Engineered Bacteriophage as a Delivery Vehicle for Antibacterial Protein, SASP

The difficulties in developing novel classes of antibacterials is leading to a resurgence of interest in bacteriophages as therapeutic agents, and in particular engineered phages that can be optimally designed. Here, pre-clinical microbiology assessment is presented of a Staphylococcus aureus phage engineered to deliver a gene encoding an antibacterial small acid soluble spore protein (SASP) and further, rendered non-lytic to give product SASPject PT1.2. PT1.2 has been developed initially for nasal decolonisation of S. aureus, including methicillin-resistant S. aureus. Time-kill curve assays were conducted with PT1.2 against a range of staphylococcal species, and serial passaging experiments were conducted to investigate the potential for resistance to develop. SASPject PT1.2 demonstrates activity against 100% of 225 geographically diverse S. aureus isolates, exquisite specificity for S. aureus, and a rapid speed of kill. The kinetics of S. aureus/PT1.2 interaction is examined together with demonstrating that PT1.2 activity is unaffected by the presence of human serum albumin. SASPject PT1.2 shows a low propensity for resistance to develop with no consistent shift in sensitivity in S. aureus cells passaged for up to 42 days. SASPject PT1.2 shows promise as a novel first-in-class antibacterial agent and demonstrates potential for the SASPject platform.

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Trans-Cinnamaldehyde Exhibits Synergy with Conventional Antibiotic against Methicillin-Resistant Staphylococcus aureus

International Journal of Molecular Sciences ◽

10.3390/ijms22052752 ◽

2021 ◽

Vol 22 (5) ◽

pp. 2752

Author(s):

Shu Wang ◽

Ok-Hwa Kang ◽

Dong-Yeul Kwon

Keyword(s):

Staphylococcus Aureus ◽

Methicillin Resistant Staphylococcus Aureus ◽

Synergistic Effects ◽

Regulatory Gene ◽

Western Blot Assay ◽

Methicillin Resistant ◽

Wide Range ◽

Kill Curve ◽

Conventional Antibiotics ◽

Time Kill

Methicillin-resistant Staphylococcus aureus (MRSA) is a major nosocomial pathogen worldwide and has acquired multiple resistance to a wide range of antibiotics. Hence, there is a pressing need to explore novel strategies to overcome the increase in antimicrobial resistance. The present study aims to investigate the efficacy and mechanism of plant-derived antimicrobials, trans-cinnamaldehyde (TCA) in decreasing MRSA’s resistance to eight conventional antibiotics. A checkerboard dilution test and time–kill curve assay are used to determine the synergistic effects of TCA combined with the antibiotics. The results indicated that TCA increased the antibacterial activity of the antibiotics by 2-16-fold. To study the mechanism of the synergism, we analyzed the mecA transcription gene and the penicillin-binding protein 2a level of MRSA treated with TCA by quantitative RT-PCR or Western blot assay. The gene transcription and the protein level were significantly inhibited. Additionally, it was verified that TCA can significantly inhibit the biofilm, which is highly resistant to antibiotics. The expression of the biofilm regulatory gene hld of MRSA after TCA treatment was also significantly downregulated. These findings suggest that TCA maybe is an exceptionally potent modulator of antibiotics.

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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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Exploitation of the IDO Pathway in the Therapy of Rheumatoid Arthritis

International Journal of Tryptophan Research ◽

10.4137/ijtr.s11737 ◽

2013 ◽

Vol 6s1 ◽

pp. IJTR.S11737 ◽

Cited By ~ 5

Author(s):

Richard O. Williams

Keyword(s):

Rheumatoid Arthritis ◽

Anthranilic Acid ◽

Kynurenine Pathway ◽

Gene Encoding ◽

Rate Limiting Step ◽

Tryptophan Metabolites ◽

Synthetic Derivative ◽

Rate Limiting ◽

Kinetics Of ◽

Draining Lymph Nodes

Indoleamine 2,3-dioxygenase (IDO) is the first and rate-limiting step along the kynurenine pathway and is thought to play a key role in immune homeostasis through depletion of tryptophan and accumulation of kynurenines. In this review we summarize recent research into the possibility of harnessing the IDO pathway for the therapy of rheumatoid arthritis. Inhibition of IDO activity, or knockout of the gene encoding IDO, was shown to cause an increase in the severity of collagen-induced arthritis, an animal model of rheumatoid arthritis. The increased severity of disease was associated with elevated numbers of pathogenic Th1 and Th17 cells in the joints and draining lymph nodes. In another study, analysis of the kinetics of expression of downstream kynurenine pathway enzymes during the course of arthritis revealed a potential role for tryptophan metabolites in resolution of arthritis. Furthermore, the therapeutic administration of L-kynurenine or [3,4-dimethoxycinnamonyl]-anthranilic acid (a synthetic derivative of 3-hydroxy-anthranilic acid) significantly reduced both clinical and histological progression of experimental arthritis. These findings raise the possibility of exploiting the IDO pathway for the therapy of autoimmune disease.

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Automated CUT&Tag profiling of chromatin heterogeneity in mixed-lineage leukemia

Nature Genetics ◽

10.1038/s41588-021-00941-9 ◽

2021 ◽

Author(s):

Derek H. Janssens ◽

Michael P. Meers ◽

Steven J. Wu ◽

Ekaterina Babaeva ◽

Soheil Meshinchi ◽

...

Keyword(s):

Single Cell ◽

Binding Sites ◽

Therapeutic Agents ◽

Mixed Lineage Leukemia ◽

Regulatory Proteins ◽

Chromosomal Translocations ◽

Tumor Subtype ◽

Gene Encoding ◽

Lysine Methyltransferase ◽

Chromatin Profiling

AbstractAcute myeloid and lymphoid leukemias often harbor chromosomal translocations involving the KMT2A gene, encoding the KMT2A lysine methyltransferase (also known as mixed-lineage leukemia-1), and produce in-frame fusions of KMT2A to other chromatin-regulatory proteins. Here we map fusion-specific targets across the genome for diverse KMT2A oncofusion proteins in cell lines and patient samples. By modifying CUT&Tag chromatin profiling for full automation, we identify common and tumor-subtype-specific sites of aberrant chromatin regulation induced by KMT2A oncofusion proteins. A subset of KMT2A oncofusion-binding sites are marked by bivalent (H3K4me3 and H3K27me3) chromatin signatures, and single-cell CUT&Tag profiling reveals that these sites display cell-to-cell heterogeneity suggestive of lineage plasticity. In addition, we find that aberrant enrichment of H3K4me3 in gene bodies is sensitive to Menin inhibitors, demonstrating the utility of automated chromatin profiling for identifying therapeutic vulnerabilities. Thus, integration of automated and single-cell CUT&Tag can uncover epigenomic heterogeneity within patient samples and predict sensitivity to therapeutic agents.

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A Study of the Disruptive Effect of the Acetate Fraction of Punica granatum Extract on Cryptococcus Biofilms

Frontiers in Microbiology ◽

10.3389/fmicb.2020.568258 ◽

2021 ◽

Vol 11 ◽

Author(s):

Paulo C. M. Villis ◽

Alessandra T. de Macedo ◽

Haryne L. A. Furtado ◽

Pedro H. C. Fontenelle ◽

Ingrid S. Gonçalves ◽

...

Keyword(s):

Treatment Options ◽

Folk Medicine ◽

Antimicrobial Effect ◽

Punica Granatum ◽

Cryptococcus Laurentii ◽

Disruptive Effect ◽

Worldwide Distribution ◽

Kill Curve ◽

Therapeutic Alternatives ◽

Time Kill

Cryptococcosis, caused by yeasts of the genus Cryptococcus, is an infectious disease with a worldwide distribution. Cryptococcus neoformans and Cryptococcus gattii are the species that commonly cause this disease in humans; however, infections caused by Cryptococcus laurentii, especially in immunocompromised patients, are increasingly being reported. Owing to the increase in the resistance of fungi to antifungals, and a lack of treatment options, it is important to seek new therapeutic alternatives such as natural products. Among these are plant species such as Punica granatum, which is used in folk medicine to treat various diseases. This study aimed to evaluate the activity of the acetate fraction of P. granatum leaf extract against environmental and clinical isolates of Cryptococcus. Three environmental isolates of C. laurentii, PMN, PMA, and PJL II, isolated from soils of different municipalities in the state of Maranhão, a clinical isolate, C. gattii, from a patient with neurocryptococcosis, and a standard strain of C. gattii (ATCC 32068) were used. The minimum and fractional inhibitory concentrations (MIC and FIC, respectively) and time-kill curve of the extract and fluconazole were determined to assess the susceptibility profile of the fungal isolates. Larvae of Tenebrio molitor were infected with Cryptococcus strains, and the effects of acetate fraction of P. granatum extract and fluconazole on the survival and fungal burden were determined. The extract activity was tested against pre-formed biofilms. The acetate fraction of P. granatum extract showed promising antifungal activity against all the species of Cryptococcus evaluated in this study, with an MIC value lower than that of fluconazole. The indices obtained in the FIC test indicated that the antimicrobial effect of the combination of the extract and antifungal was indifferent for 80% of the isolates. The P. granatum acetate fraction reduced the pre-formed biofilm of some isolates, showing better activity than fluconazole, which is consistent with results from fluorescence microscopy. This is the first study on the use of P. granatum and its ability to inhibit Cryptococcus biofilms; therefore, further studies and tests are needed to investigate the components and mechanism of action of P. granatum against cryptococcosis agents.

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Synergistic bactericide and antibiotic effects of dimeric, tetrameric, or palindromic peptides containing the RWQWR motif against Gram-positive and Gram-negative strains

RSC Advances ◽

10.1039/c9ra00708c ◽

2019 ◽

Vol 9 (13) ◽

pp. 7239-7245 ◽

Cited By ~ 8

Author(s):

Yerly Vargas-Casanova ◽

Andrea Verónica Rodríguez-Mayor ◽

Karen Johanna Cardenas ◽

Aura Lucía Leal-Castro ◽

Liliana Constanza Muñoz-Molina ◽

...

Keyword(s):

Gram Positive ◽

Gram Negative ◽

Kill Curve ◽

Blue Line ◽

Time Kill

Time-kill curve plot. Peptide LfcinB (20–25)4againstS. aureusATCC 25923. The peptide was tested at concentrations corresponding to MIC (blue line), 2 MIC (pink line) and 4 MIC (orange line) values.

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Pharmacodynamic Analysis of the Activity of Quinupristin-Dalfopristin against Vancomycin-ResistantEnterococcus faecium with Differing MBCs via Time-Kill-Curve and Postantibiotic Effect Methods

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.42.9.2188 ◽

1998 ◽

Vol 42 (9) ◽

pp. 2188-2192 ◽

Cited By ~ 24

Author(s):

Jeffrey R. Aeschlimann ◽

Michael J. Rybak

Keyword(s):

Antibacterial Activities ◽

Water Soluble ◽

Postantibiotic Effect ◽

Curve Method ◽

Highly Correlated ◽

Kill Curve ◽

The Individual ◽

Time Kill

ABSTRACT Quinupristin-dalfopristin (Q-D) is a new water-soluble, semisynthetic antibiotic that is derived from natural streptogramins and that is combined in a 30:70 ratio. A number of studies have described the pharmacodynamic properties of this drug, but most have investigated only staphylococci or streptococci. We evaluated the relationship between Q-D, quinupristin (Q), and/or dalfopristin (D) susceptibility parameters and antibacterial activities against 22 clinical isolates of vancomycin-resistant Enterococcus faecium (VREF) by using the concentration-time-kill-curve method and by measuring postantibiotic effects. Q-D, Q, and D MICs and minimum bactericidal concentrations (MBCs) ranged from 0.125 to 1 and 0.25 to 64, 8 to 512 and >512, and 2 to 8 and 8 to 512 μg/ml, respectively. There were no significant relationships between susceptibilities to the individual components and the susceptibilities to the Q-D combination product. In the time-kill-curves studies, Q-D at a concentration of 6 μg/ml was at least bacteriostatic against all VREF tested. There was increased activity against more susceptible isolates when the isolates were grouped either by Q-D MBCs or by Q MICs. By multivariate regression analyses, the percent change in the inoculum from that at the baseline was significantly correlated with the Q MIC (R = 0.74; P = 0.008) and the Q-D concentration-to-MBC ratio (R = 0.58;P = 0.02) and was inversely correlated with the Q-D MBC-to-MIC ratio (R = 0.68; P = 0.003). A strong correlation existed between the killing rate and the Q-D concentration-to-MBC ratio (R = 0.99;P < 0.0001). Time to 99.9% killing was best correlated with the Q-D MBC (R = 0.96;P < 0.0001). The postantibiotic effect ranged from 0.2 to 3.2 h and was highly correlated with the Q-D concentration-to-MBC ratio (R = 0.96;P < 0.0001) and was less highly correlated with the Q MIC (R = 0.42; P = 0.04). Further study of these relationships with in vitro or in vivo infection models that simulate Q-D pharmacokinetics should further define the utility of these pharmacodynamic parameters in the prediction of Q-D activity for the treatment of VREF infections in humans.

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In vitro antimicrobial combination testing of and evolution of resistance to the first-in-class spiropyrimidinetrione zoliflodacin combined with six therapeutically relevant antimicrobials for Neisseria gonorrhoeae

Journal of Antimicrobial Chemotherapy ◽

10.1093/jac/dkz376 ◽

2019 ◽

Vol 74 (12) ◽

pp. 3521-3529 ◽

Cited By ~ 4

Author(s):

Sunniva Foerster ◽

George Drusano ◽

Daniel Golparian ◽

Michael Neely ◽

Laura J V Piddock ◽

...

Keyword(s):

Neisseria Gonorrhoeae ◽

Single Agent ◽

Curve Analysis ◽

Phase Iii ◽

Transmitted Infection ◽

Kill Curve ◽

Time Kill ◽

Selection Of ◽

Selection Of Resistance

Abstract Objectives Resistance in Neisseria gonorrhoeae to all gonorrhoea therapeutic antimicrobials has emerged. Novel therapeutic antimicrobials are imperative and the first-in-class spiropyrimidinetrione zoliflodacin appears promising. Zoliflodacin could be introduced in dual antimicrobial therapies to prevent the emergence and/or spread of resistance. We investigated the in vitro activity of and selection of resistance to zoliflodacin alone and in combination with six gonorrhoea therapeutic antimicrobials against N. gonorrhoeae. Methods The international gonococcal reference strains WHO F (WT) and WHO O, WHO V and WHO X (strains with different AMR profiles) were examined. Zoliflodacin was evaluated alone or combined with ceftriaxone, cefixime, spectinomycin, gentamicin, tetracycline, cethromycin or sitafloxacin in chequerboard assays, time–kill curve analysis and selection-of-resistance studies. Results Zoliflodacin alone or in combination with all six antimicrobials showed rapid growth inhibition against all examined strains. The time–kill curve analysis indicated that tetracycline or cethromycin combined with zoliflodacin can significantly decrease the zoliflodacin kill rate in vitro. The frequency of selected zoliflodacin-resistance mutations was low when evaluated as a single agent and further reduced for all antimicrobial combinations. All resistant mutants contained the GyrB mutations D429N, K450T or K450N, resulting in zoliflodacin MICs of 0.5–4 mg/L. Conclusions Zoliflodacin, alone or in combination with sexually transmitted infection therapeutic antimicrobials, rapidly kills gonococci with infrequent resistance emergence. Zoliflodacin remains promising for gonorrhoea oral monotherapy and as part of dual antimicrobial therapy with low resistance emergence potential. A Phase III trial evaluating efficacy and safety of zoliflodacin for uncomplicated gonorrhoea treatment is planned in 2019.

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