Broth Microdilution Technique Research Articles

Antimicrobial combination effects against multidrug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa strains: A cross-sectional study.

Emergence of multidrug resistance in non-fermenting Gram-negative bacilli is a threat to public health. Combination therapy is a strategy for the treatment of antibiotic-resistant infections. In this cross-sectional study, a total of 63 nonduplicate clinical isolates of Acinetobacter baumannii and Pseudomonas aeruginosawere collected from various specimens. Identification of bacterial isolates was performed by phenotypic and molecular tests. Antibiotic susceptibility patterns and detection of β-lactamase genes were determined using the broth microdilution and polymerase chain reaction (PCR) techniques, respectively. Then, the combined effects analysis was determined by the checkerboard method. Based on the status of resistance to carbapenems (imipenem and meropenem), 25 isolates of each genus were selected for further investigation. For A. baumannii, bla OXA-23, bla OXA-58, and bla OXA-48 genes were positive in 21 (84%), 17 (68%), and 11 (44%) of isolates, respectively. In P. aeruginosa isolates, bla VIM was the most common gene (44%) and other genes including bla IMP, bla NDM, and bla OXA-23 were found in nine (36%), six (24%), and three (12%) isolates, respectively. Meropenem (MER)-tigecycline (TIG) had a significant synergistic effect against 20 (80%) A. baumannii (pvalue < 0.001). This combination was also efficient against 5 (20%) P. aeruginosa isolates. Moreover, the other combination, tigecycline-amikacin (TIG-AMK) was effective against 10 (40%) A. baumannii isolates. The combination of colistin (COL) and MER showed a significant synergistic effect against 21 (84%) A. baumannii (pvalue < 0.001) and 17 (68%) P. aeruginosa isolates (pvalue < 0.001). The MER-TIG and COL-MER combinations are promising options against resistant bacteria. Our study could be helpful for the development of a new treatment recommendation.

Biological Activities of Virgin Coconut and Virgin Olive Oil Mixture against Oral Primary Colonizers: An In Vitro Study.

This study aimed to explore the potential synergistic interaction of virgin coconut oil (VCO) and virgin olive oil (VOO) mixture against Streptococcus sanguinis, Streptococcus mutans, and Lactobacillus casei in a single and mixture species through the minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), antiadherence, and antibiofilm activities. The broth microdilution technique was used to individually determine the MIC of both oils and an oil mixture (in the ratio of 1:1) in a 96-well microtiter plate. As for the MBC, the subcultured method was used. The fractional inhibitory concentration index (ΣFIC) was determined to identify the interaction types between both oils. The oil mixture at its MIC was then tested on its antibiofilm and antiadherence effect. The MIC of the oil mixture against the tested microbiota was 50-100%. The oil mixture was bactericidal at 100% concentration for all the mentioned microbes except S. mutans. The ΣFIC value was 2 to 4, indicating that the VCO and VOO acted additively against the microbiota. Meanwhile, the oil mixture at MIC (50% for S. sanguinis and L. casei; 100% for S. mutans and mixture species) exhibited antiadherence and antibiofilm activity toward the microbiota in mixture species. The oil mixture possesses antibacterial, antibiofilm, and antiadherence properties toward the tested microbiota, mainly at 50-100% concentration of oil mixture. There was no synergistic interaction found between VCO and VOO. Children and individuals with special care may benefit from using the oil mixture, primarily to regulate the biofilm formation and colonization of the bacteria. Furthermore, the oil mixture is natural and nontoxic compared to chemical-based oral healthcare products. How to cite this article: Ng YM, Sockalingam SNMP, Shafiei Z, et al. Biological Activities of Virgin Coconut and Virgin Olive Oil Mixture against Oral Primary Colonizers: An In Vitro Study. J Contemp Dent Pract 2024;25(3):260-266.

Evaluation of ZnO nanoparticles from ‘Monsooned Malabar Robusta Coffee’ husk as a potential antioxidant and biocidal candidate: A sustainable valorization approach

Nanotechnology has become a sustainable strategy to combat drug resistance. As agro-waste management has become a concern, efficient management of produced waste has been an imposing global issue. This study evaluated antioxidant, as well as antibacterial and antibiofilm potential of zinc oxide nanoparticles (ZnO NPs) synthesized by hydrothermal approach using ethanolic extract of ‘Monsooned Malabar Robusta coffee’ husk against multi-drug-resistant (MDR) strains of enteroaggregative Escherichia coli, Salmonella Enteritidis, S. Typhimurium and methicillin-resistant Staphylococcus aureus. The fabrication of ZnO NPs was confirmed by spectroscopy, whereas thermogravimetric analysis-differential thermogravimetric analysis confirmed stability of ZnO NPs, while the X-ray diffraction pattern confirmed the wurtzite crystalline structure. The agglomerated nature of ZnO NPs with a nearly spherical shape was evident with scanning electron microscopy, whereas transmission electron microscopy revealed a poly-crystalline nature with a mean diameter of 26.33 ± 3.778 nm. The microbroth dilution technique revealed a minimum inhibitory concentration (MIC) of 250 μg/mL and a minimum bactericidal concentration (MBC) of 500 μg/mL for the ZnO NPs. Furthermore, ZnO NPs exhibited significant antibiofilm activity against the MDR-test strains. Moreover, ZnO NPs were tested safe at MIC and MBC doses in chicken erythrocytes, and commensal gut microflora tested were not inhibited. Besides, a dose-dependent antioxidant property was exhibited by ZnO NPs. Additionally, in vitro time-kill kinetic assay of MDR-test strains treated with ZnO NPs revealed a complete bacterial clearance at 24 h. Overall, the synthesis of ZnO NPs from coffee husk demonstrated a simple, eco-friendly and valorization approach that could be devised as a potential delivery molecule.

In vitro inhibitory activity of Riparins against Candida spp. strains and in silico interaction with multi-drug-resistance proteins

Candida spp. includes opportunistic pathogens that are responsible for superficial and invasive infections. These strains have mechanisms of resistance against antimicrobials. Riparin compounds isolated from the green fruit of the plant Aniba riparia (Nees) Mez, are alkaloids with alkamide function with notable antimicrobial properties. We evaluated the inhibitory effect (in vitro) of Riparins isolated and combined with Fluconazole against strains of Candida albicans, C. tropicalis, and C. krusei; and in an in silico test, the anti-Candida activity of the factors MdR1, CdR1, and CdR2. Riparins were identified by 1D NMR spectroscopy (1 H and 13 C), electrospray ionization mass spectrometry (ESI-MS), and compared with previously reported spectral data. For the antifungal activity profile, the broth microdilution technique was used (1–1024 µg/mL). For combined antifungal activity, Riparins were tested at fixed subinhibitory concentrations (RI, RII, and RIV of 256 µg/mL and RIII, 64 µg/mL), and FCZ, ranging from 1 to 1024 µg/mL. Minimum Fungicide Concentration was performed by microculture in Petri dishes. For molecular docking, protein sequences were obtained from GenBank; targets were analyzed for homology, and model reliability was assessed using the Ramachandran plot. Composite stratification was performed by the MolDock Score, Rerank Score, PLANTS Score, and Vina Score. The results demonstrated that Riparins I, II, III, and IV, isolated or combined with FCZ have an inhibitory effect on Candida spp. cells, and a high probability of inhibiting antifungal resistance mechanisms, MdR1, CdR1, and CdR2.

Zein nanoparticles containing ceftazidime and tobramycin: antibacterial activity against Gram-negative bacteria.

Aims: This work describes the encapsulation of ceftazidime and tobramycin in zein nanoparticles (ZNPs) and the characterization of their antibacterial and antibiofilm activities against Gram-negative bacteria. Materials & methods: ZNPs were synthesized by nanoprecipitation. Cytotoxicity was assessed by MTT assay and antibacterial and antibiofilm assays were performed by broth microdilution and violet crystal techniques. Results: ZNPs containing ceftazidime (CAZ-ZNPs) and tobramycin (TOB-ZNPs) showed drug encapsulation and thermal stability. Encapsulation of the drugs reduced their cytotoxicity 9-25-fold. Antibacterial activity, inhibition and eradication of biofilm by CAZ-ZNPs and TOB-ZNPs were observed. There was potentiation when CAZ-ZNPs and TOB-ZNPs were combined. Conclusion: CAZ-ZNPs and TOB-ZNPs present ideal physical characteristics for in vivo studies of antibacterial and antibiofilm activities.

The Antioxidant, Antibacterial and Cell-Protective Properties of Bioactive Compounds Extracted from Rowanberry (Sorbus aucuparia L.) Fruits In Vitro.

Considering that Sorbus aucuparia fruits have been underutilized despite their tremendous potential, this study aimed to correlate the in vitro antioxidant, antibacterial and cell-protective abilities of fruit extracts derived from Sorbus aucuparia Romanian cultivars with their phytochemical composition. Therefore, following the preparation of ethanolic and carotenoid extracts, phytochemical screening was performed using UV-Vis and HPLC-DAD-ESI-MS methods. The antioxidant activity was analyzed using DPPH and FRAP tests. As the results revealed high contents of bioactive compounds (polyphenols 1.11 mg GAE/g DM, flavonoids 430.06 µg QE/g DM and carotenoids 95.68 µg/g DM) and an important antiradical action (DPPH 24.51 mg/mL and FRAP 0.016 µM TE/mL), we chose to further examine the fruits' biological properties. The antibacterial capacity was assessed employing agar well diffusion and broth microdilution techniques, with fruits displaying an intense activity against MSSA, MRSA and Enterococcus faecalis, but also E. coli and Pseudomonas aeruginosa. The cell-protective activity was analyzed on gentamicin-stressed renal cells, through MTT and Annexin V-FITC assays. Importantly, a significant increase in viability was registered on stressed cells following extract administration in low doses; nevertheless, viability was noticed to decline when exposed to elevated concentrations, potentially due to the cumulative actions of the extract and gentamicin. These findings offer novel light on the antibacterial activity of Sorbus aucuparia Romanian cultivars, as well as their cell-protective ability in renal cell injury.

Use of a combined antibacterial synergy approach and the ANNOgesic tool to identify novel targets within the gene networks of multidrug-resistant Klebsiella pneumoniae.

Since the 1980s, the development of new drug classes for the treatment of multidrug-resistant Klebsiella pneumoniae has become limited, highlighting the urgent need for novel antibiotics. To address this challenge, this study aimed to explore the synergistic interactions between chemical compounds and representative antibiotics, such as carbapenem and colistin. The primary objective of this study was not only to mitigate the adverse impact of multidrug-resistant K. pneumoniae on public health but also to establish a sustainable balance among humans, animals, and the environment. Phenotypical measurements were conducted using the broth microdilution technique to determine the drug sensitivity of bacterial strains. Additionally, a genotypical approach was employed, involving traditional RNA sequencing analysis to identify differentially expressed genes and the computational ANNOgesic tool to detect noncoding RNAs. This study revealed the existence of various pathways and regulatory RNA elements that form a functional network. These pathways, characterized by the expression of specific genes, contribute to the combined treatment effect and bacterial survival strategies. The connections between pathways are facilitated by regulatory RNA elements that respond to environmental changes. These findings suggest an adaptive response of bacteria to harsh environmental conditions.IMPORTANCENoncoding RNAs were identified as key players in post-transcriptional regulation. Moreover, this study predicted the presence of novel small regulatory RNAs that interact with target genes, as well as the involvement of riboswitches and RNA thermometers in conjunction with associated genes. These findings will contribute to the discovery of potential antimicrobial therapeutic candidates. Overall, this study offers valuable insights into the synergistic effects of chemical compounds and antibiotics, highlighting the role of regulatory RNA elements in bacterial response, and survival strategies. The identification of novel noncoding RNAs and their interactions with target genes, riboswitches, and RNA thermometers holds promise for the development of antimicrobial therapies.

Evaluation of the antibacterial, modulatory and anti-adherent properties of oregano (Origanum vulgare) essential oil against food pathogenic bacteria

There are over 250 types of foodborne diseases, the majority of which are infections caused by bacteria. Klebsiella pneumoniae, Pseudomonas aeruginosa, and Staphylococcus saprophyticus are considered contaminants of meat products. The use of natural products as antimicrobials to combat these diseases can be an effective and economical approach. This study proposes to assess the antibacterial, modulatory, and anti-adherent activity of the essential oil of Origanum vulgare against strains of Klebsiella pneumoniae, Pseudomonas aeruginosa, and Staphylococcus saprophyticus isolated from meat products. The assay was conducted in duplicate. The minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) were determined using the broth microdilution technique. MIC represents the lowest concentration of the product capable of inhibiting the growth of the bacterial strain, whereas MBC represents the lowest concentration capable of inhibiting total growth. The study of association of the product with antimicrobials was undertaken by disk diffusion using ampicillin, gentamicin, ceftazidime, and ciprofloxacin, resulting in synergistic, antagonistic, or indifferent effects. Anti-adherent activity was determined in the presence of sucrose, as the lowest concentration of the agent in contact with sucrose that prevented adherence to the glass tube. Oregano oil exhibited strong inhibitory and bactericidal activity against Klebsiella pneumoniae, Pseudomonas aeruginosa, and Staphylococcus saprophyticus, with MIC values ranging from 32 to 512 μg mL-1 and MBC values ranging from 128 to 512 μg mL-1. Origanum vulgare oil showed varied interactions when associated with antimicrobials, with modulations for synergism (37.5%), indifference (50%), and antagonism (12.5%). Regarding anti-adherent activity, the test product effectively inhibited the adherence of P. aeruginosa bacterial strains in the presence of sucrose (1:8) but had no effect against K. pneumoniae or S. saprophyticus. Therefore, oregano oil proves to be an antibacterial and modulating agent against different bacteria isolated from meat products. Additionally, it displays anti-adherent properties against P. aeruginosa, making it a natural product that could serve as an interesting alternative in efforts to combat foodborne diseases.

Glandular trichomes as a source of antifungal metabolites in a karyologically characterized population of Argyrochosma flava: Morphology and histochemistry

Argyrochosma, a xeric-adapted fern genus mostly from the New World, includes both farinose and non-farinose species. The yellow farina appears to be a synapomorphy of A. flava. The aims of this research were to study the morphology, location, distribution, and density of glandular trichomes; perform a histochemical analysis of the farina secretion; evaluate the biological activity of farinas against phytopathogenic fungi; and characterize the species genetically by making chromosome counts and building the karyotype of A. flava. Usual methods in plant anatomy and histochemistry were performed. Minimum Inhibitory Concentrations (MICs) and Minimum Fungicidal Concentrations (MFCs) of farina against four phytopathogenic fungi were determined by the broth microdilution technique. Chromosome counts and karyotyping metaphase plates were obtained from fiddleheads. The distribution of glandular trichomes in pinnules changes along the ontogenetic development of the frond. The histochemical study of the trichomes revealed the complex nature of the secreted material, including flavonoids and terpenes along with other metabolites. The farina was active against the four pathogenic fungi evaluated, with MICs between 31.25 and 125 μg/mL showing the highest inhibition for Monilinia fructicola. Botrytis cinerea displayed a MIC value of only one-fold dilutions more than the commercial fungicide Carbendazim. Cytogenetically, the base number proposed for the genus is x = 27, which is derived from aneuploidy of x = 29. The genus has diploid and tetraploid forms. The results obtained support the important role of glandular trichomes and farinous exudate secretion in the protection against phytopathogenic fungi. The obtained chromosome numbers support the taxonomic placement of A. flava.

Antimicrobial activity of selected nutraceutical plants used in Northern Uganda

Background: Nutraceutical plants (NP) play a vital role as supportive treatment with antiretroviral drugs (ARVs). However, there is limited scientific evidence on the efficacy of NP to justify their extensive use. This study aimed to evaluate the antibacterial activity of three nutraceutical plants which are commonly used as antimicrobials.&#x0D; Methodology: Leaves of Cajanus cajan L. Millsp. and Eucalyptus globulus Labill., and stem bark of Mangifera indica L. were collected from Northern Uganda. The three samples of each NP were extracted with acetone and the minimum inhibitory concentration (MIC) values of the extracts against Staphylococcus aureus, Streptococcus pneumoniae, Pseudomonas aeruginosa and Klebsiella pneumoniae were determined using the serial broth microdilution technique [1]. The mean MIC values of the extracts against each bacterial species were recorded.&#x0D; Results: The three NP extracts were active against all the four bacteria species with MIC ranging from 0.08 to 2.5 mg/ml. The extract of Cajanus cajan was very active against Klebsiella pneumoniae with the lowest recorded MIC value of 0.08 mg/ml. The extract of Mangifera indica bark was very active against Pseudomonas aeruginosa with the lowest MIC of 0.08 mg/ml.&#x0D; Conclusion: The results of the present study support the traditional use of the nutraceutical plants as antimicrobials.

Phytochemical screening of Podocarpus lambertii Klotzch ex Endl. leaf extracts and potential antimicrobial, antioxidant and antibiofilm activity.

Species of the genus Podocarpus L'Hér. ex Pers.present biological activities, such as analgesic, antioxidant, antifungal, acting in the fight against anemia, depurative and fortifying. Podocarpus lambertii Klotzch ex Endl. is a Brazilian native species popularly known as maritime pine and lacks information about its phytochemical profile and possible biological activities. The study was conducted to determine the phytochemical composition of soluble plant extracts of acetone (EA), ethyl acetate (EAE) and hexane (HE) from leaves of P. lambertii; evaluate the antimicrobial potential by the broth microdilution technique; antioxidant potential by the DPPH method, as well as to evaluate the biofilm inhibition capacity by the crystal violet assay and reduction of the yellow tetrazolium salt (MTT). Phytochemical screening detected the presence of flavonoids, triterpenoids, steroids, tannins, alkaloids and saponins. All extracts showed antimicrobial activity on the microorganisms tested, and the EA showed the best results. High free radical scavenging potential was observed only in EAE (96.35%). The antibiofilm potential was observed in the EAE extract. The results contribute to the knowledge of the species and indicate the potential of P. lambertii extracts as a source of plant bioactives for the development of new alternative strategies to control resistant microorganisms.

Investigation of phytochemical composition and bioactivity evaluation of extracts from Myrsine umbellata Mart.

The objective of the study was to carry out phytochemical prospection through colorimetric tests to determine the groups of secondary metabolites and also to determine the total content of phenolic compounds (TPC) present in plant extracts methanol (ME), ethyl acetate (EAE), hexane (HE) and dichloromethane (DE) from the leaves of Myrsine umbellata, as well as to investigate the antimicrobial activity against twelve standard ATCC strains by the broth microdilution technique; the antioxidant potential by the DPPH method and the ABTS method and the antibiofilm potential on the biofilm biomass of standard bacteria by the crystal violet technique and tetrazolium salt reduction (MTT) assay. Phytochemical prospection detected the presence of saponins, steroids, alkaloids, anthocyanins, anthocyanidins, flavonoids, and tannins. The results of the quantitative phytochemical estimation revealed a higher content of total phenolics in DE (280.24 ± 0.037 µM GAE g ext. -1) followed by ME (159.01 ± 0.031 µM GAE g ext. -1). The ME showed the best biological activities when compared to the other extracts tested. We observed antimicrobial activity against Gram-positive Staphylococcus epidermidis strain (MIC 3.12 and MBC 6.25), antioxidant percentage of 92.58% against the DPPH radical and 420.31 µM Trolox g ext. -1 against the ABTS radical, finally showed antibiofilm action against Gram-positive strain Staphylococcus aureus, with eradication of the biomass in 92.58%. The results suggest that EM from M. umbellata represents an alternative source of plant bioactives for the development of natura products.

Utilizing the photodynamic properties of curcumin to disrupt biofilms in Cutibacterium acnes: A promising approach for treating acne

BackgroundThe treatment of acne vulgaris is often challenging due to the antibiotic resistance frequently observed in Cutibacterium acnes (C.acnes), a prevalent bacterium linked to this condition. ObjectiveThe objective of this research was to examine the impact of curcumin photodynamic therapy (PDT) on the survival of C.acnes and activity of biofilms produced by this microorganism. MethodsFollowing the Clinical and Laboratory Standards Institute (CLSI) guidelines, we assessed the drug sensitivity of 25 clinical C.acnes strains to five antibiotics (erythromycin, clindamycin, tetracycline, doxycycline, minocycline) and curcumin by implementing the broth microdilution technique. In addition, we established C.acnes biofilms in a laboratory setting and subjected them to curcumin-PDT(curcumin combined with blue light of 180 J/cm2). Afterwards, we evaluated their viability using the XTT assay and observed them using confocal laser scanning microscopy. ResultsThe result revealed varying resistance rates among the tested antibiotics and curcumin, with erythromycin, clindamycin, tetracycline, doxycycline, minocycline, and curcumin exhibiting resistance rates of 72 %, 44 %, 36 %, 28 %, 0 %, and 100 %, respectively. In the curcumin-PDT inhibition tests against four representative antibiotic-resistant strains, it was found that the survival rate of all strains of planktonic C. acnes was reduced, and the higher the concentration of curcumin, the lower the survival rate. Furthermore, in the biofilm inhibition tests, the vitality and three-dimensional structure of the biofilms were disrupted, and the inhibitory effect became more significant with higher concentrations of curcumin. ConclusionThe results emphasize the possibility of using curcumin PDT as an alternative approach for the treatment of C.acnes, especially in instances of antibiotic-resistant variations and infections related to biofilms.

Fungicidal and antibiofilm activities of 2-Bromo-N-Phenylacetamide Against Fluconazole-Resistant Candida spp.: a promising antifungal agent

Purpose: Amides are proven to possess antimicrobial properties against both fungi and bacteria. The synthetic amide 2-bromo-N-phenylacetamide exhibits antifungal activity according to scientific literature, however, data remains scarce. Given the clinical and epidemiological importance as well as the increasing antifungal resistance in Candida spp., here we aimed to better explore the antifungal potential of 2-bromo-N-phenylacetamide against fluconazole-resistant Candida albicans, Candida glabrata, Candida parapsilosis and for the first time, against Candida tropicalis as well. Methods: Minimum Inhibitory and Minimum Fungicidal Concentrations (MIC/MFC) were established by a Broth Microdilution Technique. Mode of action was explored by Ergosterol and Sorbitol assays. For investigating the combined effect between 2-bromo-N-phenylacetamide and the standard antifungals Amphotericin B and Fluconazole a Checkerboard Technique was performed. Finally, the antibiofilm properties of 2-bromo-N-phenylacetamide were assessed by a Crystal Violet Staining Assay. Results: 2-bromo-N-phenylacetamide exhibited a fungicidal effect, MIC of 32 µg.mL-1 (87.5% of the strains) and MFC of 64 µg.mL-1 (81.25% of the strains). It also demonstrated an activity against Candida albicans mature biofilms as effective as that for Amphotericin B at all tested concentrations. When associated with Fluconazole and Amphotericin B it exerted indifferent effects. Mode of action remained unclear. Conclusion: The synthetic amide 2-bromo-N-phenylacetamide possesses a strong antifungal activity against Candida albicans, Candida tropicalis, Candida glabrata and Candida parapsilosis and an antibiofilm property against Candida albicans as effective as that exhibited for Amphotericin B.

Antibacterial and Antiviral Properties of Pinus densiflora Essential Oil.

The Korean mountains are home to the Korean red pine (Pinus densiflora). Pine needle oil has been used as a food additive and a traditional herbal medicine; however, any health-related properties of its trunk oil remain unknown. Herein, we assessed antibacterial and antiviral properties of essential oil extracted from the trunk of P. densiflora. Th extracted oil was hydrodistilled using a Clevenger apparatus and analyzed using gas chromatography-mass spectrometry. The antimicrobial activity of the oil was tested using the microbroth dilution technique against 10 bacterial species (6 g-positive and 4 g-negative) and fungi. The extract exerted strong antimicrobial activity against Vibrio parahaemolyticus, Bacillus cereus, Listeria monocytogenes, Propionibacterium acnes, and Malassezia furfur (minimum inhibitory concentration = 10 mL/L). Additionally, it exhibited dose-dependent activity against influenza virus A and feline coronavirus. Furthermore, among 20 identified constituents accounting for 98.7% of the oil contents, the major components included 3-cyclohexene-1-methanol (10.12%), 2-(4-methylcyclohexyl)-2-propanol (9.09%), fenchone (8.14%), O-isopropyltoluene (6.35%), and isothymol methyl ether (6.14%). The P. densiflora trunk essential oil showed antibacterial and antiviral activities that depended on its chemical composition and the microbial strains tested herein. The essential oil can be used as an antimicrobial agent and disinfectant.

Molecular Characterization of Escherichia coli Producing Extended-Spectrum ß-Lactamase and MCR-1 from Sick Pigs in a Greek Slaughterhouse.

The first prospective surveillance of ESBL and colistin-resistant Escherichia coli recovered from sick pigs from a slaughterhouse in Central Greece aimed to investigate the spread of relevant genetic elements. In February 2021, 25 E. coli isolates were subjected to antimicrobial susceptibility testing using disk diffusion and broth microdilution techniques. PCR screening was conducted to identify ESBLs and mcr genes. Additional assays, encompassing mating-out procedures, molecular typing utilizing Pulsed-Field Gel Electrophoresis, multilocus sequence typing analysis, and plasmid typing, were also conducted. A 40% prevalence of ESBLs and an 80% prevalence of MCR-1 were identified, with a co-occurrence rate of 32%. The predominant ESBL identified was CTX-M-3, followed by SHV-12. Resistance to colistin, chloramphenicol, cotrimoxazol, and ciprofloxacin was detected in twenty (80%), fifteen (60%), twelve (48%), and four (16%) isolates, respectively. All blaCTX-M-3 harboring plasmids were conjugative, belonging to the incompatibility group IncI1, and approximately 50 kb in size. Those carrying blaSHV-12 were also conjugative, classified into incompatibility group IncI2, and approximately 70 kb in size. The mcr-1 genes were predominantly located on conjugative plasmids associated with the IncX4 incompatibility group. Molecular typing of the ten concurrent ESBL and MCR-1 producers revealed seven multilocus sequence types. The heterogeneous population of E. coli isolates carrying resistant genes on constant plasmids implies that the dissemination of resistance genes is likely facilitated by horizontal plasmid transfer.

Antibacterial Activity of Surfactin and Synergistic Effect with Conventional Antibiotics Against Methicillin-Resistant Staphylococcus aureus Isolated from Patients with Diabetic Foot Ulcers.

The prevalence of diabetic foot ulcers (DFUs) is increasing, leading to a huge financial burden and human suffering. Furthermore, antibiotic resistance is an urgent problem in the realm of clinical practice. Antimicrobial peptides are an effective and feasible strategy for combating infections caused by drug-resistant bacteria. Therefore, we investigated the in vitro antimicrobial ability of the lipopeptide surfactin, either alone or in combination with conventional antibiotics, against the standard and clinical strains of Staphylococcus aureus, including methicillin-resistant S. aureus (MRSA), isolated from patients with DFUs. The minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of surfactin on the selected strains were evaluated by a microbroth dilution technique. The growth curves of the selected strains with and without surfactin were measured, and transmission electron microscopy was used to observe the structure of surfactin-treated bacterial cells. The biofilm inhibitory abilities of surfactin were assessed by crystal violet staining. The antimicrobial interactions between surfactin and conventional antibiotics were established using a checkerboard assay, as well as determining the mutant prevention concentration. The inhibitory effect of surfactin on penicillinase was tested by iodometry. The MIC and MBC values of surfactin ranged from 512 to 1024 µg/mL and 1024 to 2048 µg/mL, respectively. Moreover, surfactin significantly prevented the S. aureus biofilm formation and displayed limited toxicity on human red blood cells. The synergies between surfactin and ampicillin, oxacillin, and tetracycline against S. aureus were revealed. In vitro resistance was not readily produced by surfactin. The action of surfactin may be by disrupting bacterial cell membranes and inhibiting penicillinase. Surfactin appears to be a potential option for the treatment of DFUs infected with MRSA, as it is capable of improving antimicrobial activities and can be used alone or in combination with conventional antibiotics to prevent or postpone the emergence of resistance.

The Essential Oil of Sclerorhachis leptoclada Rech.f.: Chemical Composition and Biological Activity

Objective/Background: Sclerorhachis leptoclada Rech.f. belong to the Asteraceae family and is a very aromatic plant applied as a traditional herbal tea to treat digestive illnesses and also to flavor many food products. This study explored the chemical structure, antioxidant, and antibacterial actions of the essential oils (EOs) of S. leptoclada. Methods: The oil conformation was recognized using gas chromatography-mass spectrometry examination. 2,2-Diphenyl-1-picrylhydrazyl (DPPH) test was utilized to assess the antioxidant action of the EOs. In vitro antimicrobial action of the EOs of S. leptoclada was experienced against two Gram-positive ( Klebsiella pneumoniae, Staphylococcus aureus) and two Gram-negative ( Escherichia coli, Pseudomonas aeruginosa) bacteria by the broth microdilution technique. Results: The EOs of S. leptoclada involved a large number of monoterpenes. α-Terpineol (14.6%) and (E)-nerolidol (13.2%) were the key component of the oil. The IC50 value for antioxidant activity of EOs from S. leptoclada was 25.83 µg/mL. The results indicated a good antimicrobial action of S. leptoclada oil against Gram-positive compared with Gram-negative bacteria. Conclusion: The findings revealed a great number of monoterpenes and phenolic composites in S. leptoclada EO as well as the plant's antimicrobial and antioxidant activities.

Synthesis, Antimicrobial, Antioxidant Evaluation, and DFT Estimation of some New Cyclohexenone Derivatives Derived from Benzyloxy Chalcones.

New cyclohexenone derivatives candidates were designed to discover their antioxidant and antibacterial activity potentials, respectively. Aldehydes with the diverse functional group were prepared from 4-hydroxy benzaldehyde and benzyl bromide and converted to chalcones by reaction with 4-substituted benzophenones and 2-acetyl naphthalene. When chalcone derivatives were subjected to ethyl acetoacetate, it produced new cyclohexenone derivatives through NaOH- catalyzed addition-ring closure reaction. The new compound structures were strengthened by their spectral information. The new compounds are examined for in vitro antifungal and antibacterial actions through broth microdilution technique, and they exhibited potential responses against various bacteria and fungi. As a result, composites with (-F) group as a substituent on para position were established to be the forceful derivatives against S. aureus, E. coli organism, and C Albicans since this compound could inhibit the microbial and Fungai growth at lower concentrations compared to the standards (Ciprofloxacin HCl, and Fluconazole). DPPH radical scavenging experiments were used to evaluate proton donating antioxidant effects; it was exposed that compound 7c has the most significant antioxidant activity, while less than the standard ascorbic acid. Finally, HOMO-LUMO was calculated, which represents the quantum mechanical calculations of energies and conducted by the theory of density functional (DFT) method based on the level of (B3LYP) with 6-31G (d, p) basis set.

Evaluation of the Antibacterial and Anti-Adherent Activities of Eucalyptus globulus and Eucalyptus citriodora Essential Oils against Pseudomonas aeruginosa Strains

Biofilm can be defined as a complex sessile microbiological ecosystem formed by one or more species of bacteria, fungi or protozoa. Among the microorganisms capable of forming biofilms is Pseudomonas aeruginosa, a gram-negative bacterium with extensive virulence factors and high resistance to antimicrobials, making it difficult to treat infections caused by it. In this sense, phytotherapy and essential oils are promising therapeutic alternatives to multi-resistant microorganisms. The aim of this study was to evaluate the antibacterial and anti-adherent activities of Eucalyptus globulus and Eucalyptus citriodora essential oils against clinical strains of P. aeruginosa, and to compare their antibiofilm effects with 0.12% chlorhexidine digluconate. To determine the Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) of the oils, the broth microdilution technique was used in 96-well plates. To determine the Minimum Adherence Inhibitory Concentration (MAIC), the inclined glass tube technique was used in the presence of 5% sucrose, using proportions corresponding to the pure essential oil up to a dilution of 1:1024, and its positive control was 0.12% chlorhexidine digluconate. It was therefore concluded that the essential oils under study had an antibacterial effect against the strains tested, varying between bactericidal and bacteriostatic action, as well as demonstrating an effective anti-adherence effect that was superior to the positive control.