Antimicrobial therapy for bacillus anthracis-induced polymicrobial infection in (60)Co gamma-irradiated mice

Abstract

Challenge with both nonlethal ionizing radiation and toxigenic Bacillus anthracis spores increases the rate of mortality from a mixed bacterial infection. If biological weapons, such as B. anthracis spores, and nuclear weapons were used together, casualties could be more severe than they would be from the use of either weapon alone. We previously discovered that a polymicrobial infection developed in B6D2F(1)/J mice after nonlethal (7-Gy) (60)Co gamma irradiation and intratracheal challenge with B. anthracis Sterne spores 4 days after irradiation. In this present study, we investigated the survival of mice and the response of the polymicrobial infection during the course of antimicrobial therapy with penicillin G procaine, ofloxacin, trovafloxacin, or gatifloxacin. Survival was prolonged, but not ensured, when the mice were treated with either broad-spectrum ofloxacin or narrow-spectrum penicillin G for 7 days beginning 6 or 24 h after challenge. Survival was not prolonged when therapy was delayed more than 24 h after challenge. When these two antimicrobial agents were given for 21 days, the survival rate was increased from 0% for the controls to 38 to 63% after therapy. Therapy with trovafloxacin or gatifloxacin reduced the incidence of mixed infection and improved the rate of survival to 95% (trovafloxacin) or 79% (gatifloxacin), whereas the rate of survival for the controls was 5%. We conclude that the mixed infection induced by B. anthracis in irradiated mice complicates effective therapy with a single antimicrobial agent. To limit mortality following nonlethal irradiation and challenge with B. anthracis spores, antimicrobial therapy needs to be initiated within a few hours after challenge and continued for up to 21 days.

FIG. 1.

General experimental design. Mice were irradiated and were then challenged by i.t. inoculation of B. anthracis Sterne spores 4 days later and given antimicrobial therapy daily for 7, 14, or 21 days beginning 6, 7.5, 24, 48, or 72 h after bacterial spore challenge.

FIG. 2.

Time of administration of PEN G (62.5 mg/kg i.m.) (A) or OFX (40 mg/kg i.p.) (B) for 7 days and rates of survival from B. anthracis Sterne challenge (7.8 × 10⁸ CFU i.t.) in ⁶⁰Co γ-irradiated (7 Gy) mice. Therapy was started 6, 24, 48, or 72 h after challenge. Significant comparisons were as follows: P < 0.01 for the control versus PEN G started at 6 h, OFX started at 6 h, PEN G started at 24 h, and OFX started at 24 h; P < 0.05 for the control versus OFX started at 48 h; P < 0.05 for PEN G started at 48 h versus PEN G started at 24 h, OFX started at 24 h versus OFX started at 6 h, OFX started at 48 h versus OFX started at 24 h, and OFX started at 72 h versus OFX started at 48 h.

FIG. 3.

Survival of mice treated with PEN G (125 mg/kg s.c.) or OFX (40 mg/kg p.o.) for 21 days after ⁶⁰Co γ irradiation (7 Gy) and challenge with B. anthracis Sterne spores (3.3 × 10⁸ CFU i.t.). Therapy was started either 7.5 or 24 h after challenge. Significant comparisons were as follows: P < 0.01 for control versus PEN G started at 7.5 h, OFX started at 7.5 h, PEN G started at 24 h, and OFX started at 24 h.

FIG. 4.

Survival of B6D2F₁/J mice treated with antimicrobial agents (20 mg of TVA/kg p.o. or s.c., 40 mg of OFX/kg p.o., or 125 mg of PEN G/kg s.c.) for 21 days after ⁶⁰Co γ irradiation (7 Gy) and challenge with B. anthracis Sterne spores (4.1 × 10⁸ CFU i.t.). Therapy was started 24 h after challenge. Significant comparisons were as follows: P < 0.01 for control versus TVA s.c., TVA p.o., PEN G plus OFX, and OFX p.o., for PEN G plus OFX versus TVA p.o., and for PEN G + OFX versus TVA s.c.; P < 0.05 for control versus PEN G s.c., PEN G s.c. versus PEN G plus OFX, and OFX p.o. versus PEN G plus OFX.

FIG. 5.

Survival of B6D2F₁/J mice treated with GAT and TVA (20 mg/kg p.o. or s.c.) for 21 days after ⁶⁰Co γ irradiation (7 Gy) and challenge with B. anthracis Sterne spores (4.5 × 10⁸ CFU i.t.). Therapy was started 24 h after challenge. Significant comparisons were as follows: P < 0.01 for control versus TVA s.c., TVA p.o., GAT s.c., and GAT p.o.

FIG. 6.

Survival of B6D2F₁/J mice treated with macrolide antimicrobial agents (50 mg of AZM/kg p.o. or s.c., 150 mg of CLR/kg p.o., 500 mg of ERY/kg p.o., and 20 mg of TVA/kg p.o.) for 14 days after ⁶⁰Co γ irradiation (7 Gy) and challenge with B. anthracis Sterne spores (1.8 × 10⁸ CFU i.t.). Therapy was started 24 h after challenge and continued for 14 days. Significant comparisons were as follows: ERY versus TVA (P = 0.0001).