Acyclic nucleoside phosphonates with 2-aminothiazole base as inhibitors of bacterial and mammalian adenylate cyclases.

A series of novel acyclic nucleoside phosphonates (ANPs) was synthesized as potential adenylate cyclase inhibitors, where the adenine nucleobase of adefovir (PMEA) was replaced with a 5-substituted 2-aminothiazole moiety. The design was based on the structure of MB05032, a potent and selective inhibitor of fructose 1,6-bisphosphatase and a good mimic of adenosine monophosphate (AMP). From the series of eighteen novel ANPs, which were prepared as phosphoroamidate prodrugs, fourteen compounds were potent (single digit micromolar or submicromolar) inhibitors of Bordetella pertussis adenylate cyclase toxin (ACT), mostly without observed cytotoxicity in J774A.1 macrophage cells. Selected phosphono diphosphates (nucleoside triphosphate analogues) were potent inhibitors of ACT (IC50 as low as 37 nM) and B. anthracis edema factor (IC50 as low as 235 nM) in enzymatic assays. Furthermore, several ANPs were found to be selective mammalian AC1 inhibitors in HEK293 cell-based assays (although with some associated cytotoxicity) and one compound exhibited selective inhibition of mammalian AC2 (only 12% of remaining adenylate cyclase activity) but no observed cytotoxicity. The mammalian AC1 inhibitors may represent potential leads in development of agents for treatment of human inflammatory and neuropathic pain.

The use of ozone gas for the inactivation of Bacillus anthracis and Bacillus subtilis spores on building materials.

A study was conducted to assess the efficacy of ozone gas in inactivating spores of both Bacillus anthracis and Bacillus subtilis inoculated onto six building materials (glass, wood, carpet, laminate, galvanized metal, and wallboard paper). Testing conditions consisted of ozone gas concentrations ranging from 7,000-12,000 parts per million (ppm), contact times from 4 to 12 h, and two relative humidity (RH) levels of 75 and 85%. Results showed that increasing the ozone concentration, contact time, and RH generally increased decontamination efficacy. The materials in which the highest decontamination efficacy was achieved for B. anthracis spores were wallboard paper, carpet, and wood with ≥ 6 log10 reduction (LR) occurring with 9,800 ppm ozone, 85% RH, for 6 h. The laminate and galvanized metal materials were generally more difficult to decontaminate, requiring 12,000 ppm ozone, 85% RH, and 9-12 h contact time to achieve ≥6 LR of B. anthracis. Lastly, overall, there were no significant differences in decontamination efficacy between the two species.

Functional characterization and evaluation of protective efficacy of EA752-862 monoclonal antibody against B. anthracis vegetative cell and spores.

The most promising means of controlling anthrax, a lethal zoonotic disease during the early infection stages, entail restricting the resilient infectious form, i.e., the spores from proliferating to replicating bacilli in the host. The extractible antigen (EA1), a major S-layer protein present on the vegetative cells and spores of Bacillus anthracis, is highly immunogenic and protects mice against lethal challenge upon immunization. In the present study, mice were immunized with r-EA1C, the C terminal crystallization domain of EA1, to generate a neutralizing monoclonal antibody EA752-862, that was evaluated for its anti-spore and anti-bacterial properties. The monoclonal antibody EA752-862 had a minimum inhibitory concentration of 0.08 mg/ml, was bactericidal at a concentration of 0.1 mg/ml and resulted in 100% survival of mice against challenge with B. anthracis vegetative cells. Bacterial cell lysis as observed by scanning electron microscopy and nucleic acid leakage assay could be attributed as a possible mechanism for the bactericidal property. The association of mAb EA752-862 with spores inhibits their subsequent germination to vegetative cells in vitro, enhances phagocytosis of the spores and killing of the vegetative cells within the macrophage, and subsequently resulted in 90% survival of mice upon B. anthracis Ames spore challenge. Therefore, owing to its anti-spore and bactericidal properties, the present study demonstrates mAb EA752-862 as an efficient neutralizing antibody that hinders the establishment of early infection before massive multiplication and toxin release takes place.

Rapid Discovery and Characterization of Synthetic Neutralizing Antibodies against Anthrax Edema Toxin.

Anthrax, a lethal, weaponizable disease caused by Bacillus anthracis, acts through exotoxins that are primary mediators of systemic toxicity and also targets for neutralization by passive immunotherapy. The ease of engineering B. anthracis strains resistant to established therapy and the historic use of the microbe in bioterrorism present a compelling test case for platforms that permit the rapid and modular development of neutralizing agents. In vitro antigen-binding fragment (Fab) selection offers the advantages of speed, sequence level molecular control, and engineering flexibility compared to traditional monoclonal antibody pipelines. By screening an unbiased, chemically synthetic phage Fab library and characterizing hits in cell-based assays, we identified two high-affinity neutralizing Fabs, A4 and B7, against anthrax edema factor (EF), a key mediator of anthrax pathogenesis. Engineered homodimers of these Fabs exhibited potency comparable to that of the best reported neutralizing monoclonal antibody against EF at preventing EF-induced cyclic AMP production. Using internalization assays in COS cells, B7 was found to block steps prior to EF internalization. This work demonstrates the efficacy of synthetic alternatives to traditional antibody therapeutics against anthrax while also demonstrating a broadly generalizable, rapid, and modular screening pipeline for neutralizing antibody generation.

Revisiting an old antibiotic: bacitracin neutralizes binary bacterial toxins and protects cells from intoxication.

The antibiotic bacitracin (Bac) inhibits cell wall synthesis of gram-positive bacteria. Here, we discovered a totally different activity of Bac: the neutralization of bacterial exotoxins. Bac prevented intoxication of mammalian cells with the binary enterotoxins Clostridium botulinum C2, C. perfringens ι, C. difficile transferase (CDT), and Bacillus anthracis lethal toxin. The transport (B) subunits of these toxins deliver their respective enzyme (A) subunits into cells. Following endocytosis, the B subunits form pores in membranes of endosomes, which mediate translocation of the A subunits into the cytosol. Bac inhibited formation of such B pores in lipid bilayers in vitro and in living cells, thereby preventing translocation of the A subunit into the cytosol. Bac preserved the epithelial integrity of toxin-treated CaCo-2 monolayers, a model for the human gut epithelium. In conclusion, Bac should be discussed as a therapeutic option against infections with medically relevant toxin-producing bacteria, including C. difficile and B. anthracis, because it inhibits bacterial growth and neutralizes the secreted toxins.-Schnell, L., Felix, I., Müller, B., Sadi, M., von Bank, F., Papatheodorou, P., Popoff, M. R., Aktories, K., Waltenberger, E., Benz, R., Weichbrodt, C., Fauler, M., Frick, M., Barth, H. Revisiting an old antibiotic: bacitracin neutralizes binary bacterial toxins and protects cells from intoxication.

Differential regulation of actin-activated nucleotidyl cyclase virulence factors by filamentous and globular actin.

Several bacterial pathogens produce nucleotidyl cyclase toxins to manipulate eukaryotic host cells. Inside host cells they are activated by endogenous cofactors to produce high levels of cyclic nucleotides (cNMPs). The ExoY toxin from Pseudomonas aeruginosa (PaExoY) and the ExoY-like module (VnExoY) found in the MARTX (Multifunctional-Autoprocessing Repeats-in-ToXin) toxin of Vibrio nigripulchritudo share modest sequence similarity (~38%) but were both recently shown to be activated by actin after their delivery to the eukaryotic host cell. Here, we further characterized the ExoY-like cyclase of V. nigripulchritudo. We show that, in contrast to PaExoY that requires polymerized actin (F-actin) for maximum activation, VnExoY is selectively activated by monomeric actin (G-actin). These two enzymes also display different nucleotide substrate and divalent cation specificities. In vitro in presence of the cation Mg2+, the F-actin activated PaExoY exhibits a promiscuous nucleotidyl cyclase activity with the substrate preference GTP>ATP≥UTP>CTP, while the G-actin activated VnExoY shows a strong preference for ATP as substrate, as it is the case for the well-known calmodulin-activated adenylate cyclase toxins from Bordetella pertussis or Bacillus anthracis. These results suggest that the actin-activated nucleotidyl cyclase virulence factors despite sharing a common activator may actually display a greater variability of biological effects in infected cells than initially anticipated.

Role of a Small Molecule in the Modulation of Cell Death Signal Transduction Pathways.

Inflammasomes activate caspase-1 in response to molecular signals from pathogens and other dangerous stimuli as a part of the innate immune response. A previous study discovered a small-molecule, 4-fluoro- N'-[1-(2-pyridinyl)ethylidene]benzohydrazide, which we named DN1, that reduces the cytotoxicity of anthrax lethal toxin (LT). We determined that DN1 protected cells irrespectively of LT concentration and reduced the pathogenicity of an additional bacterial exotoxin and several viruses. Using the LT cytotoxicity pathway, we show that DN1 does not prevent LT internalization and catalytic activity or caspase-1 activation. Moreover, DN1 does not affect the proteolytic activity of host cathepsin B, which facilitates the cytoplasmic entry of toxins. PubChem Bioactivities lists two G protein-coupled receptors (GPCR), type-1 angiotensin II receptor and apelin receptor, as targets of DN1. The inhibition of phosphatidylinositol 3-kinase, phospholipase C, and protein kinase B, which are downstream of GPCR signaling, synergized with DN1 in protecting cells from LT. We hypothesize that DN1-mediated antagonism of GPCRs modulates signal transduction pathways to induce a cellular state that reduces LT-induced pyroptosis downstream of caspase-1 activation. DN1 also reduced the susceptibility of Drosophila melanogaster to toxin-associated bacterial infections. Future experiments will aim to further characterize how DN1 modulates signal transduction pathways to inhibit pyroptotic cell death in LT-sensitive macrophages. DN1 represents a novel chemical probe to investigate host cellular mechanisms that mediate cell death in response to pathogenic agents.

In-cell production of a genetically-encoded library based on the θ-defensin RTD-1 using a bacterial expression system.

We report the high-yield heterologous expression of bioactive θ-defensin RTD-1 inside Escherichia coli cells by making use of intracellular protein trans-splicing in combination with a high efficient split-intein. RTD-1 is a small backbone-cyclized polypeptide with three disulfide bridges and a natural inhibitor of anthrax lethal factor protease. Recombinant RTD-1 was natively folded and able to inhibit anthrax lethal factor protease. In-cell expression of RTD-1 was very efficient and yielded ≈0.7mg of folded RTD-1 per gram of wet E. coli cells. This approach was used to generate of a genetically-encoded RTD-1-based peptide library in live E. coli cells. These results clearly demonstrate the possibility of using genetically-encoded RTD-1-based peptide libraries in live E. coli cells, which is a critical first step for developing in-cell screening and directed evolution technologies using the cyclic peptide RTD-1asa molecular scaffold.

Small molecule inhibitors of anthrax edema factor.

Anthrax is a highly lethal disease caused by the Gram-(+) bacteria Bacillus anthracis. Edema toxin (ET) is a major contributor to the pathogenesis of disease in humans exposed to B. anthracis. ET is a bipartite toxin composed of two proteins secreted by the vegetative bacteria, edema factor (EF) and protective antigen (PA). Our work towards identifying a small molecule inhibitor of anthrax edema factor is the subject of this letter. First we demonstrate that the small molecule probe 5'-Fluorosulfonylbenzoyl 5'-adenosine (FSBA) reacts irreversibly with EF and blocks enzymatic activity. We then show that the adenosine portion of FSBA can be replaced to provide more drug-like molecules which are up to 1000-fold more potent against EF relative to FSBA, display low cross reactivity when tested against a panel of kinases, and are nanomolar inhibitors of EF in a cell-based assay of cAMP production.

Diagnosis of cutaneous anthrax in resource-poor settings in West Arsi Province, Ethiopia.

INTRODUCTION: Cutaneous anthrax is a zoonotic disease caused by the spore-forming bacterium Bacillus anthracis, which typically presents with ulcers after contact with animals or animal products, and is rarely seen in high-income countries but is common in those with low- and middle-incomes. Objective. The aim of this study is to show the main clinical characteristics of cutaneous anthrax in endemic areas. MATERIAL AND METHODS: The study describes the main clinical characteristics of cutaneous anthrax in eight patients (six female and two male, age range 1 - 56 years) admitted to the rural General Hospital of Gambo, West Arsi Province of Ethiopia from 2010-2013. RESULTS: In all cases, lesions began as an erythematous papule located on exposed sites (n=7 head; n=1 thigh) and subsequently became a necrotic black eschar surrounded by an edematous halo. Two patients presented with painful ipsilateral adenopathy near the black eschar. Four patients developed a malignant pustule on the suborbital region of the face. Patients responded positively to treatment, and the lesions resolved, leaving eschars. However, one patient suffered the loss of an eyeball, and another died 12 hours after starting treatment. CONCLUSIONS: Physicians working in rural areas of resource-poor settings should be trained in the clinical identification of cutaneous anthrax. Early antibiotic treatment is essential for decreasing morbidity and mortality.

Tetrazole-Based trans-Translation Inhibitors Kill Bacillus anthracis Spores To Protect Host Cells.

Bacillus anthracis, the causative agent of anthrax, remains a significant threat to humans, including potential use in bioterrorism and biowarfare. The capacity to engineer strains with increased pathogenicity coupled with the ease of disseminating lethal doses of B. anthracis spores makes it necessary to identify chemical agents that target and kill spores. Here, we demonstrate that a tetrazole-based trans-translation inhibitor, KKL-55, is bactericidal against vegetative cells of B. anthracis in culture. Using a fluorescent analog, we show that this class of compounds colocalizes with developing endospores and bind purified spores in vitro KKL-55 was effective against spores at concentrations close to its MIC for vegetative cells. Spore germination was inhibited at 1.2× MIC, and spores were killed at 2× MIC. In contrast, ciprofloxacin killed germinants at concentrations close to its MIC but did not prevent germination even at 32× MIC. Because toxins are released by germinants, macrophages infected by B. anthracis spores are killed early in the germination process. At ≥2× MIC, KKL-55 protected macrophages from death after infection with B. anthracis spores. Ciprofloxacin required concentrations of ≥8× MIC to exhibit a similar effect. Taken together, these data indicate that KKL-55 and related tetrazoles are good lead candidates for therapeutics targeting B. anthracis spores and suggest that there is an early requirement for trans-translation in germinating spores.

Targeting Bacillus anthracis toxicity with a genetically selected inhibitor of the PA/CMG2 protein-protein interaction.

The protein-protein interaction between the human CMG2 receptor and the Bacillus anthracis protective antigen (PA) is essential for the transport of anthrax lethal and edema toxins into human cells. We used a genetically encoded high throughput screening platform to screen a SICLOPPS library of 3.2 million cyclic hexapeptides for inhibitors of this protein-protein interaction. Unusually, the top 3 hits all contained stop codons in the randomized region of the library, resulting in linear rather than cyclic peptides. These peptides disrupted the targeted interaction in vitro; two act by binding to CMG2 while one binds PA. The efficacy of the most potent CMG2-binding inhibitor was improved through the incorporation of non-natural phenylalanine analogues. Cell based assays demonstrated that the optimized inhibitor protects macrophages from the toxicity of lethal factor.

Combined virtual screening, MMPBSA, molecular docking and dynamics studies against deadly anthrax: An in silico effort to inhibit Bacillus anthracis nucleoside hydrolase.

Anthrax is a deadly disease caused by Bacillus anthracis, a dangerous biological warfare agent employed for both military and terrorist purposes. A critical selective target for chemotherapy against this disease is nucleoside hydrolase (NH), an enzyme still not found in mammals. In the current study, we have performed molecular docking and dynamics studies, aiming to propose the new potent inhibitors of B. anthracis NH among National Cancer Institute (NCI) Diversity Set. We also analyzed the principal interactions of proposed compounds with the active site residues of NH and the relevant factors to biological activity. Additionally, the physic-chemical properties of free and inhibitor bound NH were evaluated and discussed. Our data showed that compound NSC79887 is a good candidate to inhibit NH and also for biological tests and further development. Also, ADMET prediction revealed that all physic-chemical parameters are within the acceptable range defined for human use.

The Poly-γ-D-Glutamic Acid Capsule of Bacillus licheniformis, a Surrogate of Bacillus anthracis Capsule Induces Interferon-Gamma Production in NK Cells through Interactions with Macrophages.

The poly-γ-D-glutamic acid (PGA) capsule, a major virulence factor of Bacillus anthracis, provides protection of the bacterium from phagocytosis and allows its unimpeded growth in the host. We investigated crosstalk between murine natural killer (NK) cells and macrophages stimulated with the PGA capsule of Bacillus licheniformis, a surrogate of the B. anthracis capsule. PGA induced interferon-gamma production from NK cells cultured with macrophages. This effect was dependent on macrophage-derived IL-12 and cell-cell contact interaction with macrophages through NK cell receptor NKG2D and its ligand RAE-1. The results showed that PGA could enhance NK cell activation by inducing IL-12 production in macrophages and a contact-dependent crosstalk with macrophages.

S-nitrosylation of peroxiredoxin 1 contributes to viability of lung epithelial cells during Bacillus anthracis infection.

BACKGROUND: Using Bacillus anthracis as a model gram-positive bacterium, we investigated the effects of host protein S-nitrosylation during bacterial infection. B. anthracis possesses a bacterial nitric oxide synthase (bNOS) that is important for its virulence and survival. However, the role of S-nitrosylation of host cell proteins during B. anthracis infection has not been determined. METHODS: Nitrosoproteomic analysis of human small airway epithelial cells (HSAECs) infected with toxigenic B. anthracis Sterne was performed, identifying peroxiredoxin 1 (Prx1) as one predominant target. Peroxidase activity of Prx during infection was measured using 2-Cys-Peroxiredoxin activity assay. Chaperone activity of S-nitrosylated Prx1 was measured by insulin aggregation assay, and analysis of formation of multimeric species using Native PAGE. Griess assay and DAF-2DA fluorescence assay were used to measure NO production. Cell viability was measured using the Alamar Blue assay and the ATPlite assay (Perkin Elmer). RESULTS: S-nitrosylation of Prx1 in Sterne-infected HSAECs leads to a decrease in its peroxidase activity while enhancing its chaperone function. Treatment with bNOS inhibitor, or infection with bNOS deletion strain, reduces S-nitrosylation of Prx1 and decreases host cell survival. Consistent with this, siRNA knockdown of Prx1 lowers bNOS-dependent protection of HSAEC viability. CONCLUSIONS: Anthrax infection results in S-nitrosylation of multiple host proteins, including Prx1. The nitrosylation-dependent decrease in peroxidase activity of Prx1 and increase in its chaperone activity is one factor contributing to enhancing infected cell viability. GENERAL SIGNIFICANCE: These results provide a new venue of mechanistic investigation for inhalational anthrax that could lead to novel and potentially effective countermeasures.

A simple decontamination approach using hydrogen peroxide vapour for Bacillus anthracis spore inactivation.

AIMS: To evaluate the use of relatively low levels of hydrogen peroxide vapour (HPV) for the inactivation of Bacillus anthracis spores within an indoor environment. METHODS AND RESULTS: Laboratory-scale decontamination tests were conducted using bacterial spores of both B. anthracis Ames and Bacillus atrophaeus inoculated onto several types of materials. Pilot-scale tests were also conducted using a larger chamber furnished as an indoor office. Commercial off-the-shelf (COTS) humidifiers filled with aqueous solutions of 3 or 8% hydrogen peroxide (H2 O2 ) were used to generate the HPV inside the mock office. The spores were exposed to HPV for periods ranging from 8 h up to 1 week. CONCLUSIONS: Four- to seven-day exposures to low levels of HPV (average air concentrations of approx. 5-10 parts per million) were effective in inactivating B. anthracis spores on multiple materials. The HPV can be generated with COTS humidifiers and household H2 O2 solutions. With the exception of one test/material, B. atrophaeus spores were equally or more resistant to HPV inactivation compared to those from B. anthracis Ames. SIGNIFICANCE AND IMPACT OF THE STUDY: This simple and effective decontamination method is another option that could be widely applied in the event of a B. anthracis spore release.

[PERSPECTIVES OF DEVELOPMENT OF LIVE RECOMBINANT ANTHRAX VACCINES BASED ON OPPORTUNISTIC AND APATHOGENIC MICROORGANISMS].

Live genetic engineering anthrax vaccines on the platform of avirulent and probiotic micro-organisms are a safe and adequate alternative to preparations based on attenuated Bacillus anthracis strains. Mucosal application results in a direct contact of the vaccine preparations with mucous membranes in those organs arid tissues of the macro-organisms, that are exposed to the pathogen in the first place, resulting in a development of local and systemic immune response. Live recombinant anthrax vaccines could be used both separately as well as in a prime-boost immunization scheme. The review focuses on immunogenic and protective properties of experimental live genetic engineering prearations, created based on members of geni of Salmonella, Lactobacillus and adenoviruses.

Expresión y renaturalización del antígeno protector de Bacillus anthracis: un modelo para evaluar el replegado de proteínas antigénicas recombinantes a gran escala / Expression and refolding of the protective antigen of Bacillus anthracis: A model for high-throughput screening of antigenic recombinant protein refolding

Bacillus anthracis protective antigen (PA) is a well known and relevant immunogenic protein that is the basis for both anthrax vaccines and diagnostic methods. Properly folded antigenic PA is necessary for these applications. In this study a high level of PA was obtained in recombinant Escherichia coli. The protein was initially accumulated in inclusion bodies, which facilitated its efficient purification by simple washing steps; however, it could not be recognized by specific antibodies. Refolding conditions were subsequently analyzed in a high-throughput manner that enabled nearly a hundred different conditions to be tested simultaneously. The recovery of the ability of PA to be recognized by antibodies was screened by dot blot using a coefficient that provided a measure of properly refolded protein levels with a high degree of discrimination. The best refolding conditions resulted in a tenfold increase in the intensity of the dot blot compared to the control. The only refolding additive that consistently yielded good results was L-arginine. The statistical analysis identified both cooperative and negative interactions between the different refolding additives. The high-throughput approach described in this study that enabled overproduction, purification and refolding of PA in a simple and straightforward manner, can be potentially useful for the rapid screening of adequate refolding conditions for other overexpressed antigenic proteins.

El antígeno protector de Bacillus anthracis (protective antigen, PA) es una importante proteína inmunogénica, en la que se basan tanto las vacunas contra el ántrax/carbunclo como varios métodos diagnósticos. Para estas aplicaciones es esencial que el PA mantenga sus propiedades antigénicas, para lo cual debe estar correctamente plegado. En este estudio se obtuvieron altos niveles del PA en Escherichia coli recombinante. Inicialmente, la proteína se acumuló desnaturalizada en cuerpos de inclusión, lo que facilitó su eficiente purificación en simples pasos de lavado, pero no fue reconocida por anticuerpos específicos. Se analizaron las condiciones de replegado con un sistema de alto rendimiento, evaluando simultáneamente casi un centenar de condiciones diferentes. La recuperación de la capacidad del PA de ser reconocido por los anticuerpos se evaluó por dot blot utilizando un coeficiente que proporcionó una medida de los niveles de proteína correctamente plegada, con un alto grado de discriminación. Las mejores condiciones de renaturalización permitieron un aumento de diez veces en la intensidad de los dot blots con respecto del control. El único aditivo que produjo buenos resultados de forma constante fue la L-arginina. El análisis estadístico de las interacciones entre los diferentes aditivos de replegado permitió identificar tanto interacciones cooperativas como negativas. El enfoque de alto rendimiento descripto en este trabajo, que permitió la sobreproducción, purificación y plegado del PA de una manera sencilla y directa, puede ser potencialmente útil para el rápido screening de las condiciones adecuadas de replegado cuando se sobreexpresan otras proteínas antigénicas.

The efficacy and safety of nine South African medicinal plants in controlling Bacillus anthracis Sterne vaccine strain.

BACKGROUND: Anthrax is a zoonotic disease caused by Bacillus anthracis, a Gram-positive spore-forming bacterium. The presence of the bacteria and the toxins in the blood of infected hosts trigger a cascade of pathological events leading to death. Nine medicinal plants with good activities against other bacteria were selected to determine their in vitro antibacterial activity against Bacillus anthracis Sterne strain. The cytotoxicity of the extracts on Vero kidney cells was also determined. RESULTS: The minimum inhibitory concentration (MIC) values of the extracts against Bacillus anthracis Sterne strain ranged from 0.02 to 0.31 mg/ml. Excellent MIC values were observed for the following plant species: Maesa lanceolata (0.02 mg/ml), Bolusanthus speciosus, Hypericum roeperianum, Morus mesozygia (0.04 mg/ml) and Pittosporum viridiflorum (0.08 mg/ml). The total antibacterial activity of the extracts ranged from 92 to 5562 ml/g. Total activity presents the volume to which the extract from 1 g of plant material can be diluted and still inhibit microbial growth. Maesa lanceolata and Hypericum roeperianum had the highest total activity with values of 5562 and 2999 ml/g respectively. The extracts of Calpurnia aurea had the lowest total activity (92 ml/g). The cytotoxicity determined on Vero cells indicated that most of the extracts were relatively non-toxic compared to doxorubicin (LC50 8.3 ± 1.76 µg/ml), except for the extracts of Maesa lanceolata, Elaeodendron croceum and Calpurnia aurea with LC50 values at 2.38 ± 0.25, 5.20 ± 0.24 and 13 ± 2.26 µg/ml respectively. The selectivity index (SI) ranged from 0.02 to 1.66. Hypericum roeperianum had the best selectivity index, (SI = 1.66) and Elaeodendron croceum had lowest value (SI = 0.02). CONCLUSIONS: The crude acetone extracts of the selected plant species had promising antibacterial activity against Bacillus anthracis. Maesa lanceolata extracts could be useful as a disinfectant and Hypericum roeperianum could be useful to protect animals based on its high total activity and selectivity index. Further investigation of these plant extracts may lead to the development of new therapeutic agents to protect humans or animals against anthrax.