Determination Of Minimum Inhibitory Concentration Research Articles

Hytochemical Analysis and Determination of MIC and MFC of Cacao Leaves Extract (Theobroma cacao L.) against Malassezia furfur

Pityriasis versicolor is a disease caused by Malassezia furfur. One of the plants that can potentially act as antifungal is cacao leaves (Theobroma cacao L.), which contain several secondary metabolite compounds, including alkaloids, flavonoids, tannins, quinones, terpenoids, and saponins. This study aims to determine the phytochemical analysis and the Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal Concentration (MFC) of cacao leaves ethanolic extract against M. furfur. Extracts were obtained by maceration using 70% ethanol and phytochemicals analysis using Herborne’s method. Furthermore, the extracts were made into concentrations’ variations of 3.125%; 6.25%; 12.5%; 25%; 50%; and 100% for testing antifungal activity, with 10% DMSO as negative control and 2% Ketoconazole and Itraconazole as positive controls. MIC was performed with liquid macrodilution method, and MFC was performed with spread method. The results of phytochemical analysis showed that the extract contained alkaloids, flavonoids, phenols, tannins, quinones, terpenoids and saponins. MIC value was obtained at concentration of 50% and MFC value was obtained at 100%. Cacao leaves ethanolic extract with concentration of 50% can inhibit M. furfur’s growth and with 100% can eliminate M. furfur.

Prevalence and genetic basis of Mycobacterium tuberculosis resistance to pretomanid in China

BackgroundPretomanid is a key component of new regimens for the treatment of drug-resistant tuberculosis (TB) which are being rolled out globally. However, there is limited information on the prevalence of pre-existing resistance to the drug.MethodsTo investigate pretomanid resistance rates in China and its underlying genetic basis, as well as to generate additional minimum inhibitory concentration (MIC) data for epidemiological cutoff (ECOFF)/breakpoint setting, we performed MIC determinations in the Mycobacterial Growth Indicator Tube™ (MGIT) system, followed by WGS analysis, on 475 Mycobacterium tuberculosis (MTB) isolated from Chinese TB patients between 2013 and 2020.ResultsWe observed a pretomanid MIC distribution with a 99% ECOFF equal to 0.5 mg/L. Of the 15 isolates with MIC values > 0.5 mg/L, one (MIC = 1 mg/L) was identified as MTB lineage 1 (L1), a genotype previously reported to be intrinsically less susceptible to pretomanid, two were borderline resistant (MIC = 2–4 mg/L) and the remaining 12 isolates were highly resistant (MIC ≥ 16 mg/L) to the drug. Five resistant isolates did not harbor mutations in the known pretomanid resistant genes.ConclusionsOur results further support a breakpoint of 0.5 mg/L for a non-L1 MTB population, which is characteristic of China. Further, our data point to an unexpected high (14/475, 3%) pre-existing pretomanid resistance rate in the country, as well as to the existence of yet-to-be-discovered pretomanid resistance genes.

Biogenic Synthesis of Selenium and Copper Oxide Nanoparticles and Inhibitory Effect against Multi-Drug Resistant Biofilm-Forming Bacterial Pathogens

Antimicrobial resistance (AMR), caused by microbial infections, has become a major contributor to morbid rates of mortality worldwide and a serious threat to public health. The exponential increase in resistant pathogen strains including Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) poses significant hurdles in the health sector due to their greater resistance to traditional treatments and medicines. Efforts to tackle infectious diseases caused by resistant microbes have prompted the development of novel antibacterial agents. Herein, we present selenium and copper oxide monometallic nanoparticles (Se-MMNPs and CuO-MMNPs), characterized using various techniques and evaluated for their antibacterial potential via disc diffusion, determination of minimum inhibitory concentration (MIC), antibiofilm, and killing kinetic action. Dynamic light scattering (DLS), scanning electron microscopy (SEM/EDX), and X-ray diffraction (XRD) techniques confirmed the size-distribution, spherical-shape, stability, elemental composition, and structural aspects of the synthesized nanoparticles. The MIC values of Se-MMNPs and CuO-MMNPs against S. aureus and E. coli were determined to be 125 μg/mL and 100 μg/mL, respectively. Time–kill kinetics studies revealed that CuO-MMNPs efficiently mitigate the growth of S. aureus and E. coli within 3 and 3.5 h while Se-MMNPs took 4 and 5 h, respectively. Moreover, CuO-MMNPs demonstrated better inhibition compared to Se-MMNPs. Overall, the proposed materials exhibited promising antibacterial activity against S. aureus and E. coli pathogens.

Total Phenolic Compound and Antimicrobial Activity of Spirogyra spp.

Spirogyra spp. is widely distributed in freshwater habitats and is included in the category of natural resources whose significance and advantages have been researched. In this study, 6 samples of Spirogyra spp. were collected: Spirogyra ellipsospora Transeua from Bueng Kan province, Spirogyra sp.1 from Sakon Nakhon Province and S. neglecta (Hassall) Kützing, S. ellipsospora Transeua, Spirogyra sp.2, and Spirogyra sp.3 from Nakhon Phanom province. The highest value of total phenolic compound was measured at 18.53±0.23 mg GAE/mg in Spirogyra sp. 2. The evaluation of antimicrobial activity of algal extracts against fungal Candida sp. and bacteria Staphylococcus aureus and Escherichia coli was performed by using the disk diffusion method along with determination of minimum inhibitory concentration (MIC), minimum fungicidal concentration (MFC) and minimum bacterial concentration (MBC). The result of disk diffusion and MIC determination showed the efficiency of all extract samples against all tested strains at a concentration of 10 mg/mL. The most effective algal extract against Candida sp. showed with the MFC’s ranged from 10 to 20 mg/mL, whereas MBC of all extracts destroyed the two tested bacterial strains ranged from 10 to 20 mg/mL within 24 hours and up to 30 mg/mL after 48 hours. The results of this study support that Spirogyra spp. contains bioactive compounds that are the basis for further application in cosmeceuticals.

ISOLATION AND CHARACTERIZATION OF CYCLOEUCALENOL TRANSFERULATE FROM LEAVES OF Euphorbia tithymaloides

The latex, leaves, stems, and roots of Euphorbia tithymaloides (Euphorbiaceae) have traditionally been used to treat earaches, insect stings, ringworm, toothaches, skin cancer, umbilical hernia, and warts. Approximately 1 kg of the pulverized leaves was macerated successively with hexane and ethyl acetate, and resultant Crude extracts were phytochemically screened using standard established procedures. Ethyl acetate extract was loaded onto a column containing slurry of silica gel. Gradients of hexane in ethyl acetate mobile phase were used to elute the column, affording a total of 130 fractions. Fractions 41-47 gave tiny deposit on evaporation labelled UMPT (m.p: 269-270 oC).Their thin layer chromatography gave a single spot of Rf 0.61.These were combined and labelled UMPT44. Phytochemical screening of UMPT44 gave positive test for phenols and triterpenoids. Antimicrobial screening of UMPT44 showed that it was active against some fungi and bacteria. The zone of inhibition measured 30 mm when tested against Coniophora puteana, 29 mm against methicillin-resistant Staphylococcus aureus, 27 mm against Salmonella typhi, and 26 mm against Staphylococcus aureus, Fusarium oxysporum, and Escherichia coli. Additionally, there was a 25 mm zone of inhibition against Serpula lacrymans. The determination of minimum inhibitory concentration and minimum bactericidal/fungicidal concentration indicated that UMPT44 holds promise as a potential antimicrobial agent. Based on evaluation of NMR spectral data and comparison with literature reports, UMPT44 was identified as cycloeucalenol transferulate. This is the first time this compound is isolated from the plant

Synthesis, characterization, and antibacterial activity studies of two Co(II) complexes with 2-[(E)-(3-acetyl-4-hydroxyphenyl)diazenyl]-4-(2-hydroxyphenyl)thiophene-3-carboxylic acid as a ligand

Two new Cobalt(II) complexes 12 and 13 have been synthesized from 2-[(E)-(3-acetyl-4-hydroxyphenyl)diazenyl]-4-(2-hydroxyphenyl)thiophene-3-carboxylic acid (11) as a novel ligand. These three new compounds were characterized on the basis of their powder X-Ray Diffraction, UV–Vis, IR, NMR, elemental analysis and MS spectral data. DFT/B3LYP mode of calculations were carried out to determine some theorical parameters of the molecular structure of the ligand. The purity of the azoic ligand and the metal complexes were ascertained by TLC and melting points. The analysis of the IR spectra of the polyfunctionalized azo compound 11 and its metal complexes 12 and 13, reveals that the coordination patterns of the ligand are hexadentate and tetradentate respectively. Based on the UV–Vis electronic spectral data and relevant literature reports, the ligand and derived complexes were assigned the E (trans) isomer form. Likewise, octahedral and square-planar geometries were respectively assigned to the cobalt(II) complexes. The broth microdilution method was used for antibacterial assays through the determination of minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). The ligand 11 displayed moderate antibacterial activity (MIC = 32–128 μg/mL) against Staphylococcus aureus ATCC25923, Escherichia coli ATCC25922, Pseudomonas aeruginosa and Klebsiella pneumoniae 22. The octahedral cobalt(II) complex 12 showed moderate activity against Pseudomonas aeruginosa (MIC = 128 μg/mL) and Klebsiella pneumoniae 22 (MIC = 64 μg/mL) and none against Staphylococcus aureus ATCC25923 and Escherichia coli ATCC25922, whereas the square-planar complex 13 displayed moderate activity only on Klebsiella pneumoniae 22 (MIC = 64 μg/mL).Graphical abstract

Chemical Characterization and Evaluation of the Antimicrobial Activity of Extracts from Two Cultivars of Cannabis sativa L. (Tisza and Kompolti) Grown in Sardinia

The present work was aimed at the chemical characterization and antimicrobial activity of some extracts of aerial parts (essential oils from leaves and inflorescences and resins from inflorescences) of two legal hemp (Cannabis sativa) varieties, Tisza and Kompolti, grown in Sardinia. Chemical characterization was carried out by gas chromatography (GC) and gas chromatography–mass spectrometry (GC-MS) techniques. The main constituent was myrcene (11.75% in Tisza and 18.21% in Kompolti); delta-9-tetrahydrocannabinol (THC) was not found, while cannabidiol was present up to 0.36% in Tisza and up to 2.80% in Kompolti. The antimicrobial activity of these extracts against a panel of microorganisms was also determined via minimum inhibitory concentration (M.I.C.) determination. While the results showed minor or negligible antimicrobial activity of the extracts against the Gram+ and Candida strains (M.I.C. values equal to or greater than 4 mg/mL), good antibacterial activity (especially of resins) was recorded against S. aureus (M.I.C. 0.015–0.031 mg/mL); no substantial differences were detected between the chemical compositions of the two Cannabis varieties.

Chemical Composition and Antimicrobial Activity of Essential Oils and Hydrosols from Oregano, Sage and Pennyroyal against Oral Pathogens

In this study, the chemical composition of essential oils (EOs) extracted from Origanum vulgare ssp. hirtum Lamiaceae, (oregano), Salvia officinalis Lamiaceae (sage), Mentha pulegium Lamiaceae (pennyroyal), and respective hydrosols (HSs) has been investigated by Gas Chromatography–Mass Spectrometry (GC-MS). The antimicrobial activity was assessed against two oral pathogens: Gram-positive bacterium Streptococcus mutans and the fungus Candida albicans by determination of Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal/Fungicidal concentration (MBC/MFC). Three-fold diluted solutions were dispensed into each well of a 96-well microtiter plate and, after incubation, MIC was determined by visual monitoring. The MBC/MFC was determined by transferring a small quantity of sample contained in each replicate well of the microtiter plates to appropriate culture media using a microplate replicator. The EOs of the tested herbs showed antimicrobial properties, especially the EO oil of O. vulgare, which exerted the highest antimicrobial activity. HSs of S. officinalis and M. pulegium exerted no antimicrobial activity, in contrast to oregano HS, which displayed strong antimicrobial activity. In all cases, a higher number of compounds were detected in EOs than in the corresponding HSs. The major compounds of sage EO were detected to be α-thujone (25.1%), 1,8-cineole (15.8%) and β-pinene (10.0%), while the HS was characterized by the presence of 1,8-cineole (32.6%), borneol (22.6%) and α-thujone (22.4%). Pennyroyal EO and HS consists mainly of pulegone (62.1 and 50.6%, respectively). Carvacrol was the major component present in EO (63%) and HS (97.3%) of oregano, probably contributing to the antimicrobial activity. Further research is needed in order to elucidate the antimicrobial mechanisms of specific compounds present in essential oils and hydrosols of Lamiaceae grown in Greece against oral pathogens.

Novel synergistic interactions between monolaurin, a mono-acyl glycerol and β lactam antibiotics against Staphylococcus aureus: an in vitro study

BackgroundA major worldwide health issue is the rising frequency of resistance of bacteria.Drug combinations are a winning strategy in fighting resistant bacteria and might help in protecting the existing drugs.Monolaurin is natural compound extracted from coconut oil and has a promising antimicrobial activity against Staphylococcus.aureus. This study aims to examine the efficacy of monolaurin both individually and in combination with β-lactam antibiotics against Staphylococcus aureus isolates.MethodsAgar dilution method was used for determination of minimum inhibitory concentration (MIC) of monolaurin against S.aureus isolates. Scanning electron microscope (SEM) was used to detect morphological changes in S.aureus after treatment with monolaurin. Conventional and Real-time Polymerase chain reaction (RT-PCR) were performed to detect of beta-lactamase (blaZ) gene and its expressional levels after monolaurin treatment. Combination therapy of monolaurin and antibiotics was assessed through fractional inhibitory concentration and time-kill method.ResultsThe antibacterial activity of monolaurin was assessed on 115 S.aureus isolates, the MIC of monolaurin were 250 to 2000 µg/ml. SEM showed cell elongation and swelling in the outer membrane of S.aureus in the prescence of 1xMIC of monolaurin. blaZ gene was found in 73.9% of S.aureus isolates. RT-PCR shows a significant decrease in of blaZ gene expression at 250 and 500 µg/ml of monolaurin. Synergistic effects were detected through FIC method and time killing curve. Combination therapy established a significant reduction on the MIC value. The collective findings from the antibiotic combinations with monolaurin indicated synergism rates ranging from 83.3% to 100%.In time-kill studies, combination of monolaurin and β-lactam antibiotics produced a synergistic effect.ConclusionThis study showed that monolaurin may be a natural antibacterial agent against S. aureus, and may be an outstanding modulator of β-lactam drugs. The concurrent application of monolaurin and β-lactam antibiotics, exhibiting synergistic effects against S. aureus in vitro, holds promise as potential candidates for the development of combination therapies that target particularly, patients with bacterial infections that are nearly incurable.

Antiviral Drug Candidate Repositioning for Streptococcus suis Infection in Non-Tumorigenic Cell Models.

The increasing prevalence of antimicrobial resistance against zoonotic bacteria, including Streptococcus (S.) suis, highlights the need for new therapeutical strategies, including the repositioning of drugs. In this study, susceptibilities of bacterial isolates were tested toward ten different 3-amidinophenyalanine (Phe(3-Am)) derivatives via determination of minimum inhibitory concentration (MIC) values. Some of these protease inhibitors, like compounds MI-432, MI-471, and MI-476, showed excellent antibacterial effects against S. suis. Their drug interaction potential was investigated using human liver microsomal cytochrome P450 (CYP450) measurements. In our work, non-tumorigenic IPEC-J2 cells and primary porcine hepatocytes were infected with S. suis, and the putative beneficial impact of these inhibitors was investigated on cell viability (Neutral red assay), on interleukin (IL)-6 levels (ELISA technique), and on redox balance (Amplex red method). The antibacterial inhibitors prevented S. suis-induced cell death (except MI-432) and decreased proinflammatory IL-6 levels. It was also found that MI-432 and MI-476 had antioxidant effects in an intestinal cell model upon S. suis infection. Concentration-dependent suppression of CYP3A4 function was found via application of all three inhibitors. In conclusion, our study suggests that the potential antiviral Phe(3-Am) derivatives with 2',4' dichloro-biphenyl moieties can be considered as effective drug candidates against S. suis infection due to their antibacterial effects.

Antimicrobial-resistant Neisseria gonorrhoeae in Europe in 2020 compared with in 2013 and 2018: a retrospective genomic surveillance study

Regular quality-assured whole-genome sequencing linked to antimicrobial resistance (AMR) and patient metadata is imperative to elucidate the shifting gonorrhoea epidemiology, both nationally and internationally. We aimed to examine the gonococcal population in the European Economic Area (EEA) in 2020, elucidate emerging and disappearing gonococcal lineages associated with AMR and patient metadata, compare with 2013 and 2018 whole-genome sequencing data, and explain changes in gonococcal AMR and gonorrhoea epidemiology. In this retrospective genomic surveillance study, we analysed consecutive gonococcal isolates that were collected in EEA countries through the European Gonococcal Antimicrobial Surveillance Programme (Euro-GASP) in 2020, and made comparisons with Euro-GASP data from 2013 and 2018. All isolates had linked AMR data (based on minimum inhibitory concentration determination) and patient metadata. We performed whole-genome sequencing and molecular typing and AMR determinants were derived from quality-checked whole-genome sequencing data. Links between genomic lineages, AMR, and patient metadata were examined. 1932 gonococcal isolates collected in 2020 in 21 EEA countries were included. The majority (81·2%, 147 of 181 isolates) of azithromycin resistance (present in 9·4%, 181 of 1932) was explained by the continued expansion of the Neisseria gonorrhoeae sequence typing for antimicrobial resistance (NG-STAR) clonal complexes (CCs) 63, 168, and 213 (with mtrD/mtrR promoter mosaic 2) and the novel NG-STAR CC1031 (semi-mosaic mtrD variant 13), associated with men who have sex with men and anorectal or oropharyngeal infections. The declining cefixime resistance (0·5%, nine of 1932) and negligible ceftriaxone resistance (0·1%, one of 1932) was largely because of the progressive disappearance of NG-STAR CC90 (with mosaic penA allele), which was predominant in 2013. No known resistance determinants for novel antimicrobials (zoliflodacin, gepotidacin, and lefamulin) were found. Azithromycin-resistant clones, mainly with mtrD mosaic or semi-mosaic variants, appear to be stabilising at a relatively high level in the EEA. This mostly low-level azithromycin resistance might threaten the recommended ceftriaxone-azithromycin therapy, but the negligible ceftriaxone resistance is encouraging. The decreased genomic population diversity and increased clonality could be explained in part by the COVID-19 pandemic resulting in lower importation of novel strains into Europe. European Centre for Disease Prevention and Control and Örebro University Hospital.

Optimising machine learning prediction of minimum inhibitory concentrations in Klebsiella pneumoniae.

Minimum Inhibitory Concentrations (MICs) are the gold standard for quantitatively measuring antibiotic resistance. However, lab-based MIC determination can be time-consuming and suffers from low reproducibility, and interpretation as sensitive or resistant relies on guidelines which change over time. Genome sequencing and machine learning promise to allow in silico MIC prediction as an alternative approach which overcomes some of these difficulties, albeit the interpretation of MIC is still needed. Nevertheless, precisely how we should handle MIC data when dealing with predictive models remains unclear, since they are measured semi-quantitatively, with varying resolution, and are typically also left- and right-censored within varying ranges. We therefore investigated genome-based prediction of MICs in the pathogen Klebsiella pneumoniae using 4367 genomes with both simulated semi-quantitative traits and real MICs. As we were focused on clinical interpretation, we used interpretable rather than black-box machine learning models, namely, Elastic Net, Random Forests, and linear mixed models. Simulated traits were generated accounting for oligogenic, polygenic, and homoplastic genetic effects with different levels of heritability. Then we assessed how model prediction accuracy was affected when MICs were framed as regression and classification. Our results showed that treating the MICs differently depending on the number of concentration levels of antibiotic available was the most promising learning strategy. Specifically, to optimise both prediction accuracy and inference of the correct causal variants, we recommend considering the MICs as continuous and framing the learning problem as a regression when the number of observed antibiotic concentration levels is large, whereas with a smaller number of concentration levels they should be treated as a categorical variable and the learning problem should be framed as a classification. Our findings also underline how predictive models can be improved when prior biological knowledge is taken into account, due to the varying genetic architecture of each antibiotic resistance trait. Finally, we emphasise that incrementing the population database is pivotal for the future clinical implementation of these models to support routine machine-learning based diagnostics.

Determination of minimum inhibitory concentrations using machine-learning-assisted agar dilution.

This research uses modern artificial intelligence and machine-learning approaches to standardize and automate the interpretation of agar dilution minimum inhibitory concentration testing. Artificial intelligence is currently of significant topical interest to researchers and clinicians. In our manuscript, we demonstrate a use-case in the microbiology laboratory and present validation data for the model's performance against manual interpretation.

IN- VITRO ANTIBACTERIAL ACTIVITY OF SPILANTHES ACMELLA (AKARKARA) EXTRACT ON PORPHYROMONAS GINGIVALIS AND AGGREGATIBACTOR ACTINOMYCETEMCOMITANS

Periodontal diseases are caused by certain bacteria found in the bacterial plaque. Usage of plant-derived antimicrobial agents could serve as an effective alternative treatment against periodontal infections due to continuous rise seen in antibiotic resistance. Spilanthes acmella (S. acmella), a vital medicinal plant has been used for its various properties such as anti- inflammatory, antibacterial, antifungal, antinociceptive, anti-cancerous and hastening wound healing. This study was conducted to assess its antibacterial efficacy against common periodontal pathogens.Objective: The present study was conducted to assess the antibacterial activity of S. acmella plant extract against Porphyromonas gingivalis (Pg), and Aggregatibactor actinomycetemcomitans (Aa) and determine the presence of various phytochemicals in it.Materials and Methodology: An extract was prepared using dried S. acmella plant powder and mixed with methanol in 1:1 (w/v) ratio. Determination of minimal inhibitory concentration (MIC) was done by using tube dilution technique and time- kill assay was performed against Pg and Aa. Presence of phytochemicals was checked by thin layer chromatography (TLC) method.Results: MIC of S. acmella was found to be 40 μg/ml for Pg and 20 μg/ml for Aa within 2 h interval. Various phytochemicals were found in S. acmella extract which may be responsible for its anti- bacterial property.Conclusion: S. acmella extract shows a significant antibacterial effect against the major periodontal pathogens and hence may be a potential natural alternative for controlling the growth of these bacteria.

Antibacterial and Antibiofilm Activity of Ethanol Extract of Batak Onion Bulbs (Allium chinense G.Don.) against Streptococcus mutans and Enterococcus faecalis.

Background: S.mutans is the main pathogen causing caries, while E.faecalis is the dominant microorganism in dental root canals. Batak onion is one of Indonesia's biological resources frequently used as condiments by Batak tribes and possesses antibacterial compounds such as alkaloids, saponins, and flavonoids. Aims: To analyze antibacterial activity test based on Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC) and antibiofilm from 70% ethanol extract of Batak onion bulbs against S.mutans and E.faecalis. Material and Methods: This type of research is a laboratory experiment with a post-test only control group design. Determination of MIC by Kirby Baurer disc diffusion method, determination of MBC by streaking method, and colony tests are calculated using colony counters, determination of antibiofilm by Static Microtiter Plate Assays method, and checking the optical density at around 600nm. Data analysis of MIC, MBC, and antibiofilm using Oneway Anova and Post Hoc LSD tests. Results: MIC on S.mutans and E.faecalis was at a concentration of 0.78% with an average of one of inhibition of 9.00 ± 0.43 mm and 8.06 ± 0.20 mm; no MBC was found because no group was able to reduce bacteria 98-98%, Batak onion bulb extract has antibiofilm ability starting at a concentration of 0.78% with the ability to reduce S.mutans bacteria by 84.55% and 85.73% on E.faecalis. Conclusion: 70% ethanol extract of Batak onion bulbs can inhibit bacterial growth and biofilm of S.mutans and E.faecalis. The recommended dose for antibacterial and antibiofilm is a 6.25-50% concentration.

Aktivitas Antibakteri dan Antioksidan dari Senyawa Bioaktif Asal Bakteri Endofit Tanaman Nyatoh (Palaquium amboinense B.)

Nyatoh plant (Palaquium amboinense B.) is native to Indonesia which is known to have the ability to produce potential secondary metabolites. The aim of this study was to evaluate the antibacterial and antioxidant activities from bioactive compounds produced by endophytic bacteria from nyatoh leaves. Five endophytic bacteria were tested against 4 targeted bacteria including Escherichia coli ATCC 8739, Pseudomonas aeruginosa ATCC 15442, Bacillus subtilis ATCC 19659, and Staphylococcus aureus ATCC 6538. Bacterial isolate which had the strongest antibacterial activity was then fermented and extracted. The corresponding extract was further tested on disc diffusion method, determination of Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) values, antioxidant evaluation, and analyzed for its bioactive compound components via GC-MS analysis. The results showed that one isolate, namely D1 can produce secondary metabolite which act as antibacterial and antioxidant potentials. This corresponding extract has the strongest MIC and MBC against B. subtilis with value of 39.5 and >78.1 µg/ml and exhibited antioxidant activity against 2,2-difenil-1-pikrilhidrazil (DPPH) with inhibition value of 85.37%. Based on GC-MS analysis, secondary metabolites derived from this potential extract contained some major active constituents including Tributyl acetylcitrate and bis (2-ethylhexyl) ester which might act as antibacterial and antioxidant agents.

Chemical composition, antioxidant, antimicrobial, and wound healing effects of Trachyspermum roxburghianum (DC.) H. Wolff essential oil: An in vivo and in silico approach

Ethnopharmacological relevanceTrachyspermum roxburghianum (DC.) H. Wolff, commonly known as ‘Ajamoda,’ is a neglected Indian spice highly used in Ayurveda and folklore remedies as an antimicrobial for chronic wounds and discharges, along with many other disease conditions. Aim of the studyThe objective of the study was to explore chemical composition and to investigate the antioxidant, antimicrobial, analgesic, and wound healing activities of T. roxburghianum fruit essential oil from India. Materials and methodsThe phytochemical characterization of the oil was determined through standard qualitative procedures and the gas chromatography-mass spectrometry (GC-MS) technique. The in vitro antioxidant aptitude was assessed by scavenging DPPH and ABTS radicals. The antimicrobial potential of the oil was investigated using the disc diffusion method, followed by the determination of minimum inhibitory concentration against Gram-positive and Gram-negative bacterial and fungal strains. The analgesic potential was evaluated using thermal and chemically induced pain models in Swiss albino mice. Wound healing was assessed in vivo, including determining wound contraction rates, histopathology, and hydroxyproline estimation, using the excision wound model in Swiss albino mice. ResultsGC-MS analysis identified 55 compounds with major terpenoids, including thymol (13.8%), limonene (11.5%), and others. Substantial radical-scavenging activity was exhibited by T. roxburghianum fruit essential oil (TREO) (IC50 94.41 ± 2.00 μg/mL in DPPH assay and 91.28 ± 1.94 μg/mL in ABTS assay). Microorganisms were inhibited with low MIC (2 μL/mL for the inhibition of Staphylococcus aureus and Bacillus subtilis; 4 μL/mL against Salmonella typhi and 16 μL/mL against Candida albicans). In the cytotoxicity study, no cytotoxicity was observed on the Monkey Normal Kidney Cell line (Vero). Significant antinociceptive effects were observed (25.47 ± 1.10 % of inhibition at 100 mg/kg and 44.31 ± 1.69 % at 200 mg/kg). A remarkable rate of wound closure and epithelization, along with a marked increase in hydroxyproline content, were observed for the oil during wound healing in mice. ConclusionsThe results suggested that oil could be utilized as a potential source of wound healing therapeutics.

Antilisterial Effectiveness of Origanum vulgare var. hirtum and Coridothymus capitatus Essential Oils and Hydrolates Alone and in Combination.

The antimicrobial activity of Origanum vulgare var. hirtum (O) and Coridothymus capitatus (C) essential oils (EOs) and hydrolates (HYs) of the same botanical species was evaluated on sixteen L. monocytogenes strains from food and clinical origins. The antimicrobial activity was assessed by Minimum Inhibitory Concentration (MIC) determination, viable cell enumeration over time up to 60 min, and evaluation of the cellular damage through Confocal Laser Scanning Microscope (CLSM) analysis. EOs exhibited antimicrobial activity with MIC values ranging from 0.3125 to 10 µL/mL. In contrast, HYs demonstrated antimicrobial effectiveness at higher concentrations (125-500 µL/mL). The effect of HYs was rapid after the contact with the cells, and the cell count reduction over 60 min of HY treatment was about 1.2-1.7 Log CFU/mL. L. monocytogenes cells were stressed by HY treatment, and red cell aggregates were revealed through CLSM observation. Moreover, the combinations of EOs and HYs had an additive antilisterial effect in most cases and allowed the concentration of use to be reduced, while maintaining or improving the antimicrobial effectiveness. The combined use of EOs and HYs can offer novel opportunities for applications, thereby enhancing the antimicrobial effectiveness and diminishing the concentration of use. This provides the added benefit of reducing toxicity and mitigating any undesirable sensory effects.

LiaX is a surrogate marker for cell envelope stress and daptomycin non-susceptibility in Enterococcus faecium.

Daptomycin (DAP) is often used as a first-line therapy to treat vancomycin-resistant Enterococcus faecium infections, but emergence of DAP non-susceptibility threatens the effectiveness of this antibiotic. Moreover, current methods to determine DAP minimum inhibitory concentrations (MICs) have poor reproducibility and accuracy. In enterococci, DAP resistance is mediated by the LiaFSR cell membrane stress response system, and deletion of liaR encoding the response regulator results in hypersusceptibility to DAP and antimicrobial peptides. The main genes regulated by LiaR are a cluster of three genes, designated liaXYZ. In Enterococcus faecalis, LiaX is surface-exposed with a C-terminus that functions as a negative regulator of cell membrane remodeling and an N-terminal domain that is released to the extracellular medium where it binds DAP. Thus, in E. faecalis, LiaX functions as a sentinel molecule recognizing DAP and controlling the cell membrane response, but less is known about LiaX in E. faecium. Here, we found that liaX is essential in E. faecium with an activated LiaFSR system. Unlike E. faecalis, E. faecium LiaX is not detected in the extracellular milieu and does not appear to alter phospholipid architecture. We further postulated that LiaX could be used as a surrogate marker for cell envelope activation and non-susceptibility to DAP. For this purpose, we developed and optimized a LiaX enzyme-linked immunosorbent assay (ELISA). We then assessed 86 clinical E. faecium bloodstream isolates for DAP MICs and used whole genome sequencing to assess for substitutions in LiaX. All DAP-resistant clinical strains of E. faecium exhibited elevated LiaX levels. Strikingly, 73% of DAP-susceptible isolates by standard MIC determination also had elevated LiaX ELISAs compared to a well-characterized DAP-susceptible strain. Phylogenetic analyses of predicted amino acid substitutions showed 12 different variants of LiaX without a specific association with DAP MIC or LiaX ELISA values. Our findings also suggest that many E. faecium isolates that test DAP susceptible by standard MIC determination are likely to have an activated cell stress response that may predispose to DAP failure. As LiaX appears to be essential for the cell envelope response to DAP, its detection could prove useful to improve the accuracy of susceptibility testing by anticipating therapeutic failure.

Culturable Streptomyces spp. from high-altitude, oligotrophic North Western Himalaya: a comprehensive study on the diversity, bioactivity and insights into the proteome of potential species.

The increasing global concern of antimicrobial resistance and shortage of new antimicrobials necessitates exploring untapped terrestrial environments for new bioactive microbiome diversity. The low-temperature and oligotrophic North Western Himalaya (NWH) region has a vast diversity of Streptomyces with potential antimicrobial properties that remain largely unexplored. This study evaluates the diversity of culturable Streptomyces from high-altitude NWH and their potential as a source of new antimicrobials through genus-specific isolation and identification. The results demonstrate a distinct phylogenetic clustering of Streptomyces from different sampling regions of NWH, site-specific variation in culturable β-diversity and species commonness with varying intersite bioactivity among different sites. Further, the study optimized the media selection for large-scale culture cultivation in antibiotic production processes and demonstrated the antimicrobial efficacy of Streptomyces against a range of pathogens through in vitro bioassays using minimum inhibitory concentration determination and antibiofilm activity. Untargeted label-free proteomic profiling also revealed variable expression of stress-response proteins and antibiotic regulators as a competitive survival strategy for selective antagonistic Streptomyces. The findings highlight the potential of NWH in augmenting antimicrobial discovery and combating antimicrobial resistance through the isolation and study of novel bioactive Streptomyces.