Short communicationIn vitro resistance of Bacillus anthracis Sterne to doxycycline, macrolides and quinolones
Introduction
Bacillus anthracis, the causative agent of anthrax, is the primary biological warfare threat to US military forces. According to United Nations Special Commission (UNSCOM) reports, B. anthracis spores were weaponized for aerosol delivery by Iraq during the 1991 Gulf War [1], [2]. Penicillin G, doxycycline and the quinolone ciprofloxacin are the drugs of choice for the therapy of anthrax [1], [3], [4]. Selection of antimicrobial resistance against ciprofloxacin, alatrofloxacin and gatifloxacin has not been previously reported.
Our preliminary studies showed that B. anthracis developed resistance to ofloxacin after multiple, serial passages in vitro in subinhibitory concentrations of this antimicrobial agent [5]. Our current work extended these studies by investigating the potential for B. anthracis to develop resistance to three other quinolones (ciprofloxacin, alatrofloxacin, and gatifloxacin), which may also be used to treat anthrax.
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Materials and methods
We grew B. anthracis Sterne in Schaeffer's sporulation medium [6] from live B. anthracis spores veterinary vaccine (Colorado Serum Co., Denver, CO). The resulting bacterial spores were harvested and stored at 35 °C for 24 h. For our study, we used seven antimicrobial agents. These included three quinolones: ciprofloxacin (Bayer Corp., West Haven, CT), alatrofloxacin (Pfizer Roerig, New York, NY) and gatifloxacin (Bristol-Meyers Squibb Co, Princeton, NJ). Three macrolides: erythromycin
Resistance development in sequential subculturing of quinolones and doxycycline
The increase in MICs of ciprofloxacin, gatifloxacin alatrofloxacin and doxycycline after 21 passages of sequential subculturing of B. anthracis Sterne in sub-inhibitory concentrations of these antibiotics is shown in Table 1. The number of passages required for selection of resistance varied from six to 17.
Resistance development in sequential subculturing of macrolides and ciprofloxacin
The increase in MICs of ciprofloxacin, azithromycin, clarithromycin and erythromycin after 15 passages of sequential subculturing of B. anthracis Sterne in sub-inhibitory concentrations of
Discussion
Our study showed the development of resistance in vitro to several antimicrobial agents by B. anthracis Sterne. The method of sequential subcultures using sub-inhibitory concentrations of antimicrobials selected for bacterial resistance [7]. Serial subculturing using three quinolones (ciprofloxacin, alatrofloxacin and gatifloxacin) and three macrolides (erythromycin, azithromycin and clarythromycin) led to selection of a subpopulation of bacteria with increasing MICs not only for the initial
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