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Expanded options for treatments directed against pathogens that can be used for bioterrorism are urgently needed. Treatment regimens directed against such pathogens can be identified only by using data derived from in vitro and animal studies. It is crucial that these studies reliably predict the efficacy of proposed treatments in humans. The objective of this study was to identify a levofloxacin treatment regimen that will serve as an effective therapy for Bacillus anthracis infections and postexposure prophylaxis. An in vitro hollow-fiber infection model that replicates the pharmacokinetic profile of levofloxacin observed in humans (half-life [t 1/2 ], 7.5 h) or in animals, such as the mouse or the rhesus monkey (t 1/2 , ϳ2 h), was used to evaluate a proposed indication for levofloxacin (500 mg once daily) for the treatment of Bacillus anthracis infections. The results obtained with the in vitro model served as the basis for the doses and the dose schedules that were evaluated in the mouse inhalational anthrax model. The effects of levofloxacin and ciprofloxacin treatment were compared to those of no treatment (untreated controls). The main outcome measure in the in vitro hollow-fiber infection model was a persistent reduction of culture density (>4 log 10 reduction) and prevention of the emergence of levofloxacin-resistant organisms. In the mouse inhalational anthrax model the main outcome measure was survival. The results indicated that levofloxacin given once daily with simulated human pharmacokinetics effectively sterilized Bacillus anthracis cultures. By using a simulated animal pharmacokinetic profile, a once-daily dosing regimen that provided a human-equivalent exposure failed to sterilize the cultures. Dosing regimens that "partially humanized" levofloxacin exposures within the constraints of animal pharmacokinetics reproduced the antimicrobial efficacy seen with human pharmacokinetics. In a mouse inhalational anthrax model, once-daily dosing was significantly inferior (survival end point) to regimens of dosing every 12 h or every 6 h with identical total daily levofloxacin doses. These results demonstrate the predictive value of the in vitro hollow-fiber infection model with respect to the success or the failure of treatment regimens in animals. Furthermore, the model permits the evaluation of treatment regimens that "humanize" antibiotic exposures in animal models, enhancing the confidence with which animal models may be used to reliably predict the efficacies of proposed antibiotic treatments in humans in situations (e.g., the release of pathogens as agents of bioterrorism or emerging infectious diseases) where human trials cannot be performed. A treatment regimen effective in rhesus monkeys was identified.

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Prevention of inhalational anthrax after Bacillus anthracis spore exposure requires a prolonged course of antibiotic prophylaxis. In response to the 2001 anthrax attack in the United States, Ϸ10,000 people were offered 60 days of antibiotic prophylaxis to prevent inhalational anthrax, but adherence to this regimen was poor. We sought to determine whether a short course of antibiotic prophylaxis after exposure could protect non-human primates from a high-dose spore challenge if vaccination was combined with antibiotics. Two groups of 10 rhesus macaques were exposed to Ϸ1,600 LD 50 of spores by aerosol. Both groups were given ciprofloxacin by orogastric tube twice daily for 14 days, beginning 1-2 h after exposure. One group also received three doses of the licensed human anthrax vaccine (anthrax vaccine adsorbed) after exposure. In the ciprofloxacin-only group, four of nine monkeys (44%) survived the challenge. In contrast, all 10 monkeys that received 14 days of antibiotic plus anthrax vaccine adsorbed survived (P ‫؍‬ 0.011). Thus postexposure vaccination enhanced the protection afforded by 14 days of antibiotic prophylaxis alone and completely protected animals against inhalational anthrax. These data provide evidence that postexposure vaccination can shorten the duration of antibiotic prophylaxis required to protect against inhalational anthrax and may impact public health management of a bioterrorism event.Bacillus anthracis ͉ treatment ͉ vaccine B acillus anthracis infection in humans occurs as cutaneous, gastrointestinal, or inhalational anthrax depending upon the route of exposure. Cutaneous anthrax is rarely fatal and can be effectively treated with antibiotics. Inhalational anthrax, the form likely to occur after a bioterrorist attack, on the other hand, is difficult to diagnose early, and despite antibiotic therapy, has a high fatality rate. Anthrax is rare in industrialized countries, and vaccination with anthrax vaccine adsorbed (AVA) is confined to those who could be potentially exposed to anthrax, such as veterinary workers, woolen mill employees, and laboratory workers (1). Military personnel in the United States are also vaccinated due to the potential threat of B. anthracis being used as a bioweapon.Past experiments have shown that the rhesus macaque is the animal model that most closely mimics inhalational anthrax in humans (2). In both humans and macaques, inhalational anthrax begins with the deposition of 1-to 5-m spores in the alveolar spaces, where spores are thought to be ingested by alveolar phagocytic cells. Some spores survive inside the phagocyte and are transported to the draining pulmonary and mediastinal lymph nodes where germination occurs. Although most spores probably germinate within a few days after inhalation, germination is not synchronous (3). Some spores remain dormant and do not germinate for prolonged periods (4, 5). It is the delayed germination of retained spores into vegetative bacilli that necessitates the prolonged use of prophylactic antibiotics after an inhalational ...

Petrobactin is the primary siderophore synthesized by Bacillus anthracis str Sterne and is required for virulence of this organism in a mouse model. The siderophore's biosynthetic machinery was recently defined and gene homologues of this operon exist in several other Bacillus strains known to be mammalian pathogens, but are absent in several known to be harmless such as B. subtilis and B. lichenformis. Thus, a common hypothesis regarding siderophore production in Bacillus species is that petrobactin production is exclusive to pathogenic isolates. In order to test this hypothesis, siderophores produced by 106 strains of an in-house library of the Bacillus cereus sensu lato group were isolated and identified using a MALDI-TOF-MS assay. Strains were selected from a previously defined phylogenetic tree of this group in order to include both known pathogens and innocuous strains. Petrobactin is produced by pathogenic strains and innocuous isolates alike, and thus is not itself indicative of virulence.

Cerulenin is a potent inhibitor of fatty acid synthase (FAS) in a variety of prokaryotic and eukaryotic cells. Using a standardized mycobacterial susceptibility test, we have observed that cerulenin inhibits the growth of several species of mycobacteria, including tuberculous species such as Mycobacterium tuberculosis (H37Rv and clinical isolates) and Mycobacterium bovis BCG (hereafter called BCG), as well as several non-tuberculous species: Mycobacterium smegmatis, the Mycobacterium avium-intracellulare complex (MAC), Mycobacterium kansasii and others. All species and strains tested, including multi-drug resistant isolates of M. tuberculosis, were susceptible to cerulenin with MICs ranging from 1.5 to 12.5 mg/L. Two-dimensional thin-layer chromatography revealed different inhibition patterns of lipid synthesis between tuberculous and non-tuberculous mycobacteria. Cerulenin treatment resulted in a relative increase in phospholipids and mycolic acids in MAC and M. smegmatis, whereas in cerulenin-treated BCG, phospholipids and mycolic acids diminished relative to controls. In addition, long-chain extractable lipids (intermediate in polarity), triglycerides and glycopeptidolipids decreased with cerulenin treatment in all three species of mycobacteria tested. Qualitative changes in several of these lipid classes indicate inhibition in the synthesis of intermediate and long-chain fatty acids. Our results suggest that cerulenin's primary effect may be inhibition of intermediate and long-chain lipid synthesis, with little effect on the synthesis of other lipid classes. In addition, the BCG-specific reduction in phospholipids and mycolic acids suggests the presence of a unique cerulenin-sensitive FAS system in tuberculous mycobacteria. Since pathogenic mycobacteria produce novel long-chain fatty acids, inhibition of fatty acid synthesis offers a potential target for the development of antimycobacterial drugs.

In vitro susceptibilities to 28 antibiotics were determined for 11 strains of Burkholderia mallei by the broth microdilution method. The B. mallei strains demonstrated susceptibility to aminoglycosides, macrolides, quinolones, doxycycline, piperacillin, ceftazidime, and imipenem. For comparison and evaluation, 17 antibiotic susceptibilities were also determined by the E-test. E-test values were always lower than the broth dilution values. Establishing and comparing antibiotic susceptibilities of specific B. mallei strains will provide reference information for assessing new antibiotic agents.