Virulence difference between the prototypic Schu S4 strain (A1a) and Francisella tularensis A1a, A1b, A2 and type B strains in a murine model of infection

Abstract

Background: The use of prototypic strains is common among laboratories studying infectious agents as it promotes consistency for data comparability among and between laboratories. Schu S4 is the prototypic virulent strain of Francisella tularensis and has been used extensively as such over the past six decades. Studies have demonstrated virulence differences among the two clinically relevant subspecies of F. tularensis, tularensis (type A) and holarctica (type B) and more recently between type A subpopulations (A1a, A1b and A2). Schu S4 belongs to the most virulent subspecies of F. tularensis, subspecies tularensis.

Methods: In this study, we investigated the relative virulence of Schu S4 in comparison to A1a, A1b, A2 and type B strains using a temperature-based murine model of infection. Mice were inoculated intradermally and a hypothermic drop point was used as a surrogate for death. Survival curves and the length of temperature phases were compared for all infections. Bacterial burdens were also compared between the most virulent type A subpopulation, A1b, and Schu S4 at drop point.

Results: Survival curve comparisons demonstrate that the Schu S4 strain used in this study resembles the virulence of type B strains, and is significantly less virulent than all other type A (A1a, A1b and A2) strains tested. Additionally, when bacterial burdens were compared between mice infected with Schu S4 or MA00-2987 (A1b) significantly higher burdens were present in the blood and spleen of mice infected with MA00-2987.

Conclusions: The knowledge gained from using Schu S4 as a prototypic virulent strain has unquestionably advanced the field of tularemia research. The findings of this study, however, indicate that careful consideration of F. tularensis strain selection must occur when the overall virulence of the strain used could impact the outcome and interpretation of results.

Figure 1

Survival curve comparison of mice infected with A1a strains OK01-2528, MO02-4195, and Schu S₄. C57BL/6 J mice (n = 7/strain) were challenged intradermally with 10–20 CFU of F. tularensis A1a strains OK01-2528 (▬ III ▬; red), MO02-4195 (▬; blue) and Schu S₄ (▬ ▬; orange) and survival curves were modeled according to a Weibull distribution. The drop point for each mouse is shown in orange circles for mice infected with strain Schu S₄, red triangles for mice infected with OK01-2528 and blue squares for mice infected with MO02-4195.

Figure 2

Comparison of fitted survival curves for mice infected with F. tularensis A1a, A1b, A2, type B and Schu S₄. Survival curves were modeled previously [16,18] according to a Weibull distribution for mice (n = 7/strain) infected intradermally with 10–20 CFU of two strains each of A1a (▬; black), A1b (▬ I ▬; blue), A2 (▬ ▬; red), and type B (▬ III ▬; green), and are compared to the survival curve of Schu S₄ infected mice (III ; orange). The time at which each mouse reached drop point is shown below the graph as black squares for A1a infected mice, blue circles for A1b infected mice, red triangles for A2 infected mice, green diamonds for type B infected mice and orange upside-down triangles for Schu S₄ infected mice.

Figure 3

Bacterial burden within the blood, spleen, liver and lungs of infected mice with A1a strain Schu S₄ and A1b strain MA00-2987. C57BL/6 J mice (n = 7/strain) were challenged intradermally with 10–20 CFU of F. tularensis strain MA00-2987 (A1b) and Schu S₄ (A1a). When mice reached drop point, organs were harvested and blood was taken from each mouse. Significant differences in bacterial burden between the two infecting strains were calculated using MANOVA with an overall Type I error rate of α =0.05 and are designated with an asterisk. The mean bacterial burden is shown as a bar.