Repurposing Toremifene for Treatment of Oral Bacterial Infections

ABSTRACT The spread of antibiotic resistance and the challenges associated with antiseptics such as chlorhexidine have necessitated a search for new antibacterial agents against oral bacterial pathogens. As a result of failing traditional approaches, drug repurposing has emerged as a novel paradigm to find new antibacterial agents. In this study, we examined the effects of the FDA-approved anticancer agent toremifene against the oral bacteria Porphyromonas gingivalis and Streptococcus mutans. We found that the drug was able to inhibit the growth of both pathogens, as well as prevent biofilm formation, at concentrations ranging from 12.5 to 25 μM. Moreover, toremifene was shown to eradicate preformed biofilms at concentrations ranging from 25 to 50 μM. In addition, we found that toremifene prevents P. gingivalis and S. mutans biofilm formation on titanium surfaces. A time-kill study indicated that toremifene is bactericidal against S. mutans. Macromolecular synthesis assays revealed that treatment with toremifene does not cause preferential inhibition of DNA, RNA, or protein synthesis pathways, indicating membrane-damaging activity. Biophysical studies using fluorescent probes and fluorescence microscopy further confirmed the membrane-damaging mode of action. Taken together, our results suggest that the anticancer agent toremifene is a suitable candidate for further investigation for the development of new treatment strategies for oral bacterial infections.

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Bactericidal Activity, Absence of Serum Effect, and Time-Kill Kinetics of Ceftazidime-Avibactam against β-Lactamase-Producing Enterobacteriaceae and Pseudomonas aeruginosa

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02894-14 ◽

2014 ◽

Vol 58 (9) ◽

pp. 5297-5305 ◽

Cited By ~ 55

Author(s):

Tiffany R. Keepers ◽

Marcela Gomez ◽

Chris Celeri ◽

Wright W. Nichols ◽

Kevin M. Krause

Keyword(s):

Pseudomonas Aeruginosa ◽

Human Serum ◽

Bactericidal Activity ◽

Minimum Bactericidal Concentration ◽

Wild Type ◽

Content Type ◽

Positive Isolate ◽

New Treatment ◽

Time Kill ◽

Kinetics Of

ABSTRACTAvibactam, a non-β-lactam β-lactamase inhibitor with activity against extended-spectrum β-lactamases (ESBLs), KPC, AmpC, and some OXA enzymes, extends the antibacterial activity of ceftazidime against most ceftazidime-resistant organisms producing these enzymes. In this study, the bactericidal activity of ceftazidime-avibactam against 18Pseudomonas aeruginosaisolates and 15Enterobacteriaceaeisolates, including wild-type isolates and ESBL, KPC, and/or AmpC producers, was evaluated. Ceftazidime-avibactam MICs (0.016 to 32 μg/ml) were lower than those for ceftazidime alone (0.06 to ≥256 μg/ml) against all isolates except for 2P. aeruginosaisolates (1blaVIM-positive isolate and 1blaOXA-23-positive isolate). The minimum bactericidal concentration/MIC ratios of ceftazidime-avibactam were ≤4 for all isolates, indicating bactericidal activity. Human serum and human serum albumin had a minimal effect on ceftazidime-avibactam MICs. Ceftazidime-avibactam time-kill kinetics were evaluated at low MIC multiples and showed time-dependent reductions in the number of CFU/ml from 0 to 6 h for all strains tested. A ≥3-log10decrease in the number of CFU/ml was observed at 6 h for allEnterobacteriaceae, and a 2-log10reduction in the number of CFU/ml was observed at 6 h for 3 of the 6P. aeruginosaisolates. Regrowth was noted at 24 h for some of the isolates tested in time-kill assays. These data demonstrate the potent bactericidal activity of ceftazidime-avibactam and support the continued clinical development of ceftazidime-avibactam as a new treatment option for infections caused byEnterobacteriaceaeandP. aeruginosa, including isolates resistant to ceftazidime by mechanisms dependent on avibactam-sensitive β-lactamases.

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Toxin-Antitoxin Genes of the Gram-Positive Pathogen Streptococcus pneumoniae: So Few and Yet So Many

Microbiology and Molecular Biology Reviews ◽

10.1128/mmbr.00030-12 ◽

2012 ◽

Vol 76 (4) ◽

pp. 773-791 ◽

Cited By ~ 36

Author(s):

Wai Ting Chan ◽

Inma Moreno-Córdoba ◽

Chew Chieng Yeo ◽

Manuel Espinosa

Keyword(s):

Streptococcus Pneumoniae ◽

Biofilm Formation ◽

Bacterial Infections ◽

Host Cells ◽

Molecular Characteristics ◽

Type Ii ◽

Pneumococcal Infections ◽

Interesting Phenomenon ◽

Content Type ◽

Toxin Protein

SUMMARYPneumococcal infections cause up to 2 million deaths annually and raise a large economic burden and thus constitute an important threat to mankind. Because of the increase in the antibiotic resistance ofStreptococcus pneumoniaeclinical isolates, there is an urgent need to find new antimicrobial approaches to triumph over pneumococcal infections. Toxin-antitoxin (TA) systems (TAS), which are present in most living bacteria but not in eukaryotes, have been proposed as an effective strategy to combat bacterial infections. Type II TAS comprise a stable toxin and a labile antitoxin that form an innocuous TA complex under normal conditions. Under stress conditions, TA synthesis will be triggered, resulting in the degradation of the labile antitoxin and the release of the toxin protein, which would poison the host cells. The three functional chromosomal TAS fromS. pneumoniaethat have been studied as well as their molecular characteristics are discussed in detail in this review. Furthermore, a meticulous bioinformatics search has been performed for 48 pneumococcal genomes that are found in public databases, and more putative TAS, homologous to well-characterized ones, have been revealed. Strikingly, several unusual putative TAS, in terms of components and genetic organizations previously not envisaged, have been discovered and are further discussed. Previously, we reported a novel finding in which a unique pneumococcal DNA signature, the BOX element, affected the regulation of the pneumococcalyefM-yoeBTAS. This BOX element has also been found in some of the other pneumococcal TAS. In this review, we also discuss possible relationships between some of the pneumococcal TAS with pathogenicity, competence, biofilm formation, persistence, and an interesting phenomenon called bistability.

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ComparativeIn VitroActivities of Oritavancin, Dalbavancin, and Vancomycin against Methicillin-Resistant Staphylococcus aureus Isolates in a Nondividing State

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00169-16 ◽

2016 ◽

Vol 60 (7) ◽

pp. 4342-4345 ◽

Cited By ~ 8

Author(s):

Adam Belley ◽

David Lalonde Seguin ◽

Francis Arhin ◽

Greg Moeck

Keyword(s):

Staphylococcus Aureus ◽

Bactericidal Activity ◽

Antibacterial Agents ◽

Methicillin Resistant Staphylococcus Aureus ◽

Methicillin Resistant ◽

Content Type ◽

Skin Structure ◽

Persistent Infections ◽

Time Kill

ABSTRACTAntibacterial agents that kill nondividing bacteria may be of utility in treating persistent infections. Oritavancin and dalbavancin are bactericidal lipoglycopeptides that are approved for acute bacterial skin and skin structure infections in adults caused by susceptible Gram-positive pathogens. Using time-kill methodology, we demonstrate that oritavancin exerts bactericidal activity against methicillin-resistantStaphylococcus aureus(MRSA) isolates that are maintained in a nondividing statein vitro, whereas dalbavancin and the glycopeptide vancomycin do not.

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Murein Hydrolase LytF of Streptococcus sanguinis and the Ecological Consequences of Competence Development

Applied and Environmental Microbiology ◽

10.1128/aem.01709-17 ◽

2017 ◽

Vol 83 (24) ◽

Cited By ~ 5

Author(s):

Nyssa Cullin ◽

Sylvio Redanz ◽

Kirsten J. Lampi ◽

Justin Merritt ◽

Jens Kreth

Keyword(s):

Dental Caries ◽

Oral Cavity ◽

Cell Morphology ◽

Biofilm Formation ◽

Oral Bacteria ◽

Tooth Surface ◽

Content Type ◽

Streptococcus Sanguinis ◽

Murein Hydrolase ◽

Biofilm Development

ABSTRACT The overall health of the oral cavity is dependent on proper homeostasis between health-associated bacterial colonizers and bacteria known to promote dental caries. Streptococcus sanguinis is a health-associated commensal organism, a known early colonizer of the acquired tooth pellicle, and is naturally competent. We have shown that LytF, a competence-controlled murein hydrolase, is capable of inducing the release of extracellular DNA (eDNA) from oral bacteria. Precipitated LytF and purified LytF were used as treatments against planktonic cultures and biofilms. Larger amounts of eDNA were released from cultures treated with protein samples containing LytF. Additionally, LytF could affect biofilm formation and cellular morphology. Biofilm formation was significantly decreased in the lytF-complemented strain, in which increased amounts of LytF are present. The same strain also exhibited cell morphology defects in both planktonic cultures and biofilms. Furthermore, the LytF cell morphology phenotype was reproducible in wild-type cells using purified LytF protein. In sum, our findings demonstrate that LytF can induce the release of eDNA from oral bacteria, and they suggest that, without proper regulation of LytF, cells display morphological abnormalities that contribute to biofilm malformation. In the context of the oral biofilm, LytF may play important roles as part of the competence and biofilm development programs, as well as increasing the availability of eDNA. IMPORTANCE Streptococcus sanguinis, a commensal organism in the oral cavity and one of the pioneer colonizers of the tooth surface, is associated with the overall health of the oral environment. Our laboratory showed previously that, under aerobic conditions, S. sanguinis can produce H2O2 to inhibit the growth of bacterial species that promote dental caries. This production of H2O2 by S. sanguinis also induces the release of eDNA, which is essential for proper biofilm formation. Under anaerobic conditions, S. sanguinis does not produce H2O2 but DNA is still released. Determining how S. sanguinis releases DNA is thus essential to understand biofilm formation in the oral cavity.

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Thermoregulation of Pseudomonas aeruginosa Biofilm Formation

Applied and Environmental Microbiology ◽

10.1128/aem.01584-20 ◽

2020 ◽

Vol 86 (22) ◽

Author(s):

Suran Kim ◽

Xi-Hui Li ◽

Hyeon-Ji Hwang ◽

Joon-Hee Lee

Keyword(s):

Pseudomonas Aeruginosa ◽

Biofilm Formation ◽

Bacterial Infections ◽

Effect Of Temperature ◽

Cyclic Diguanylate ◽

Content Type ◽

Protection Mechanism ◽

Formation Pattern ◽

Temperature Ranges ◽

Different Strains

ABSTRACT We investigated the effect of temperature on the biofilm formation of Pseudomonas aeruginosa and revealed that the biofilm formation increased rapidly at temperatures lower than 25°C. P. aeruginosa formed the most robust biofilm of a conspicuous mushroom-like structure at 20°C. However, when the temperature increased to 25°C, the biofilm formation rapidly decreased. Above 25°C, as the temperature rose, the biofilm formation increased again little by little despite its less-structured form, indicating that 25°C is the low point of biofilm formation. The intracellular 3′,5′-cyclic diguanylate (c-di-GMP) levels also decreased rapidly as the temperature rose from 20 to 25°C. The expression levels of pelA, algD, and pslA encoding Pel, alginate, and Psl, respectively, were also dramatically affected by temperature, with pelA being regulated in a pattern similar to that of the intracellular c-di-GMP levels, and the pattern seen for algD regulation was the most similar to the actual biofilm formation pattern. Total exopolysaccharide production was thermoregulated and followed the regulation pattern of c-di-GMP. Interestingly, the thermoregulation patterns in biofilm formation were different depending on the strain of P. aeruginosa. Unlike PAO1, another strain, PA14, showed a gradual decrease in biofilm formation and c-di-GMP in the range of 20 to 37°C, and P. aeruginosa clinical isolates also showed slightly different patterns in biofilm formation in conjunction with temperature change, suggesting that different strains may sense different temperature ranges for biofilm formation. However, it is obvious that P. aeruginosa forms more biofilms at lower temperatures and that temperature is an important factor in determining the biofilm formation. IMPORTANCE Biofilm formation is an important protection mechanism used by most microorganisms and provides cells with many advantages, like high infectivity, antibiotic resistance, and strong survivability. Since most persistent bacterial infections are believed to be associated with biofilms, biofilm control is an important issue in medicine, environmental engineering, and industry. Biofilm formation is influenced by various environmental factors. Temperature is the most direct environmental cue encountered by microorganisms. Here, we investigated the effect of temperature on the biofilm formation of P. aeruginosa, a notorious pathogen, and found that temperature is an important factor determining the amount and structure of biofilms. Low temperatures greatly increase biofilm formation and give biofilms a highly conspicuous structure. Although thermoregulation of biofilm formation is mainly mediated by c-di-GMP, some c-di-GMP-independent regulations were also observed. This study shows how biofilms are formed at various temperatures and provides new insights to control biofilms using temperature.

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Multiple Drug-Induced Stress Responses Inhibit Formation of Escherichia coli Biofilms

Applied and Environmental Microbiology ◽

10.1128/aem.01113-20 ◽

2020 ◽

Vol 86 (21) ◽

Cited By ~ 1

Author(s):

Nataliya A. Teteneva ◽

Sergey V. Mart’yanov ◽

María Esteban-López ◽

Jörg Kahnt ◽

Timo Glatter ◽

...

Keyword(s):

Escherichia Coli ◽

Stress Responses ◽

Biofilm Formation ◽

Molecular Mechanisms ◽

Drug Repurposing ◽

Bacterial Attachment ◽

Bacterial Biofilm ◽

Multiple Drug ◽

Content Type ◽

Approved Drugs

ABSTRACT In most ecosystems, bacteria exist primarily as structured surface-associated biofilms that can be highly tolerant to antibiotics and thus represent an important health issue. Here, we explored drug repurposing as a strategy to identify new antibiofilm compounds, screening over 1,000 compounds from the Prestwick Chemical Library of approved drugs for specific activities that prevent biofilm formation by Escherichia coli. Most growth-inhibiting compounds, which include known antibacterial but also antiviral and other drugs, also reduced biofilm formation. However, we also identified several drugs that were biofilm inhibitory at doses where only a weak effect or no effect on planktonic growth could be observed. The activities of the most specific antibiofilm compounds were further characterized using gene expression analysis, proteomics, and microscopy. We observed that most of these drugs acted by repressing genes responsible for the production of curli, a major component of the E. coli biofilm matrix. This repression apparently occurred through the induction of several different stress responses, including DNA and cell wall damage, and homeostasis of divalent cations, demonstrating that biofilm formation can be inhibited through a variety of molecular mechanisms. One tested drug, tyloxapol, did not affect curli expression or cell growth but instead inhibited biofilm formation by suppressing bacterial attachment to the surface. IMPORTANCE The prevention of bacterial biofilm formation is one of the major current challenges in microbiology. Here, by systematically screening a large number of approved drugs for their ability to suppress biofilm formation by Escherichia coli, we identified a number of prospective antibiofilm compounds. We further demonstrated different mechanisms of action for individual compounds, from induction of replicative stress to disbalance of cation homeostasis to inhibition of bacterial attachment to the surface. Our work demonstrates the potential of drug repurposing for the prevention of bacterial biofilm formation and suggests that also for other bacteria, the activity spectrum of antibiofilm compounds is likely to be broad.

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Screening of the Dichloromethane: Methanolic Extract of Centella asiatica for Antibacterial Activities against Salmonella typhi, Escherichia coli, Shigella sonnei, Bacillus subtilis, and Staphylococcus aureus

The Scientific World JOURNAL ◽

10.1155/2020/6378712 ◽

2020 ◽

Vol 2020 ◽

pp. 1-8

Author(s):

Berick Moturi Sieberi ◽

George Isanda Omwenga ◽

Rachael Kitondo Wambua ◽

Judith Chemutai Samoei ◽

Mathew Piero Ngugi

Keyword(s):

Escherichia Coli ◽

Staphylococcus Aureus ◽

Bacillus Subtilis ◽

Bacterial Infections ◽

Antibacterial Agents ◽

Methanolic Extract ◽

Centella Asiatica ◽

Salmonella Typhi ◽

Phytochemical Screening ◽

Time Kill

Bacterial infections are responsible for a large number of deaths every year worldwide. On average, 80% of the African population cannot afford conventional drugs. Moreover, many synthetic antibiotics are associated with side effects and progressive increase in antimicrobial resistance. Currently, there is growing interest in discovering new antibacterial agents from ethnomedicinal plants. About 60% of the population living in developing countries depends on herbal drugs for healthcare needs. This study involved the screening of Centella asiatica commonly used by herbal medicine practitioners in Kisii County to treat symptoms related to bacterial infections. Standard bioassay methods were applied throughout the study. They included preliminary screening of dichloromethane: methanolic extract of Centella asiatica against human pathogenic bacteria including Salmonella typhi ATCC 19430, Escherichia coli ATCC 25922, Shigella sonnei ATCC 25931, Bacillus subtilis ATCC 21332, and Staphylococcus aureus ATCC 25923 using agar disc diffusion, broth microdilution method, and time-kill kinetics with tetracycline as a positive control. Phytochemical screening was carried out to determine the different classes of compounds in the crude extracts. Data were analyzed using one way ANOVA and means separated by Tukey’s test. Dichloromethane: methanolic extract of Centella asiatica was screened against the selected bacterial strains. Time-kill kinetic studies of the extracts showed dose- and time-dependent kinetics of antibacterial properties. Phytochemical screening of the DCM-MeOH extract revealed the presence of alkaloids, flavonoids, phenolics, terpenoids, cardiac glycosides, saponins, steroids, and tannins. The present study indicates that the tested plant can be an important source of antibacterial agents and recommends that the active phytoconstituents be isolated, identified, and screened individually for activities and also subjected further for in vivo and toxicological studies.

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Antibacterial and Antibiofilm Activities of a Novel Synthetic Cyclic Lipopeptide against Cariogenic Streptococcus mutans UA159

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00776-17 ◽

2017 ◽

Vol 61 (8) ◽

Cited By ~ 12

Author(s):

Kyung R. Min ◽

Adriana Galvis ◽

Brandon Williams ◽

Ramanjaneyulu Rayala ◽

Predrag Cudic ◽

...

Keyword(s):

Streptococcus Mutans ◽

Biofilm Formation ◽

Minimum Bactericidal Concentration ◽

The Novel ◽

Content Type ◽

Cyclic Lipopeptide ◽

Nonspecific Cytotoxicity ◽

Antimicrobial Treatments ◽

Time Kill

ABSTRACT Despite continuous efforts to control cariogenic dental biofilms, very few effective antimicrobial treatments exist. In this study, we characterized the activity of the novel synthetic cyclic lipopeptide 4 (CLP-4), derived from fusaricidin, against the cariogenic pathogen Streptococcus mutans UA159. We determined CLP-4's MIC, minimum bactericidal concentration (MBC), and spontaneous resistance frequency, and we performed time-kill assays. Additionally, we assessed CLP-4's potential to inhibit biofilm formation and eradicate preformed biofilms. Our results demonstrate that CLP-4 has strong antibacterial activity in vitro and is a potent bactericidal agent with low spontaneous resistance frequency. At a low concentration of 5 μg/ml, CLP-4 completely inhibited S. mutans UA159 biofilm formation, and at 50 μg/ml, it reduced the viability of established biofilms by >99.99%. We also assessed CLP-4's cytotoxicity and stability against proteolytic digestion. CLP-4 withstood trypsin or chymotrypsin digestion even after treatment for 24 h, and our toxicity studies showed that CLP-4 effective concentrations had negligible effects on hemolysis and the viability of human oral fibroblasts. In summary, our findings showed that CLP-4 is a potent antibacterial and antibiofilm agent with remarkable stability and low nonspecific cytotoxicity. Hence, CLP-4 is a promising novel antimicrobial peptide with potential for clinical application in the prevention and treatment of dental caries.

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Inhibition of Protein Secretion inEscherichia coliand Sub-MIC Effects of Arylomycin Antibiotics

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01253-18 ◽

2018 ◽

Vol 63 (2) ◽

pp. e01253-18 ◽

Cited By ~ 4

Author(s):

Shawn I. Walsh ◽

David S. Peters ◽

Peter A. Smith ◽

Arryn Craney ◽

Melissa M. Dix ◽

...

Keyword(s):

Mass Spectrometry ◽

Biofilm Formation ◽

Bacterial Infections ◽

Protein Identification ◽

Secretory Pathway ◽

Signal Peptidase ◽

Type I ◽

Content Type ◽

Multidimensional Protein Identification Technology ◽

Bacterial Type

ABSTRACTAt sufficient concentrations, antibiotics effectively eradicate many bacterial infections. However, during therapy, bacteria are unavoidably exposed to lower antibiotic concentrations, and sub-MIC exposure can result in a wide variety of other effects, including the induction of virulence, which can complicate therapy, or horizontal gene transfer (HGT), which can accelerate the spread of resistance genes. Bacterial type I signal peptidase (SPase) is an essential protein that acts at the final step of the general secretory pathway. This pathway is required for the secretion of many proteins, including many required for virulence, and the arylomycins are a class of natural product antibiotics that target SPase. Here, we investigated the consequences of exposingEscherichia colicultures to sub-MIC levels of an arylomycin. Using multidimensional protein identification technology mass spectrometry, we found that arylomycin treatment inhibits the proper extracytoplasmic localization of many proteins, both those that appear to be SPase substrates and several that do not. The identified proteins are involved in a broad range of extracytoplasmic processes and include a number of virulence factors. The effects of arylomycin on several processes required for virulence were then individually examined, and we found that, at even sub-MIC levels, the arylomycins potently inhibit flagellation, motility, biofilm formation, and the dissemination of antibiotic resistance via HGT. Thus, we conclude that the arylomycins represent promising novel therapeutics with the potential to eradicate infections while simultaneously reducing virulence and the dissemination of resistance.

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Azidothymidine Produces Synergistic Activity in Combination with Colistin against Antibiotic-Resistant Enterobacteriaceae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01630-18 ◽

2018 ◽

Vol 63 (1) ◽

Cited By ~ 11

Author(s):

Yanmin Hu ◽

Yingjun Liu ◽

Anthony Coates

Keyword(s):

Bacterial Infections ◽

Therapeutic Potential ◽

Infection Model ◽

Synergistic Activity ◽

Antibiotic Resistant ◽

Content Type ◽

E Coli ◽

Broth Microdilution Method ◽

Peritoneal Infection ◽

Time Kill

ABSTRACT Bacterial infections remain a leading killer worldwide, which is worsened by the continuous emergence of antibiotic resistance. In particular, antibiotic-resistant Enterobacteriaceae are prevalent and extremely difficult to treat. Repurposing existing drugs and improving the therapeutic potential of existing antibiotics represent an attractive novel strategy. Azidothymidine (AZT) is an antiretroviral drug which is used in combination with other antivirals to prevent and to treat HIV/AIDS. AZT is also active against Gram-negative bacteria but has not been developed for that purpose. Here, we investigated the in vitro and in vivo efficacy of AZT in combination with colistin against antibiotic-resistant Enterobacteriaceae, including strains producing extended-spectrum beta-lactamases (ESBLs) or New Delhi metallo-beta-lactamase 1 (NDM) or carrying mobilized colistin resistance (mcr-1). The MIC was determined using the broth microdilution method. The combined effect of AZT and colistin was examined using the checkerboard method and time-kill analysis. A murine peritoneal infection model was used to test the therapeutic effect of the combination of AZT and colistin. The fractional inhibitory concentration index from the checkerboard assay demonstrated that AZT synergized with colistin against 61% and 87% of ESBL-producing Escherichia coli and Klebsiella pneumoniae strains, respectively, 100% of NDM-1-producing strains, and 92% of mcr-1-producing E. coli strains. Time-kill analysis demonstrated significant synergistic activities when AZT was combined with colistin. In a murine peritoneal infection model, AZT in combination with colistin showed augmented activities of both drugs in the treatment of NDM-1 K. pneumoniae and mcr-1 E. coli infections. The AZT and colistin combination possesses a potential to be used coherently to treat antibiotic-resistant Enterobacteriaceae infections.

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