Gentamicin Eluting 3D-Printed Implants for Preventing Post-Surgical Infections in Bone Fractures.

A surgically implantable device is an inevitable treatment option for millions of people worldwide suffering from diseases arising from orthopedic injuries. A global paradigm shift is currently underway to tailor and personalize replacement or reconstructive joints. Additive manufacturing (AM) has provided dynamic outflow to the customized fabrication of orthopedic implants by enabling need-based design and surface modification possibilities. Surgical grade 316L Stainless Steel (316L SS) is promising with its cost, strength, composition, and corrosion resistance to fabricate 3D implants. This work investigates the possibilities of application of the laser powder bed fusion (L-PBF) technique to fabricate 3D-printed (3DP) implants, which are functionalized with a multilayered antimicrobial coating to treat potential complications arising due to postsurgical infections (PSIs). Postsurgical implant-associated infection is a primary reason for implantation failure and is complicated mainly by bacterial colonization and biofilm formation at the installation site. PLGA (poly-d,l-lactide-co-glycolide), a biodegradable polymer, was utilized to impart multiple layers of coating using the airbrush spray technique on 3DP implant surfaces loaded with gentamicin (GEN). Various PLGA-based polymers were tested to optimize the ideal lactic acid: glycolic acid ratio and molecular weight suited for our investigation. 3D-Printed PLGA-GEN substrates sustained the release of gentamicin from the surface for approximately 6 weeks. The 3DP surface modification with PLGA-GEN facilitated cell adhesion and proliferation compared to control surfaces. The cell viability studies showed that the implants were safe for application. The 3DP PLGA-GEN substrates showed good concentration-dependent antibacterial efficacy against the common PSI pathogen Staphylococcus aureus (S. aureus) and Staphylococcus epidermidis (S. epidermidis). The GEN-loaded substrates demonstrated antimicrobial longevity and showed significant biofilm growth inhibition compared to control. The substrates offered great versatility regarding the in vitro release rates, antimicrobial properties, and biocompatibility studies. These results radiate great potential in future human and veterinary clinical applications pertinent to complications arising from PSIs, focusing on personalized sustained antibiotic delivery.

Dissociating antibacterial from ototoxic effects of gentamicin C-subtypes.

Gentamicin is a potent broad-spectrum aminoglycoside antibiotic whose use is hampered by ototoxic side-effects. Hospital gentamicin is a mixture of five gentamicin C-subtypes and several impurities of various ranges of nonexact concentrations. We developed a purification strategy enabling assaying of individual C-subtypes and impurities for ototoxicity and antimicrobial activity. We found that C-subtypes displayed broad and potent in vitro antimicrobial activities comparable to the hospital gentamicin mixture. In contrast, they showed different degrees of ototoxicity in cochlear explants, with gentamicin C2b being the least and gentamicin C2 the most ototoxic. Structure-activity relationships identified sites in the C4'-C6' region on ring I that reduced ototoxicity while preserving antimicrobial activity, thus identifying targets for future drug design and mechanisms for hair cell toxicity. Structure-activity relationship data suggested and electrophysiological data showed that the C-subtypes both bind and permeate the hair cell mechanotransducer channel, with the stronger the binding the less ototoxic the compound. Finally, both individual and reformulated mixtures of C-subtypes demonstrated decreased ototoxicity while maintaining antimicrobial activity, thereby serving as a proof-of-concept of drug reformulation to minimizing ototoxicity of gentamicin in patients.

Hair cell uptake of gentamicin in the developing mouse utricle.

Intratympanic injection of gentamicin has proven to be an effective therapy for intractable vestibular dysfunction. However, most studies to date have focused on the cochlea, so little is known about the distribution and uptake of gentamicin by the counterpart of the auditory system, specifically vestibular hair cells (HCs). Here, with a combination of in vivo and in vitro approaches, we used a gentamicin-Texas Red (GTTR) conjugate to investigate the mechanisms of gentamicin vestibulotoxicity in the developing mammalian utricular HCs. In vivo, GTTR fluorescence was concentrated in the apical cytoplasm and the cellular membrane of neonatal utricular HCs, but scarce in the nucleus of HCs and supporting cells. Quantitative analysis showed the GTTR uptake by striolar HCs was significantly higher than that in the extrastriola. In addition, the GTTR fluorescence intensity in the striola was increased gradually from 1 to 8 days, peaking at 8-9 days postnatally. In vitro, utricle explants were incubated with GTTR and candidate uptake conduits, including mechanotransduction (MET) channels and endocytosis in the HC, were inhibited separately. GTTR uptake by HCs could be inhibited by quinine, a blocker of MET channels, under both normal and stressed conditions. Meanwhile, endocytic inhibition only reduced GTTR uptake in the CoCl2 hypoxia model. In sum, the maturation of MET channels mediated uptake of GTTR into vestibular HCs. Under stressed conditions, MET channels play a pronounced role, manifested by channel-dependent stress enhanced GTTR permeation, while endocytosis participates in GTTR entry in a more selective manner.

Pyridoxal-5'-phosphate-dependent enzyme GenB3 Catalyzes C-3',4'-dideoxygenation in gentamicin biosynthesis.

BACKGROUND: The C-3',4'-dideoxygenation structure in gentamicin can prevent deactivation by aminoglycoside 3'-phosphotransferase (APH(3')) in drug-resistant pathogens. However, the enzyme catalyzing the dideoxygenation step in the gentamicin biosynthesis pathway remains unknown. RESULTS: Here, we report that GenP catalyzes 3' phosphorylation of the gentamicin biosynthesis intermediates JI-20A, JI-20Ba, and JI-20B. We further demonstrate that the pyridoxal-5'-phosphate (PLP)-dependent enzyme GenB3 uses these phosphorylated substrates to form 3',4'-dideoxy-4',5'-ene-6'-oxo products. The following C-6'-transamination and the GenB4-catalyzed reduction of 4',5'-olefin lead to the formation of gentamicin C. To the best of our knowledge, GenB3 is the first PLP-dependent enzyme catalyzing dideoxygenation in aminoglycoside biosynthesis. CONCLUSIONS: This discovery solves a long-standing puzzle in gentamicin biosynthesis and enriches our knowledge of the chemistry of PLP-dependent enzymes. Interestingly, these results demonstrate that to evade APH(3') deactivation by pathogens, the gentamicin producers evolved a smart strategy, which utilized their own APH(3') to activate hydroxyls as leaving groups for the 3',4'-dideoxygenation in gentamicin biosynthesis.

A novel human donor cornea preservation cocktail incorporating a thermo-reversible gelation polymer (TGP), enhancing the corneal endothelial cell density maintenance and explant culture of corneal limbal cells.

PURPOSE: McCarey-Kaufman's (MK) medium and Optisol-GS medium are the most commonly employed media for human donor corneal preservation. In this study, we evaluated the preservation efficacy of discarded human donor corneas using a Thermo-reversible gelation polymer (TGP) added to these two media. METHODS: Thirteen human corneal buttons collected from deceased donors, which were otherwise discarded due to low endothelial cell density (ECD) were used. They were stored in four groups: MK medium, MK medium with TGP, Optisol-GS and Optisol-GS with TGP at 4 °C for 96 h. Slit lamp examination and specular microscopy were performed. Corneal limbal tissues from these corneas were then cultured using explant methodology one with and the other without TGP scaffold, for 21 days. RESULTS: MK + TGP and Optisol-GS + TGP preserved corneas better than without TGP, which was observed by maintenance of ECD which was significantly higher in Optisol-GS + TGP than MK + TGP (p-value = 0.000478) and corneal thickness remaining the same for 96 h. Viable corneal epithelial cells could be grown from the corneas stored only in MK + TGP and Optisol-GS + TGP. During culture, the TGP scaffold helped maintain the native epithelial phenotype and progenitor/stem cell growth was confirmed by RT-PCR characterization. CONCLUSION: TGP reconstituted with MK and Optisol-GS media yields better preservation of human corneal buttons in terms of relatively higher ECD maintenance and better in vitro culture outcome of corneal limbal tissue. This method has the potential to become a standard donor corneal transportation-preservation methodology and it can also be extended to other tissue or organ transportation upon further validation.

Preparation of Cross-linked Chitosan Quaternary Ammonium Salt Hydrogel Films Loading Drug of Gentamicin Sulfate for Antibacterial Wound Dressing.

Hydrogels, possessing high biocompatibility and adaptability to biological tissue, show great usability in medical applications. In this research, a series of novel cross-linked chitosan quaternary ammonium salt loading with gentamicin sulfate (CTMCSG) hydrogel films with different cross-linking degrees were successfully obtained by the reaction of chitosan quaternary ammonium salt (TMCS) and epichlorohydrin. Fourier transform infrared spectroscopy (FTIR), thermal analysis, and scanning electron microscope (SEM) were used to characterize the chemical structure and surface morphology of CTMCSG hydrogel films. The physicochemical property, gentamicin sulphate release behavior, cytotoxicity, and antibacterial activity of the CTMCSG against Escherichia coli and Staphylococcus aureus were determined. Experimental results demonstrated that CTMCSG hydrogel films exhibited good water stability, thermal stability, drug release capacity, as well as antibacterial property. The inhibition zone of CTMCSG hydrogel films against Escherichia coli and Staphylococcus aureus could be up to about 30 mm. Specifically, the increases in maximum decomposition temperature, mechanical property, water content, swelling degree, and a reduction in water vapor permeability of the hydrogel films were observed as the amount of the cross-linking agent increased. The results indicated that the CTMCSG-4 hydrogel film with an interesting physicochemical property, admirable antibacterial activity, and slight cytotoxicity showed the potential value as excellent antibacterial wound dressing.

In situ drug release measuring in α-TCP cement by electrochemical impedance spectroscopy.

The use of drug delivery systems is a good technique to leave the right quantity of medicine in the patient's body in a suitable dose, because the drug application is delivered directly to the affected region. The current techniques such as HPLC and UV-Vis for the drug delivery calculation has some disadvantages, as the accuracy and the loss of the sample after characterization. With the aim of reducing the amount of material used during the characterization and have a non-destructive test with instantaneous results, the present paper shows the possibility of using electrochemical impedance spectroscopy (EIS) to have a drug delivery measurement during the release phenomena for a calcium phosphate cement (CFC) delivery system with gentamicin sulfate (GS) and lidocaine hydrochloride (LH), at a ratio of 1% and 2%, respectively. The equivalent circuit and the chemical mechanism involved during the measurements have been proposed as a tool to determine the drug delivery profile. The method has been compared with the UV-Vis technique. XRD was realized to verify conditions, before and after release. It was possible to verify the potential for using EIS as an instant technique to quantify drug delivery.

Analysis of histamine and sisomicin in gentamicin: Search for the causative agents of adverse effects.

In 1998, the aminoglycoside antibiotic gentamicin sulfate caused several cases of deaths in the United States, after the switch from twice- to once-daily application. Endotoxins were discussed as the cause for the adverse effects and sisomicin was identified as the lead impurity; batches containing sisomicin were contaminated with more impurities and were responsible for the fatalities. In 2016, anaphylactic reactions in horses, and later in humans with one fatality, were observed after application of gentamicin sulfate contaminated with histamine. To determine whether histamine was responsible for the 1990s death cases as well, histamine was quantified by means of liquid chromatography-tandem mass spectrometry (LC-MS/MS) in 30 samples of gentamicin sulfate analyzed in previous studies. Furthermore, a relative quantification of sisomicin was performed to check for a correlation between histamine and the lead impurity. A maximum amount of 11.52 ppm histamine was detected, which is below the limit for anaphylactic reactions of 16 ppm, and no correlation of the two impurities was observed. However, the European Medicines Agency recommends a stricter limit with regard to the maximum single dose of gentamicin sulfate to reach a greater gap between the maximum histamine exposition of 4.3 µg and the quantity known to cause hypotension of 7 µg. The low amounts of histamine and the fact that there is no connection with the contamination with sisomicin showed that histamine was not the cause for the death cases in the United States in 1998, and endotoxins remain the most probable explanation.

Biomediated synthesis, characterization, and biological applications of nickel oxide nanoparticles derived from Toona ciliata, Ficus carica and Pinus roxburghii.

Biomediated ecofriendly method for the synthesis of nickel oxide nanoparticles using plants extracts (Toona ciliata, Ficus carica and Pinus roxburghii) has been reported. The nanoparticles so obtained were characterized by various techniques such as ultraviolet-visible, powder X-ray diffraction, Fourier transform infrared spectroscopy, attenuated total reflectance spectroscopy, scanning electron microscopy, transmission electron microscopy, energy dispersive X-ray spectroscopy, thermogravimetric analysis and fluorescence spectroscopy. Formation of nickel oxide nanoparticles was confirmed by Fourier transform infrared spectroscopy and X-ray diffraction where the former technique ascertains the formation of bond between nickel and oxygen. The nickel oxide nanoparticles were found to be crystalline cubic face centered and show intense photoluminescence emission at 416, 414 and 413 nm, respectively. The antibacterial activity was studied against gram positive and gram negative bacterial species by agar well diffusion method. The nickel oxide nanoparticles show better activity against some bacterial strains with reference to the standard drugs Ciprofloxacin and Gentamicin. The anthelmintic activity against Pheretima posthuma of nanomaterials obtained from Pinus roxburghii was found to be greater than that derived from Toona ciliata and Ficus carica using the standard drug Albendazole. This method takes the advantage of the sustainable and economic approach for the synthesis of metal oxide nanoparticles.

A promising technology for wound healing; in-vitro and in-vivo evaluation of chitosan nano-biocomposite films containing gentamicin.

Aim: This paper aims to study in-vitro and in-vivo evaluation of chitosan (CHI) biocomposite of gentamicin nanoparticles (GNPs) for wound healing. Methods: In this study, CHI nanoparticles (NPs) were prepared using the ionic gelation technique. GNP biocomposites were examined on the excision wound model in Wistar rats to determine the in-vivo efficiency. Results: The diameter and zeta potential of NPs were between 151-212.9 nm and 37.2 - 51.1 mV, respectively. The entrapment efficiency was in an acceptable range of 36.6-42.7% w/w. The release test information was fitted to mathematical models (Zero, First order, Higuchi, and Korsmeyer-Peppas), and according to calculations, the kinetics of drug release followed the Korsmeyer-Peppas model. A comparison of thermograms revealed that the drug was present in the formulation in a non-crystalline form. Conclusion: Histological studies of the wound showed that the rate of skin tissue repair was higher in the GNP biocomposite treatment group than in the others.

Facile Synthesis of Bio-Antimicrobials with "Smart" Triiodides.

Multi-drug resistant pathogens are a rising danger for the future of mankind. Iodine (I2) is a centuries-old microbicide, but leads to skin discoloration, irritation, and uncontrolled iodine release. Plants rich in phytochemicals have a long history in basic health care. Aloe Vera Barbadensis Miller (AV) and Salvia officinalis L. (Sage) are effectively utilized against different ailments. Previously, we investigated the antimicrobial activities of smart triiodides and iodinated AV hybrids. In this work, we combined iodine with Sage extracts and pure AV gel with polyvinylpyrrolidone (PVP) as an encapsulating and stabilizing agent. Fourier transform infrared spectroscopy (FT-IR), Ultraviolet-visible spectroscopy (UV-Vis), Surface-Enhanced Raman Spectroscopy (SERS), microstructural analysis by scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), and X-Ray-Diffraction (XRD) analysis verified the composition of AV-PVP-Sage-I2. Antimicrobial properties were investigated by disc diffusion method against 10 reference microbial strains in comparison to gentamicin and nystatin. We impregnated surgical sutures with our biohybrid and tested their inhibitory effects. AV-PVP-Sage-I2 showed excellent to intermediate antimicrobial activity in discs and sutures. The iodine within the polymeric biomaterial AV-PVP-Sage-I2 and the synergistic action of the two plant extracts enhanced the microbial inhibition. Our compound has potential for use as an antifungal agent, disinfectant and coating material on sutures to prevent surgical site infections.

Ion Exchange Nanoparticles for Ophthalmic Drug Delivery.

We report here on ion-exchange polymeric nanoparticles from a linear copolymer of maleic anhydride methyl vinyl ether esterified with 30% octadecanol. The side chains for the polymer structure were optimized through metadynamics simulations, which revealed the use of octadecanol esters generates ideal free energy surfaces for drug encapsulation and release. Nanoparticles were synthesized using a solvent evaporation-precipitation method by mixing the polymer solution in acetone into water; upon acetone evaporation, a nanodispersion with an average particle size of ∼150 nm was obtained. Gentamicin sulfate, possessing five amino groups, was spontaneously entrapped in the nanocarrier by ionic interactions. Encapsulation efficiency increases significantly with the increase in pH and ionic strength. In vivo results demonstrate high gentamicin (GM) content in the enteric chamber (AUC 8207 ± 1334 (µg min)/mL) compared to 3% GM solution (AUC 2024 ± 438 (µg min)/mL). The formulation was also able to significantly extend the release of gentamicin when applied to rabbit cornea. These anionic nanoparticles can be used for extended-release of other cationic drugs.

The bifunctional enzyme, GenB4, catalyzes the last step of gentamicin 3',4'-di-deoxygenation via reduction and transamination activities.

BACKGROUND: New semi-synthetic aminoglycoside antibiotics generally use chemical modifications to avoid inactivity from pathogens. One of the most used modifications is 3',4'-di-deoxygenation, which imitates the structure of gentamicin. However, the mechanism of di-deoxygenation has not been clearly elucidated. RESULTS: Here, we report that the bifunctional enzyme, GenB4, catalyzes the last step of gentamicin 3',4'-di-deoxygenation via reduction and transamination activities. Following disruption of genB4 in wild-type M. echinospora, its products accumulated in 6'-deamino-6'-oxoverdamicin (1), verdamicin C2a (2), and its epimer, verdamicin C2 (3). Following disruption of genB4 in M. echinospora ΔgenK, its products accumulated in sisomicin (4) and 6'-N-methylsisomicin (5, G-52). Following in vitro catalytic reactions, GenB4 transformed sisomicin (4) to gentamicin C1a (9) and transformed verdamicin C2a (2) and its epimer, verdamicin C2 (3), to gentamicin C2a (11) and gentamicin C2 (12), respectively. CONCLUSION: This finding indicated that in addition to its transamination activity, GenB4 exhibits specific 4',5' double-bond reducing activity and is responsible for the last step of gentamicin 3',4'-di-deoxygenation. Taken together, we propose three new intermediates that may refine and supplement the specific biosynthetic pathway of gentamicin C components and lay the foundation for the complete elucidation of di-deoxygenation mechanisms.

A porphycene-gentamicin conjugate for enhanced photodynamic inactivation of bacteria.

A novel photoantimicrobial agent, namely 2-aminothiazolo[4,5-c]-2,7,12,17-tetrakis(methoxyethyl)porphycene (ATAZTMPo-gentamicin) conjugate, has been prepared by a click reaction between the red-light absorbing 9-isothiocyanate-2,7,12,17-tetrakis(methoxyethyl)porphycene (9-ITMPo) and the antibiotic gentamicin. The conjugate exhibits submicromolar activity in vitro against both Gram-positive and Gram-negative bacteria (Staphylococcus aureus and Escherichia coli, respectively) upon exposure to red light and is devoid of any cytotoxicity in the dark. The conjugate outperforms the two components delivered separately, which may be used to enhance the therapeutic index of gentamicin, broaden the spectrum of pathogens against which it is effective and reduce its side effects. Additionally, we report a novel straightforward synthesis of 2,7,12,17-tetrakis(methoxyethyl) porphycene (TMPo) that decreases the number of steps from nine to six.

Gentamicin B1 is a minor gentamicin component with major nonsense mutation suppression activity.

Nonsense mutations underlie about 10% of rare genetic disease cases. They introduce a premature termination codon (PTC) and prevent the formation of full-length protein. Pharmaceutical gentamicin, a mixture of several related aminoglycosides, is a frequently used antibiotic in humans that can induce PTC readthrough and suppress nonsense mutations at high concentrations. However, testing of gentamicin in clinical trials has shown that safe doses of this drug produce weak and variable readthrough activity that is insufficient for use as therapy. In this study we show that the major components of pharmaceutical gentamicin lack PTC readthrough activity but the minor component gentamicin B1 (B1) is a potent readthrough inducer. Molecular dynamics simulations reveal the importance of ring I of B1 in establishing a ribosome configuration that permits pairing of a near-cognate complex at a PTC. B1 induced readthrough at all three nonsense codons in cultured cancer cells with TP53 (tumor protein p53) mutations, in cells from patients with nonsense mutations in the TPP1 (tripeptidyl peptidase 1), DMD (dystrophin), SMARCAL1 (SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a-like 1), and COL7A1 (collagen type VII alpha 1 chain) genes, and in an in vivo tumor xenograft model. The B1 content of pharmaceutical gentamicin is highly variable and major gentamicins suppress the PTC readthrough activity of B1. Purified B1 provides a consistent and effective source of PTC readthrough activity to study the potential of nonsense suppression for treatment of rare genetic disorders.

Fabrication of Gentamicin-Loaded Hydroxyapatite/Collagen Bone-Like Nanocomposite for Anti-Infection Bone Void Fillers.

A gentamicin-loaded hydroxyapatite/collagen bone-like nanocomposite (GNT-HAp/Col) was fabricated and evaluated for its absorption-desorption properties, antibacterial efficacy, and cytotoxicity. The hydroxyapatite/collagen bone-like nanocomposite (HAp/Col) powder was mixed with gentamicin sulfate (GNT) in phosphate-buffered saline (PBS) at room temperature. After 6 h mixing, the GNT adsorption in all conditions reached plateau by Langmuir's isotherm, and maximum GNT adsorption amount was 34 ± 7 µg in 250 µg/mL GNT solution. Saturated GNT-loaded HAp/Col powder of 100 mg was soaked in 10 mL of PBS at 37 °C and released all GNT in 3 days. A shaking culture method for a GNT extraction from the GNT-HAp/Col and an inhibition zone assay for the GNT-HAp/Col compact showed antibacterial efficacy to Escherichia coli (E. coli) at least for 2 days. From the release profile of the GNT from the GNT-HAp/Col powder, antibacterial efficacy would affect E. coli at least for 3 days. Further, no cytotoxicities were observed on MG-63 cells. Thus, the GNT-HAp/Col is a good candidate of bioresorbable anti-infection bone void fillers by prevention initial infections, which is the primary cause of implant-associated infection even for rapid bioresorbable materials.

Gentamicin-Coated Tibia Nail in Fractures and Nonunion to Reduce Fracture-Related Infections: A Systematic Review.

The incidence of a fracture-related infection (FRI) can reach 30% of open tibia fractures (OTF). The use of antibiotic-coated implants is one of the newest strategies to reduce the risk of infection in orthopedic surgery. The aim of this study was to investigate the efficacy and safety of a gentamicin-coated tibia nail in primary fracture fixation (FF) and revision surgery (RS) of nonunion cases in terms of FRI incidence. We conducted a systematic review according to the PRISMA checklist on Pub-Med, Cochrane, and EMBASE. Of the 32 studies, 8 were included, for a total of 203 patients treated: 114 were FF cases (63% open fractures) and 89 were RS cases, of which 43% were infected nonunion. In the FF group, four FRI were found (3.8%): three OTF (Gustilo-Anderson III) and one closed fracture; bone healing was achieved in 94% of these cases. There were four relapses of infection and one new onset in the RS group; bone healing occurred in 88% of these cases. No side effects were found. There were no significant differences in terms of FRI, nonunion, and healing between the two groups. Gentamicin-coated tibia nail is an effective therapeutic option in the prophylaxis of high-risk fracture infections and in complex nonunion cases.

Antifungal Activity of Gentamicin B1 Against Systemic Plant Mycoses.

BACKGROUND: Gentamicin is a broad-spectrum aminoglycoside antibiotic produced by Micromonospora purpurea bacteria, effective against Gram-negative bacterial infections. Major fractions of the gentamicin complex (C1, C1a, C2, C2a) possess weak antifungal activity and one of the minor components (A, A1-A4, B, B1, X), gentamicin B1 was found to be a strong antifungal agent. METHODS: This work uses in vitro and in vivo dilution methods to compare the antifusarial, antiaspergillic and anticryptococcal effects of gentamicin derivatives and structurally-related congeners. RESULTS: The in vitro antifusarial activity of gentamicin B1 (minimum inhibitory concentration (MIC) 0.4 µg/mL) and structurally-related compounds (MIC 0.8-12.5 µg/mL) suggests that the purpuroseamine ring substituents are responsible for the specific antimycotic effect. The functional groups of the garoseamine and 2-deoxystreptamine rings of gentamicin derivatives are identical in gentamicin compounds and are unlikely to exert a significant antifungal effect. Among soil dermatophytes, Microsporum gypseum was more susceptible to gentamicin B1 (MIC 3.1 µg/mL) than Trichophyton gypseum (MIC 25 µg/mL). The in vitro antifungal effect of gentamicin B1 against plant pathogenic fungi was comparable to primary antifungal agents. CONCLUSION: Gentamicin is already in medical use. In vitro and preclinical in vivo synergisms of gentamicin B1 with amphotericin B suggest immediate clinical trials starting with subtoxic doses.

Synergistic Effects of Thiosemicarbazides with Clinical Drugs against S. aureus.

Antimicrobial resistance spurred by the overuse and misuse of antibiotics is a major global health concern, and of the Gram positive bacteria, S. aureus is a leading cause of mortality and morbidity. Alternative strategies to treat S. aureus infections, such as combination therapy, are urgently needed. In this study, a checkerboard method was used to evaluate synergistic interactions between nine thiosemicarbazides (4-benzoyl-1-(2,3-dichloro-benzoyl)thiosemicarbazides 1-5 and 4-aryl-1-(2-fluorobenzoyl)thiosemicarbazides 6-9) and conventional antibiotics against S. aureus ATCC 25923, which were determined as the fractional inhibitory concentration indices (FICIs). For these experiments, amoxicillin, gentamicin, levofloxacin, linezolid, and vancomycin were selected to represent the five antimicrobial classes most commonly used in clinical practice. With one exception of 7-vancomycin combination, none of the forty-five thiosemicarbazide-antibiotic combinations tested had an antagonistic effect, showing promising results with respect to a combination therapy. The synergic effect was observed for the 2-linezolid, 4-levofloxacin, 5-linezolid, 6-gentamicin, 6-linezolid, and 7-levofloxacin combinations. No interactions were seen in combination of the thiosemicarbazide with gentamicin or vancomycin, whereas all combinations with linezolid acted in additive or synergism, except for 6-gentamicin and 7-linezolid. The 4-(4-chlorophenyl)-1-(2-fluorobenzoyl)thiosemicarbazide 6 showed a clear preference for the potency; it affected synergistically in combinations with gentamicin or linezolid and additively in combinations with amoxicillin, levofloxacin, or vancomycin. In further studies, the inhibitory potency of the thiosemicarbazides against S. aureus DNA gyrase and topoisomerase IV was examined to clarify the molecular mechanism involved in their synergistic effect in combination with levofloxacin. The most potent synergist 6 at concentration of 100 µM was able to inhibit ~50% activity of S. aureus DNA gyrase, thereby suggesting that its anti-gyrase activity, although weak, may be a possible factor contributing to its synergism effect in combination with linezolid or gentamycin.

Artificial Channels in an Infectious Biofilm Created by Magnetic Nanoparticles Enhanced Bacterial Killing by Antibiotics.

The poor penetrability of many biofilms contributes to the recalcitrance of infectious biofilms to antimicrobial treatment. Here, a new application for the use of magnetic nanoparticles in nanomedicine to create artificial channels in infectious biofilms to enhance antimicrobial penetration and bacterial killing is proposed. Staphylococcus aureus biofilms are exposed to magnetic-iron-oxide nanoparticles (MIONPs), while magnetically forcing MIONP movement through the biofilm. Confocal laser scanning microscopy demonstrates artificial channel digging perpendicular to the substratum surface. Artificial channel digging significantly (4-6-fold) enhances biofilm penetration and bacterial killing efficacy by gentamicin in two S. aureus strains with and without the ability to produce extracellular polymeric substances. Herewith, this work provides a simple, new, and easy way to enhance the eradication of infectious biofilms using MIONPs combined with clinically applied antibiotic therapies.