Screening of medicinal plant extracts as novel DNA gyrase inhibitors

Bioactivities of a number of medicinal plants; Alkanna tinctoria (L.) Tausch, Alnus glutinosa (L.) Gaertn., Calamintha nepeta Willk. and C. nepeta, Centaurea iberica Trevir. ex Spreng., Citrus paradisi Macfad., C. paradisi, Citrus sinensis (L.) Osbeck, Colutea cilicica Boiss. & Balansa, Cotinus coggygria Scop., Cuscuta arvensis Beyr. ex Engelm., Equisetum palustre L., Lapsana communis L., Laurus nobilis L., Olea europea L., Plantago major L., Rhus coriaria L, Salvia verticillata L., Sambucus ebulus L., Sedum acre L, Thymus capitatus (L.) Hoffmanns. & Link, T. capitatus, Thymbra spicata L., T. spicata (n: 20), which are used for the prevention and treatment of diverse diseases, were investigated. The antimicrobial activities of extracts were evaluated using broth microdilution assay. The cytotoxicities of extracts were investigated on HeLa cell line by MTT assay. Statistical analysis was performed using GraphPad Prism (5.0). The effects of the extracts, which have the highest antimicrobial activity, on the Escherichia coli and Staphylococcus aureus DNA gyrase gene expression were determined by using quantitative reverse transcription polymerase chain reaction (qRT-PCR). The MICs (µg/ml) of extracts were determined as 32-64, 2-128, 8-128,1-128, 4-128 against Gram-positive, Gram-negative bacteria, yeasts, dermatophytes, and Mycobacterium spp., respectively. No cytotoxicity has been observed in plant extracts tested. DNA gyrase activity was determined for T. capitatus-SFE (128 µg/mL) and L. nobilis-Hx (128 µg/mL) extracts according to the inhibition of DNA gyrase gene expression. Overall, T. capitatus-SFE and L. nobilis-Hx are good candidates for further antimicrobial studies.

Comparative in vitro antimicrobial activities of CSA-142 and CSA-192, second-generation ceragenins, with CSA-13 against various microorganisms

Ceragenins, a novel family of antibiotics, are cationic derivatives of cholic acid and display broad-spectrum antimicrobial activities. Multiple ceragenins have been synthesized and studied, and most published data are with ceragenin CSA-13. This study aimed to investigate the in vitro antimicrobial properties of second-generation ceragenins, CSA-142 and CSA-192, and compare them to CSA-13. The antimicrobial activities of CSA-13, CSA-142 and CSA-192 were studied against 20 strains of Escherichia coli, Acinetobacter baumannii, Pseudomonas aeruginosa, Staphylococcus aureus and Candida albicans. Time-kill curve methods were performed to determine the bactericidal and fungicidal activities of the ceragenins against three strains of each microorganism. Overall, CSA-13 showed the lowest MIC values, but also CSA-142 and CSA-192 had good activities against tested microorganisms. The killing curves showed that ceragenins generally had bactericidal and fungicidal effects. The bactericidal and fungicidal behaviours of CSA-142 and CSA-192 may make them good alternative agents to CSA-13

Effects of Ceragenins and Antimicrobial Peptides on the A549 Cell Line and an In Vitro Co-Culture Model of A549 Cells and <i>Pseudomonas aeruginosa</i>

Pseudomonas aeruginosa is an important pathogen that can adhere to host tissues and epithelial surfaces, especially during chronic infections such as cystic fibrosis (CF) lung infections. The effect of ceragenins and antimicrobial peptides (AMP) on this colonization was investigated in a co-culture infection model. After determining the antimicrobial effects of the substances on P. aeruginosa planktonic cells, their cytotoxicity on the A549 cell line was also determined. After the A549 cell line was infected with P. aeruginosa, the effect of antimicrobials on intracellular bacteria as well as the effects in inhibiting the adhesion of P. aeruginosa were investigated. In addition, LDH release from cells was determined by performing an LDH experiment to understand the cytotoxicity of bacterial infection and antimicrobial treatment on cells. CSA-131 was determined as the antimicrobial agent with the highest antimicrobial activity, while the antimicrobial effects of AMPs were found to be much lower than those of ceragenins. The antimicrobial with the lowest IC50 value was determined as the combination of CSA-131 with Pluronic F127. CSA-13 has been determined to be the most effective antimicrobial with its effectiveness to both intracellular bacteria and bacterial adhesion. Nevertheless, further safety, efficacy, toxicity, and pharmacological studies of ceragenins are needed to evaluate clinical utility

Comparative Fungicidal Activities of N-Chlorotaurine and Conventional Antiseptics against Candida spp. Isolated from Vulvovaginal Candidiasis

N-chlorotaurine (NCT), the N-chloro derivative of the amino acid taurine, is a long-lived oxidant produced by stimulated human leucocytes. NCT has antimicrobial activities which are generally enhanced in the presence of organic material. The aim of this study was to investigate fungicidal effects of NCT and conventional antiseptics against Candida isolated from vulvovaginal candidiasis (VVC). Chlorhexidine (CHX, 1.6%), octenidine dihydrochloride (OCT, 0.08%), povidone iodine (PVP-I, 8%), boric acid (8%), and NCT (0.1% (5.5 mM)) were evaluated against forty-four Candida isolates, according to European Standard methods, at 30, 60, 90, and 120 min and 24 h in the presence of skim milk as an organic material. CHX, OCT, and PVP-I showed rapid fungicidal activity against all Candida isolates with 5&ndash;6 log10 reduction of viable counts after 30 min, whereas boric acid and NCT needed 1 h against Candida albicans and 2 h against non-albicans Candida for a significant 3 log10 reduction. NCT showed fungicidal activity (defined as &ge;4 log10 reduction) against C. albicans within 90 min and C. non&ndash;albicans within 24 h. Based upon all presently available data, including our results, NCT could be used as a new agent for treatment of local fungal infections such as VVC

In vitro activity of ceragenins against Burkholderia cepacia complex

Burkholderia cepacia complex (Bcc) species are aerobic, Gram-negative and non-fermantative bacilli. Bcc can cause clinical symptoms in patients with cystic fibrosis, ranging from asymptomatic carriage to fatal pneumonia. A pressing need exists for new antimicrobial agents that target Bcc. Ceragenins, CSA-13, CSA-131 and CSA-131 with 5% Pluronic (R) F127 (CSA-131P), were evaluated against Bcc clinical isolates (n = 42). MICs of ceragenins and conventional antibiotics were determined. Time-kill curve experiments were performed with 1x, 4x MICs of ceragenins and sulfamethoxazole-trimethoprim (SXT), levofloxacin. MIC50/ MIC90 results (mg l(-1)) of CSA-13, CSA-131 and CSA-131P were determined as 16/64, 16/128 and 16/128, respectively. CSA-13 and CSA-131 showed bactericidal activity. CSA-13 - levofloxacin combination displayed synergistic activity against Bcc. First-generation (CSA-13) and second-generation (CSA-131 and CSA-131P) ceragenins have significant antimicrobial effects on Bcc. The findings of this study demonstrate that combinations of ceragenins with currently marketed antibiotics could be synergistic in vitro against Bcc isolates. These results suggest that combination therapy with conventional antibiotics could be an alternative approach for treating Bcc infections in the future

Antimicrobial effect of the extracts from Hypericum perforatum against oral bacteria and biofilm formation

Context: One traditional medicines, Hypericum perforatum L. (Hypericaceae), possesses several beneficial effects against depression, ulcers, dyspepsia, abdominal pains, burns, bacterial infections, migraine headaches, and sciatica. Objective: The present study investigates the antimicrobial activity of the ethanol extract (HP-EtOH) of H. perforatum and its sub-extracts, namely n-hexane (HP-hexane), chloroform (HP-CHCl3), ethyl acetate (HP-EtOAc), n-butanol (HP-n-BuOH), and water (HP-H2O) extracts, against Streptococcus mutans, S. sobrinus, Lactobacillus plantarum, and Enterococcus faecalis. Materials and methods: For the evaluation of the antimicrobial activity, flowering aerial parts of H. perforatum were extracted with EtOH and then this extract was fractionated to obtain five sub-extracts in different polarities. Antimicrobial activities of HP-EtOH and its sub-extracts against Streptococcus mutans, S. sobrinus, L. plantarum, and E. faecalis were assessed by using colorimetric micro-well dilution at concentration ranges of 64-0.5 mu g/ml as well as resazurin microplate and modified microtiter-plate assays between the ranges of 100 and 0.78125 mu g/ml. Results: According to the results of the present study, HP-H2O sub-extract displayed strong antibacterial activity (MIC values 8 mu g/mL) against S. sobrinus and L. plantarum, and exerted moderate activity against S. mutans and E. faecalis at 32 and 16 mu g/mL concentrations, respectively. Other sub-extracts also demonstrated antimicrobial activity against S. sobrinus at a concentration of 16 mu g/mL. HP-EtOAc and HP-n-BuOH showed antimicrobial activity against L. plantarum and HP-EtOAc and HP-H2O were also active against E. faecalis at the same concentrations (16 mu g/mL). Conclusion: According to the results, we suggest that H. perforatum could be employed as a natural antibacterial agent in oral care products

Ceragenins exhibiting promising antimicrobial activity against various multidrug resistant Gram negative bacteria

Ceragenins are novel promising agents for the treatment of infections caused by multi-drug resistant microorganisms. Since colistin resistance has become a worldwide problem, the need for new treatment agents has been increasing steadily. Therefore, this study aimed to investigate in vitro antimicrobial activities of ceragenins (Cationic Steroid Antibiotics) (CSA-8, CSA-13, CSA-142 and CSA-192) against multidrug resistant Gram negative isolates from Turkey. Experiments were performed by using broth microdilution method against Klebsiella pneumoniae, Morganella morganii, Pseudomonas aeruginosa and Stenotrophomonas maltophilia isolates. All microorganisms except for three isolates were identified as multidrug resistant. Among tested ceragenins, CSA-13 showed the best results (MIC: 8-64 mu g/ml). Nevertheless, the antimicrobial activity of CSA-8 was not significant. In conclusion, ceragenins appear to be a good candidate as antimicrobial therapy in the presence of multidrug (including colistin) resistant microorganisms

Immunization against poly--acetylglucosamine reduces neutrophil activation and GVHD while sparing microbial diversity

Microbial invasion into the intestinal mucosa after allogeneic hematopoietic cell transplantation (allo-HCT) triggers neutrophil activation and requires antibiotic interventions to prevent sepsis. However, antibiotics lead to a loss of microbiota diversity, which is connected to a higher incidence of acute graft-versus-host disease (aGVHD). Antimicrobial therapies that eliminate invading bacteria and reduce neutrophil-mediated damage without reducing the diversity of the microbiota are therefore highly desirable. A potential solution would be the use of antimicrobial antibodies that target invading pathogens, ultimately leading to their elimination by innate immune cells. In a mouse model of aGVHD, we investigated the potency of active and passive immunization against the conserved microbial surface polysaccharide poly--acetylglucosamine (PNAG) that is expressed on numerous pathogens. Treatment with monoclonal or polyclonal antibodies to PNAG (anti-PNAG) or vaccination against PNAG reduced aGVHD-related mortality. Anti-PNAG treatment did not change the intestinal microbial diversity as determined by 16S ribosomal DNA sequencing. Anti-PNAG treatment reduced myeloperoxidase activation and proliferation of neutrophil granulocytes (neutrophils) in the ileum of mice developing GVHD. In vitro, anti-PNAG treatment showed high antimicrobial activity. The functional role of neutrophils was confirmed by using neutrophil-deficient mice that had no survival advantage under anti-PNAG treatment. In summary, the control of invading bacteria by anti-PNAG treatment could be a novel approach to reduce the uncontrolled neutrophil activation that promotes early GVHD and opens a new avenue to interfere with aGVHD without affecting commensal intestinal microbial diversity