Short communication
In vitro resistance of Bacillus anthracis Sterne to doxycycline, macrolides and quinolones

https://doi.org/10.1016/S0924-8579(01)00464-2Get rights and content

Abstract

Bacillus anthracis is a potential biological warfare agent. Its ability to develop resistance to antimicrobial agents currently recommended for the treatment of anthrax infection is a major concern. B. anthracis Sterne was grown from a live veterinary vaccine and used it to test for the development of resistance after 21 sequential subcultures in sub-inhibitory concentrations of doxycycline and three quinolones (ciprofloxacin, alatrofloxacin and gatifloxacin) and 15 sequential subcultures in sub-inhibitory concentrations of three macrolides (erythromycin, azithromycin and clarithromycin). After 21 subcultures the minimal inhibitory concentrations (MICs) increased from 0.1 to 1.6 mg/l for ciprofloxacin, from 1.6 to 12.5 mg/l for alatrofloxacin, from 0.025 to 1.6 mg/l for gatifloxacin and from 0.025 to 0.1 mg/l for doxycycline. After 15 passages of sequential subculturing with macrolides, the MICs increased from 12.5 to 12.5 or 50.0 mg/l for azithromycin, from 0.2 to 1.6 or 0.4 mg/l for clarithromycin and from 6.25 to 6.25 or 50 mg/l for erythromycin. After sequential passages with a single quinolone or doxycycline, each isolate was cross-tested for resistance using the other drugs. All isolates selected for resistance to one quinolone were also resistant to the other two quinolones, but not to doxycycline. The doxycycline-resistant isolate was not resistant to any quinolone.

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.

Section snippets

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

References (10)

  • J.C. Pile et al.

    Anthrax as a potential biological warfare agent

    Arch. Intern. Med.

    (1998)
  • United Nations. Eighth Report of the Secretary-General on the Status of the Implementation of the Plan for the Ongoing...
  • G.B. Knudson

    Treatment of anthrax in man: history and current concepts

    Mil. Med.

    (1986)
  • J.G. Bartlett
  • C.H. Choe et al.

    In vitro development of resistance to ofloxacin and doxycycline in Bacillus anthracis Sterne

    Antimicrob. Agents Chemother.

    (2000)
There are more references available in the full text version of this article.

Cited by (0)

View full text