Antibiofilm Assays Research Articles

Implementation of Silver Nanoparticles Green Synthesized with Leaf Extract of Coccinia grandis as Antimicrobial Agents Against Head and Neck Infection MDR Pathogens.

Head and neck infections (HNI) associated with multidrug resistance (MDR) offer several health issues on a global scale due to inaccurate diagnosis. This study aimed to identify the bacteria and Candidal isolates and implement the silver nanoparticles green synthesized with leaf extract of Coccinia grandis (Cg-AgNPs) as a therapeutic approach against HNI pathogens. The Cg-AgNPs were characterized by the UV-visible spectrophotometer, FT-IR analysis, Zeta particle size, Zeta potential, and field emission scanning electron microscope (FESEM) analysis to validate the synthesis of nanoparticles. Additionally, the antimicrobial activity of Cg-AgNPs was presented by the zone of inhibition (ZOI), minimum inhibitory concentration (MIC), minimum bactericidal/fungicidal concentration (MBC/MFC), and antibiofilm assay. Moreover, the cell wall rupture assay was visualized on SEM for the morphological study of antimicrobial activities, and the in-vivo toxicity was performed in a swiss mice model to evaluate the impact of Cg-AgNPs on various biological parameters. Different bacterial strains (Staphylococcus aureus, Acinetobacter baumannii, Klebsiella pneumoniae, and Pseudomonas aeruginosa) and Candida sp. (Candida albicans, Candida tropicalis, Candida orthopsilosis, and Candida glabrata) were identified. The MIC, MBC, and antibiofilm potential of Cg-AgNPs were found to be highest against A. baumannii: 1.25 μg/ml, 5 μg/ml, and 85.01±5.19% respectively. However, C. albicans and C. orthopsilosis revealed 23 mm and 21 mm of ZOI. Subsequently, the micromorphology of the cell wall rupture assay confirmed the efficacy of Cg-AgNPs, and no significant alterations were seen in biochemical and hematological parameters on the swiss mice model in both acute and subacute toxicity studies. The green synthesized Cg-AgNPs have multifunctional activities like antibacterial, anticandidal, and antibiofilm activity with no toxicity and can be introduced against the HNI pathogens.

Biosynthesis; Characterization; and Antibacterial, Antioxidant, and Docking Potentials of Doped Silver Nanoparticles Synthesized from Pine Needle Leaf Extract

The current study focused on the synthesis of doped silver nanoparticles (doped AgNPs) with yttrium (Y), gadolinium (Gd), and chromium (Cr) from pine needle leaf extract (PNLE). X-ray diffraction (XRD) was performed to assess the phase formation, detecting 61.83% from Ag and 38.17% for secondary phases of AgCl, AgO, Y, Cr-, and Gd phases. The size and shape of the NPs were determined by transmission electron microscopy (TEM), showing a spherical shape with an average particle size of 26.43 nm. X-ray photoelectron spectroscopy (XPS) detected the oxidation state of the presented elements. The scanning electron microscope (SEM) and the energy-dispersive X-ray analysis (EDX) determined the morphology and elemental composition of the NPs, respectively. Fourier transform infrared spectroscopy (FTIR) determined the different functional groups indicating the presence of Ag, Y, Gd, Cr, and other groups. Photoluminescence (PL) spectroscopy showed the optical properties of the NPs. A vibrating sample magnetometer (VSM) revealed the ferromagnetic behavior of the doped AgNPs. The antibacterial activity of the doped AgNPs was tested against six uro-pathogenic bacteria (Staphylococcus aureus, Staphylococcus haemolyticus, Enterococcus faecalis, Escherichia coli, Klebsiella pneumonia, and Pseudomonas aeruginosa) using the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) microdilution assays, agar well diffusion assay, time–kill test, and antibiofilm screening assays, revealing significant activity, with MICs ranging between 0.0625 and 0.5 mg/mL and antibiofilm activity between 40 and 85%. The antioxidant activity was determined by the 1,1, diphenyl 1-2 picrylhydrazyl (DPPH) radical scavenging assay with a potential of 61.3%. The docking studies showed that the doped AgNPs had the potential to predict the inhibition of crucial enzymes such as penicillin-binding proteins, LasR-binding proteins, carbapenemase, DNA gyrase, and dihydropteroate synthase. The results suggest that the doped AgNPs can be applied in different medical domains.

Anti-biofilm and anti-glucosyltransferase effects of nano liposomal plant extracts against Streptococcus mutans

The role of Streptococcus mutans in the initiation of caries is related to its acidogenicity, aciduricity, and polysaccharides extracellular layer production by glucosyltransferases in dental biofilms. Therefore, inhibition of glucosyltransferase activity impairs the virulence of cariogenic biofilms, which can be used to prevent dental caries. We evaluated the anti-bacterial, anti-biofilm, and anti-glucosyltransferases effects of nanoliposomal herbal aqueous extracts of Liquorice (Glycyrrhiza glabra; G. glabra), Ginger (Zingiber officinale; Z. officinale), Pomegranate (Punica granatum; P. granatum), and Rose (Rosa damascene; R. damascene) via minimum bactericidal concentration and minimum inhibitory concentration against Streptococcus mutans strain ATCC 35,668. An anti-biofilm assay was performed using a minimum biofilm inhibitory concentration test. Among herbs, only P. granatum showed an antibacterial effect. Therefore, a nanoliposomal formulation of P. granatum was developed and characterized. Its effect on S.mutans glucosyltransferases was assessed by measuring glucan amount. The nanoliposomal formulation of P.granatum showed a significantly higher anti-biofilm effect than P. granatum aqueous extract. Their similar potential in blocking glucosyltransferases showed that the nanoliposomal formulation of P.granatum blocked other pathways rather than blocking glucosyltransferases for its anti-biofilm effect. Collectively, the nanoliposomal formulation of P.granatum, due to its anti- Streptococcus mutans characteristics, would be a production which open a new horizon for the oral pharmaceutical industry.

Characterization of two Friunavirus phages and their inhibitory effects on biofilms of extremely drug resistant Acinetobacter baumannii in Dakar, Senegal

BackgroundAcinetobacter baumannii is a gram-negative, opportunistic pathogen, that is responsible for a wide variety of infections and is a significant cause of hospital-acquired infections. A. baumannii is listed by the World Health Organization (WHO) as a critical priority pathogen because of its high level of antibiotic resistance and the urgent need for alternative treatment solutions. To address this challenge, bacteriophages have been used to combat bacterial infections for more than a century, and phage research has regained interest in recent years due to antimicrobial resistance (AMR). However, although the vast majority of deaths from the AMR crisis will occur in developing countries in Africa and Asia, few phages’ studies have been conducted in these regions. In this study, we present a comprehensive characterization of the bacteriophages vAbBal23 and vAbAbd25, actives against extremely drug-resistant (XDR) A. baumannii.MethodsPhages were isolated from environmental wastewaters in Dakar, Senegal. The host-range, thermal and pH stabilities, infection kinetics, one step growth assay, antibiofilm activity assay, sequencing, and genomic analysis, were performed to characterize the isolated phages.ResultsComparative genomic and phylogenetic analyses revealed that vAbBal23 and vAbAbd25 belong to the Caudoviricetes class, Autographiviridae family and Friunavirus genus. Both phages demonstrated activity against strains with capsular type KL230. They were stable over a wide pH range (pH 3 to 9) and at temperatures ranging from 25 °C to 40 °C. Additionally, the phages exhibited notable activity against both planktonic and biofilm cells of targeted extremely drug resistant A. baumannii. The results presented here indicate the lytic nature of vAbBal23 and vAbAbd25. This is further supported by the absence of genes encoding toxins, resistance genes and bacterial virulence factors, highlighting their potential for future phage applications.ConclusionPhages vAbBal23 and vAbAbd25 are promising biological agents that can infect A. baumannii, making them suitable candidates for use in phage therapies.

Enhancing Gentamicin Antibacterial Activity by Co-Encapsulation with Thymoquinone in Liposomal Formulation

Background and Purpose. Gentamicin (GEN) is a broad-spectrum antibiotic that cannot be prescribed freely because of its toxicity. Thymoquinone (THQ), a phytochemical, has antibacterial, antioxidant, and toxicity-reducing properties. However, its hydrophobicity and light sensitivity make it challenging to utilize. This incited the idea of co-encapsulating GEN and THQ in liposomes (Lipo-GEN-THQ). Method. Lipo-GEN-THQ were characterized using the zeta-potential, dynamic light scattering, Fourier transform infrared spectroscopy, and transmission electron microscope (TEM). The liposomes’ stability was evaluated under different storage and biological conditions. Lipo-GEN-THQ’s efficacy was investigated by the minimum inhibitory/bactericidal concentrations (MICs-MBCs), time–kill curves, and antibiofilm and antiadhesion assays. Bacterial interactions with the empty and GEN-THQ-loaded liposomes were evaluated using TEM. Results. The Lipo-GEN-THQ were spherical, monodispersed, and negatively charged. The Lipo-GEN-THQ were relatively stable and released GEN sustainably over 24 h. The liposomes exhibited significantly higher antibacterial activity than free GEN, as evidenced by the four-fold lower MIC and biofilm eradication in resistant E. coli strain (EC-219). TEM images display how the empty liposomes fused closely to the tested bacteria and how the loaded liposomes caused ultrastructure damage and intracellular component release. An antiadhesion assay showed that the Lipo-GEN-THQ and free GEN (0.125 mg/L) similarly inhibited Escherichia coli (EC-157) adhesion to the A549 cells (68% vs. 64%). Conclusions. The Lipo-THQ-GEN enhanced GEN by combining it with THQ within the liposomes, reducing the effective dose. The reduction in the GEN dose after adding THQ may indirectly reduce the toxicity and aid in developing an enhanced and safer form of GEN.

Innovative Microencapsulation of Polymyxin B for Enhanced Antimicrobial Efficacy via Coated Spray Drying.

This study aims to develop an innovative microencapsulation method for coated Polymyxin B, utilizing various polysaccharides such as hydroxypropyl β-cyclodextrin, alginate, and chitosan, implemented through a three-fluid nozzle (3FN) spray drying process. High-performance liquid chromatography (HPLC) analysis revealed that formulations with a high ratio of sugar cage, hydroxypropyl β-cyclodextrin (HPβCD), and sodium alginate (coded as ALGHCDHPLPM) resulted in a notable 16-fold increase in Polymyxin B recovery compared to chitosan microparticles. Morphological assessments using fluorescence labeling confirmed successful microparticle formation with core/shell structures. Alginate-based formulations exhibited distinct layers, while chitosan formulations showed uniform fluorescence throughout the microparticles. Focused beam reflectance and histograms from fluorescence microscopic measurements provided insights into physical size analysis, indicating consistent sizes of 6.8 ± 1.2 μm. Fourier-transform infrared (FTIR) spectra unveiled hydrogen bonding between Polymyxin B and other components within the microparticle structures. The drug release study showed sodium alginate's sustained release capability, reaching 26 ± 3% compared to 94 ± 3% from the free solution at the 24 h time point. Furthermore, the antimicrobial properties of the prepared microparticles against two Gram-negative bacteria, Escherichia coli and Pseudomonas aeruginosa, were investigated. The influence of various key excipients on the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values was evaluated. Results demonstrated effective bactericidal effects of ALGHCDHPLPM against both E. coli and P. aeruginosa. Additionally, the antibiofilm assay highlighted the potential efficacy of ALGHCDHPLPM against the biofilm viability of E. coli and P. aeruginosa, with concentrations ranging from 3.9 to 500 μg/m. This signifies a significant advancement in antimicrobial drug delivery systems, promising improved precision and efficacy in combating bacterial infections.

Structural studies and functional validation of Eugenol and Ferulic acid against Enterococcus faecalis Sortase A.

Enterococcus faecalis (E. faecalis) is commonly occurring pathogen associated with nosocomial infections. Infections are difficult to treat because of their multidrug-resistant (MDR) nature and their tendency to form biofilms. Therefore, it is essential to find alternative medicinal approaches of treatment. In this regard, targeting an important protein for drug development can be an alternative approach. Sortase A (SrtA) is an important enzyme involved in anchoring cell surface-exposed proteins to the cell envelope. SrtA is present in Gram-positive bacteria which catalyses the attachment of several virulence factors and other proteins to the cell membrane. It is involved in bacterial pathogenesis, therefore, it's a promising drug target for the development of anti-microbial drugs targeting cell adhesion, evasion, and biofilm development.To identify SrtA potential inhibitors, we havepurified E. faecalis Sortase A (EfSrtAΔN59). Structural studiesalong with molecular docking of protein with selected ligand molecules were done andconfirmed by MD simulation experiments. We have also performed functional validation of these compounds on bacterial growth, anti-biofilm assays and inhibition assay of selected ligands were also done against E. faecalis individually and in synergistic combinations. Results indicated that both Eugenol and Ferulic acid bind to EfSrtAΔN59 with significant interactions and show promising results.

Assessment of the potential susceptibility of planktonic cells and bacterial biofilms by diosgenin

Bacterial biofilms are formed by a complex community of microorganisms, held together by a matrix that provides structural support and stability to microbial communities, making infections associated with biofilms more difficult to treat. In the face of growing research in the health area in search of antibiotics with action against biofilms, natural products present an effective alternative strategy. In this context, the aim of this study was to evaluate bacterial biofilm formation, assess the susceptibility of planktonic cells to diosgenin, and examine its efficacy on inhibiting and treating biofilms. The broth microdilution method was used to determine the Minimum Inhibitory Concentration (MIC). Six bacterial strains were tested: Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 9027, Pseudomonas aeruginosa 24, Enterococcus faecalis INCQS 0017, Enterococcus faecalis INCQS 00018, and Enterococcus faecalis INCQS 654. The crystal violet method was employed to evaluate the potential of diosgenin in biofilm inhibition and eradication. Chlorhexidine gluconate served as a reference standard for the antibiofilm assays, along with the antibiotics norfloxacin, ampicillin, and gentamicin. MIC determination revealed that diosgenin exhibited intrinsic antimicrobial activity, with an MIC of 406 μg/mL against S. aureus ATCC 25923. The antibiofilm activity showed significant inhibition for E. faecalis 17 and moderate inhibition against E. faecalis 654 and S. aureus 25923. Furthermore, diosgenin effectively eradicated preformed biofilms of S. aureus 25923 and P. aeruginosa 9027, indicating its ability to interfere with biofilm biomass. These results suggest that diosgenin not only inhibits bacterial biofilm formation but also eradicates established biofilms, highlighting it as a promising candidate for the development of products aimed at combating bacterial biofilms.

Synergistic antimicrobial combination of third-generation cephalosporins and polymyxin B against carbapenem-polymyxin-resistant Klebsiella pneumoniae: an in vitro and in vivo analysis.

Antibiotic combination therapy is a promising approach to address the urgent need for novel treatment options for infections caused by carbapenem-polymyxin-resistant Klebsiella pneumoniae (CPR-Kp). The present study aimed to investigate the synergistic potential of four cephalosporins in combination with polymyxin B (PMB). A checkerboard assay was performed to evaluate the synergistic effects of cephalexin (CLX), cefixime, cefotaxime (CTX), and cefmenoxime (CMX) in combination with PMB. Subsequently, experiments evaluating the use of CTX or CMX in combination with PMB (CTX-PMB or CMX-PMB, respectively), including growth curve and SynergyFinder analysis, antibiofilm activity assays, cell membrane integrity assays, and scanning electron microscopy, were performed. Safety assessments were also conducted, including hemolysis and toxicity evaluations, using Caenorhabditis elegans. Furthermore, an in vivo model in C. elegans was adopted to assess the treatment efficacy against CPR-Kp infections. CTX-PMB and CMX-PMB exhibited low fractional inhibitory concentration indexes ranging from 0.19 to 0.50 and from 0.25 to 1.5, respectively, and zero interaction potency scores of 37.484 and 15.076, respectively. The two combinations significantly reduced growth and biofilm formation in CPR-Kp. Neither CTX-PMB nor CMX-PMB compromised bacterial cell integrity. Safety assessments revealed a low hemolysis percentage and high survival rates in the C. elegans toxicity evaluations. The in vivo model revealed that the CTX-PMB and CMX-PMB treatments improved the survival rates of C. elegans. The synergistic effects of the CTX-PMB and CMX-PMB combinations, both in vitro and in vivo, indicate that these antibiotic pairings could represent effective therapeutic options for infections caused by CPR-Kp.

Green Synthesis of Yttrium Derivatives Nanoparticles Using Pine Needle Leaf Extract: Characterization, Docking, Antibacterial, and Antioxidant Potencies

Green nanoparticles are synthesized using environmentally friendly methods, and natural materials hold significant importance. This makes the process environmentally sustainable and reduces the production of harmful waste by-products. Green nanoparticles exhibit reduced toxicity which is crucial for biomedical applications. The current study suggested that yttrium nanoparticles (YNPs) should be synthesized, characterized, and evaluated for their diverse biological applications due to the rise in antibacterial resistance. The YNPs were prepared using a pine needle leaf extract (PNLE). The structural and morphological features have been investigated using X-ray diffraction (XRD), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDX), transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS), photoluminescence (PL), Fourier transform infrared spectroscopy (FTIR), ultraviolet–visible spectroscopy (UV–vis), and vibrating sample magnetometry (VSM). The XRD pattern demonstrated the presence of yttrium oxide and yttrium nitrate phases. The crystallite size and particle size of the synthesized YNPs measured 1.696 nm and 24.55 nm, respectively. The XPS peaks showed two components with binding energies at 530.940 eV and 532.18 eV due to the bond between O–Y and OH–Y, respectively. Additionally, the ferromagnetic nature of the YNPs was confirmed by VSM analysis. The YNPs were tested for antibacterial activity on six uropathogenic bacteria (S. aureus, S. haemolyticus, E. faecalis, E. coli, K. pneumonia, and P. aeruginosa) using the microdilution assays, to find the minimum inhibitory concentration (MIC) as well as the minimum bactericidal concentration (MBC), the agar well diffusion assay, and antibiofilm screening assays, where they showed bacteriostatic action against all isolates (0.5–1 mg/mL MIC) and significant inhibition of biofilm formation (80% inhibition rate). The antioxidant capacity assessed by 1,1, diphenyl-2-picrylhydrazyl (DPPH) radical scavenging revealed 50% DPPH scavenging. Moreover, docking studies exhibited that YNPs inhibit crucial bacterial enzymes, including DNA gyrase, penicillin-binding proteins, carbapenemase, LasR-binding protein, and dihydropteroate synthase. These findings may explain the mechanisms responsible for the observed antibacterial effects of YNPs. Overall, these findings underscore YNPs as promising candidates for antioxidant and antibacterial applications.

Isolation and Molecular Characterization of Therapeutic Probiotic Lactobacillus from Different Sources in Ernakulam-Kerala

This study aims to present knowledge on therapeutic potential bacteriocin producing bacterial strains. Samples (n=35) from diverse Ernakulam sources yielded 26 lactic acid bacteria (LAB) isolates on De Man Rogosa Sharpe (MRS) medium. Among the isolates potent antimicrobial activity observed against known pathogens in one of the isolates from goat milk sample number 3, labelled as GO3. Microscopic and biochemical test revealed the isolate is Gram-positive rod, non-sporulating and resilience to pH and bile salts, with adaptability to gastric enzymes and surfactants. Genetic and physiological traits of the positive strains were confirmd by 16S rRNA technique. Blast analysis revealed that sequence of the strain GO3 from goat milk identified as Lactobacillus casei, exhibited antibiotic sensitivity, auto-aggregation, co-aggregation, and antibiofilm assays highlighted inhibitory properties as a promising therapeutic potential probiotic bacterium. LAB isolates can be further investigated at the genetic level to enhance their probiotic characteristics such as resistance and cellular adhesion. Additionally, the identification of gene responsible for expressing bacteriocins can be conducted on these isolates.

Ecofriendly biosynthesis of copper nanoparticles from novel marine S. rhizophila species for enhanced antibiofilm, antimicrobial and antioxidant potential

Marine microorganisms offer a promising avenue for the eco-friendly synthesis of nanoparticles due to their unique biochemical capabilities and adaptability to various environments. This study focuses on exploring the potential of a marine bacterial species, Stenotrophomonas rhizophila BGNAK1, for the synthesis of biocompatible copper nanoparticles and their application for hindering biofilms formed by monomicrobial species. The study begins with the isolation of the novel marine S. rhizophila species from marine soil samples collected from the West coast region of Kerala, India. The isolated strain is identified through 16S rRNA gene sequencing and confirmed to be S. rhizophila species. Biosynthesis of copper nanoparticles using S. rhizophila results in the formation of nanoparticles with size of range 10–50 nm. The nanoparticles exhibit a face-centered cubic crystal structure of copper, as confirmed by X-Ray Diffraction analysis. Furthermore, the synthesized nanoparticles display significant antimicrobial activity against various pathogenic bacteria and yeast. The highest inhibitory activity was against Staphylococcus aureus with a zone of 27 ± 1.00 mm and the least activity was against Pseudomonas aeruginosa with a zone of 22 ± 0.50 mm. The zone of inhibition against Candida albicans was 16 ± 0.60 mm. The antibiofilm activity against biofilm-forming clinical pathogens was evidenced by the antibiofilm assay and SEM images. Additionally, the copper nanoparticles exhibit antioxidant activity, as evidenced by their scavenging ability against DPPH, hydroxyl, nitric oxide, and superoxide radicals, as well as their reducing power in the FRAP assay. The study highlights the potential of the marine bacterium S. rhizophila BGNAK1 for the eco-friendly biosynthesis of copper nanoparticles with diverse applications. Synthesized nanoparticles exhibit promising antibiofilm, antimicrobial, and antioxidant properties, suggesting their potential utility in various fields such as medicine, wastewater treatment, and environmental remediation.

Hybrid nanoparticles combining nanoselenium-mediated Carica papaya extract and trimethyl chitosan for combating clinical multidrug-resistant bacteria

Multidrug-resistant bacterial infections pose a significant threat to human health, prompting the exploration of innovative solutions. In this study, a new series of antibacterial hybrid nanoparticles (HNPs) were developed. The HNPs are based on a combination of selenium nanoparticles (SeNPs), synthesized using Carica papaya leaf extract, and chitosan (CS/SeHNPs) or trimethyl chitosan (TMC/SeHNPs), respectively. Comprehensive characterization using UV‐Vis, FTIR, XRD, SEM-EDX, DLS, TEM, and DSC confirmed the structure and properties of the developed HNPs. SeNPs, CS/SeHNPs, and TMC/SeHNPs showed average hydrodynamic size of 78.8, 91.3, and 122 nm, and zeta potentials of −6.35 mV, +32.8 mV, and +54.8 mV, respectively. Biological assessments were conducted, including antibacterial and antibiofilm assays against clinical strains (E. coli, S. aureus, and K. pneumoniae), along with antioxidant activity. TMC/SeHNPs demonstrated superior performance compared to SeNPs and CS/SeHNPs with the lowest minimum inhibition concentrations (MIC) against S. aureus and K. pneumoniae (3.9 μg/mL) and 62.5 μg/mL against E. coli in addition to robust antibiofilm activity. Furthermore, the TMC/SeHNPs exhibited potent DPPH free radical scavenging ability and demonstrated good biocompatibility, as evidenced by cell viability assays on HFB4 cells. Overall, TMC/SeHNPs emerged as promising candidates in nanomedicine, offering high antioxidant, antibacterial, and antibiofilm activities alongside excellent biocompatibility.

Bioactive compound from marine seagrass Streptomyces argenteolus TMA13: combatting fish pathogens with time-kill kinetics and live-dead cell imaging.

Actinobacteria, pervasive in aquatic and terrestrial environments, exhibit a filamentous morphology, possess DNA with a specific G + C content and production of numerous secondary metabolites. This study, focused on actinobacteria isolated from marine seagrass, investigating their antibacterial activity against fish pathogens. Among 28 isolates, Streptomyces argenteolus TMA13 displayed the maximum zone of inhibition against fish pathogens-Aeromonas hydrophila (10mm), Aeromonas caviae (22mm), Edwardsiella tarda (17mm), Vibrio harveyi (22mm) and Vibrio anguillarum (12mm) using the agar plug method. Optimization of this potent strain involved with various factors, including pH, temperature, carbon source and salt condition to enhance both yield production and antibacterial efficacy. In anti-biofilm assay shows the maximum percentage of inhibition while increasing concentration of TMA13 extract. Minimal Inhibitory Concentration (MIC) and Minimal Bactericidal Concentration (MBC) assays with TMA13 crude extract demonstrated potent activity against fish pathogens at remarkably low concentrations. Time-kill kinetics assay showcased growth curve variations over different time intervals for all fish pathogens treated with a 100µg/ml concentration of crude extract, indicating a decline in cells viability and progression into the death phase. Additionally, fluorescence microscopic visualization of bacterial cells exposed to the extracts emitting green and red fluorescence, enabling live-dead cell differentiation was also studied. Further characterization of the crude extract through GC-MS and FT-IR analyses performed and identified secondary metabolites with functional groups exhibiting significant antibacterial activity. This study elucidates the capacity of Streptomyces argenteolus TMA13 to enhance the production of antibiotic compounds effective against fish pathogens.

In vitro and In silico Network Pharmacology Analysis of Eugenol from Syzygium aromaticum Against Escherichia coli

ABSTRACT Background: In this era, Escherichia coli is an ambivalent microbe and commonly causes fecal contamination. Eugenol from clove oil has also demonstrates antibacterial effects that may aid in combating against the microorganisms that cause tooth infections. Aim and Objectives: The current study provides a comprehensive analysis of the pharmacological characteristics of eugenol, a significant bioactive substance extracted from clove (Syzygium aromaticum). The present study aimed to target the genes from E. coli identified through network pharmacology analysis of eugenol and antibiofilm effect of eugenol from clove oil against E. coli. Materials and Methods: In the present study, eugenol was isolated from clove identified through Fourier transform infrared spectroscopy analysis. The antibacterial and antibiofilm activity of eugenol was performed against E. coli were investigated. Targets were identified for Eugenol from clove oil through string database and mapped with E. coli pathway. Molecular docking analysis was carried out for the identified targets and eugenol. Results: The best antibacterial activity of eugenol from clove oil was found against E. coli and evaluated by zone of inhibition and minimum inhibitory concentration analysis at 10 μL. The antibiofilm assay against resistant E. coli was determined by concentration of 10 μL and inhibited in 100% dilution. The top five active targets from E. coli was identified by network pharmacology and docking analysis shows the good binding interaction between targets and eugenol. Conclusion: The research provided in this study lays the foundation for eugenol’s anti-bacterial property with various biofilm forming targets namely Acetyl-CoA acetyltransferase, Acetyl-CoA carboxylase-B, Acetyl-CoA carboxylase-C, Enoyl-[acyl-carrier-protein] reductase [NADH], and 3-oxoacyl-[acyl-carrier-protein] synthase 1. Additional in vivo approach validation is required in future to support the current study.

Revealing the Phenolic Composition and the Antioxidant, Antimicrobial and Antiproliferative Activities of Two Euphrasia sp. Extracts.

The species of the genus Euphrasia present important medicinal potential according to their traditional uses. However, few studies aim to sustain this fact by scientific evidence. The present study aimed to explore the phytochemical profile and investigate the antioxidant, antimicrobial and antiproliferative potential of E. officinalis subsp. pratensis Fr. (EO) and E. stricta J.P.Wolff ex J.F.Lehm (ES). The tested samples consisted of ethanolic extracts. The identification and quantification of phenolic compounds were performed using spectrophotometric and LC-MS/MS methods. The antioxidant capacity was evaluated using the DPPH, FRAP and xanthine oxidase methods. Antimicrobial properties were screened using disk diffusion, broth microdilution and anti-biofilm assays, while antiproliferative potential was assessed on a colorectal adenocarcinoma human cancer cell line (DLD-1). The LC-MS/MS analysis showed chlorogenic acid and rutin as the dominant constituents in the tested extracts. The antioxidant activity assays showed important capacity for both samples; in vitro antimicrobial and anti-biofilm properties were exhibited, especially against Gram-positive bacteria, and an important inhibitory potential was observed on the proliferation of the DLD-1 cell line. The findings in the present study contribute to the recommendation of EO and ES for the prevention and treatment of oxidative stress-related pathologies, cancer and microbial infections.

Ceftazidime and Usnic Acid Encapsulated in Chitosan-Coated Liposomes for Oral Administration against Colorectal Cancer-Inducing Escherichia coli.

Escherichia coli has been associated with the induction of colorectal cancer (CRC). Thus, combined therapy incorporating usnic acid (UA) and antibiotics such as ceftazidime (CAZ), co-encapsulated in liposomes, could be an alternative. Coating the liposomes with chitosan (Chi) could facilitate the oral administration of this nanocarrier. Liposomes were prepared using the lipid film hydration method, followed by sonication and chitosan coating via the drip technique. Characterization included particle size, polydispersity index, zeta potential, pH, encapsulation efficiency, and physicochemical analyses. The minimum inhibitory concentration and minimum bactericidal concentration were determined against E. coli ATCC 25922, NCTC 13846, and H10407 using the microdilution method. Antibiofilm assays were conducted using the crystal violet method. The liposomes exhibited sizes ranging from 116.5 ± 5.3 to 240.3 ± 3.5 nm and zeta potentials between +16.4 ± 0.6 and +28 ± 0.8 mV. The encapsulation efficiencies were 51.5 ± 0.2% for CAZ and 99.94 ± 0.1% for UA. Lipo-CAZ-Chi and Lipo-UA-Chi exhibited antibacterial activity, inhibited biofilm formation, and preformed biofilms of E. coli. The Lipo-CAZ-UA-Chi and Lipo-CAZ-Chi + Lipo-UA-Chi formulations showed enhanced activities, potentially due to co-encapsulation or combination effects. These findings suggest potential for in vivo oral administration in future antibacterial and antibiofilm therapies against CRC-inducing bacteria.

The influence of the chlorine atom on the biological activity of 2′-hydroxychalcone in relation to the lipid phase of biological membranes - Anticancer and antimicrobial activity

The study investigates the effect of the presence of a chlorine atom in the 2′-hydroxychalcone molecule on its interaction with model lipid membranes, in order to discern its potential pharmacological activity. Five chlorine derivatives of 2′-hydroxychalcone were synthesized and evaluated against liposomes composed of POPC and enriched with cationic (DOTAP) or anionic (POPG) lipids. The physicochemical properties of the compounds were initially simulated using SwissAdame software, revealing high lipophilicity (ilogP values: 2.79–2.90). The dynamic light scattering analysis of liposomes showed that chloro chalcones induce minor changes in the diameter of liposomes of different surface charges. Fluorescence quenching assays with a TMA-DPH probe demonstrated the strong ability of the compounds to interact with the lipid bilayer, with varying quenching capacities based on chlorine atom position. FTIR studies indicated alterations in carbonyl, phosphate, and choline groups, suggesting a transition area localization rather than deep penetration into the hydrocarbon chains. Additionally, dipole potential reduction was observed in POPC and POPC-POPG membranes, particularly pronounced by derivatives with a chlorine atom in the B ring. Antibacterial and antibiofilm assays revealed enhanced activity of derivatives with a chlorine atom compared to 2′-hydroxychalcone, especially against Gram-positive bacteria. The MIC and MBIC50 values showed increased efficacy in the presence of chlorine with 3′-5′-dichloro-2′-hydroxychalcone demonstrating optimal antimicrobial and antibiofilm activity. Furthermore, antiproliferative assays against breast cancer cell lines indicated higher activity of B-ring chlorine derivatives, particularly against MDA-MB-231 cells. In general, the presence of a chlorine atom in 2′-hydroxychalcone improves its pharmacological potential, with derivatives showing improved antimicrobial, antibiofilm, and antiproliferative activities, especially against aggressive breast cancer cell lines. These findings underscore the importance of molecular structure in modulating biological activity and highlight chalcones with a chlorine as promising candidates for further drug development studies.

Antibacterial and Antifungal Activities of Cimbopogon winterianus and Origanum syriacum Extracts and Essential Oils against Uropathogenic Bacteria and Foodborne Fungal Isolates.

This study focused on testing the antibacterial and antifungal activity of Origanum syriacum (O. syriacum) and Cimbopogon winterianus (C. winterianus) extracts and their essential oils (EOs). The bacteria were isolated from urine samples and identified by a VITEK assay, and the fungi were isolated from spoiled food samples and further identified by MALDI-TOF. The susceptibility of the microbial isolates was assessed by determining the bacteriostatic and bactericidal/fungicidal effects by the minimum inhibitory concentration (MIC) and minimum bactericidal/fungicidal concentration (MBC/MFC) broth microdilution assay and time-kill test. The antibiofilm activities were assessed by the antibiofilm screening assays. The bacterial isolates included three Gram-negative isolates (Escherichia coli, Klebsiella pneumonia, and Citrobacter freundii) and two Gram-positive isolates (Staphylococcus aureus and Streptococcus intermedius). The fungal isolates included Candida albicans and Aspergillus niger. The O. syriacum and C. winterianus extracts exhibited bacteriostatic and fungistatic activities (MIC 1.25-2.5 mg/mL for the bacterial isolates and 2.5-5 mg/mL for the fungal isolates). However, their EOs exhibited bactericidal (MBC 5-20%) and fungicidal (MFC 1.25-10%) activities, meaning that the EOs had a better antimicrobial potential than the extracts. The antibiofilm activities of the mentioned extracts and their EOs were relatively weak. The O. syriacum extract inhibited S. aureus, S. intermedius, and K. pneumonia biofilms at a concentration of 0.3125 mg/mL and C. albicans and A. niger biofilms at 0.625 mg/mL. No antibiofilm activity was recorded for C. winterianus extract. In addition, the packaging of grapes with C. winterianus extract preserved them for about 40 days. The results reflect the significant antimicrobial activity of O. syriacum and C. winterianus extracts and their EOs, thus suggesting their potential in food packaging and preservation.

Evaluation of antibiofilm activity of metal oxides nanoparticles and carbon nanotubes coated styrofoam on the bacterium Jeotgalicoccus huakuii

The co-existence of metal oxide nanoparticles (MONPs), carbon-based nanomaterials and microplastics (MPs) in the natural environment are expected to be of growing global concern due to their increasing abundance and persistence in the environment worldwide. Knowledge of the interaction of the above compounds particularly under light irradiation in water remains limited. In the present study, the possible individual and combined toxic effects of MONPs, carbon nanotubes (CNTs) through styrofoam (SF) on the environmental bacterium Jeotaglicoccus huakuii were systematically investigated. The fabricated MONPs and CNTs were characterized using the following techniques: FT-IR (functional groups), XRD (crystallinity), SEM, and EDX (topological morphology). The objective of this study was to investigate and identify naturally occurring bacteria capable of mitigating and detoxifying toxic pollutants under adverse conditions. Moreover, the assessment of minimum inhibitory concentration (MIC) was made through an agar well plate method, resazurin (ELISA measurement), growth kinetics and bacterial viability were assessed employing live and dead assay and biofilm combating ability was analyzed using an antibiofilm assay. Further, the biotransformation of f-MWCNTs by J. huakuii was evaluated employing RT-PCR and SEM analysis. The results demonstrated that the toxicity of Pb3O4@f-MWCNTs was comparatively higher than the remaining Pb3O4 NPs and SF coated NPs.. Interestingly, J. huakuii showed resistance against f-MWCNTs at very high concentrations and able to utilize f-MWCNTs as a sole carbon source suggesting J. huakuii as a suitable aquatic bioremediation tool for both MONPs and CNTs transfer via MPs. The results also enhanced our understanding of the affinity of MPs towards MONPs and CNTs under extreme environmental conditions.