virulence similar to that of 10403S (intermediate virulence). Red and green circles indicate isolates with significantly higher and lower virulences, respectively. (C) CD1 mice were infected with one erythromycin-resistant 10403S strain and one erythromycin-susceptible untagged clinical isolate at a 1:1 ratio. The clinical isolates were chosen on the basis of their virulence scores in panel B: 3 hypervirulent (red circles) and 3 hypovirulent (green circles) strains. Competitive indices (isolate/10403S) were calculated for bacteria recovered from the spleen at 48 hpi. The control group was infected with two 10403S strains that differed in their susceptibility to erythromycin (10403S/E; black circles). Each group contained 5 mice from 2 separate experiments.