Developing Penicillium digitatum Management Strategies on Post-Harvest Citrus Fruits with Metabolic Components and Colonization of Bacillus subtilis L1-21

Citrus is among the most important plants in the fruit industry severely infected with pathogens. Citrus green mold caused by Penicillium digitatum is one of the most devastating diseases during post-harvest stages of citrus fruit. In this study, a potential endophyte Bacillus subtilis L1-21, isolated from healthy citrus plants, was assessed for its biocontrol activity against the pathogen P. digitatum. Based on an in vitro crosstalk assay, we suggested that B. subtilis L1-21 inhibits the pathogen with an inhibition zone of 3.51 ± 0.08 cm. Biocontrol efficacy was highest for the fermented culture filtrate of B. subtilis L1-21. Additionally, using GC-MS analysis, 13 compounds were detected in the extract of this endophyte. The culture filtrate in Landy medium could enlarge and deform pathogen spores and prevent them from developing into normal mycelium. Accordingly, the Landy culture filtrate of B. subtilis L1-21 was stable in the temperature range of 4–90 °C and pH of 3–11. Further, MALDI-TOF-MS for B. subtilis L1-21 detected surfactin, fengycin, bacillaene and bacilysin as potential antifungal compounds. GFP-tagged B. subtilis L1-21 easily colonized in citrus fruit peel and pulp, suggesting its role in eliminating the fungal pathogen. Altogether, it is highly expected that the production of antifungal compounds, and the colonization potential of B. subtilis L1-21 are required against the post-harvest P. digitatum pathogen on citrus fruit.

Synthesis, Antibacterial, Thrombolytic and Cytotoxic Evaluation of Substituted and Un-substituted Selenium-n-heterocyclic Carbene Adducts

N-heterocyclic carbene salts bearing alkyl substituents (1-8) and their selenium N-heterocyclic carbene adducts (9-12) were synthesized and characterized by elemental analysis, FT-IR, NMR (1HNMR, 13CNMR) spectroscopic techniques. All the adducts were found to be stable in air and moisture at room temperature. Compounds (5-12) were evaluated against Bacillus subtilis Macrococcus brunensis and Bacillus cereus in vitro. The biological assay revealed that antibacterial activity of Selenium-N-heterocyclic carbene adducts are comparatively better than the salts. MIC and inhibition zone values showed that Bacillus subtilis is more active to selenium adducts (9-12) than Macrococcus brunensis and Bacillus cereus whereas opposite in the salts (5-8). In vitro studies of hemolysis and thrombolysis demonstrated that the synthesized compounds are innocuous for pre-clinical trials to mouse blood.

Cymbopogon citratus Essential Oil: Its Application as an Antimicrobial Agent in Food Preservation

Antimicrobial in vitro and in situ efficacies of Cymbopogon citratus essential oil (lemongrass, LGEO) against 17 spoilage microorganisms (bacteria, yeasts and fungi) were evaluated. Additionally, its chemical composition, and antioxidant and antibiofilm activities were investigated. The LGEO exhibited a strong antioxidant activity (84.0 ± 0.1%), and its main constituents were citral (61.5%), geraniol (6.6%) and 1,8-cineole (6.4%). An in vitro antimicrobial evaluation revealed the lowest inhibition zone (1.00 ± 0.00 mm) in Pseudomonas fluorescens, and the highest inhibition zone (18.00 ± 2.46 mm) in Candida krusei. The values for the minimal inhibitory concentration were determined to be the lowest for Salmonella enteritidis and the highest for C. albicans. Furthermore, the concentration of ≥250 µL/L of LGEO suppressed the growth of Penicillium aurantiogriseum, Penicillium expansum, Penicillium chrysogenum and Penicillium italicum. The changes in the molecular structure of the biofilms produced by Pseudomonas fluorescens and Salmonella enteritidis, after their treatment with LGEO, confirmed its action on both biofilm-forming bacteria. Moreover, an in situ antimicrobial activity evaluation displayed the most effective inhibitory effectiveness of LGEO against Micrococcus luteus, Serratia marcescens (250 µL/L) and Penicillium expansum (125, 250 and 500 µL/L) growing on a carrot. Our results suggest that LGEO, as a promising natural antimicrobial agent, can be applied in the innovative packaging of bakery products and different types of vegetables, which combines commonly used packing materials with the addition of LGEO.

Phytochemical compositions and antimicrobial activity of leaf extracts of Calotropis procera against food spoilage microorganisms

Background: The purpose of this study was to determine the phytochemical composition and antimicrobial potential of crude n-hexane, ethyl-acetate, methanol and aqueous extracts of Calotropis procera leaves against food spoilage microorganisms. Methods: Standard protocols were employed for the analysis of qualitative phytochemical compositions of the extracts, and antimicrobial activities against Staphylococcus aureus, Bacillus cereus, Pseudomonas aerugenosa and Aspergillus niger. Results: Phytochemical analysis revealed the presence of tannin, saponin, alkaloids, flavonoids, reducing sugar and phenolics. Terpenoids were absent in ethyl acetate and n-hexane extracts while cardiac glycoside was absent in all extracts. All extracts produced antimicrobial activity at a varying zone of inhibition. The widest inhibition zone was produced by methanol extract (21.35±0.43 mm) on staphylococcus aureus while the lowest inhibition zone (12.05±0.45 mm) was observed in the n-hexane fraction. Similarly, the widest inhibition zone (17.24±0.95 mm) was produced by methanol on A. niger while the lowest inhibition zone (5.45±0.42 mm) was produced on n-hexane on A. niger. However, the ethyl acetate extract showed no visible inhibitory zone on all the tested microorganisms. The minimum inhibitory concentration ranged from 32 mg/ml (S. aureus and B. cereus) for ethanol extracts to 128 mg/ml (B. cereus, P. aerugenosa and A. niger) for n-hexane extract. Conclusion: The result shows that the plant is a good source of bioactive compounds that can be used as a natural alternative to a chemical agent in preserving and controlling food poisoning organisms.

DESIGNING OF NOVEL TOPICAL IN SITU POLYMERIC FILM-FORMING SOLUTION SPRAY FORMULATION OF ANTIFUNGAL AGENT: IN VITRO ACTIVITY AND IN VIVO CHARACTERIZATION

Objective: Voriconazole (VCZ) is a broad-spectrum antifungal medication that works by inhibiting fungal Cytochrome P450, preventing fungi growth. The current study aims at developing and characterizing an antifungal in situ film-forming polymeric solution spray containing VCZ for use in topical drug delivery systems. Methods: Optimized VCZ in situ polymeric film formulation was evaluated for Fourier transform infrared spectroscopy (FTIR), differential scanning calorimeter (DSC), X-ray diffractometry (XRD), Scanning electron microscope (SEM), in vitro and in vivo, ex-vivo investigation using abdominal rat skin and stability studies. The in vivo antifungal activity of the advanced in situ film was examined in albino Wistar rats. Results: The optimized batch contained 22% Eudragit RS 100 (ERS) and 4% Sorbitol. Based on FTIR, XRD, SEM, and rheological studies. Formulation ingredients of VCZ loaded topical in situ polymeric film spray were observed to be compatible and showed no evidence of precipitation, deformation, or discoloration. Diffusion test (in vitro %), and ex-vivo drug diffusion % obtained 99.22%, and 97.45% respectively. The maximum inhibition zone was measured at 13±0.07 mm. The Wistar rat was employed as an animal model for skin irritation and antifungal studies. A study of short-term stability observed no significant modifications in the physical properties. Conclusion: The findings of the optimized VCZ topical in situ polymeric film spray formulation were satisfactory, demonstrating comparable improvement in superficial antifungal treatment.

ANTIBACTERIAL ACTIVITY OF CHLOROGENIC ACID PHYTOVESICLES AGAINST RESISTANT BACTERIA: DEVELOPMENT, OPTIMIZATION AND EVALUATION

Objective: To investigate the in vitro antibacterial activity of a naturally occurring polyphenol chlorogenic acid (CGA) and compares it with formulated chlorogenic acid phytovesicles against 4 different bacterial strains; two gram positive [Staphylococcous aureus and Bacillus subtilis] and two gram negative strains [Klebsiella pneumonia and Escherichia coli]. Methods: CGA phytovesicles were developed and optimized using central composite design to improvise CGA’s physicochemical properties. Bactericidal activity was evaluated using agar diffusion, minimum inhibitory concentration (MIC) and time kill assay. The effect of pH and temperature on the antimicrobial activity was determined. Results: The optimized CGA phytovesicles showed entrapment of 96.89% with 30 times better lipophilic solubility than the plain drug. The inhibition zone sizes for CGA phytovesicle ranged from 17-25 mm as compared to 15-20 mm of plain CGA while the MIC values ranged 200-250 µg/ml as compared to 500-550 µg/ml of plain CGA. CGA phytovesicles exhibited a strong bactericidal effect at MIC with a log reduction in the range of 0.90-2.04 in Colony forming units (CFUs) at 24h for different strains as compared to 1.38-2.17 of plain CGA. Furthermore, the antibacterial effect was found to augment with increasing temperature but decreased with alkaline pH. Conclusion: Results strongly supports the hypothesis of potential use of CGA phytovesicles as a mode of drug delivery for its antibacterial use against different resistant bacteria.

Determination of Biocontrol Potential of Bacillus spp. and Stenotrophomonas sp. against Macrophomina phaseolina in Sunflower

Macrophomina phaseolina is a soil pathogen known as charcoal rot and can cause up to 90% yield loss in sunflower under suitable conditions. The serious damage caused by chemicals used in the control of soil-borne pathogens to the environment and health has become one of the most important concerns in agriculture. Therefore, in our study, it was aimed to determine the in vitro antagonistic effects of various bacterial species against M phaseolina. A total of 38 bacterial strains were isolated from soil samples in the rhizosphere of Malva sylvestris (hibiscus), Vicia sativa (vetch), Cicer arietinum (chickpea), Papaver rhoeas (weasel), Carlina marianum (thistle), Glebionis coronaria (crown daisy) and Vicia faba collected from Urla district of İzmir. All bacterial strains exhibited antibiosis effect under in vitro conditions, but it was determined that 5 bacterial isolates among them showed a high inhibition zone and showed an average inhibition potential ranging between 55% and 74%. The most effective bacteria identified at species and genus level by Maldi biotyping (MALDI-TOF MS) were identified as Bacillus amyloliquefaciens, Stenotrophomonas sp. and Bacillus cereus (3 isolates), and these species showed that they can be important biocontrol agents in biological control against M. phaseolina.

Green Synthesis of Mn + Cu Bimetallic Nanoparticles Using Vinca rosea Extract and Their Antioxidant, Antibacterial, and Catalytic Activities

This article outlines the preparation of manganese-doped copper nanoparticles (Mn + Cu NPs) using Vinca rosea (L.) leaf extract as a convenient and environmentally friendly substance. UV–vis, FT–IR, XRD, SEM–EDAX, and DLS instrumental techniques were employed to describe the physical and chemical properties of synthesized V. rosea extract-mediated Vr-Mn + Cu NPs. The synthesized Vr-Mn + Cu NPs were observed to be monodispersed and spherical, with an average size of 412 nm. The plant extract includes a variety of phytochemical components. The Vr-Mn + Cu NPs also have potential antioxidant and antibacterial properties against selected pathogens. The green synthesized Vr-Mn + Cu NPs showed a maximum inhibition zone of 16.33 ± 0.57 mm against E. coli. For dye degradation, MR, EBT, and MO showed the highest degradation percentage capabilities with Vr-Mn + Cu NP-based adsorbents, which were determined to be 78.54 ± 0.16, 87.67 ± 0.06, and 69.79 ± 0.36. The results clearly show that biosynthesized Vr-Mn + Cu NPs may be employed as an antioxidant, antibacterial, photocatalytic dye degradation, and catalytic agent, as well as being ecologically benign.

Tunisian Native Mentha pulegium L. Extracts: Phytochemical Composition and Biological Activities

Mint species (Lamiaceae family) have been used as traditional remedies for the treatment of several diseases. In this work, we aimed to characterize the biological activities of the total phenolic and flavonoid contents of Mentha pulegium L. extracts collected from two different regions of Tunisia. The highest amounts of total phenols (74.45 ± 0.01 mg GAE/g DW), flavonoids (28.87 ± 0.02 mg RE/g DW), and condensed tannins (4.35 ± 0.02 mg CE/g DW) were found in the Bizerte locality. Methanolic leaf extracts were subjected to HPLC-UV analysis in order to identify and quantify the phenolic composition. This technique allowed us to identify seven phenolic compounds: two phenolic acids and five flavonoid compounds, such as eriocitrin, hesperidin, narirutin, luteolin, and isorhoifolin, which were found in both extracts with significant differences between samples collected from the different regions (p < 0.05). Furthermore, our results showed that the methanolic extract from leaves collected from Bizerte had the highest antioxidant activities (DPPH IC50 value of 16.31 μg/mL and 570.08 μmol Fe2+/g, respectively). Both extracts showed high radical-scavenging activity as well as significant antimicrobial activity against eight tested bacteria. The highest antimicrobial activities were observed against Gram-positive bacteria with inhibition zone diameters and MIC values ranging between 19 and 32 mm and 40 and 160 µg/mL, respectively. Interestingly, at 10 μg/mL, the extract had a significant effect on cell proliferation of U87 human glioblastoma cells. These findings open perspectives for the use of Mentha pulegium L. extract in green pharmacy, alternative/complementary medicine, and natural preventive therapies for the development of effective antioxidant, antibacterial, and/or antitumoral drugs.

Green Synthesis of Silver Nanoparticles from Alhagi graecorum Leaf Extract and Evaluation of Their Cytotoxicity and Antifungal Activity

Green synthesis of silver nanoparticles (AgNPs) using different plant parts has shown a great potential in medicinal and industrial applications. In this study, AgNPs were in vitro green synthesized using A. graecorum, and its antifungal and antitumoractivities were investigated. Scanning electron microscopy (SEM) image result indicated spherical shape of AgNPs with a size range of 22-36 nm indicated by using Image J program. The functional groups indicated by Fourier-transform infrared spectroscopy (FTIR) represented the groups involved in the reduction of silver ion into nanoparticles. Alhagi graecorum AgNPs inhibited MCF-7 breast cancer cell line growth in increased concentration depend manner, significant differences shown at 50, 100, and 150 μg/ml concentrations compared to the control. Strong antifungal activity against Candida species (C. albicans., C. glabrata, C. parapsilosis, C. tropicales, and C. krusei) was observed and the inhibition zone range from 14-22 mm at a concentration of 0.01 mmol/ml and from 17-27 mm at a concentration of 0.02 mmol/ml. Based on our findings, it is concluded that synthesized silver nanoparticles from A. graecorum can be used as a potential antitumor and antifungal agent for various therapeutical applications.