Pentamidine inhibits Coxiella burnetii growth and 23S rRNA intron splicing in vitro

Abstract

Coxiella burnetii is the bacterial agent of Q fever in humans. Acute Q fever generally manifests as a flu-like illness and is typically self-resolving. In contrast, chronic Q fever usually presents with endocarditis and is often life-threatening without appropriate antimicrobial therapy. Unfortunately, available options for the successful treatment of chronic Q fever are both limited and protracted (>18 months). Pentamidine, an RNA splice inhibitor used to treat fungal and protozoal infections, was shown to reduce intracellular growth of Coxiella by ca. 73% at a concentration of 1 microM (ca. 0.6 microg/mL) compared with untreated controls, with no detectable toxic effects on host cells. Bacterial targets of pentamidine include Cbu.L1917 and Cbu.L1951, two group I introns that disrupt the 23S rRNA gene of Coxiella, as demonstrated by the drug's ability to inhibit intron RNA splicing in vitro. Since both introns are highly conserved amongst all eight genotypes of the pathogen, pentamidine is predicted to be efficacious against numerous strains of C. burnetii. To our knowledge, this is the first report describing antibacterial activity for this antifungal/antiprotozoal agent.

Fig. 1

Effect of varying concentrations of pentamidine on Coxiella burnetii growth. Coxiella genomes were quantified using quantitative polymerase chain reaction (qPCR) [8] after 96 h of growth in infected Vero cells in the presence of 0–10 μM pentamidine. Values represent the mean ± standard deviation (n = 3). * Statistically significant difference from results of the untreated control (P < 0.05 by Student's t test).

Fig. 2

Effect of pentamidine on Vero host cell numbers during 96 h of growth. Quantitative polymerase chain reaction (qPCR) was used to compare Vero cell genome numbers using a primer set specific to its cytochrome oxidase (GenBank accession no. AY972804). Values represent mean cycle threshold (C_T) ± standard deviation (n = 6) following 96 h growth in media containing the indicated concentrations of pentamidine. C_T values did not show a statistically significant difference from the untreated control (P > 0.05 by Student's t-test).

Fig. 3

Effect of pentamidine on RNA splicing by Coxiella burnetii group I intron RNAs in vitro. Values represent mean ³⁵S-guanosine-5'-triphosphate (³⁵S-GTP) incorporation into spliced RNAs [counts per min (CPMs) as a percent of the untreated control ± standard error of the mean]. Black and white bars denote Cbu.L1917 (n = 3) and Cbu.L1951 (n = 4), respectively. * Statistically significant difference from the untreated control (P < 0.05 by Student's t-test).