Decrease In Bacterial Survival Research Articles

A novel ruthenium-silver based antimicrobial potentiates aminoglycoside activity against Pseudomonas aeruginosa.

The rapid dissemination of antibiotic resistance combined with the decline in the discovery of novel antibiotics represents a major challenge for infectious disease control that can only be mitigated by investments in novel treatment strategies. Alternative antimicrobials, including silver, have regained interest due to their diverse mechanisms of inhibiting microbial growth. One such example is AGXX, a broad-spectrum antimicrobial that produces highly cytotoxic reactive oxygen species (ROS) to inflict extensive macromolecular damage. Due to the connections identified between ROS production and antibiotic lethality, we hypothesized that AGXX could potentially increase the activity of conventional antibiotics. Using the gram-negative pathogen Pseudomonas aeruginosa, we screened possible synergistic effects of AGXX on several antibiotic classes. We found that the combination of AGXX and aminoglycosides tested at sublethal concentrations led to a rapid exponential decrease in bacterial survival and restored the sensitivity of a kanamycin-resistant strain. ROS production contributes significantly to the bactericidal effects of AGXX/aminoglycoside treatments, which is dependent on oxygen availability and can be reduced by the addition of ROS scavengers. Additionally, P. aeruginosa strains deficient in ROS detoxifying/repair genes were more susceptible to AGXX/aminoglycoside treatment. We further demonstrate that this synergistic interaction was associated with a significant increase in outer and inner membrane permeability, resulting in increased antibiotic influx. Our study also revealed that AGXX/aminoglycoside-mediated killing requires an active proton motive force across the bacterial membrane. Overall, our findings provide an understanding of cellular targets that could be inhibited to increase the activity of conventional antimicrobials. IMPORTANCE The emergence of drug-resistant bacteria coupled with the decline in antibiotic development highlights the need for novel alternatives. Thus, new strategies aimed at repurposing conventional antibiotics have gained significant interest. The necessity of these interventions is evident especially in gram-negative pathogens as they are particularly difficult to treat due to their outer membrane. This study highlights the effectiveness of the antimicrobial AGXX in potentiating aminoglycoside activities against P. aeruginosa. The combination of AGXX and aminoglycosides not only reduces bacterial survival rapidly but also significantly re-sensitizes aminoglycoside-resistant P. aeruginosa strains. In combination with gentamicin, AGXX induces increased endogenous oxidative stress, membrane damage, and iron-sulfur cluster disruption. These findings emphasize AGXX's potential as a route of antibiotic adjuvant development and shed light on potential targets to enhance aminoglycoside activity.

USP8 inhibition regulates autophagy flux and controls Salmonella infection.

Ubiquitination is an important protein modification that regulates various essential cellular processes, including the functions of innate immune cells. Deubiquitinases are enzymes responsible for removing ubiquitin modification from substrates, and the regulation of deubiquitinases in macrophages during infection with Salmonella Typhimurium and Yersinia enterocolitica remains unknown. To identify deubiquitinases regulated in human macrophages during bacterial infection, an activity-based proteomics screen was conducted. The effects of pharmacological inhibition of the identified deubiquitinase, USP8, were examined, including its impact on bacterial survival within macrophages and its role in autophagy regulation during Salmonella infection. Several deubiquiitnases were differentially regulated in infected macrophages. One of the deubiquitinases identified was USP8, which was downregulated upon Salmonella infection. Inhibition of USP8 was associated with a decrease in bacterial survival within macrophages, and it was found to play a distinct role in regulating autophagy during Salmonella infection. The inhibition of USP8 led to the downregulation of the p62 autophagy adaptor. The findings of this study suggest a novel role of USP8 in regulating autophagy flux, which restricts intracellular bacteria, particularly during Salmonella infection.

Does Ultraviolet Radiation Exhibit Antimicrobial Effect against Oral Pathogens Attached on Various Dental Implant Surfaces? A Systematic Review

Background: Dental implant therapy is currently identified as the most effective treatment for edentulous patient. However, peri-implant inflammations were found to be one of the most common complications that leads to the loss and failure of dental implantation. Ultraviolet (UV) radiation has been proposed to enhance bone integration and reduce bacterial attachment. In this study, we aimed to systematically review the current evidence regarding the antimicrobial effect of UV on different dental implant surfaces. Methods: Five databases including PubMed, Scopus, Web of science, VHL, and Cochran Library were searched to retrieve relevant articles. All original reports that examined the effect of the application of UV radiation on dental implants were included in our study. Results: A total of 16 in vitro studies were included in this systematic review. Polymethyl methacrylate UV radiation has induced a significant decrease in bacterial survival in PMMA materials, with an increased effect by modification with 2.5% and 5% TiO2 nanotubes. UV-C showed a superior effect to UV-A in reducing bacterial attachment and accumulation. UV wavelength of 265 and 285 nm showed powerful bactericidal effects. UV of 365 nm for 24 h had the highest inhibition of bacterial growth in ZnO coated magnesium alloys. In UV-irradiated commercially pure titanium surfaces treated with plasma electrolytic oxidation, silver ion application, heat or alkali had shown significant higher bactericidal effect vs non-irradiated treated surfaces than the treatment with any of them alone. UVC and gamma-ray irradiation increased the hydrophilicity of zirconia surface, compared to the dry heat. Conclusion: UV radiation on Ti surfaces exhibited significant antibacterial effects demonstrated through the reduction in bacterial attachment and biofilm formation with suppression of bacterial cells growth. Combination of UV and treated surfaces with alkali, plasma electrolytic oxidation, silver ion application or heat enhance the overall photocatalytic antimicrobial effect.

Indirect Bactericidal Properties of Recombinant Human Bone Morphogenetic Protein 2 In Vitro

Bone morphogenetic proteins (BMPs) are secreted cytokines and are involved in various metabolic functions and inflammatory processes in different organs. The purpose of this study was to investigate whether BMPs also possess antimicrobial properties in direct or indirect ways. Antibacterial properties of recombinant human BMP2 (rhBMP2) were tested on 4 bacteria species (Staphylococcus aureus, Escherichia coli, Streptococcus mitis, Streptococcus constellatus) to examine the potential synergism of rhBMP2 with antibiotics. Indirect antibacterial properties were tested by infecting neutrophils with rhBMP2 and bacteria to investigate bacterial survival. Reactive oxidative species (ROS) production in neutrophils in the presence of rhBMP2 also was tested. RhBMP2 in cardboard disks or sponge collagen as carriers did not show antibacterial activity against all tested bacteria. Further, synergism of rhBMP2 with antibiotics was not evident. Survival of bacteria inoculated with neutrophils and rhBMP2 led to a marked decrease in bacterial survival compared with neutrophils without rhBMP2. Although rhBMP2 inoculation of neutrophils alone did not induce ROS, its presence with the bacterial infection showed augmented ROS production for all tested bacteria. RhBMP2 did not show direct antibacterial properties but did exhibit an indirect bactericidal effect in the presence of neutrophils. ROS production indicated that rhBMP2 has a role as a priming agent for neutrophils by augmenting their bactericidal capabilities and suggests the importance of its presence in contaminated surgical bone augmentation sites.

Photodynamic antimicrobial activity of new porphyrin derivatives against methicillin resistant Staphylococcus aureus.

Methicillin resistant Staphylococcus aureus (MRSA) with multiple drug resistance patterns is frequently isolated from skin and soft tissue infections that are involved in chronic wounds. Today, difficulties in the treatment of MRSA associated infections have led to the development of alternative approaches such as antimicrobial photodynamic therapy. This study aimed to investigate photoinactivation with cationic porphyrin derivative compounds against MRSA in in-vitro conditions. In the study, MRSA clinical isolates with different antibiotic resistance profiles were used. The newly synthesized cationic porphyrin derivatives (PM, PE, PPN, and PPL) were used as photosensitizer, and 655 nm diode laser was used as light source. Photoinactivation experiments were performed by optimizing energy doses and photosensitizer concentrations. In photoinactivation experiments with different energy densities and photosensitizer concentrations, more than 99% reduction was achieved in bacterial cell viability. No decrease in bacterial survival was observed in control groups. It was determined that there was an increase in photoinactivation efficiency by increasing the energy dose. At the energy dose of 150 J/cm2 a survival reduction of over 6.33 log10 was observed in each photosensitizer type. While 200 μM PM concentration was required for this photoinactivation, 12.50 μM was sufficient for PE, PPN, and PPL. In our study, antimicrobial photodynamic therapy performed with cationic porphyrin derivatives was found to have potent antimicrobial efficacy against multidrug resistant S. aureus which is frequently isolated from wound infections.

Carnosine-graphene oxide conjugates decorated with hydroxyapatite as promising nanocarrier for ICG loading with enhanced antibacterial effects in photodynamic therapy against Streptococcus mutans

Antimicrobial photodynamic therapy (aPDT) has been emerged as a noninvasive strategy to remove bacterial contaminants such as S. mutans from the tooth surface. Photosensitizer (PS), like indocyanine green (ICG), plays a key role in this technique which mainly suffers from the poor stability and concentration-dependent aggregation. An appropriate nanocarrier (NC) with enhanced antibacterial effects could overcome these limitations and improve the efficiency of ICG as a PS. In this study, various ICG-loaded NCs including graphene oxide (GO), GO-carnosine (Car) and GO-Car/Hydroxyapatite (HAp) were synthesized and characterized by Fourier Transform Infrared Spectroscopy (FT-IR), X-ray Diffraction (XRD), Filed Emission Scanning Electron Microscopy (FE-SEM), Energy Dispersive Spectroscopy (EDS), Zeta Potential and Ultraviolet-Visible spectrometry (UV–Vis). The colony forming unit and crystal violet assays were performed to evaluate the antimicrobial and anti-biofilm properties of PSs against S. mutans. The quantitative real-time PCR approach was also applied to determine the expression ratio of the gtfB gene in S. mutans. The zeta potential analysis and UV–Vis spectrometry indicated successful loading of ICG onto/into NCs. GO-Car/HAp showed highest amount of ICG loading (57.52%) and also highest aqueous stability after one week (94%). UV–Vis spectrometry analyses disclosed a red shift from 780 to 800 nm for the characteristic peak of ICG-loaded NCs. In the lack of aPDT, GO-Car@ICG showed the highest decrease in bacterial survival (86.4%) which indicated that Car could significantly promote the antibacterial effect of GO. GO@ICG, GO-Car@ICG and GO-Car/HAp@ICG mediated aPDT, dramatically declined the count of S. mutans strains to 91.2%, 95.5% and 93.2%, respectively (P < 0.05). The GO@ICG, GO-Car@ICG, GO-Car/HAp@ICG significantly suppressed the S. mutans biofilm formation by 51.4%, 63.8%, and 56.8%, respectively (P < 0.05). The expression of gtfB gene was considerably reduced to 6.0, 9.0 and 7.9-fold after aPDT in the presence of GO@ICG, GO-Car@ICG, GO-Car/HAp@ICG, respectively (P < 0.05). It could be concluded that the multi-functionalized GO as a novel nanocarrier could significantly enhance the ICG loading, stability, and improve its inhibitory effects as a photosensitizer in aPDT against S. mutans. These findings might provide opportunity for efficient treatment of local dental infections.

Pharmacodynamic Evaluation of Fosfomycin against Escherichia coli and Klebsiella spp. from Urinary Tract Infections and the Influence of pH on Fosfomycin Activities.

Fosfomycin is widely used for the treatment of uncomplicated urinary tract infection (UTI), and it has recently been recommended that fosfomycin be used to treat infections caused by multidrug-resistant (MDR) Gram-negative bacilli. Whether urine acidification can improve bacterial susceptibility to fosfomycin oral dosing regimens has not been analyzed. The MIC of fosfomycin for 245 Gram-negative bacterial isolates, consisting of 158 Escherichia coli isolates and 87 Klebsiella isolates which were collected from patients with urinary tract infections, were determined at pH 6.0 and 7.0 using the agar dilution method. Monte Carlo simulation of the urinary fosfomycin area under the concentration-time curve (AUC) after a single oral dose of 3,000 mg fosfomycin and the MIC distribution were used to determine the probability of target attainment (PTA). Fosfomycin was effective against E. coli (MIC90 ≤ 16 μg/ml) but not against Klebsiella spp. (MIC90 > 512 μg/ml). Acidification of the environment increased the susceptibility of 71% of the bacterial isolates and resulted in a statistically significant decrease in bacterial survival. The use of a regimen consisting of a single oral dose of fosfomycin against an E. coli isolate with an MIC of ≤64 mg/liter was able to achieve a PTA of ≥90% for a target pharmacodynamic index (AUC/MIC) of 23 in urine; PTA was not achieved when the MIC was higher than 64 mg/liter. The cumulative fractions of the bacterial responses (CFR) were 99% and 55% against E. coli and Klebsiella spp., respectively, based on simulated drug exposure in urine with an acidic pH of 6.0. A decrease of the pH from 7.0 to 6.0 improved the PTA and CFR of the target pharmacodynamic index in both E. coli and Klebsiella isolates.

Liposomal nanoformulations of rhodamine for targeted photodynamic inactivation of multidrug resistant gram negative bacteria in sewage treatment plant

The antimicrobial photodynamic therapy is an alternative method for killing bacterial cells in view of the rising problem of antibiotic resistance microorganisms. The present study examined the effect of a water soluble photosensitizer, Rhodamine 6G (R6G) in stealth liposomes on multidrug resistant Pseudomonas aeruginosa in the presence of visible light. Liposomes were prepared with cholesterol and phospholipids that extracted from hen eggs in a cost effective way and characterized by light microscopy, particle size analyzer, electron microscopy, steady state spectrophotometry and spectrofluorometry. The photoefficacies of R6G in polymer encapsulated liposomes and positively charged liposomes are much higher compared to the free R6G (R6G in water) in terms of singlet oxygen quantum yield. This high potential of producing more reactive oxygen species (ROS) by liposomal nanoformulated R6G leads to efficient photodynamic inactivation of multidrug resistant gram negative bacteria in waste water. Though the singlet oxygen quantum yield of polymer coated liposomal R6G was higher than the cationic liposomal formulation, a faster decrease in bacterial survival was observed for positively charged liposomal R6G treated bacteria due to electrostatic charge interactions. Therefore, it can be concluded that the positively charged liposomal nanoformulations of laser dyes are efficient for photodynamic inactivation of multiple drug resistant gram negative microorganisms.

Enhanced Survival of Rifampin- and Streptomycin-Resistant Escherichia coli Inside Macrophages.

The evolution of multiple-antibiotic-resistant bacteria is an increasing global problem. Even though mutations causing resistance usually incur a fitness cost in the absence of antibiotics, the magnitude of such costs varies across environments and genomic backgrounds. We studied how the combination of mutations that confer resistance to rifampin (Rifr) and streptomycin (Strr) affects the fitness of Escherichia coli when it interacts with cells from the immune system, i.e., macrophages (Mϕs). We found that 13 Rifr Strr doubly resistant genotypes, of the 16 tested, show a survival advantage inside Mϕs, indicating that double resistance can be highly beneficial in this environment. Our results suggest that there are multiple paths to acquire multiple-drug resistance in this context, i.e., if a clone carrying Rifr allele H526 or S531 acquires a second mutation conferring Strr, the resulting double mutant has a high probability of showing increased survival inside Mϕs. On the other hand, we found two cases of sign epistasis between mutations, leading to a significant decrease in bacterial survival. Remarkably, infection of Mϕs with one of these combinations, K88R+H526Y, resulted in an altered pattern of gene expression in the infected Mϕs. This indicates that the fitness effects of resistance may depend on the pattern of gene expression of infected host cells. Notwithstanding the benefits of resistance found inside Mϕs, the Rifr Strr mutants have massive fitness costs when the bacteria divide outside Mϕs, indicating that the maintenance of double resistance may depend on the time spent within and outside phagocytic cells.

The AirSR two-component system contributes to Staphylococcus aureus survival in human blood and transcriptionally regulates sspABC operon.

To date, genes identified and transcriptionally regulated by the AirSR TCS have been involved in energy production and cellular homeostasis of the staphylococcal cell. It is well accepted that the state of cellular metabolism impacts the expression of virulence factors in Staphylococcus aureus. For this reason, we conducted experiments to determine if the AirSR TCS contributes to the pathogenesis of S. aureus using an antisense RNA interference technology, an inducible overexpression system, and gene deletions. Depletion of AirSR by antisense RNA expression or deletion of the genes, results in significant decrease in bacterial survival in human blood. Conversely, overexpression of AirR significantly promotes survival of S. aureus in blood. AirR promotes the secretion of virulence factors that inhibits opsonin-based phagocytosis. This enhanced survival is partially linked to the transcriptional regulation of the sspABC operon, encoding V8 protease (SspA), staphopain B (SspB) and staphostatin B (SspC). SspA and SspB are known virulence factors which proteolytically digest opsonins and inhibit killing of S. aureus by professional phagocytes. This is the first evidence linking the AirSR TCS to pathogenesis of S. aureus.

Photodynamic inactivation of multidrug-resistant bacteria in hospital wastewaters: influence of residual antibiotics

One environmental concern related to hospital effluents is discharge of them without preliminary treatment. Antimicrobial photodynamic inactivation (PDI) may represent an alternative to the traditional expensive, unsafe and not always effective disinfection methods. The main goal of this work was to assess the efficiency of PDI on clinical multidrug-resistant (MDR) bacteria in hospital wastewaters in order to evaluate its potential use in treating hospital effluents. The efficiency of PDI was assessed using a cationic porphyrin as the photosensitizer (PS), four MDR bacteria either in phosphate buffered saline or in filtrated hospital wastewaters. The synergistic effect of PDI and antibiotics (ampicillin and chloramphenicol) was also evaluated, as well as the effect of the surfactant sodium dodecyl sulfate (SDS). The results show the efficient inactivation of MDR bacteria in PBS (reduction of 6-8 log after 270 min of irradiation at 40 W m(-2) with 5.0 μM of PS). In wastewater, the inactivation of the four MDR bacteria was again efficient and the decrease in bacterial survival starts even sooner. A faster decrease in bacterial survival occurred when PDI was combined with the addition of antibiotics, at sub-inhibitory and inhibitory concentrations, but the SDS did not affect the PDI efficiency. It can be concluded that PDI has potential to be an effective alternative for the inactivation of MDR bacteria in hospital wastewaters and that the presence of antibiotics may enhance its effectiveness.

Photodynamic inactivation of Gram-positive bacteria employing natural resources

The aim of this paper was to investigate a collection of plant extracts from Argentina as a source of new natural photosensitizers (PS) to be used in Photodynamic Inactivation (PDI) of bacteria. A collection of plants were screened for phototoxicity upon the Gram-positive species Staphylococcus epidermidis.Three extracts turned out to be photoactive: Solanum verbascifolium flower, Tecoma stans flower and Cissus verticillata root. Upon exposure to a light dose of 55J/cm2, they induced 4, 2 and 3logs decrease in bacterial survival, respectively.Photochemical characterisation of S. verbascifolium extract was carried out. PDI reaction was dependent mainly on singlet oxygen and to a lesser extent, on hydroxyl radicals, through type II and I reactions. Photodegradation experiments revealed that the active principle of the extract was not particularly photolabile.It is noticeable that S. verbascifolium –PDI was more efficient under sunlight as compared to artificial light (total eradication vs. 4 logs decrease upon 120min of sunlight). The balance between oxidant and antioxidant compounds is likely to be masking or unmasking potential PS of plant extracts, but employing the crude extract, the level of photoactivity of S. verbascifolium is similar to some artificial PS upon exposure to sunlight, demonstrating that natural resources can be employed in PDI of bacteria.

Photodynamic therapy against common bacteria causing wound and skin infections

Photodynamic therapy (PDT) uses the combination of a photosensitising (PS) agent and visible light. Historically, various injectable PS agents have been used with medical grade lasers to treat neoplasia. The objective of this in vitro study was to determine whether PDT using topical aminolevulinic acid (ALA) with a non-coherent light source would kill common wound infecting bacteria, namely, Staphylococcus intermedius, Streptococcus canis, Pseudomonas aeruginosa, and Escherichia coli. Bacterial strains were sensitised to light with ALA before exposing to the non-coherent light source. Colony counts were performed in triplicate and compared to controls. When compared with controls there was a significant decrease in bacterial survival following PDT for all organisms except E. coli. A single treatment required 2-3h of light exposure. These data suggest that PDT may be a possible treatment option for wound infections but repeated treatments or alterations in the PS or its carrier will be needed to decrease treatment times.

Lactate-Dependent Killing of Escherichia coli by Nitrite plus Hydrogen-Peroxide: A Possible Role of Nitrogen Dioxide

The killing of Escherichia coli by nitrite plus hydrogen peroxide was observed in lactate, but not in phosphate or acetate. Although nitrite or hydrogen peroxide alone caused a slight decrease in bacterial survival, nitrite plus hydrogen peroxide killed bacteria synergistically in time-, dose-, and pH-dependent manners. The killing was increased with decreasing pH. The plot of viable cells versus [nitrous acid] was linear, Among the hydroxyl radical scavengers used, only benzoate and formate at concentrations higher than that of lactate inhibited the killing by nitrite plus hydrogen peroxide, whereas dimethyl sulfoxide enhanced it. The generation of peroxynitrous acid during the reaction of nitrite and hydrogen peroxide was confirmed by the formation of malondialdehyde using deoxyribose as a hydroxyl radical-like oxidant detector. The nitration of glycyl-tryosine was observed only in lactate buffer, but not in phosphate and acetate buffers. Benzoate and formate inhibited the nitration, whereas dimethyl sulfoxide and ethanol enhanced it. No evidence for the formation of nitric oxide and superoxide during the reaction of nitrite and hydrogen peroxide was found. These data suggest that nitrogen dioxide from the decomposition of peroxynitrous acid or secondary oxidants formed from the reaction of peroxynitrous acid with lactate is responsible for the lactate-dependent killing of E. coli induced by the reaction of protonated nitrite and hydrogen peroxide.

Stimulation of nonspecific resistance to infection induced by muramyl dipeptide analogs substituted in the gamma-carboxyl group and evaluation of N alpha-muramyl dipeptide-N epsilon-stearoyllysine

Stimulation of resistance to infection induced by the analogs of muramyl dipeptide (MDP) having substituted functions in the gamma-carboxyl group of D-isoglutamyl residue was examined in experimental Escherichia coli infections in mice. An MDP analog which is an efficient strengthener of resistance to infection, N alpha-MDP-N epsilon-stearoyllysine [MDP-Lys(L18)], was selected through the comparative assessment of a number of compounds in three categories: (i) gamma-alkylamides, (ii) gamma-esters, and (iii) N alpha-MDP-N epsilon-acyllysine derivatives. Furthermore, the antiinfectious activity of MDP-Lys(L18) was evaluated bacteriologically in comparison with that of MDP. The effect of MDP-Lys(L18) on the susceptibility of mice to infections with various species of microorganisms was studied. Protective activity was greatest against E. coli and staphylococcal infections, considerable against Pseudomonas and Candida infections, and least against Klebsiella infection. The effects of bacterial inoculum size and MDP treatment timing, dose, and route of administration on protective activity were studied. The efficacy of MDP-Lys(L18) in protection tests was demonstrated for all administration routes, even the oral. Its high potency was confirmed by the smaller influence of inoculum size and particularly small value of the minimum dosage required for inducing protective activity. A decrease in bacterial survival was observed in the blood and organs of mice treated with the analog and infected with E. coli. The following two useful effects were obtained: the synergistic effect of glycopeptide and chemotherapeutic agents and the stimulation of resistance to infection in animals immunocompromised by cyclophosphamide treatment.