Efficacy of rifapentine and other rifamycins against Coxiella burnetii in vitro

Abstract

Since 1999, doxycycline and hydroxychloroquine have been the recommended treatment for chronic Q fever, a life-threatening disease caused by the bacterial pathogen, Coxiella burnetii. Despite the duration of its use, the treatment is not ideal due to the lengthy treatment time, high mortality rate, resistant strains, and the potential for contraindicated usage. A literature search was conducted to identify studies that screened large panels of drugs against C. burnetii to identify novel targets with potential efficacy against C. burnetii. Twelve candidate antimicrobials approved for use in humans by the US Food and Drug Administration were selected and minimum inhibitory concentrations (MICs) were determined against the low virulence strain Nine Mile phase II. Rifabutin and rifaximin were the best performing antibiotics tested with MICs of ≤0.01 µg mL^-1. Further screening of these top candidates was conducted alongside two drugs from the same class, rifampin, well-characterized, and rifapentine, not previously reported against C. burnetii. These were screened against virulent strains of C. burnetii representing three clinically relevant genotypes. Rifapentine was the most effective in the human monocytic leukemia cell line, THP-1, with a MIC ≤0.01 µg mL^-1. In the human kidney epithelial cell line, A-498, efficacy of rifapentine, rifampin, and rifabutin varied across C. burnetii strains with MICs between ≤0.001 and 0.01 µg mL^-1. Rifampin, rifabutin, and rifapentine were all bactericidal against C. burnetii; however, rifabutin and rifapentine demonstrated impressive bactericidal activity as low as 0.1 µg mL^-1 and should be further explored as alternative Q fever treatments given their efficacy in vitro.

Importance: This work will help inform investigators and physicians about potential alternative antimicrobial therapies targeting the causative agent of Q fever, Coxiella burnetii. Chronic Q fever is difficult to treat, and alternative antimicrobials are needed. This manuscript explores the efficacy of rifamycin antibiotics against virulent strains of C. burnetii representing three clinically relevant genotypes in vitro. Importantly, this study determines the susceptibility of C. burnetii to rifapentine, which has not been previously reported. Evaluation of the bactericidal activity of the rifamycins reveals that rifabutin and rifapentine are bactericidal at low concentrations, which is unusual for antibiotics against C. burnetii.

Keywords: Q fever; antibiotics; antimicrobial activity; bacterial infection; bactericidal activity; zoonotic infections.

Fig 1

MICs for selected antibiotics against NMI in THP-1 host cells. NMI C. burnetii was cultured in phorbol myristate acetate-differentiated THP-1 cells in the presence of increasing concentrations of antibiotics to determine the MIC. Data displayed are the mean log₁₀ genome equivalent per well ± SEM at 7 days post-exposure for each antibiotic at the respective concentration. The MIC was determined as the lowest concentration of antibiotic required to significantly inhibit growth of C. burnetii. Significant inhibition of growth was determined relative to DMSO solvent controls using one-way analysis of variance with Dunnett’s multiple comparisons post hoc test of log₁₀ transformed data. ****P < 0.0001.

Fig 2

MICs for selected antibiotics against virulent C. burnetii in A-498 host cells. C. burnetii strains (A) NMI, (B) HPF-GA1, (C) CM-SC1, and (D) GP-CO1 were cultured in A-498 cells in the presence of increasing concentrations of antibiotics to determine the MIC. Data displayed are the mean log₁₀ genome equivalent per well ± SEM at 7 (for NMI and HPF-GA1) and 14 (for CM-SC1 and GP-CO1) days post-exposure for each antibiotic at the respective concentration. The MIC was determined as the lowest concentration of antibiotic required to significantly inhibit growth of C. burnetii. Significant inhibition of growth was determined relative to DMSO solvent controls using one-way analysis of variance with Dunnett’s multiple comparisons post hoc test of log₁₀ transformed data. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Fig 3

MBCs for selected antibiotics against virulent C. burnetii in A-498 host cells. C. burnetii strains (A) NMI, (B) HPF-GA1, (C) CM-SC1, and (D) GP-CO1 were allowed to recover in A-498 cells for 21 days following a 7-day exposure to antibiotics. GE at day 0 (defined as the day that antibiotics were removed) and at 21 days post-recovery were calculated and transformed by taking the natural log (ln). Data are displayed as the ln difference of the GE per well at day 21 relative to day 0 [ln (GE per well D21) – ln (GE per well D0)]. MBC was determined as the lowest concentration of antibiotic required to render C. burnetii nonviable. C. burnetii was determined to be nonviable if the mean difference in ln transformed GE from day 21 relative to day 0 was significantly different relative to DMSO solvent controls based on one-way analysis of variance with Dunnett’s multiple comparisons post hoc test. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.