Modified poly(L-lysine)-based structures as novel antimicrobials for diabetic foot infections, an in-vitro study

Background: Wound infections occur as sequelae to skin trauma and cause significant hospitalizations, morbidity and mortality. Skin traumas arise more frequently in those with diabetes or cardiovascular disease and in these settings, may be chronic with poorer outcomes including lower limb amputation. Treatment of chronic wound infection is challenging due to antibiotic resistance and biofilm formation by bacteria including S. aureus and P. aeruginosa, which are among the most frequent causative pathogens. Managing these challenging infections requires new molecules and modalities. Methods: We evaluated antimicrobial and anti-biofilm activity of star-shaped poly(L-lysine) (PLL) polymers against S. aureus and P. aeruginosa strains and clinical isolates recovered from wounds including diabetic foot wounds (DFW) in a Dublin Hospital in 2019. A star-shaped PLL polypeptide series, specifically G2(8)PLL20, G3(16)PLL10, G4(32)PLL5 with variation in polypeptide chain length and arm-multiplicity, were compared to a linear peptide, PLL160 with equivalent number of lysine residues. Results: All PLLs, including the linear polypeptide, were bactericidal at 1mM against S. aureus 25923 and P. aeruginosa PAO1, with log reduction in colony forming units/ml between 2.7-3.6. PLL160 demonstrated similar killing potency against 20 S. aureus and five P. aeruginosa clinical isolates from DFW, mean log reductions: 3.04 ± 0.16 and 3.96 ± 0.82 respectively after 1 hour incubation. Potent anti-biofilm activity was demonstrated against S. aureus 25923 but for clinical isolates, low to moderate loss of biofilm viability was shown using PLL160 and G3(16)PLL10 at 50 mM (S. aureus) and 200 mM (P. aeruginosa) with high inter-isolate variability. In the star-shaped architecture, antimicrobial activity was retained with incorporation of 5-mer hydrophobic amino-acid modifications to the arms of the polypeptides (series G3(16)PLL20-coPLT5, G3(16)PLL20-coPLI5, G3(16)PLL20-coPLP5). Conclusion: These polypeptides offer structural flexibility for clinical applications and have potential for further development, particularly in the setting of diabetic foot and other chronic wound infections.

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Development of Cephradine-Loaded Gelatin/Polyvinyl Alcohol Electrospun Nanofibers for Effective Diabetic Wound Healing: In-Vitro and In-Vivo Assessments

Pharmaceutics ◽

10.3390/pharmaceutics13030349 ◽

2021 ◽

Vol 13 (3) ◽

pp. 349

Author(s):

Anam Razzaq ◽

Zaheer Ullah Khan ◽

Aasim Saeed ◽

Kiramat Ali Shah ◽

Naveed Ullah Khan ◽

...

Keyword(s):

Wound Healing ◽

Polyvinyl Alcohol ◽

Mtt Assay ◽

Chronic Wound ◽

Drug Loading ◽

Group Identification ◽

Wound Infections ◽

Diabetic Wound

Diabetic wound infections caused by conventional antibiotic-resistant Staphylococcus aureus strains are fast emerging, leading to life-threatening situations (e.g., high costs, morbidity, and mortality) associated with delayed healing and chronic inflammation. Electrospinning is one of the most widely used techniques for the fabrication of nanofibers (NFs), induced by a high voltage applied to a drug-loaded polymer solution. Particular attention is given to electrospun NFs for pharmaceutical applications (e.g., original drug delivery systems) and tissue regeneration (e.g., as tissue scaffolds). However, there is a paucity of reports related to their application in diabetic wound infections. Therefore, we prepared eco-friendly, biodegradable, low-immunogenic, and biocompatible gelatin (GEL)/polyvinyl alcohol (PVA) electrospun NFs (BNFs), in which we loaded the broad-spectrum antibiotic cephradine (Ceph). The resulting drug-loaded NFs (LNFs) were characterized physically using ultraviolet-visible (UV-Vis) spectrophotometry (for drug loading capacity (LC), drug encapsulation efficiency (EE), and drug release kinetics determination), thermogravimetric analysis (TGA) (for thermostability evaluation), scanning electron microscopy (SEM) (for surface morphology analysis), and Fourier-transform infrared spectroscopy (FTIR) (for functional group identification). LNFs were further characterized biologically by in-vitro assessment of their potency against S. aureus clinical strains (N = 16) using the Kirby–Bauer test and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, by ex-vivo assessment to evaluate their cytotoxicity against primary human epidermal keratinocytes using MTT assay, and by in-vivo assessment to estimate their diabetic chronic wound-healing efficiency using NcZ10 diabetic/obese mice (N = 18). Thin and uniform NFs with a smooth surface and standard size (<400 nm) were observed by SEM at the optimized 5:5 (GEL:PVA) volumetric ratio. FTIR analyses confirmed the drug loading into BNFs. Compared to free Ceph, LNFs were significantly more thermostable and exhibited sustained/controlled Ceph release. LNFs also exerted a significantly stronger antibacterial activity both in-vitro and in-vivo. LNFs were significantly safer and more efficient for bacterial clearance-induced faster chronic wound healing. LNF-based therapy could be employed as a valuable dressing material to heal S. aureus-induced chronic wounds in diabetic subjects.

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An in-vitro study of carbapenem-induced morphological changes and endotoxin release in clinical isolates of gram-negative bacilli

Journal of Antimicrobial Chemotherapy ◽

10.1093/jac/41.4.435 ◽

1998 ◽

Vol 41 (4) ◽

pp. 435-442 ◽

Cited By ~ 37

Author(s):

T Horii

Keyword(s):

Morphological Changes ◽

In Vitro Study ◽

Vitro Study ◽

Clinical Isolates ◽

Gram Negative

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Volatile Oils ofNepeta tenuifolia(Jing Jie) as an Alternative Medicine against Multidrug-Resistant Pathogenic Microbes

Canadian Journal of Infectious Diseases and Medical Microbiology ◽

10.1155/2018/8347403 ◽

2018 ◽

Vol 2018 ◽

pp. 1-8 ◽

Cited By ~ 1

Author(s):

Yi-Hsuan Lee ◽

Chao-Min Wang ◽

Po-Yu Liu ◽

Ching-Chang Cheng ◽

Zong-Yen Wu ◽

...

Keyword(s):

Essential Oils ◽

In Vitro Study ◽

Multidrug Resistant ◽

Clinical Isolates ◽

Gram Negative ◽

E Coli ◽

Wide Range ◽

Main Component ◽

Reference Strains

Essential oils from the dried spikes ofNepeta tenuifolia(Benth) are obtained by steam distillation. Pulegone was identified as the main component in the spikes ofN. tenuifoliathrough analysis, with greater than 85% purity obtained in this study. The essential oils are extremely active against all Gram-positive and some Gram-negative reference bacteria, particularlySalmonella enterica,Citrobacter freundii, andEscherichia coli. The minimum inhibitory concentration was found to be between 0.08 and 0.78% (againstS. enterica), 0.39 and 0.78% (againstC. freundii), and 0.097 and 0.39% (againstE. coli), whereas the minimum bactericidal concentration varied in range from 0.097% to 1.04%. In general, the essential oils show a strong inhibitory action against all tested reference strains and clinical isolates. However, the antibacterial activity of EOs against bothPseudomonas aeruginosareference strains and clinical isolates was relatively lower than other Gram-negative pathogens. The essential oils ofN. tenuifoliaalso displayed bactericidal activities (MBC/MIC < 4) in this study. These findings reflect the bactericidal activity of the essential oils against a wide range of multidrug-resistant clinical pathogens in an in vitro study. In addition, we propose the fragmentation pathways of pulegone and its derivatives by LC-ESI-MS/MS in this study.

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Evaluation of a Bioengineered Honey and Its Synthetic Equivalent as Novel Staphylococcus aureus Biofilm-Targeted Topical Therapies in Chronic Rhinosinusitis

American Journal of Rhinology and Allergy ◽

10.1177/1945892419874700 ◽

2019 ◽

Vol 34 (1) ◽

pp. 80-86 ◽

Cited By ~ 1

Author(s):

Dionyssia Papadopoulou ◽

Alicja Dabrowska ◽

Philip G. Harries ◽

Jeremy S. Webb ◽

Raymond N. Allan ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Antimicrobial Activity ◽

Chronic Rhinosinusitis ◽

In Vitro Study ◽

Sinus Surgery ◽

Bacterial Strains ◽

Methicillin Resistant ◽

Log Reduction ◽

Lower Minimum Inhibitory Concentration

Background Chronic rhinosinusitis (CRS) is a common condition which affects the quality of life of millions of patients worldwide and has a significant impact on health-care resources. While Staphylococcus aureus bacterial biofilms play an important role in this disease, antimicrobial therapy is rarely effective and may promote antibiotic resistance. Thus, development of novel biofilm-targeting and antibiotic-sparing therapies is highly desirable and urgently required. Objective This in vitro study evaluated the antimicrobial activity of a novel synthetic honey-equivalent product which was designed to have the same reactive oxygen release profile as the engineered honey SurgihoneyRO™. Methods Treatment efficacy was investigated by assessment of planktonic growth, biofilm viability, thickness, and biomass using 12 CRS-related S. aureus mucosal bacterial strains. Results Both SurgihoneyRO™ and the synthetic honey-equivalent product inhibited growth of planktonic methicillin-resistant and methicillin-sensitive S. aureus strains, with the synthetic honey-equivalent product exhibiting a lower minimum inhibitory concentration. Treatment of established S. aureus biofilms reduced biofilm viability with 24-hour treatment resulting in a 2-log reduction in viability of biofilms formed by methicillin-resistant strains and a 1-log reduction in biofilms formed by methicillin-sensitive strains. Conclusions This preliminary study shows that the synthetic honey-equivalent product provides marked antimicrobial activity against S. aureus biofilms, with the potential for development in the clinical setting as an adjunctive biofilm-targeted therapy in CRS. The ultimate aim of such a product would be to reduce the need for antibiotics, steroids, and invasive surgical procedures in CRS patients as well as improving clinical outcomes following endoscopic sinus surgery.

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d-Amino Acids Enhance the Activity of Antimicrobials against Biofilms of Clinical Wound Isolates of Staphylococcus aureus and Pseudomonas aeruginosa

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02468-14 ◽

2014 ◽

Vol 58 (8) ◽

pp. 4353-4361 ◽

Cited By ~ 57

Author(s):

Carlos J. Sanchez ◽

Kevin S. Akers ◽

Desiree R. Romano ◽

Ronald L. Woodbury ◽

Sharanda K. Hardy ◽

...

Keyword(s):

Amino Acids ◽

Staphylococcus Aureus ◽

Pseudomonas Aeruginosa ◽

Clinical Isolates ◽

Chronic Infections ◽

Content Type ◽

Log Reduction ◽

Viable Bacteria ◽

Biofilm Bacteria

ABSTRACTWithin wounds, microorganisms predominantly exist as biofilms. Biofilms are associated with chronic infections and represent a tremendous clinical challenge. As antibiotics are often ineffective against biofilms, use of dispersal agents as adjunctive, topical therapies for the treatment of wound infections involving biofilms has gained interest. We evaluatedin vitrothe dispersive activity ofd-amino acids (d-AAs) on biofilms from clinical wound isolates ofStaphylococcus aureusandPseudomonas aeruginosa; moreover, we determined whether combinations ofd-AAs and antibiotics (clindamycin, cefazolin, oxacillin, rifampin, and vancomycin forS. aureusand amikacin, colistin, ciprofloxacin, imipenem, and ceftazidime forP. aeruginosa) enhance activity against biofilms.d-Met,d-Phe, andd-Trp at concentrations of ≥5 mM effectively dispersed preformed biofilms ofS. aureusandP. aeruginosaclinical isolates, an effect that was enhanced when they were combined as an equimolar mixture (d-Met/d-Phe/d-Trp). When combined withd-AAs, the activity of rifampin was significantly enhanced against biofilms of clinical isolates ofS. aureus, as indicated by a reduction in the minimum biofilm inhibitory concentration (MBIC) (from 32 to 8 μg/ml) and a >2-log reduction of viable biofilm bacteria compared to treatment with antibiotic alone. The addition ofd-AAs was also observed to enhance the activity of colistin and ciprofloxacin against biofilms ofP. aeruginosa, reducing the observed MBIC and the number of viable bacteria by >2 logs and 1 log at 64 and 32 μg/ml in contrast to antibiotics alone. These findings indicate that the biofilm dispersal activity ofd-AAs may represent an effective strategy, in combination with antimicrobials, to release bacteria from biofilms, subsequently enhancing antimicrobial activity.

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In VitroAntibacterial and Antibiofilm Activities of Chlorogenic Acid against Clinical Isolates ofStenotrophomonas maltophiliaincluding the Trimethoprim/Sulfamethoxazole Resistant Strain

BioMed Research International ◽

10.1155/2013/392058 ◽

2013 ◽

Vol 2013 ◽

pp. 1-7 ◽

Cited By ~ 47

Author(s):

Arunkumar Karunanidhi ◽

Renjan Thomas ◽

Alex van Belkum ◽

Vasanthakumari Neela

Keyword(s):

Chlorogenic Acid ◽

Inhibition Zone ◽

Clinical Isolates ◽

Antibiofilm Activity ◽

Log Reduction ◽

Fold Reduction ◽

Viable Bacteria ◽

Study Support ◽

Time Kill

Thein vitroantibacterial and antibiofilm activity of chlorogenic acid against clinical isolates ofStenotrophomonas maltophiliawas investigated through disk diffusion, minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), time-kill and biofilm assays. A total of 9 clinicalS. maltophiliaisolates including one isolate resistant to trimethoprim/sulfamethoxazole (TMP/SMX) were tested. The inhibition zone sizes for the isolates ranged from 17 to 29 mm, while the MIC and MBC values ranged from 8 to 16 μg mL−1and 16 to 32 μg mL−1. Chlorogenic acid appeared to be strongly bactericidal at 4x MIC, with a 2-log reduction in viable bacteria at 10 h.In vitroantibiofilm testing showed a 4-fold reduction in biofilm viability at 4x MIC compared to 1x MIC values (0.085<0.397A 490 nm) of chlorogenic acid. The data from this study support the notion that the chlorogenic acid has promisingin vitroantibacterial and antibiofilm activities againstS. maltophilia.

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Hypertonic glucose inhibits growth and attenuates virulence factors of multidrug-resistant Pseudomonas aeruginosa

10.21203/rs.2.21617/v3 ◽

2020 ◽

Author(s):

Tao Chen ◽

Ye Xu ◽

Wenya Xu ◽

Wenli Liao ◽

Chunquan Xu ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Virulence Factors ◽

Diabetic Foot ◽

Biofilm Formation ◽

Chronic Wound ◽

Multidrug Resistant ◽

Wound Infections ◽

Hypertonic Glucose ◽

Diabetic Foot Infections ◽

Chronic Wound Infections

Abstract Background: Pseudomonas aeruginosa is the most common Gram-negative pathogen responsible for chronic wound infections, such as diabetic foot infections, and further exacerbates the treatment options and cost of such conditions. Hypertonic glucose, a commonly used prolotherapy solution, can accelerate the proliferation of granulation tissue and improve microcirculation in wounds. However, the action of hypertonic glucose on bacterial pathogens that infect wounds is unclear. In this study, we investigated the inhibitory effects of hypertonic glucose on multidrug-resistant P. aeruginosa strains isolated from diabetic foot infections. Hypertonic glucose represents a novel approach to control chronic wound infections caused by P. aeruginosa. Results: Four multidrug-resistant P. aeruginosa clinical strains isolated from diabetic foot ulcers from a tertiary hospital in China and the reference P. aeruginosa PAO1 strain were studied. Hypertonic glucose significantly inhibited the growth, biofilm formation, and swimming motility of P. aeruginosa clinical strains and PAO1. Furthermore, hypertonic glucose significantly reduced the production of pyocyanin and elastase virulence factors in P. aeruginosa. The expression of major quorum sensing genes (lasI, lasR, rhlI, and rhlR) in P. aeruginosa were all downregulated in response to hypertonic glucose treatment. In a Galleria mellonella larvae infection model, the administration of hypertonic glucose was shown to increase the survival rates of larvae infected by P. aeruginosa strains (3/5).Conclusions: Hypertonic glucose inhibited the growth, biofilm formation, and swimming motility of P. aeruginosa, as well as reduced the production of virulence factors and quorum sensing gene expression. Further studies that investigate hypertonic glucose therapy should be considered in treating chronic wound infections.

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