Broth Microdilution Technique Research Articles

Antimicrobial evaluation of Bovine Lactoferrin: A bioactive protein isolated from milk of crossbred and Indigenous cow breeds

A multifunctional iron-binding glycoprotein called lactoferrin (Lf) acts as a broad-spectrum first line of defense against fungi, protozoa, bacteria, and viruses. Bovine Lactoferrin (BLf), which was extracted from the milk of crossbred and native breeds of cattle, was investigated for its antibacterial properties. The BLf protein was purified from the milk of HF crossbred and indigenous breeds like Sahiwal and Poda thurpu cows using the cation-exchange chromatography method and Broth microdilution and disc diffusion techniques were used to investigate the antibacterial activity. The minimum inhibitory concentration (MIC) of isolated BLf from the milk of HF cross-bred cow was found at 8000 μg/mL against Staphylococcus aureus and BLf isolated from the milk of Sahiwal and Poda thurpu cattle breeds respectively showed MIC of 4000 μg/mL against S. aureus. The MIC against Bacillus spp. was at 4000 μg/mL for all the experimental cow breeds. The MIC value of isolated BLf from Holstein-Friesian (HF) cross bred against Escherichia coli and Proteus spp. was 4000 μg/mL. However, the MIC for isolated BLf) from Sahiwal and Poda thurpu breeds against E. coli and Proteus spp. in the present study was 2000 μg/mL. The antimicrobial activity was also studied by disc diffusion studies against selected gram-positive and gram-negative bacteria. The BLf isolated from the milk of all the three experimental breeds showed a smaller zone of inhibition at 2000 μg/mL concentration and a larger inhibition zone was observed at 4000μg/mL against the selected gram-positive and gram-negative bacteria. However, more potency was shown by the BLf isolated from the indigenous breed viz. Poda thurpu followed by Sahiwal.

Efficacy of Green Synthesised Iron Oxide Nanoparticles against Various Uropathogens: A Cross-sectional Study

Introduction: The shoot-up of antimicrobial resistance leading to the Multidrug Resistance (MDR) phenomenon in clinical pathogens has forced us to develop novel technologies to cease this global threat immediately. Iron oxide nanoparticles can be a breakthrough solution to this dilemma due to its magnetic properties and biocompatibility. Non toxic and biocompatible applications of magnetic nanoparticles can be enriched further by special surface coating with organic or inorganic molecules. Aim: To determine the antibacterial activity of green synthesised iron oxide nanoparticles against various clinical isolates. Materials and Methods: This was a cross-sectional study conducted from June 2021 to April 2022. This study was conducted at the Department of Microbiology, SRM Medical College Hospital and Research Centre (SRMMCH&RC), Kattankulathur, Chengalpattu, Tamil Nadu, India. Nanoparticles underwent surface modifications and characterisation using X-ray Diffraction (XRD), Scanning Electron Microscopy (SEM) and Energy Dispersive X-ray (EDX), Ultraviolet (UV) Visible Absorption Spectra, and Fourier-Transform Infrared Spectroscopy (FT-IR) followed by charge characterisation through Agarose Gel Electrophoresis. Kirby-Bauer Disc Diffusion method was used for screening the sensitivity and resistance pattern of 50 selected isolates and Minimum Inhibitory Concentration (MIC) was assessed using MIC Microbroth Dilution technique with the help of resazurin. Results: Out of the four different concentrations of bare and coated nanoparticles (0.0375 mg/mL, 0.07 mg/mL, 0.15 mg/mL, 0.3 mg/mL), bare nanoparticles inhibited the growth of Methicillin Resistant Staphylococcus aureus (MRSA) at 0.3 mg/mL while citrate-coated nanoparticles inhibited the growth at 0.15 mg/mL, 0.018 mg/mL, 0.0375 mg/mL, 0.07 mg/mL, and 0.15 mg/mL dilutions were used in case of Carbapenem-resistant Klebsiella pneumoniae (CR K. pneumoniae) and MDR Escherichia coli, from which both organisms were inhibited at 0.15 mg/mL of bare and coated nanoparticles. Conclusion: Iron nanoparticles synthesised from the marine algae Chaetomorpha antennina could be used in the future as a drug carrier or as an antimicrobial agent.

Emergence of Plasmid-mediated Quinolone-Resistant Salmonella Typhi Clinical Isolates from Punjab, Pakistan

The present study evaluated the prevalence of fluoroquinolone resistance and the underlying mechanisms among Salmonella Typhi clinical strains from Punjab, Pakistan. A total of 174 Salmonella Typhi strains were isolated from the blood culture samples. The strains identification was done by using the API 20E system and VITEK®, while serovar validation was done by agglutination assays using antisera. Molecular characterization was done by PCR using the primers targeting the fliC-d gene of Salmonellaenterica serovar Typhi. Antimicrobial susceptibility testing was performed by disc diffusion procedure and the minimum inhibitory concentration using the broth microdilution technique. Moreover,plasmid-mediated quinolone resistance genes were amplified through a polymerase chain reaction. Alarming rate of ciprofloxacin resistance (90.8%) were observed with high MICs ranging from <0.06 µg/mL to > 2.0 µg/mL.Overall, the qnrS gene was detected among every ciprofloxacin-resistant isolate, of which maximum frequency of qnrS genes was detected among isolates showing MICs <1.0 µg/mL, while 14 qnrSgene-positive isolates were showing very high MICs values 2.0 µg/mL.Moreover, the ciprofloxacin-resistantisolates, as well as the ten isolates showing intermediate resistance (MIC; 0.5 µg/mL), were negative for qnrA and qnrB genes. This study highlights that the quinolone resistance among S. Typhiis at a critical level that necessitates the need for alternative therapeutic measures and the development of new antibiotics.

Active Quinolinequinones against Methicillin-Resistant Staphylococcus spp.

Serious bacterial infections could be caused by Gram-positive microorganisms, in particular methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis. Aiming to address this challenging issue by developing the potent and selective antimicrobial lead structures against methicillin-resistant Staphylococcus spp., herein, we report in vitro evaluation of quinolinequinones (QQ1-QQ10) against the Gram-negative and Gram-positive strains using the broth microdilution technique. The design principle of the quinolinequinones was based on the variation of the structures attached to the 1,4-quinone moiety and substituent(s) within amino phenyl moiety. A series of ten quinolinequinones displayed activity mainly against the Gram-positive strains with a minimal inhibitory concentration (MIC=1.22-1250 mg/L) within the Clinical and Laboratory Standards Institute (CLSI) levels. Interestingly, QQ3, QQ5, and QQ6 displayed equal antibacterial inhibitory activity against S. aureus (MIC=1.22 mg/L), respectively, to the standard positive control Cefuroxime-Na. QQ2, QQ3, and QQ5 had the best inhibitory activity with the MIC value of 1.22 mg/L (4-fold more potent compared reference standard Cefuroxime) against S. epidermidis. On the other hand, QQ3 was the most effective quinolinequinone against fungi, in particular C. albicans. The identified lead quinolinequinones (QQ3 and QQ5) with a comprehensive analysis of structure-activity relationships and further studies showed high activity against methicillin-resistant Staphylococcus spp. It is worth noting that the isopropyl group has importance for excellent bioactivity. Remarkably, the in vitro antibiofilm and bactericidal activities (each of 32 clinically obtained strains of Gram-positive bacteria) of the selected two quinolinequinones (QQ3 and QQ5) have been evaluated for the mode of action in addition to the time-kill curve study.

Antimicrobial and cytotoxic activity of electrosprayed chitosan nanoparticles against endodontic pathogens and Balb/c 3T3 fibroblast cells

The aims of this study were to synthesize highly positively charged chitosan nanoparticles (Ch-Np) using the electrospraying technique, and to test their antimicrobial activity against endodontic pathogens, and cytotoxicity against fibroblast cells. Ch-Np were synthesized from low molecular weight chitosan (LMW-Ch) using the electrospraying technique, and characterized. The antimicrobial activity was evaluated against Streptococcus mutans, Enterococcus faecalis, and Candida albicans in their planktonic state using a Time-Kill Test performed by using broth micro-dilution technique, and against biofilm biomass using a microtiter plate biofilm assay. The cytotoxicity was evaluated using Balb/c 3T3 fibroblast cells with the standard MTT assay. Electrospraying of LMW-Ch produced Ch-Np with an average size of 200 nm, and a surface charge of 51.7 mV. Ch-Np completely eradicated S. mutans and E. faecalis in the planktonic state and showed fungistatic activity against C. albicans. Furthermore, it significantly reduced the biofilm biomass for all the tested microbial species [S. mutans (p = 0.006), E. faecalis (p < 0.0001), and C. albicans (p = 0.004)]. When tested for cytotoxicity using 3T3 cells, Ch-Np showed no cytotoxicity. In conclusion, the highly positively charged, colloidal dispersion of Ch-Np are effective as a biocompatible endodontic antimicrobial agent.

Virulence Characteristics of Biofilm-Forming Acinetobacter baumannii in Clinical Isolates Using a Galleria mellonella Model.

Acinetobacter baumannii is a Gram-negative coccobacillus responsible for severe hospital-acquired infections, particularly in intensive care units (ICUs). The current study was designed to characterize the virulence traits of biofilm-forming carbapenem-resistant A. baumannii causing pneumonia in ICU patients using a Galleria mellonella model. Two hundred and thirty patients with hospital-acquired or ventilator-associated pneumonia were included in our study. Among the total isolates, A. baumannii was the most frequently isolated etiological agent in ICU patients with pneumonia (54/165, 32.7%). All A. baumannii isolates were subjected to antimicrobial susceptibility testing by the Kirby–Bauer disk diffusion method, while the minimum inhibitory concentrations of imipenem and colistin were estimated using the broth microdilution technique. The biofilm formation activity of the isolates was tested using the microtiter plate technique. Biofilm quantification showed that 61.1% (33/54) of the isolates were strong biofilm producers, while 27.7% (15/54) and 11.1% (6/54) showed moderate or weak biofilm production. By studying the prevalence of carbapenemases-encoding genes among isolates, blaOXA-23-like was positive in 88.9% of the isolates (48/54). The BlaNDM gene was found in 27.7% of the isolates (15/54 isolates). BlaOXA-23-like and blaNDM genes coexisted in 25.9% (14/54 isolates). Bap and blaPER-1 genes, the biofilm-associated genes, coexisted in 5.6% (3/54) of the isolates. For in vivo assessment of A. baumannii pathogenicity, a Galleria mellonella survival assay was used. G. mellonella survival was statistically different between moderate and poor biofilm producers (p < 0.0001). The killing effect of the strong biofilm-producing group was significantly higher than that of the moderate and poor biofilm producers (p < 0.0001 for each comparison). These findings highlight the role of biofilm formation as a powerful virulence factor for carbapenem-resistant A. baumannii that causes pneumonia in the ICU.

Synthesis of New Series Bis-3-Chloro-β-Lactam Derivatives from Symmetrical Bis-Schiff Bases as Effective Antimicrobial Agents with Molecular Docking Studies

Endeavoring to find a new type of antimicrobial agents, a new sequence of bis-Schiff bases and bis-3-cholro-β-lactams are synthesized. An astonishing class of strained compounds is part of the symmetrical bis-Schiff bases that has widespread applications and building blocks for the combination of bis-3-cholro-β-lactams antibiotics. Bis-3-cholro-β-lactams are synthesized through (Staudinger) [2+2] ketene-imine cycloaddition reaction. Structures of the produced compounds are deduced by 1H, 13C-NMR, and FT-IR spectroscopies. All produced compounds are shown moderate to good antimicrobial activity compared to human pathogenic bacteria strains (Escherichia coli and Staphylococcus aureus), also compared to Aspergillus Niger and Trichophyton mentagrophytes fungi through the broth microdilution technique. A molecular docking study is used for showing the active sides and binding affinity of the products with the target proteins or receptors of E. coli (PDB ID: 3GI9).

FORMULATION AND DEVELOPMENT OF CURCUMIN BASED EMULGEL IN TREATMENT AND RECURRENCE OF VAGINAL CANDIDIASIS

Objective: The main causative agent of vaginal candidiasis is Candida albicans and it develops resistance against several synthetic antifungal drugs and it has a high rate of infection in women. According to WHO report, around 75% of women are infected by Candida albicans and 50 % are infected a second time by Candida albicans. Therefore, we choose Curcumin an antifungal agent that had reported antifungal properties against the various fungal species. The Curcumin-containing emulgel based microemulsion system was prepared for greater retention time and penetration across the vaginal mucosa. Methods: The screening of oil phase, surfactant, and cosurfactant for microemulsion formulation was selected based on the solubility study and followed by the construction of the pseudoternary phase diagram. The oil phase, surfactant and co-surfactant are selected from the pseudoternary phase diagram for the formulation of a stable microemulsion. The prepared Curcumin-loaded microemulsion was characterized by globule size, polydispersity index, Zeta potential, accelerated stability study, drug content, percent transmittance and antifungal assay by broth microdilution technique. The formulated microemulsion was converted into a vaginal emulgel by using Pluronic@F127. The formulated curcumin-loaded emulgel was characterized by different evaluation parameters and antifungal study by agar well diffusion method. Results: The result showed that the average globule size of emulgel was 286.3 nm, polydispersity index was 0.241, Zeta potential was+19.20 mv, conductivity was 0.0390 mS/cm, and drug content was found to be 95.58%. The texture of formulated emulgel was found to be soft and smooth, with shear-thinning, pseudoplastic behavior, and easily spreadable. The in vitro permeability study of emulgel shows slow and complete release of curcumin in 10 h. The microemulsion and developed emulgel showed promising antifungal activity against Candida albicans. Conclusion: The developed curcumin-loaded emulgel showed promising antifungal activity against Candida albicans as compared to the Fluconazole as an standard antifungal antibiotic. Our formulated Curcumin-containing emulgel can be a potential alternative as compared to the conventional dosage form for the treatment of vaginal candidiasis.

Antimicrobial Efficacy, Cytotoxicity, Acute Oral Toxicity, and Phytochemical Investigation of the Aqueous and Methanolic Stem Bark Extracts of Bridellia micrantha (Hochst.) Baill

Introduction: Microbial infections cause high morbidity and mortality in humans globally. Antimicrobial resistance, emergence of new strains, high costs of antibiotics, inaccessibility-especially in remote areas, and adverse effects, impede successful eradications of pathogens, hence the need for novel strategies. Bridellia micrantha is used in traditional medicine to treat microbial infections; however, it has not been empirically validated. Methods: Antimicrobial activity of the aqueous and methanolic stem bark extracts of Bridellia micrantha was investigated using the disk diffusion and broth microdilution techniques, described by the Clinical Laboratory Standards Institute (CLSI) guidelines. The brine shrimp lethality assay technique was used to investigate the cytotoxic effects of the studied plant extracts in exposed nauplii. Acute oral toxicity effects of the studied plant extracts in Winstar rats were investigated following the up-and-down procedure described by the Organisation for Economic Development and Co-operation (OECD). Qualitative phytochemical screening was performed following standard procedures. Results: The aqueous and methanolic extract of B. micrantha indicated varied antimicrobial activities against E. coli, S. typhimurium, S. aureus, and C. albicans, with inhibition zones ranging from 6.00mm to 19.00mm. Furthermore, the studied plant extracts exhibited low MIC values (≤100 μg/ml) on selected microbes. Since the MIC values were much lower than 1000μg/ml (the cutoff for antimicrobial efficacy appraisal), it is anticipated that, the studied plant extracts can be strong antibiotics. The aqueous and methanolic stem bark extracts of B. micrantha were cytotoxic to brine shrimp nauplii, with LC50 values of 486.67±3.15 μg/ml and 458.33±2.87 μg/ml, respectively; however, these extracts did not elicit any observable signs of toxicity in rat models. Pharmacologically active phytochemicals, including flavonoids, alkaloids, saponins, tannins, phenols, and anthraquinones were detected in the two studied extracts. Conclusions: The aqueous and methanolic stem bark extracts of B. micrantha have appreciable antimicrobial activity against E. coli, S. typhimurium, S. aureus and C. albicans. Besides, the studied plant extracts are cytotoxic to brine shrimp nauplii; but they do not cause acute oral toxicity effects in rat models. Additionally, the studied plant extracts contain bioactive phytochemicals, with antimicrobial activity.

Determination of Antifungal Activity Against Invasive Candidiasis Agents and Trace Element Content of Fig Tree Latex Samples Obtained From Trabzon Province

Candidiasis is a major health concern causing both morbidity and mortality. The increasing prevalence of antimicrobial-resistant fungi associated with life-threatening systemic mycoses, led a constant need for new antifungal agents. Herbal medicines have been tried for this purpose for centuries. The antifungal effect of fig tree latex has been reported and some trace elements such as zinc were associated with antifungal effects. The aim of this study was to determine the trace element content and in-vitro antifungal activity of fig tree latex sample against C. albicans, C. glabrata, C. tropicalis and C. Krusei. Fig tree latex samples were obtained from four different fig tree at Trabzon province in July 2019. The broth microdilution technique was performed to investigate antifungal activity against standard Candida strains and trace elements level were detected with Inductively Coupled Plasma Atomic Emission Spectroscopy (ICP-OES) analyzer. The most powerful antifungal activity was reached at a concentration of 0.5 for C. albicans and C. tropicalis, and at a concentration of 0.125 for C. krusei and C. glabrata in fig tree latex. According to trace element analysis, magnesium had the highest level, followed by calcium and phosphorus. Selenium, aluminium, lead and nickel levels were too low to be measured. As a conclusion, fig tree latex has an antifungal potential against Candida species and this may be caused by the high level of magnesium that it contains, however more studies are needed to understand the therapeutic effects of fig tree latex.

Chemical composition of essential oils of eight Tunisian Eucalyptus species and their antibacterial activity against strains responsible for otitis

BackgroundThe chemical composition and biological activity of Eucalyptus essential oils have been studied extensively (EOs). A few of them were tested for antibacterial effectiveness against otitis strains. The chemical composition and antibacterial activity of the EOs of eight Tunisian Eucalyptus species were assessed in the present study.MethodsHydrodistillation was used to extract EOs from the dried leaves of eight Eucalyptus species: Eucalyptus accedens, Eucalyptus punctata, Eucalyptus robusta, Eucalyptus bosistoana, Eucalyptus cladocalyx, Eucalyptus lesouefii, Eucalyptus melliodora and Eucalyptus wandoo. They are assessed by GC/MS and GC/FID and evaluated for antibacterial activity using agar diffusion and broth microdilution techniques against three bacterial isolates (Haemophilus influenzae, Haemophilus parainfluenzae, Klebsiella pneumoniae) and three reference bacteria strains (Pseudomonas aeruginosa, ATTC 9027; Staphylococcus aureus, ATCC 6538; and Escherichia coli, ATCC 8739). Furthermore, the selected twenty-one major compounds and all values of the inhibition zone diameters were subjected to further statistical analysis using PCA and HCA.ResultsThe EO yields of the studied Eucalyptus species range from 1.4 ± 0.4% to 5.2 ± 0.3%. Among all the species studied, E. lesouefii had the greatest mean percentage of EOs. The identification of 128 components by GC (RI) and GC/MS allowed for 93.6% – 97.7% of the total oil to be identified. 1,8-cineole was the most abundant component found, followed by α-pinene, p-cymene, and globulol. The chemical components of the eight EOs, extracted from the leaves of Eucalyptus species, were clustered into seven groups using PCA and HCA analyses, with each group forming a chemotype. The PCA and HCA analyses of antibacterial activity, on the other hand, identified five groups.ConclusionThe oils of E. melliodora, E. bosistoana, and E. robusta show promise as antibiotic alternatives in the treatment of otitis media.

Reprint of: Essential oils from 9 exotic and endemic medicinal plants from Mauritius shows in vitro antibacterial and antibiotic potentiating activities

Essential oils (EOs) extracted from botanical resources have greatly been appraised as antimicrobials against a broad spectrum of microorganisms. A panoply of experimental studies have pointed out the immense potency of EOs as natural antimicrobial agents at relatively low doses; largely attributed to the synergistic interaction of various bioactive components. In the present study, 10 EOs prepared from two endemic (Pittosporum senacia Putterl. subsp. Senacia and Syzygium coriaceum J. Bosser & J. Guého) and seven exotic (Cinnamomum camphora (L.) Nees & Eberm, Citrus aurantium L., Curcuma longa L, Morinda citrifolia L., Petroselinum crispum (Mill.) Fuss, Plectranthus amboinicus (Lour.) Sprengel and Syzygium samarangense (Blume) Merr. & L. M. Perry) aromatic medicinal plants from Mauritius, were screened for their growth inhibitory activities against eight bacteria (ATCC strains and clinical isolates) using broth microdilution techniques. The EOs were found to possess varying degree of antibacterial potency. The most active EOs were found to have minimum inhibitory concentration (MIC) of 0.25-4 mg/mL and minimum bactericidal concentration (MBC) of 0.25-16 mg/mL. In particular, at its MIC values, P. amboinicus EO showed bactericidal effects against four strains. However, all tested bacteria were insensitive to P. senecia and C. aurantium fruit peel EOs. Bacillus spizizenii was found to be the most susceptible strain to the active EOs. Additionally, P. amboinicus and the two Syzygium spp. EOs showed antibiotic potentiating activities. Mostly synergistic and partial synergistic actions of the EOs, in combination with the conventional antibiotics (streptomycin, chloramphenicol and ciprofloxacin) at a ratio of 1:1 were obtained against the tested bacteria. Thus, results from this study highlighted the antibacterial potential and the efficacy of the most active EOs as antibiotic potentiating agents that could potentially be used in combinational therapy along with conventional antibiotics for a synergistic approach in the treatment/management of clinically relevant bacterial infections.

Phytochemical Analysis and Investigation of the Antimicrobial and Cytotoxic Activities of Croton dichogamus Pax Crude Root Extracts.

Background Increasing antimicrobial resistance has led to an arduous search for new potent drugs from nature. In this search, plants have proved to be rich reservoirs of efficacious medicinal components that manage ailments. The current study is designed to investigate the phytochemical composition, antimicrobial activity, and the cytotoxicity of the crude root extracts of Croton dichogamus, a shrub that is commonly used in the eastern Africa for the management of infectious diseases. Methods The roots of Croton dichogamus were obtained, dried, ground, and extracted using three solvents (acetone, distilled water, and 50% ethanol). The antimicrobial activity was tested using agar well diffusion and microbroth dilution techniques against five human pathogens. The brine shrimp lethality assay was used to assess the toxic effect. Results The phytochemical screening indicated the presence of terpenoids, flavonoids, tannins, phenols, polyuronides, saponins, and anthracenes. The brine shrimp lethality assay indicated that all the extracts were highly cytotoxic with LC50 values below 100 μg/ml. Acetonic extract had an LC50 value of 4.148 μg/ml, hydroethanolic extract had 76.09 μg/ml, and aqueous extract had 42.61 μg/ml. All extracts showed the antibacterial activity against Gram-positive bacteria (B. cereus and S. aureus) and a fungal organism, C. albicans. The extracts showed no antibacterial effect on the Gram-negative bacterial strains (P. aeruginosa and E. coli) at a concentration of 250 mg/ml. The highest antimicrobial activity was demonstrated by the acetonic extract on B. cereus which had an MIC of 10.42 mg/ml and a zone of inhibition of 17.33 ± 0.58 at a concentration of 250 mg/ml. Conclusion In this research work, we report that C. dichogamus had the antimicrobial activity confirming the folklore claim. The results made a strong case for isolation of novel anticancer lead compounds.

Effect of Zinc Oxide Nanoparticles on Loaded Antibiotics Against Multidrug-Resistant Acinetobacter spp.

Background: Metal oxide nanoparticles (NPs) have shown promising efficacy for combating bacterial resistance due to their antibacterial properties. This research investigated the effect of zinc oxide NPs (ZnO-NPs) on the antibacterial activity of conventional antibiotics including ciprofloxacin (CIP), cefotaxime (CTX), and colistin (CST) against multidrug-resistant Acinetobacter isolates. Methods: The disc diffusion method was performed to detect the pattern of antibiotic resistance in isolates. The synthesized ZnO-NPs via the solvothermal method were characterized by field emission scanning electron microscopy (FESEM), X-ray diffraction (XRD), and energy-dispersive X-ray spectroscopy (EDS). Finally, the broth microdilution technique was conducted to demonstrate the antibacterial activity of CIP, CTX, and CST antibiotics with and without a sub-inhibitory concentration of ZnO-NPs. Results: XRD, EDS, and FESEM results confirmed the crystalline structure of ZnO-NPs, and the average size was 100±58.68 nm. All isolates were discovered to be of multidrug-resistant (MDR) type and fully susceptible to CST. The antibacterial activity of CTX and CIP was restored when combined with a sub-inhibitory level of ZnO-NPs (0.25 mg/L), and the highest activity was obtained at the concentrations of 32 µg/mL CTX and 8 µg/ mL CIP. Eventually, ZnO-NPs showed a synergistic effect on the antibacterial properties of CST against MDR Acinetobacter. Conclusions: This research indicated that the combination of ZnO-NPs with some common antibiotics can be considered as a novel strategy for reducing the spread of antibiotic-resistant bacteria.

The Synergistic Activity and Optimizing Doses of Tigecycline in Combination with Aminoglycosides against Clinical Carbapenem-Resistant Klebsiella pneumoniae Isolates.

Carbapenem-resistant Enterobacteriaceae (CRE), especially carbapenem-resistant Klebsiella pneumoniae (CRKP), are among the largest pathogenic threats to humans. The available antibiotic treatment options for combating CRKP are limited. Colistin-resistant Enterobacteriaceae (CoRE) have also been reported worldwide, including in Thailand. Therefore, this study aimed (1) to determine minimum inhibitory concentrations (MICs) and synergistic activities of antibiotics of CRKP, and (2) to determine the probability target of attainment (PTA) and cumulative fraction of response (CFR) using pharmacokinetic/pharmacodynamic (PK/PD) data. Clinical CRKP isolates were obtained from Phramongkutklao Hospital (June to November 2020). Broth microdilution and checkerboard techniques were used to determine the mono- and synergistic activities of antibiotics. Carbapenemase and mcr-1 genes were also identified by polymerase chain reaction (PCR). The optimal antibiotic regimens were evaluated using Monte Carlo simulations. Forty-nine CRKP isolates were collected, 40 of which were CoRKP strains. The MIC50 and MIC90 of tigecycline, amikacin, and gentamicin were 1 and 2 µg/mL, 4 and 16 µg/mL, and 0.25 and 4 µg/mL, respectively. None of any isolates expressed the mcr-1 gene, whereas blaOXA-48 (53.1%) and blaOXA-48 plus blaNDM (42.9%) were detected. Synergistic activity was observed in 8.2% of isolates for tigecycline combined with amikacin or gentamicin. Additive activity was observed in 75.5% of isolates for tigecycline-amikacin and 69.4% for tigecycline-gentamicin, and no antagonism was observed. High-dose antibiotic regimens achieved the PTA target. The general recommended dose of combination regimens began with 200 mg tigecycline and 25 mg/kg amikacin, or 7 mg/kg gentamicin, followed by 100 mg tigecycline every 12 h and 15 mg/kg amikacin or 5 mg/kg gentamicin every 24 h. In conclusion, tigecycline plus aminoglycosides might be a potential regimen against CRKP and CoRKP. The appropriate combination regimen based on MIC-based dose adjustment can improve optimal antibiotic dosing. Further research via clinical studies will be necessary to confirm these results.

Safety profile, antimicrobial and antibiofilm activities of a nanostructured lipid carrier containing oil and butter from Astrocaryum vulgare: in vitro studies

Ethnopharmacological relevance: Tucumã (Astrocaryum vulgare)is a fruit native to the Amazon region. Extracts from the peel and pulp are thought of as promising treatments for bacterial infections. The primary constituents of Tucumã oil and butter possess unsaturated carbon chains that are susceptible to oxidation by light or heat. The oils have high volatility and low aqueous solubility that limits their use without a vehicle. Nanotechnology refers to techniques to solve these problems. Nanostructured lipid carriers (NLC), for example, protect fixed oils degradation by heat or light, as well as from oxidation and evaporation, ensuring greater stability and function, thereby prolonging the useful life of the final product. Study objectives: The objective of this study was to evaluate the hemolytic, cytotoxic, antimicrobial and antibiofilm properties of an NLC containing Tucumã butter and oil soasto improve the solubility and photosensitivity of the compounds, generating better pharmacological efficacy. Materials and methods: The NLC was assessed for stability for 60 days. The cytotoxicity of nanoparticles in peripheral blood mononucleated cells was determined in culture using assays for cell viability, DNA damage, oxidative metabolism and damage to human erythrocytes. Antimicrobial activity was determined using the broth microdilution technique and antibiofilm activity according to standardized protocols. Results: The Tucumã NLC remained stable throughout the evaluated period, with pH between 5.22–5.35, monodisperse distribution (PDI&lt;0.3) and average particle size of 170.7 ± 3nm. Cytotoxicity studies revealed that the NLC is safe and modulates inflammatory processes, demonstrated by increased cell viability and nitric oxide levels. There was low hemolytic activity of the NLC against human erythrocytes almost concentrations tested. Conclusion: Taken together, the data suggest that NLC containing Tucumã oil and butter showed antimicrobial and antibiofilm activity against organisms that cause morbidity and mortality in humans. They may be alternative solutions to public health problems related to bacterial infections.

Antimicrobial, Cytotoxicity, Acute Oral Toxicity, and Qualitative Phytochemical Screening of the Aqueous and Methanolic Stem-Bark Extracts of Croton megalocarpus Hutch. (Euphorbiaceae)

Microbial infections are feared to cause over 10 million deaths by the year 2050, whereby 50% of the global burden squarely lies in less developed countries of Africa and Asian continents. The current drugs have suffered resistance by previously susceptible strains, are associated with severe side effects, among other therapeutic and economic drawbacks, hence the need for alternatives. Despite the widespread usage of medicinal plants by over 80% of the global population to treat common ailments, including microbial infections, only a few have been empirically validated. Croton megalocarpus is used to treat microbial-associated infections like pneumonia and typhoid among the Agikuyu community of Kenya. However, its healing claims and safety have not been evaluated empirically to date, hence this study. We investigated the antimicrobial, cytotoxicity, acute oral toxicity, and qualitative phytochemical composition of the aqueous and methanolic stem bark extracts of C. megalocarpus. The disk diffusion and broth microdilution techniques described by the Clinical Laboratory Standards Institute (CLSI) were adopted for antimicrobial assays. The acute oral toxicity effects of the studied plant extracts were evaluated according to the Organisation of Economic Co-operation and Development (OECD) guideline document number 425. The brine shrimp lethality assay technique was used to appraise the cytotoxic effects of the studied plant extracts. Qualitative phytochemical screening was performed following standard procedures. The results revealed that all the studied plant extracts had varied antimicrobial effects on selected microbial strains and showed MIC values of &lt;1000 µg/ml indicating their antimicrobial potential. Moreover, the studied plant extracts had LC50 values of &gt;100 µg/ml and &gt;2000 mg/Kg bw in the brine shrimp lethality and acute oral toxicity assays, respectively, demonstrating their safety. Antimicrobial- associated phytocompounds were detected in the studied plant extracts suggesting they were responsible for the reported bioactivity. Further studies to establish the specific mode(s) of antimicrobial action, toxicological, and safety should be performed. Furthermore, antimicrobial investigations of the studied plant extracts on other clinically significant microbial strains and the isolation, characterization, and optimization of antimicrobials from the studied plant extracts should be done.

Comparative Evaluation of the Antibacterial Effects of Nigella sativa and Moringa oleifera on some Clinical Bacteria Isolates

Aim: In this study, the antibacterial effect of the n-hexane, methanol, and ethanol extracts of two traditional medicinal plants, Nigella sativa and Moringa oleifera was carried out to comparatively assess the antibacterial efficacy of these plants against some selected multi-drug resistant diarrheagenic bacterial isolates such as Escherichia coli, Salmonella typhi, Shigella spp, Enterobacter aerogenes, Klebsiella pneumonia, and Pseudomonas aeruginosa.&#x0D; Methodology: The inhibitory effects of extracts were assessed using disk diffusion method at six different concentrations i.e. 12.5 mg/ml, 25 mg/ml, 50 mg/ml, 100 mg/ml, 200 mg/ml, and 400 mg/ml. The minimum inhibitory concentrations and minimum bactericidal concentrations of the plant seed extracts were determined by the micro-broth dilution technique. Synergism and antagonism studies were similarly done to determine the combined effects of both N. sativa and M. oleifera seed oil. Further, resistant bacteria were subjected to sodium dodecyl sulphate treatment for plasmid curing and continued with the antibiotic susceptibility test.&#x0D; Results: The oil extracts of N. sativa were found to be more effective than that of M. oleifera (at MIC and MCC of 50 mg/ml and 100 mg/ml respectively). It was observed that the plasmids were cured in all the E. coli isolates. Among the pathogens, E. coli was highly susceptible to both seed oils.&#x0D; Conclusion: Higher antibacterial activity was observed when the extracts were used in combination, showing a strong synergistic effect against all the pathogens tested.

Phytochemical Analysis and Anti-Tuberculosis Activity of Extracts of Detarium senegalense Bark and Root

Tuberculosis [TB] remains one of the major global health threats leading to morbidity and mortality. Detarium senegalense J. F. Gmelin belongs to a group of medicinal plants that are used by traditional medicine healers for the treatment of venereal diseases, urogenital infections, hemorrhoids, rheumatism, stomach-ache, intestinal worms, diarrhea, bronchitis, tuberculosis, convulsion, malaria and leprosy. The present study was aimed to investigate the in vitro anti-tuberculosis effect of different solvent extracts of Detarium senegalense bark and root against attenuated strain of Mycobacterium bovis [BCG]. The crude extracts [n-hexane, ethyl-acetate, methanol] of the plant parts were successively extracted and tested in vitro by broth microdilution technique against M. bovis. The result shows that the crude extracts inhibited the growth of M. bovis at concentrations ranging from 7.8 – 250 μg/mL. The ethyl-acetate extract of Detarium Senegalense bark was the most effective in inhibiting the growth of M. bovis with a minimum inhibitory concentration [MIC] of 7.8 µg/mL while the ethyl-acetate and methanol extract of the plant’s root had MICs of 250 µg/mL. This study demonstrates the efficacy of extracts of Detarium senegalense bark and root as potential agents in the management of the tuberculosis disease.

Volatile and phenolic profiling of a traditional medicinal plant, Hypericum empetrifolium with in vitro biological activities

Ethnopharmacological relevanceHypericum empetrifolium Willd is a member of the Hypericaceae family, mainly known in southern Greece, and western Turkey. Being a centuries-old medicinal plant, the aerial parts of the plant have been used for the treatment of herpes, kidney stones, gastric ulcer, and also for their anti-helminthic, and diuretic purposes traditionally. Aim of the studyThe current study aimed to investigate the phytochemical profiles of the essential oil, and two ethanol extracts prepared from the aerial parts (H. empetrifolium aerial parts extract – HEA), and roots of the plant (H. empetrifolium roots extract – HER), and to provide data on antioxidant, anticholinesterase, antityrosinase, antiurease, cytotoxic, and antimicrobial activities of the extracts. Materials and methodsIn this study, volatile and phenolic compounds of the HEA and HER were analyzed by GC-MS and LC-MS/MS, respectively. Antioxidant potential of the extracts was clarified by using DPPH radical scavenging assay, ABTS cation radical assay, and the CUPRAC assay. Acetylcholinesterase, butyrylcholinesterase, urease, and tyrosinase inhibitory activity assays were used to determine enzyme inhibition capacity of the extracts. Cytotoxic activity of the extracts was established by using XTT assay. The antimicrobial activity of the extracts was determined by the microbroth dilution technique. ResultsThe major compounds of the essential oil were revealed as alloaromodendrene (24.7%), α-pinene (14.7%), β-pinene (10.7%), and α-terpineol (7.7%) by the GC-MS analysis. According to the LC-MS/MS analysis results, quinic acid was the most abundant constituent in both extracts with 20612.42 ± 169.02 μg/g extract in HEA extract, and with 2254.34 ± 18.49 μg/g extract in HER extract, respectively. The HEA extract was also found to be rich in terms of chlorogenic acid (5583.14 ± 38.52 μg/g extract), isoquercitrin (3076.77 ± 40.92 μg/g extract), and malic acid (2822.90 ± 31.90 μg/g extract). HEA extract exhibited a significant antioxidant effect with an IC50 value of 11.98 ± 0.22 μg/mL according to the DPPH radical scavenging assay. Similar results were obtained with the ABTS cation radical assay, and the CUPRAC assay. HER extract showed a strong butyrylcholinesterase inhibitory effect with 88.69 ± 0.62% at 200 μg/mL concentration. Both extracts were considered to have moderate anti-tyrosinase activity compared to the standard at 200 μg/mL. The best antimicrobial activity was obtained for HEA against C. parapsilosis with an MIC value of 4.88 μg/mL. HEA also exhibited antifungal activity against C. tropicalis with 19.53 μg/mL. Only HER exhibited low cytotoxicity on A498 renal cell lines with 60.6% cell viability. ConclusionUnquestionably, H. empetrifolium Willd has ethnopharmacological importance according to these results, and further investigations are required to evaluate other therapeutic properties of the plant.