Characterization of Silver Carbonate Nanoparticles Biosynthesized Using Marine Actinobacteria and Exploring of Their Antimicrobial and Antibiofilm Activity.

Bacterial resistance to different antimicrobial agents is growing with alarming speed, especially when bacterial cells are living in biofilm. Hybrid nanoparticles, synthesized through the green method, hold promise as a potential solution to this challenge. In this study, 66 actinomycete strains were isolated from three distinct marine sources: marine sediment, the algae Codium bursa, and the marine sponge Chondrosia reniformis. From the entirety of the isolated strains, one strain, S26, identified as Saccharopolyspora erythrea, was selected based on its taxonomic position and significant antimicrobial activity. Using the biomass of the selected marine Actinobacteria, the green synthesis of eco-friendly silver carbonate nanoparticles (BioAg2CO3NPs) is reported for the first time in this pioneering study. The BioAg2CO3NPs were characterized using different spectroscopic and microscopic analyses; the synthesized BioAg2CO3NPs primarily exhibit a triangular shape, with an approximate size of 100 nm. Biological activity evaluation indicated that the BioAg2CO3NPs exhibited good antimicrobial activity against all tested microorganisms and were able to remove 58% of the biofilm formed by the Klebsiella pneumoniae kp6 strain.

Selected antimicrobial essential oils to eradicate multi-drug resistant bacterial biofilms involved in human nosocomial infections.

Biofilms are the primary source of contamination linked to nosocomial infections by promoting bacterial resistance to antimicrobial agents, including disinfectants. Using essential oils, this study aims to inhibit and eradicate the biofilm of enterobacteria and staphylococci responsible for nosocomial infections at Guelma Hospital, northeastern Algeria. Thymbra capitata, Thymus pallescens and Artemesia herba-alba essential oils were evaluated against clinical strains of Klebsiella pneumoniae, Escherichia coli, and Staphylococcus aureus. The antimicrobial activity of the essential oils under consideration was assessed using an agar disc diffusion assay and the determination of minimum inhibitory concentrations (MICs). In addition, the crystal violet method and scanning electron microscopy (SEM) evaluated biofilm inhibition and eradication by those antimicrobial agents. The results indicate that T. pallescens essential oil was the most effective antimicrobial agent against pathogenic bacteria, with large zones of inhibition (up to 50 mm against S. aureus), low MICs (0.16 to 0.63 mg/mL), and powerful biofilm eradication up to 0.16 mg/mL in both 24 h and 60-min exposure times. Thus, Algerian thyme and oregano could be used in various ways to combat the biofilm that causes nosocomial infection in local hospitals.

Efficacy of endemic Algerian essential oils against single and mixed biofilms of Candida albicans and Candida glabrata.

OBJECTIVES: This study investigated the in vitro effect of Origanum glandulosum, Ammoides verticillata, and Saccocalyx satureioides essential oils against planktonic cells and biofilms formed by single and mixed species of Candida albicans and Candida glabrata oral isolates. The effect of the essential oils in combination with amphotericin B on planktonic cells was also studied. DESIGN: The antifungal susceptibility of planktonic cells was evaluated by disc diffusion and broth microdilution methods. Resazurin assay and scanning electron microscopy (SEM) were employed to determine the antibiofilm activity. The combinatory effect was evaluated by the checkerboard method. Essential oils were characterized by gas chromatography-mass spectrometry (GC-MS). RESULTS: The minimal inhibitory concentrations (MICs) and the minimum fungicidal concentrations (MFCs) of the studied essential oils were ranged between 250 and 2000 µg/mL. Biofilms were inhibited and eradicated by the essential oils at sub-inhibitory concentrations of 500 and 1000 µg/mL, respectively. SEM studies revealed a reduction in the preformed biofilm as a result of Origanum glandulosum essential oil treatment for single and mixed biofilms. Synergistic activity was found when Origanum glandulosum essential oil was combined with amphotericin B against Candida albicans. GC-MS analysis revealed that thymol was the major compound in Origanum glandulosum (38.36 %) and Ammoides verticillata (48.99 %) essential oils, while Saccocalyx satureioides essential oil was dominated by borneol (27.36 %). CONCLUSIONS: The studied essential oils showed significant antifungal and antibiofilm activities, which support their effectiveness as promising candidates for the management of oral Candida infections.

Taxonomic Characterization, Antiviral Activity and Induction of Three New Kenalactams in Nocardiopsis sp. CG3.

Exploration of secondary metabolites secreted by new Actinobacteria taxa isolated from unexplored areas, can increase the possibility to obtain new compounds which can be developed into new drugs for the treatment of serious diseases such as hepatitis C. In this context, one actinobacterial strain, CG3, has been selected based on the results of polyphasic characterization, which indicate that it represents a new putative species within the genus Nocardiopsis. Two fractions (F2 and F3), prepared from the culture of strain CG3 in soybean medium, exhibited a pronounced antiviral activity against the HCV strain Luc-Jc1. LC-HRESIMS analysis showed different bioactive compounds in both active fractions (F2 and F3), including five polyenic macrolactams (kenalactams A-E), three isoflavone metabolites, along with mitomycin C and one p-phenyl derivative. Furthermore, feeding with 1% of methionine, lysine or alanine as a unique nitrogen source, induced the production of three novel kenalactam derivatives.

Occurrence of Extended Spectrum Cephalosporin-, Carbapenem- and Colistin-Resistant Gram-Negative Bacteria in Fresh Vegetables, an Increasing Human Health Concern in Algeria.

The aim of this study was to screen for extended spectrum cephalosporin-, carbapenem- and colistin-resistant Gram-negative bacteria in fresh vegetables in Batna, Algeria. A total of 400 samples of fresh vegetables were collected from different retail stores. Samples were immediately subjected to selective isolation, then the representative colonies were identified using matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI-TOF-MS). Phenotypic and genotypic analyses were carried out in terms of species identification and relative antibiotic resistance. Transferability of the carbapenemase and mcr-bearing plasmids was verified by conjugation. The clonal relationships of carbapenemase and mcr-positive Escherichia coli isolates were studied by multi-locus sequence typing (MLST). Sixty-seven isolates were characterised and were mostly isolated from green leafy vegetables, where the dominant species identified included Citrobacter freundii, Klebsiella pneumoniae, Enterobacter cloacae, Stenotrophomona maltophilia, E. coli and Citrobacter braakii. PCR and sequencing results showed that E. coli was the bacterial species presenting the highest antibiotic resistance level in parallel to blaTEM (n = 16) and blaCTX-M-15 (n = 11), which were the most detected genes. Moreover, five isolates carried carbapenemase genes, including the blaOXA-48 and/or blaVIM-4 genes. The mcr-1 gene was detected in two E. coli isolates. MLST analysis revealed three different E. coli sequence types: ST101 (n = 1), ST216 (n = 1) and ST2298 (n = 1). Conjugation assays confirmed the transferability of the blaOXA-48 and mcr-1 genes. In this study we report, for the first time, the detection of the blaOXA-48 gene in E. coli and C. braakii isolates and the blaVIM-4 gene in vegetables. To the best of our knowledge, this is the first report on the detection of mcr-1 genes from vegetables in Algeria.

Metabolic Profile, Biotransformation, Docking Studies and Molecular Dynamics Simulations of Bioactive Compounds Secreted by CG3 Strain.

Actinobacteria isolated from untapped environments and exposed to extreme conditions such as saltpans are a promising source of novel bioactive compounds. These microorganisms can provide new molecules through either the biosynthetic pathway or the biotransformation of organic molecules. In the present study, we performed a chemical metabolic screening of secondary metabolites secreted by the new strain CG3, which was isolated from a saltpan located in the Sahara of Algeria, via high-performance liquid chromatography coupled with high-resolution mass spectrometry (HPLC-ESI-HRMS). The results indicated that this strain produced five new polyene macrolactams, kenalactams A-E, along with two known compounds, mitomycin C and 6″-hydroxy-4,2',3',4″ tetramethoxy-p-terphenyl. Furthermore, the CG3 isolate could have excellent properties for converting the aglycone isoflavone glycitein to the compounds 6,7-dimethoxy-3-(4-methoxyphenyl)chromen-4-one (50) and 6,7-dimethoxy-3-phenylchromen-4-one (54), and the isoflavone genistein can be converted to 5,7-dimethoxy-3-(4-methoxyphenyl)chromen-4-one (52). Docking studies and molecular dynamics simulations indicated that these three isoflavones, generated via biotransformation, are potent inhibitors of the target protein aromatase (CYP19A1); consequently, they can be used to prevent breast cancer risk in postmenopausal women.

Vegetables and Fruit as a Reservoir of ß-Lactam and Colistin-Resistant Gram-Negative Bacteria: A Review.

Antibacterial resistance is one of the 2019 World Health Organization's top ten threats to public health worldwide. Hence, the emergence of ß-lactam and colistin resistance among Gram-negative bacteria has become a serious concern. The reservoirs for such bacteria are increasing not only in hospital settings but in several other sources, including vegetables and fruit. In recent years, fresh produce gained important attention due to its consumption in healthy diets combined with a low energy density. However, since fresh produce is often consumed raw, it may also be a source of foodborne disease and a reservoir for antibiotic resistant Gram-negative bacteria including those producing extended-spectrum ß-lactamase, cephalosporinase and carbapenemase enzymes, as well as those harboring the plasmid-mediated colistin resistance (mcr) gene. This review aims to provide an overview of the currently available scientific literature on the presence of extended-spectrum ß-lactamases, cephalosporinase, carbapenemase and mcr genes in Gram-negative bacteria in vegetables and fruit with a focus on the possible contamination pathways in fresh produce.

Diversity of Actinobacteria Isolated from Date Palms Rhizosphere and Saline Environments: Isolation, Identification and Biological Activity Evaluation.

The diversity of cultural Actinobacteria in two types of Algerian Sahara environments, including saline environments and date palms rhizosphere, was investigated. In this study, a total of 40 strains of actinomycetes was isolated from different soil samples, using a rehydration and centrifugation method. Molecular identification, based on 16S rRNA gene sequence analysis, revealed that these isolates were affiliated to six clusters corresponding to eight genera, including Streptomyces, Nocardiopsis, Saccharopolyspora, Actinomadura, Actinocorallia, Micromonospora, Couchioplanes, and Planomonospora. A taxonomic analysis, based on the morphological, physiological, biochemical, and molecular investigation, of selected strains, which belong to the rare Actinobacteria, was undertaken. Four strains (CG3, A111, A93, and A79) were found to form distinct phyletic lines and represent new actinobacterial taxa. An assessment of antimicrobial proprieties of the 40 obtained actinomycetes strains, showed moderate to strong antimicrobial activities against fungi and bacteria. This study demonstrated the richness of Algerian Sahara with rare Actinobacteria, which can provide novel bioactive metabolites, to solving some of the most challenging problems of the day, such as multi-drug resistance.

Essential Oil of Algerian Eryngium campestre: Chemical Variability and Evaluation of Biological Activities.

The chemical composition of essential oils extracted from aerial parts of Eryngium campestre collected in 37 localities from Western Algeria was characterized using GC-FID and GC/MS analyses. Altogether, 52 components, which accounted for 70.1 to 86.8% of the total composition oils were identified. The main compounds were Germacrene D (0.4-53.4%), Campestrolide (1.6-35.3%), Germacrene B (0.2-21.5%), Myrcene (0.1-8.4%), α-Cadinol (0.2-7.6%), Spathulenol (0.1-7.6%), Eudesma-4(15)-7-dien-1-ß-ol (0.1-7.6%) and τ-Cadinol (0.3-5.5%). The chemical compositions of essential oils obtained from separate organs and during the complete vegetative cycle of the plant were also studied. With the uncommon 17-membered ring lactone named Campestrolide as the main component, Algerian E. campestre essential oils exhibited a remarkable chemical composition. A study of the chemical variability using statistical analysis allowed the discrimination of two main clusters according to the geographical position of samples. The study contributes to the better understanding of the relationship between the plant and its environment. Moreover, the antimicrobial activity of the essential oil was assessed against twelve strains bacteria and two yeasts involved in foodborne and nosocomial infections using paper disc diffusion and dilution agar assays. The in vitro study demonstrated a strong activity against Gram-positive strains such as S. aureus, B. cereus, and E. faecalis. The cytotoxicity and antiparasitic activities (on Lmm and Tbb) of the collective essential oil and one sample rich in campestrolide, as well as some enriched fractions or fractions containing other terpenic compounds, were also analyzed. Campestrolide seems to be one compound responsible for the cytotoxic and antileishmanial effect, while myrcene or/and trans-ß-farnesene have a more selective antitrypanosomal activity.

Kenalactams A-E, Polyene Macrolactams Isolated from Nocardiopsis CG3.

In our screening program for new biologically active secondary metabolites, a new strain, Nocardiopsis CG3 (DSM 106572), isolated from the saltpan of Kenadsa, was found to produce five new polyene macrolactams, the kenalactams A-E (1-5). Their structures were elucidated by spectral methods (NMR and HRESIMS), and the absolute configuration was derived by chemical derivatization (Mosher's method). Through a feeding experiment, alanine was proven to be the nitrogen-bearing starter unit involved in biosynthesis of the polyketide kenalactam A (1). Kenalactam E (5) was cytotoxic against human prostate cancer PC-3 cells with an IC50 value of 2.1 µM.

Characterization of uropathogenic ESBL-producing Escherichia coli isolated from hospitalized patients in western Algeria.

INTRODUCTION: The aim of this study is to assess the prevalence and molecular characterization of uropathogenic Extended spectrum ß-lactamases (ESBLs) producing Escherichia coli. METHODOLOGY: During 3 years, all hospitalized patients at the University-affiliated hospital of Tlemcen and presenting urinary tract infections caused by E. coli were considered as potential study participants. These E. coli isolates were examined phenotypically for ESBL production. ESBL strains were subjected to antimicrobial susceptibility testing and were investigated for the presence of plasmid mediated quinolone resistance genes, 16SrRNA methylase genes and virulence genes by using conventional PCR and DNA sequencing. The molecular characterization of ESBL strains was established by phylogenetic grouping method and ERIC-PCR. RESULTS: The overall prevalence of ESBL was 32.5%. The blaCTX-M-15 was the most frequently detected in ESBL isolates, followed by blaCTX-M-14, blaCTX-M-28, blaCTX-M-1 and blaSHV-12 respectively. The plasmid-mediated quinolone resistance genes were detected in the 15 ESBL strains with the aac(6')-Ib-cr gene was the most detected followed by qnrB1 and qnrA1 gene respectively. Among the 22 ESBL isolates resistant to gentamicin and amikacin, the 16SrRNA methylase genes were detected in 4 isolates. The sfa and pap virulent genes were founds in 26% and 22% of isolates receptively. The genotyping analysis of all strains revealed that almost were belonged to phylogenetic groups A1 and A0 and fourteen distinct clones. CONCLUSION: The emergence of uropathogenic ESBL isolates and the high rate of blaCTX-M are alarming in Algeria. Strict measure must be required to control the further spread of these strains in Algerian hospitals.

Chemical Composition and Biological Investigations of Eryngium triquetrum Essential Oil from Algeria.

The chemical composition, antibacterial and antioxidant activities of the essential oil obtained from Eryngium triquetrum from Algeria were studied. The chemical composition of sample oils from 25 locations was investigated using GC-FID and GC/MS. Twenty-four components representing always more than 87% were identified in essential oils from total aerial parts of plants, stems, flowers and roots. Falcarinol is highly dominant in the essential oil from the roots (95.5%). The relative abundance of falcarinol in the aerial parts correlates with the phenological stages of the plant. Aerial parts of E. triquetrum produce an essential oil dominated by falcarinol during the early flowering stage, and then there is a decrease in falcarinol and rebalancing of octanal during the flowering stage. To our knowledge, the present study is the first report of the chemical composition of E. triquetrum essential oil. Evaluation of the antibacterial activity by means of the paper disc diffusion method and minimum inhibitory concentration assays, showed a moderate efficiency of E. triquetrum essential oil. Using the DPPH method, the interesting antioxidant activity of E. triquetrum essential oil was established. These activities could be attributed to the dominance of falcarinol. The outcome of our literature search on the occurrence of falcarinol in essential oils suggests that E. triquetrum from Algeria could be considered as a possible source of natural falcarinol.

Chemical Variability and Biological Activities of Essential Oils of Micromeria inodora (Desf.) Benth. from Algeria.

The chemical composition of the essential oils isolated from the aerial parts of Micromeria inodora (Desf.) Benth. collected in 24 Algerian localities was investigated from the first time using GC-FID, GC/MS and 13 C-NMR. Altogether, 83 components which accounted for 94.7% of the total oil composition were identified. The main compounds were trans-sesquisabinene hydrate (1; 20.9%), α-terpinyl acetate (2; 19.8%), globulol (3; 4.9%), caryophyllene oxide (4; 4.3%), ß-bisabolol (5; 2.9%) and trans-7-epi-sesquisabinene hydrate (6; 2.6%). Comparison with the literature highlighted the originality of the Algerian M. inodora oil and indicated that 1 might be used as taxonomical marker. The study of the chemical variability allowed the discrimination of two main clusters confirming that there is a relation between the essential-oil compositions and the soil nature of the harvest locations. Biological activity of M. inodora essential oil was assessed against fourteen species of microorganisms involved in nosocomial infections using paper disc diffusion and dilution agar assays. The in vitro study demonstrated a good activity against Gram-positive strains such as Staphylococcus aureus, Bacillus cereus, Bacillus subtilis, and Enterococcus faecalis, and moderate activity against Candida albicans. These results might be useful for the future commercial valorization of M. inodora essential oil as a promising source of natural products with potential against various nosocomial community and toxinic infections.

Chemical composition and biological activity of the essential oil from Thymus lanceolatus.

Thymus lanceolatus is a rare species, which grows wild in Algeria and Tunis. It is used traditionally as a drink and to flavor and preserve meat and poultry. The composition of the essential oil was determined by GLC/FID and GLC/MS. Forty-nine components were identified and quantified, accounting for 96.75% of the total detected components in the oil. The oxygenated monoterpenes (74.85%) constitute the major class of volatile secondary metabolites in the oil. Thymol was the most abundant constituent (69.61%) followed by γ-terpinene (8.38%). The antioxidant activity was evaluated using both diphenylpicrylhydrazyl (DPPH˙) reduction and 2-deoxyribose (2-DR) degradation prevention methods. The oil showed a very potent antioxidant activity with IC(50) values of 0.20 ± 0.07 and 4.96 ± 0.39 µg/mL for the DPPH˙ and 2-DR methods, respectively. The antimicrobial activity of the oil was assessed using the agar diffusion method, and the in vitro cytotoxicity on five different cancer cells was examined using the MTT assay. The oil revealed promising inhibitory activity against Gram positive bacteria, especially Bacillus subtilis and Streptococcus pyogenes with an MIC value of 62.5 µg/mL. Additionally, the highest cytotoxic activity was observed against the HL-60 cells with an IC(50) of 113.5 µg/mL. These results validate some of their traditional uses in food preservation.

Antimicrobial activity of some essential oils against oral multidrug-resistant Enterococcus faecalis in both planktonic and biofilm state.

OBJECTIVE: To evaluate some essential oils in treatment of intractable oral infections, principally caused by biofilm of multidrug-resistant Enterococcus faecalis (E. faecalis), such as persistent endodontic infections in which their treatment exhibits a real challenge for dentists. METHODS: Ten chemically analyzed essential oils by gas chromatography-mass spectrometry were evaluated for antimicrobial activity against sensitive and resistant clinical strains of E. faecalis in both planktonic and biofilm state using two methods, disk diffusion and broth micro-dilution. RESULTS: Studied essential oils showed a good antimicrobial activity and high ability in E. faecalis biofilm eradication, whether for sensitive or multidrug-resistant strains, especially those of Origanum glandulosum and Thymbra capitata with interesting minimum inhibitory concentration, biofilm inhibitory concentration, and biofilm eradication concentration values which doesn't exceed 0.063%, 0.75%, and 1.5%, respectively. CONCLUSIONS: Findings of this study indicate that essential oils extracted from aromatic plants can be used in treatment of intractable oral infections, especially caused by biofilm of multidrug-resistant E. faecalis.

Chemical compositions of the essential oils of the aerial parts of Chamaemelum mixtum (L.) Alloni.

The chemical compositions of the aerial parts essential oils of Chamaemelum mixtum (L.) Alloni from Corsica and Sardinia were investigated employing gas chromatography and gas chromatography-mass spectrometry (GC-MS). The structure of (Z)-heptadeca-9,16-dien-7-one, a natural compound not previously described, was elucidated by GC-MS (electron impact and chemical ionization) and one-dimensional and two-dimensional nuclear magnetic resonance spectroscopy. The variation in C. mixtum essential oil was studied, and statistical analysis showed the clustering of oil samples into three groups according to the amount of oxygenated compounds; these groups correlated to the harvest area. The strong biological activity of the oxygenated fraction (minimum inhibitory concentration of <0.1 mg/mL) of the Corsican oil against Candida albicans , Citrobacter frendii , Enterococcus faecalis , Escherichia coli , Klebsiella pneumoniae , Listeria monocytogenes , and Staphyllococcus aureus can be attributed to the presence of irregular monoterpene alcohols and (Z)-heptadeca-9,16-dien-7-one.

Bicyclo[4.4.0]decane oxygenated sesquiterpenes from Eryngium maritimum essential oil.

Investigation of the essential oil of the aerial parts of Eryngium maritimum L. from Corsica led to the isolation of one known sesquiterpene (1) and three new oxygenated sesquiterpenes with a muurolane or cadinane skeleton (2-4). Structure assignments of 4 ßH -muurol-9-en-15-al (1), 4 ßH -cadin-9-en-15-al (2), 4 ßH -muurol-9-en-15-ol (3) and 4 ßH -cadin-9-en-15-ol (4) were carried out by GC/MS (EI and CI) and comprehensive 1D and 2D NMR spectroscopy. Antibacterial activity of Eryngium maritimum L. oil and column chromatography fractions have been investigated for the first time. The oxygenated fraction, which contains the four sesquiterpenes, was efficient against Listeria monocytogenes and Echerichia coli.

Chemical composition of fatty acid and unsaponifiable fractions of leaves, stems and roots of Arbutus unedo and in vitro antimicrobial activity of unsaponifiable extracts.

The chemical composition of the fatty acid and unsaponifiable fractions of the leaves, stems and roots of Arbutus unedo L. were determined using gas chromatography and gas chromatography-mass spectrometry. The fatty acid fractions of the leaves, stems and roots contained 38.5%, 31.3% and 14.1% palmitic acid, respectively, along with other long-chain fatty acids (up to C22). The chemical composition of the unsaponifiable fractions differed: the leaf and stem fractions contained high levels of aliphatic (32.1% and 62.6%, respectively) and terpenic compounds (49.6% and 25.7%, respectively), and the root fraction mainly contained esters, of which the most abundant was benzyl cinnamate (36.6%). The antimicrobial activities of the unsaponifiable fractions against nine species of microorganisms were assessed. The unsaponifiable leaf and stem extracts inhibited the growth of Klebsiella pneumoniae, Enterococcus faecalis and Candida albicans.

Inhibition of rat platelet aggregation by Urtica dioica leaves extracts.

Platelet hyperactivity plays an important role in arterial thrombosis and atherosclerosis. The present study was undertaken to investigate the effects of different extracts of Urtica dioica leaves on platelet aggregation. Rat platelets were prepared and incubated in vitro with different concentrations of the tested extracts and aggregation was induced by different agonists including thrombin (0.5 U/mL), ADP (10 microm), epinephrine (100 microm) and collagen (5 mg/mL). The crude aqueous extract inhibited thrombin-induced platelet aggregation in a dose-dependent manner. At 1 mg/mL, the percent inhibition was 17.1 +/- 4.2%. Soxhlet extraction of the plant leaves with different successive solvents showed that the ethyl acetate extract exhibited the most antiaggregant effect with an inhibition of 76.8 +/- 6.1% at 1 mg/mL. Flavonoids isolated from the plant leaves, produced a strong inhibitory effect on thrombin-induced platelet aggregation with an IC(50) of 0.25 +/- 0.05 and 0.40 +/- 0.04 mg/mL for genins and heterosidic flavonoids, respectively. Flavonoids also markedly inhibited platelet aggregation induced by ADP, collagen and epinephrine. It is concluded that Urtica dioica has an antiplatelet action in which flavonoids are mainly implicated. These results support the traditional use of Urtica dioica in the treatment and/or prevention of cardiovascular disease.