Figure 1. Lip2 is required for pathogenicity.
(A) lip2DOX-OFF is defective for systemic virulence in the presence of doxycycline. Female BALB/c mice were treated with 0.25 mg/ml doxycycline via drinking water for seven days prior to retro-orbital injection with 1×105 CFU of a 1:1 mixture of WT and lip2DOX-OFF; relative strain abundance in kidneys was determined using qPCR. Statistical significance was determined by a paired two-tailed t-test; *** p<0.001.
(B) lip2 mutants exhibit reduced lethality compared to WT or a lip2+LIP2 gene addback strain. Groups of female BALB/c mice were injected with 1×105 CFU WT (n=16), lip2 (isolate 1, n=8), lip2 (isolate 2, n=8) or lip2+LIP2 (n=8). Statistical significance between WT and lip2 #1 or lip2 #2 was determined by Mantel-Cox test *** p<0.001.
(C) lip2 fails to persist in infected kidneys. Groups of female BALB/c mice were infected with 1×105 CFU of WT, lip2, or lip2+LIP2, followed by euthanasia of three animals per group at the indicated time points. CFUs were determined by plating right kidney homogenates onto Sabouraud agar (supplemented gentamicin and ampicillin) and counting after two days. Data represent the means and standard deviations. Statistical significance between WT and lip2 was determined by an unpaired two-tailed t-test. ns nonsignificant, ** p<0.01; *** p<0.001. (Note that points without visible error bars displayed SEMs smaller than the circle.)
(D) lip2 forms normal-appearing hyphae in host kidneys. 400x image of PAS-stained left kidneys from experiment described in (C).
(E) lip2 causes minimal damage to kidneys. Kim1 mRNA was measured in the right kidney homogenates and normalized to Gadph. Statistical significance was determined by an unpaired two-tailed t-test. * p<0.05; ** p<0.01; *** p<0.001; **** p<0.0001. (Note that points without visible error bars displayed SEMs smaller than the circle.)