Time-kill Curve Assays Research Articles

A new N-acylhydrazone oxadiazole derivative with activity against mycobacteria.

Aim: To evaluate the anti-Mycobacterium tuberculosis (Mtb) potential of the hybrid oxadiazol-4-methoxynaphthalene (6n) derived from N-acylhydrazone (4k).Materials & methods: The study determined the minimal inhibitory concentration of (6n) against Mtb H37Rv and Mtb clinical isolates, potential combination of (6n) with anti-tuberculosis drugs and carried out time kill curve assay of Mtb H37Rv. Additional contribution for the analysis of (6n) was explored by in silico pharmacokinetics, and in vitro and in vivo cytotoxicity determinations.Results: The newly synthesized molecule (6n) demonstrated anti-Mtb activity, low cytotoxicity and selectivity for Mtb.Conclusion: The derivative (6n) emerges as a potential anti-TB drug candidate.

Formulation and evaluation of Phaseolus lunatus seed coat mediated silver nanoparticles mouthwash: A comprehensive study on biomedical properties and toxicological assessment

Silver nanoparticles have promising therapeutic potential in the field of dentistry, as newly emerging oral therapeutics, in the form of mouthwashes based on silver nanoparticles, demonstrate significant potential for enhancing oral management thus the present investigation aims to formulate silver nanoparticles-based mouthwash from an aqueous extract of Phaseolus lunatus seed coat (PLSC) and evaluate its biomedical properties. The green synthesized AgNPs in the mouthwash were characterized using UV–visible spectroscopy, scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and x-ray diffraction (XRD). The formulated mouthwash was assessed for its anti-microbial activity using the agar well diffusion technique and time-kill curve assay. Its anti-inflammatory and anti-oxidant activities are assessed through egg albumin assay and hydrogen peroxide assays. The Cytotoxic effect of formulated mouthwash was assessed through a brine shrimp lethality assay and MTT assay over the human osteoblast cell line (MG-63). Furthermore, the study also assessed the toxicity effect of formulated mouthwash through zebrafish embryos. The results suggest that the green synthesized AgNPs were spherical and had an average size of 42 nm and the formulated mouthwash exhibited significant anti-microbial activity, anti-inflammatory, and antioxidant properties, making them a better candidate for oral health therapeutics. The Zebrafish embryo toxicology studies of mouthwash revealed a consistent lack of abnormalities and a good viability and hatchability rate and the cytotoxic effect shows less toxicity over brine shrimp and human osteoblast cells. In conclusion, PLSC-AgNPs mouthwash is a potential oral therapeutic option with minimal toxicity for better oral management.

In vitro Synergistic and Bactericidal Effects of Aztreonam in Combination with Ceftazidime/ Avibactam, Meropenem/Vaborbactam and Imipenem/Relebactam Against Dual-Carbapenemase-Producing Enterobacterales.

Our aim was to elucidate the resistance mechanisms and assess the combined synergistic and bactericidal activities of aztreonam in combination with ceftazidime/avibactam (CZA), meropenem/vaborbactam (MEV), and imipenem/relebactam (IMR) against Enterobacterales strains producing dual carbapenemases. Species identification, antimicrobial susceptibility testing and determination of carbapenemase type were performed for these strains. Plasmid sizes, plasmid conjugation abilities and the localization of carbapenemase genes were investigated. Whole-genome sequencing was performed for all strains and their molecular characteristics were analyzed. In vitro synergistic and bactericidal activities of the combination of aztreonam with CZA, MEV and IMR against these strains were determined using checkerboard assay and time-kill curve assay. A total of 12 Enterobacterales strains producing dual-carbapenemases were collected, including nine K. pneumoniae, two P. rettgeri, and one E. hormaechei. The most common dual-carbapenemase gene pattern observed was bla (KPC-2+NDM-5) (n=4), followed by bla KPC-2+IMP-26 (n=3), bla (KPC-2+NDM-1) (n=2), bla (KPC-2+IMP-4) (n=1), bla (NDM-1+IMP-4) (n=1) and bla (KPC-2+KPC-2) (n=1). In each strain, the carbapenemase genes were found to be located on two distinct plasmids which were capable of conjugating from the original strain to the receipt strain E. coli J53. The results of the checkerboard synergy analysis consistently revealed good synergistic effects of the combination of ATM with CZA, MEV and IMR. Except for one strain, all strains exhibited significant synergistic activity and bactericidal activity between 2 and 8 hours. Dual-carbapenemase-producing Enterobacterales posed a significant threat to clinical anti-infection treatment. However, the combination of ATM with innovative β-lactam/β-lactamase inhibitor compounds had proven to be an effective treatment option.

Evaluation of Antibacterial Effects of an Oral Rinse Containing Ocimum tenuiflorum and Ocimum gratissimum on Streptococcus mutans and Lactobacillus Species.

Dental caries, primarily caused by Streptococcus mutans and Lactobacillus spp., is a major global health issue. There is a growing need for effective, natural antimicrobial treatments. Ocimum tenuiflorum and Ocimum gratissimum are known for their medicinal properties, including antimicrobial activity. This study investigates the antibacterial efficacy of a herbal oral rinse derived from these plants. This study aimed to evaluate the antibacterial activity of an herbal formulation-based oral rinse prepared from Ocimum tenuiflorum and Ocimum gratissimum against Streptococcus mutans and Lactobacillus spp. Fresh leaves of Ocimum tenuiflorum and Ocimum gratissimum were shade-dried, powdered, and extracted in distilled water. The extract was incorporated into an oral rinse formulation. The antibacterial activity was assessed using the agar well diffusion method, protein leakage and cytoplasmic leakage assays, and time-kill curve analysis. A commercial oral rinse was used as a standard. The herbal oral rinse exhibited significant antibacterial activity against both Streptococcus mutans and Lactobacillus spp. The zones of inhibition for Streptococcus mutans were 10 mm, 13 mm, and 15 mm at concentrations of 25 µg/mL, 50 µg/mL, and 100 µg/mL, respectively. For Lactobacillus spp., the inhibition zones were 10 mm, 12 mm, and 14 mm at the same concentrations. The protein leakage and cytoplasmic leakage analysis supported these findings, demonstrating the formulation's efficacy at low concentrations. Time-kill curve assays showed rapid bactericidal action, particularly at higher concentrations. The Ocimum tenuiflorum and Ocimum gratissimum-based herbal oral rinse demonstrates strong antibacterial activity against key oral pathogens, suggesting that it could be a natural alternative to conventional oral rinses.

Discovery of new antimicrobial thiophene derivatives with activity against drug-resistant Gram negative-bacteria.

Our aim is to identify new small molecules with antimicrobial potential, especially against colistin-resistant (Col-R) Acinetobacter baumannii and Escherichia coli. After initial hits identification by fingerprint similarity, MIC of 24 heterocyclic derivatives for A. baumannii and E. coli reference strains, and bactericidal activity of selected thiophenes against Col-R strains were determined. We analyzed changes in bacterial membrane permeability and the OMPs profile. Additionally, we determined bacterial adherence to host cells and performed molecular docking studies to assess their binding to bacterial targets. The compounds' MICs ranged from 4 to >64mg/L. Thiophene derivatives 4, 5 and 8 exhibited MIC50 values between 16 and 32mg/L for Col-R A. baumannii and 8 and 32mg/L for Col-R E. coli. The time-kill curve assay demonstrated that thiophenes 4 and 8 had bactericidal effects against Col-R A. baumannii and E. coli. Furthermore, treatment with them resulted in increased membrane permeabilization and reduced adherence of these isolates to host cells. Finally, the docking studies showed a stronger binding affinity to CarO1 and Omp33 of A. baumannii and OmpW and OmpC of E. coli. These findings indicate that thiophene derivatives possess antibacterial activity against Col-R A. baumannii and E. coli, suggesting that they may enhance the repertoire of drug treatments against bacteria.

Synergistic Activity of Cefiderocol in Combination with Avibactam, Sulbactam or Tazobactam against Carbapenem-Resistant Gram-Negative Bacteria.

We investigated the activity of cefiderocol/β-lactamase inhibitor combinations against clinical strains with different susceptibility profiles to cefiderocol to explore the potentiality of antibiotic combinations as a strategy to contain the major public health problem of multidrug-resistant (MDR) pathogens. Specifically, we evaluated the synergistic activity of cefiderocol with avibactam, sulbactam, or tazobactam on three of the most "Critical Priority" group of MDR bacteria (carbapenem-resistant Enterobacterales, Pseudomonas aeruginosa, and Acinetobacter baumannii). Clinical isolates were genomically characterized by Illumina iSeq 100. The synergy test was conducted with time-kill curve assays. Specifically, cefiderocol/avibactam, /sulbactam, or /tazobactam combinations were analyzed. Synergism was assigned if bacterial grow reduction reached 2 log10 CFU/mL. We reported the high antimicrobial activity of the cefiderocol/sulbactam combination against carbapenem-resistant Enterobacterales, P. aeruginosa, and A. baumannii; of the cefiderocol/avibactam combination against carbapenem-resistant Enterobacterales; and of the cefiderocol/tazobactam combination against carbapenem-resistant Enterobacterales and P. aeruginosa. Our results demonstrate that all β-lactamase inhibitors (BLIs) tested are able to enhance cefiderocol antimicrobial activity, also against cefiderocol-resistant isolates. The cefiderocol/sulbactam combination emerges as the most promising combination, proving to highly enhance cefiderocol activity in all the analyzed carbapenem-resistant Gram-negative isolates, whereas the Cefiderocol/tazobactam combination resulted in being active only against carbapenem-resistant Enterobacterales and P. aeruginosa, and cefiderocol/avibactam was only active against carbapenem-resistant Enterobacterales.

Antifungal Activity of Ocimum tenuiflorum and Ocimum gratissimum Herbal Formulation-Based Oral Rinse Against Candida albicans.

Background Candida albicans is a common fungal pathogen responsible for oral infections, posing significant health challenges. Traditional antifungal treatments often come with side effects and resistance issues, highlighting the need for effective natural alternatives. O. tenuiflorum and Ocimum gratissimum are known for their medicinal properties, including antifungal activity. Objective This study aimed to evaluate the antifungal effectiveness of an O. tenuiflorum and O. gratissimum herbal formulation-based oral rinse against C. albicans. Methods Antifungal activity was measured using agar well diffusion, time-kill curve assays, and analyses of cytoplasmic and protein leakage. The herbal rinse was tested at concentrations of 25 µg/mL, 50 µg/mL, and 100 µg/mL, and compared to a commercial oral rinse. Results The herbal rinse demonstrated strong antifungal effects that increased with concentration. At 100 µg/mL, it produced a 13 mm zone of inhibition, outperforming the commercial rinse's 11 mm. The time-kill assay revealed that the 100 µg/mL concentration reduced fungal counts to 103 CFU/mL within 5 hours, on par with the commercial rinse. Cytoplasmic leakage analysis showed an optical density of 0.42 at 100 µg/mL, close to the commercial rinse's 0.45. Protein leakage analysis indicated an optical density of 0.52 at 100 µg/mL, slightly higher than the commercial rinse's 0.51. Conclusion The O. tenuiflorum and O. gratissimum herbal formulation-based oral rinse exhibit potent antifungal activity against C. albicans, rivaling and even surpassing commercial rinses at higher concentrations. This study underscores the potential of this natural oral rinse as a powerful alternative for managing oral fungal infections, meriting further research and clinical trials to confirm its long-term safety and efficacy.

Anticariogenic, Antidiabetic, and Toxicology Evaluation of the Ethanolic Extract of Croton bonplandianum: An In Vitro Study.

Background Herbal medicine has gathered increasing attention in contemporary healthcare practices, offering natural remedies for a wide range of ailments such asskin diseases, liver disorders,bronchitis, and asthma. Among the plethora of medicinal plants, Croton bonplandianum, commonly known as "Ban Tulsi," holds significant medicinal value owing to its diverse pharmacological properties. This study investigated the cytotoxicity, embryotoxicity, antidiabetic, and anticariogenic effects of an ethanolic extract derived from C. bonplandianum.The research objectives were to explore the preparation of an ethanolic extract of C. bonplandianum and employ a multifaceted approach by evaluating its cytotoxicity, embryotoxicity, anticariogenic, and antidiabetic potentials. Materials and methods In this study, the β-glucosidase inhibitory and the α-amylase inhibitory assays were utilized to evaluate the antidiabetic activity of theC. bonplandianum ethanolic extract. The in vitro cytotoxicity activity was assessed by using the brine shrimp lethality assay (BSLA), and embryotoxicity was evaluated using zebrafish embryos and larvae. Through the agar well diffusion method and the time-kill curve analysis, the anticariogenic activity was evaluated. Results In α-amylase andβ-glucosidase inhibitory assays, the ethanolic extract ofC. bonplandianum showed potent antidiabetic properties, near those ofstandard acarbose. The cytotoxicity evaluation using the BSLA showed less toxicity. The anticariogenic activity of the ethanolic extract of C. bonplandianumwas assessed by comparing the standard (Amoxyrite) in terms of its zone of inhibition against oral pathogens such asStreptococcus mutans and Lactobacillus species (spp.). The antibacterial efficiency was validated using a time-kill curve assay in which the study depends on the concentration of the bacterial pathogenic organisms, namely, Lactobacillus spp. and S. mutans. In embryotoxicity evaluation, there were no morphological malformations in zebrafish larvae or embryos when exposed to high concentrations of C. bonplandianum ethanolic extract. Conclusion The ethanolic extract ofC. bonplandianum exhibited promising antidiabetic and anticariogenic effects, supporting its conventional usage inalternative medicine. The outcomes of these research analyses suggest the plantpotential as a natural source of compounds with bioactive qualities and can be utilized in the healthcare and pharmaceutical industries.

In vitro and in vivo activity of ceftazidime/avibactam and aztreonam alone or in combination against mcr-9, serine- and metallo-β-lactamases-co-producing carbapenem-resistant Enterobacter cloacae complex.

Enterobacteriaceae carrying mcr-9, in particularly those also co-containing metallo-β-lactamase (MBL) and TEM type β-lactamase, present potential transmission risks and lack adequate clinical response methods, thereby posing a major threat to global public health. The aim of this study was to assess the antimicrobial efficacy of a combined ceftazidime/avibactam (CZA) and aztreonam (ATM) regimen against carbapenem-resistant Enterobacter cloacae complex (CRECC) co-producing mcr-9, MBL and TEM. The in vitro antibacterial activity of CZA plus ATM was evaluated using a time-kill curve assay. Furthermore, the in vivo interaction between CZA plus ATM was confirmed using a Galleria mellonella (G. mellonella) infection model. All eight clinical strains of CRECC, co-carrying mcr-9, MBL and TEM, exhibited high resistance to CZA and ATM. In vitro time-kill curve analysis demonstrated that the combination therapy of CZA + ATM exerted significant bactericidal activity against mcr-9, MBL and TEM-co-producing Enterobacter cloacae complex (ECC) isolates with a 100% synergy rate observed in our study. Furthermore, in vivo survival assay using Galleria mellonella larvae infected with CRECC strains co-harboring mcr-9, MBL and TEM revealed that the CZA + ATM combination significantly improved the survival rate compared to the drug-treatment alone and untreated control groups. To our knowledge, this study represents the first report on the in vitro and in vivo antibacterial activity of CZA plus ATM against CRECC isolates co-harboring mcr-9, MBL and TEM. Our findings suggest that the combination regimen of CZA + ATM provides a valuable reference for clinicians to address the increasingly complex antibiotic resistance situation observed in clinical microorganisms.

Investigation of the Antibacterial Synergistic Activity of Fosfomycin-Teicoplanin Combination in Methicillin-Resistant Staphylococcus aureus Strains Isolated from Various Clinical Sample

The aim of this study was to investigate the detection of teicoplanin and fosfomycin antibiotic susceptibility of methicillin-resistant Staphylococcus aureus (MRSA) strains by different methods and to evaluate the antibacterial synergistic effect of teicoplanin-fosfomycin combination by using checkerboard assay and time kill curve assay. Forty-five MRSA strains isolated from clinical samples in routine medical microbiology laboratory of Göztepe Prof. Dr. Süleyman Yalçın City Hospital were included in the study. In the first stage of the combination study, minimum inhibitory concentrations (MIC) were investigated by broth microdilution for teicoplanin and by both broth microdilution and agar dilution methods for fosfomycin. The combination of teicoplanin and fosfomycin was tested by the checkerboard method in 45 MRSA strains and combination effect was determined according to fractional inhibitory concentration index (ΣFIC) calculation. The synergistic effect and bactericidal activity of antibiotic combination were studied against a randomly selected strain from the strains used in the study by using time-kill method for 24 hours. As a result of teicoplanin and fosfomycin antibiotic susceptibility studies, all isolates were found to be susceptible to both antibiotics according to the susceptibility breakpoints determined by the European Committee on Antimicrobial Susceptibility Testing (EUCAST). A synergistic effect was found in 22 (49%), additive effect in 22 (49%) and indifferent effect in one (2%) of the 45 strains studied with the checkerboard method. The mean ΣFIC of 45 isolates was found to be 0.5. In the combination study of the antibiotics of the isolate that was studied with time-kill method, synergism was detected for 1/8 MIC concentrations at 12th hour and 24th hour and synergism at 1/4 MIC concentration at sixth hour, 12th hour and 24th hour. In the combination study of 1/4 MIC concentrations of antibiotics, bactericidal effect was detected at sixth hour and this effect was observed to disappear at 12th and 24th hours. High rate of synergistic antibacterial effect of teicoplanin-fosfomycin combination on MRSA isolates was demonstrated as a result of in vitro tests. Such studies conducted on antibiotic-resistant bacterial infections will provide clinicians different treatment options and will contribute to increasing survival. As a result of this study, provided that it is supported by future clinical studies, it can be stated that the teicoplanin-fosfomycin combination may be an effective treatment option in community and hospital-acquired infections caused by MRSA.

Free Radical Scavenging, Anti-inflammatory and Antibacterial Activity of Acorus calamus Leaves Extract Against Pseudomonas aeruginosa and Staphylococcus aureus.

Background Herbal medicine, or phytotherapy, has been used for centuries in traditional healing practices to harness the therapeutic properties of different plant-derived elements. Acorus calamus, a perennial herbaceous plant, has significant historical importance in traditional medicine, specifically in Ayurveda, where it is referred to as "Vacha."This study investigates the antioxidant, anti-inflammatory, and antimicrobial characteristics of the A. calamus dimethyl sulfoxide (DMSO) extract. The objectives of the research are to provide valuable knowledge about the preparation of A. calamus DMSO extract and to explore its potential anti-inflammatory, antioxidant, and antimicrobial effects. Materials and methods The A. calamus DMSO extract was derived from leaves, and its antioxidant activity was evaluated through the use of the 2,2-diphenyl-1-picryl hydrazyl (DPPH) assay, hydroxyl radical scavenging assay (H2O2 assay), and ferric reducing antioxidant power (FRAP) assay. The anti-inflammatory activity was assessed using the Bovine serum albumin (BSA) denaturation assay, egg albumin (EA) denaturation assay, and membrane stabilization assays. The antimicrobial activity was analyzed using the agar well diffusion technique and the time-kill curve assay. Results In DPPH and H2O2 tests, the DMSO extract of A. calamus showed significant antioxidant activity, near that ofstandard ascorbic acid. The FRAP assay demonstrated a correlation between the dose and the activity of reducing ferric ions. The A. calamusDMSO extract exhibited significant anti-inflammatory properties in BSA and EA denaturation assays, similar to the standard diclofenac sodium. The anti-inflammatory potential of the A. calamus DMSO extract was further confirmed through the membrane stabilization assay. The DMSO extract of A. calamus exhibited a significant inhibition zone against the pathogens Streptococcus mutans and Pseudomonas aeruginosa during the antimicrobial evaluation, surpassing the efficacy of the standard antibiotic. The time-kill curve assay validated the antibacterial efficacy, which was dependent on the concentration. Conclusion The A. calamus DMSO extract exhibited promising antioxidant, anti-inflammatory, and antimicrobial properties, supporting its traditional use in alternative medicine. The findings suggest its potential as a natural resource of compounds with bioactive properties for use in pharmaceutical and nutraceutical applications.

Facile synthesis and optical characterization of selenium nanoparticles synthesized using Clitoria ternatea and Zingiber officinale: in vitro biomedical evaluation of antioxidant potential and antibacterial activity against caries-causing microbes

ABSTRACT In this study, selenium nanoparticles will be produced with Clitoria ternatea and Zingiber officinale, and their efficacy as a natural anti-cariogenic agent will be assessed. To do this, a variety of analysis and evaluation techniques have been used, such as UV-Vis spectroscopy to characterize the synthesized selenium nanoparticles that has absorption band at 440 nm. XRD of the Se nanoparticle were analysed and were found to be crystalline in nature with particles size of 36 nm. SEM images confirmed that Se nanoparticles were flaky structured. The antibacterial activity test to assess the selenium nanoparticles activity against dental caries causing bacteria and the zone of inhibition of the nanoparticles were found. Cytoplasmic leakage assay was done to govern the range of damage instigated by the synthesized selenium nanoparticles to bacterial cell membranes and time kill curve assay represents the time taken by the synthesized selenium nanoparticles to kill the cariogenic bacteria.

Green Synthesis of Euphorbia tirucalli-Mediated Titanium Dioxide Nanoparticles Against Wound Pathogens.

Background Wound infections caused by pathogens present a considerable global health challenge, resulting in extended healing durations, elevated healthcare expenses, and potential fatalities. Conventional approaches to managing wound pathogens have limitations such as antibiotic resistance, toxicity and allergic reactions. Consequently, there is a rising interest in exploring alternative strategies for preventing and treating wound infections. Titanium dioxide nanoparticles (TiO2NPs) have gained attention for their potential in wound healing, attributed to their distinctive properties, including antimicrobial and anti-inflammatory capabilities. Methods TiO2NPssynthesized through Euphorbia tirucalli were examined for their antibacterial potential against wound pathogens, using the Kirby-Bauer agar-well diffusion method and time-kill curve assay. Furthermore, the cytotoxic effect of the synthesized nanoparticles was evaluated through a brine shrimp lethality assay. Results Green-synthesized TiO2NPs demonstrated potent antimicrobial activity against tested wound pathogens, displaying a zone of inhibition against Pseudomonas aeruginosa (11 mm) and Escherichia coli (10 mm) at the highest concentration of 100 μg/mL. In the time-kill curve assay, the prepared TiO2NPs showed significant bactericidal activity against Pseudomonas aeruginosa followed by Escherichia coli.In the brine shrimp lethality assay, at the lowest concentration of 5 μg/mL of the prepared nanoparticles, 100% of the nauplii remained alive after 48 hours. Conclusion The results indicate that TiO2NPs synthesized using Euphorbia tirucalli extract exhibit potent antimicrobial activity against the tested wound pathogens. Moreover, the prepared nanoparticles exhibit lower toxicity, suggesting their potential use as an alternative to commercially available synthetic drugs.

Antimicrobial Activity of Zinc Oxide Nanoparticles Synthesized Using Ocimum Tenuiflorum and Ocimum Gratissimum Herbal Formulation Against Oral Pathogens.

Background This study deals with the antimicrobial efficacy of zinc oxide nanoparticles (ZnONPs) synthesized through green methods employing extracts from Ocimum tenuiflorum and Ocimum gratissimumand assessed for their antimicrobial properties against a range of oral pathogens. Methods Zinc oxide nanoparticles (ZnONPs) were synthesized using extracts from Ocimum tenuiflorumand Ocimum gratissimumthrough a green synthesis approach. Antimicrobial activity was determined using the agar-well diffusion assay to evaluate the consistency of inhibition zones against oral pathogens. Variations in sensitivity were assessed through the time-kill curve assay, quantifying the response of oral pathogens to zinc oxide nanoparticles (ZnONPs) exposure over time. Results The agar-well diffusion assay revealed uniform 9-mm zones of inhibition against all oral pathogens, signifying consistent antimicrobial activity of zinc oxide nanoparticles (ZnONPs). In the time-kill curve assay, Candida albicans exhibited the highest sensitivity, followed by Streptococcus mutans and Staphylococcus aureus. Enterococcus faecalis and Lactobacillus speciesdisplayed lower sensitivity, suggesting potential selectivity. Discussion The observed variation in sensitivity implies the potential selectivity of zinc oxide nanoparticles (ZnONPs) against specific oral pathogens, which may have significant implications for oral health applications. These findings underscore the versatility of green-synthesized zinc oxide nanoparticles (ZnONPs) as promising antimicrobial agents, particularly for oral health applications. Conclusion This study provides promising results for the antimicrobial potential of zinc oxide nanoparticles (ZnONPs) synthesized using Ocimum tenuiflorum and Ocimum gratissimum. The consistent antimicrobial activity and variations in sensitivity among oral pathogens highlight their promising utility in oral health care.

Antimicrobial Activity of Silver Nanoparticles Synthesized Using Ocimum tenuiflorum and Ocimum gratissimum Herbal Formulations.

Background The exploration of green synthesis for silver nanoparticles using diverse plant sources has gained significant attention. This study specifically investigates the use of Ocimum tenuiflorum and Ocimum gratissimum, known for their antibacterial properties, in synthesizing silver nanoparticles. The primary aim of this study is to evaluate the antimicrobial efficacy of silver nanoparticles synthesized using a herbal formulation composed of Ocimum tenuiflorum and Ocimum gratissimumagainst different oral pathogens. Materials and methods The process involved the combination of herbal extracts from Ocimum tenuiflorumand Ocimum gratissimumwith a silver nitrate solution leading to the synthesis of silver nanoparticles. The formation of silver nanoparticles was confirmed by ultraviolet and visible absorption spectroscopy. The obtained silver nanoparticles were used to study their antimicrobial activity. Antimicrobial activity was assessed using the agar well diffusion method against pathogens including Streptococcusmutans, Enterococcus faecalis, C. albicans, Lactobacillus acidophilus, and S. aureus. The zone of inhibition quantified antimicrobial effectiveness. A time-kill curve assay evaluated bactericidal properties and the concentration-dependent relationship between silver nanoparticles and the net growth rate of oral pathogens. Results Statistical analysis was done to compare measures such as mean, standard deviation, and percentages. The antimicrobial assessment demonstrated that 100 μg/mL of silver nanoparticles exhibited the highest efficacy against S. mutans, S. aureus, E. faecalis, and Lactobacillus sp. For C. albicans, all concentrations of silver nanoparticles and the control plant extract displayed similar antimicrobial activity. The time-kill assay illustrated effective inhibition at 100 μg/mL against all tested pathogens, including S. mutans, S. aureus, E. faecalis, C. albicans, and Lactobacillus sp.The result showed positive inhibitory activity of silver nanoparticles against all tested bacterial strains. Conclusion The significant antimicrobial efficacy of green-synthesized silver nanoparticles positions them as promising candidates for dental applications. Their demonstrated bactericidal and fungicidal activities suggest potential use as effective dental antimicrobial agents, opening avenues for innovative solutions in oral healthcare.

Clone dissemination of IncX3 plasmid carrying blaNDM-1 in ST76 carbapenem resistance Klebsiella pneumoniae and bactericidal efficiency of aztreonam combined with avibactam in vitro and in vivo

ObjectivesThe rapid spread of the New Delhi Metal-β-lactamase-1 (NDM-1) gene in Klebsiella pneumoniae poses a substantial challenge to pediatric therapeutic care. Here, we aimed to characterise the IncX3-type plasmid carrying the blaNDM-1 gene in ST76 carbapenem resistance K. pneumoniae (CRKP) strains and assess the in vitro and in vivo bactericidal efficacy of Aztreonam (ATM) combined with Avibactam (AVI) (ATM+AVI) against CRKP. MethodsThe broth microdilution method and PCR were used to detect antimicrobial susceptibility and antibiotic resistance genes. Genetic relatedness was determined using Pulsed-Field Gel Electrophoresis (PFGE) and Multilocus Sequence Typing (MLST). The plasmid conjugation assay was used to verify the transmissibility of drug-resistant plasmids. Whole-Genome Sequencing (WGS) was employed to elucidate the genomic attributes of the genes. The Fractional Inhibitory Concentration (FIC) was calculated based on the checkerboard titration assay to determine the antimicrobial effect of ATM+AVI. The time-kill curve assay and a mouse anti-infection model were used to investigate the in vitro and in vivo bactericidal efficiency of ATM+AVI. ResultsSeven blaNDM-1-producing strains were found to be highly resistant to carbapenems, and they all belonged to the same sequence type (ST76) and were classified into the same PFGE clusters with an 89.1% similarity. The conjugation assay showed that the blaNDM-1-carrying plasmid was successfully transferred to Escherichia coli 600, resulting in transconjugants with carbapenem antibiotic resistance. A 54-kb IncX3 plasmid (pNDM-XZA88) carried the blaNDM-1 gene located on a Tn125 transposon-like element structure, demonstrating the transferability of resistance genes. Genome comparative analysis revealed that pNDM-XZA88 was highly similar to pCQ17 × 3 and pRor-30818cz and had relatively conserved backbones and variable accessory regions compared to the other four plasmids (pC39-334 kb, pNDM-1-DY1928, pNDM-K725, and pNDM-Z244). The checkerboard titration and time-kill curve assays revealed that the ATM+AVI combination therapy exerted significant bactericidal efficacy against the blaNDM-1-producing strains in vitro. The ATM+AVI combination also significantly reduced the bacterial burden in a mouse infection model constructed using the blaNDM-1-producing K. pneumoniae. ConclusionThis study demonstrated the clone dissemination of blaNDM-1-harboring IncX3 plasmids among the ST76 K. pneumoniae isolated from pediatric patients. Therefore, more attention should be paid to preventing this high-risk clone from harming pediatric patients. Moreover, we deduced that the ATM+AVI combination therapy is an effective strategy for treating blaNDM-1-producing K. pneumoniae.

Evaluation Of Antimicrobial Effects Of Orthodontic Band Cement Incorporated With Zirconia, Gold, Copper Biosynthesized Nanoparticles- An In-Vitro Study

OBJECTIVE: To evaluate the antimicrobial effects of orthodontic band cement incorporated with various biosynthesized nanoparticles. MATERIALS AND METHODS: Zirconia, gold, copper Nanoparticles were green synthesized using white tea extract and dry ginger which are incorporated into orthodontic band cement, characterized using FTIR analysis. Antimicrobial activity against Streptococcus mutans and Lactobacillus acidophilus was tested in vitro by disc diffusion and Time Kill Curve Assay. RESULTS: Zirconia nanoparticles incorporated GIC exhibited better antimicrobial activity against streptococcus mutans, and gold nanoparticles incorporated GIC against lactobacillus acidophilus at varying concentrations at different time period. CONCLUSION: The antibacterial effect against streptococcus mutans and lactobacillus acidophilus were distinctly noted in Zirconia nanoparticles, gold nanoparticles followed by copper nanoparticles. Zirconia and gold nanoparticles produced more efficient antibacterial property and eventually would be effective in curtailing White Spot Lesions.

Screening and Characterization of Streptomyces spp. Isolated from Soil Producing a Potential Inhibitors of the Proteus mirabilis

Proteus mirabilis is a common cause of urinary tract infections and other nosocomial infections that can lead to severe health consequences. Despite the availability of various antibiotics, the increasing prevalence of multidrug-resistant (MDR) strains of Proteus mirabilis has necessitated the development of novel therapeutic compounds. The ability of species to produce a wide variety of secondary metabolites, such as antibiotics Streptomyces, immunosuppressants, and antivirals, is well known. In this study, a Streptomyces strain was isolated from soil near saltwater bodies, and its secondary metabolic extract was evaluated for its antimicrobial activity against Proteus mirabilis. The Streptomyces isolate demonstrated significant antimicrobial activity for Proteus mirabilis, with a minimum inhibitory concentration (MIC) of 34 µg/mL. The antimicrobial potential of the extract was confirmed through additional examination of the metabolite extract using deterministic assays, such as nucleic acid and protein release and time-kill curve assay. Molecular identification of the Streptomyces strain showed similarity with Streptomyces triticirhizae. GC-MS analysis of the extract revealed the presence of compounds such as Octadecanoic acid and n-Hexadecanoic acid, which are frequently associated with potent antimicrobial activity. HR-LCMS analysis revealed the presence of compounds such as Neomycin, Polymyxin B, and Maculosin, which also have potent antimicrobial activity. These findings strongly indicate the potential of the Streptomyces metabolite extracts as a novel therapeutic compound against Proteus mirabilis.

Evaluation of Antimicrobial Effect of Biologically Safe Selenium Nanoparticles-coated Clear Aligner—An In vitro Study

Objective To synthesize selenium nanoparticles (SeNPs) from plant extract, characterize them, coat them on clear aligners, and evaluate their cytotoxicity and antimicrobial activity. Materials and Methods The green synthesis of SeNPs utilized the leaves of Withania somnifera. The biosynthesized nanoparticles are incorporated in clear aligners using sol-gel thin film dip coating method to check for antimicrobial effect against Streptococcus mutans and Lactobacillus species using time kill curve assay. The cytotoxicity of the material was checked using brine shrimp lethality assay. Results The optical density values indicated that the SeNPs-coated clear aligners effectively inhibited the growth of Lactobacillus sp. and S. mutans. Time kill curve assessed in varying concentration of 25 µg/mL, 50 µg/mL, and 100 µg/mL revealed greater antibacterial activity at a lower concentration with increasing time. Conclusion Clear aligners coated with SeNPs had positive antibacterial effects on suspensions of Lactobacillus sp. and S. mutans and are biocompatible.

Effect of Klebsiella-specific phage on multidrug-resistant Klebsiella pneumoniae- an experimental study

BackgroundThe objective of this study was to study the effect of Klebsiella-specific phage isolated from sewage with and without the combination of antibiotics (imipenem) on the growth of clinical isolates of multidrug-resistant Klebsiella pneumoniae by time-kill assay and also to study the effect of bacteriophage and bacteriophage-antibiotic (imipenem) combination on biofilm production. MethodsA total of 40 MDR K. pneumoniae isolates were used. Klebsiella-specific phage was isolated using K. pneumoniae subspp. pneumoniae ATCC 33495 as the host from sewage. In vitro time kill curve assays were performed to evaluate the effect of Klebsiella-specific phage with and without the combination of antibiotics on the viable cell counts of MDR K. pneumoniae isolates. Microtiter plate method of O'Toole and Kolter was used to study the effect of Klebsiella-specific phage with and without the combination of antibiotics on biofilm production. For the Time kill assay, results were analyzed for significant differences using Friedman test. Tests for significant differences between the different groups were found using the Mann-Whitney U test. The correlation between the formation of biofilm was analyzed using Karl Pearson's coefficient of correlation. P value of <0.05 was considered to be statistically significant. ResultsIn vitro time-kill assay showed a 0.4 log decline and a 0.5 log decline in K. pneumoniae colony counts at 4 h, when phage was administered individually and in combination with imipenem, respectively (p < 0.001). Phage and phage-imipenem combinations reduced the ability of K. pneumoniae to produce biofilm by 38 % and 53 %, respectively. ConclusionIn conclusion, this study suggests that phage therapy has inhibiting activity against MDR K. pneumoniae. It is found to reduce bacterial cell count and biofilm formation but does not have a total cidal effect. However, in order to get a notable result, a phage cocktail or combination of phage with other antibiotic(s) is suggested.