Lahcen Hassani

Antibacterial activity of Thymus maroccanus and Thymus broussonetii essential oils against nosocomial infection - bacteria and their synergistic potential with antibiotics.

The aim of this study was to evaluate the antibacterial effect of the association between conventional antibiotics and essential oils (EOs) of endemic Moroccan thyme species, Thymus maroccanus and T. broussonetii, on antibiotic-resistant bacteria involved in nosocomial infections. Synergistic interactions between antibiotics (ciprofloxacin, gentamicin, pristinamycin, and cefixime) and EOs, and between T. maroccanus and T. Broussonetii EOs were determined by the checkerboard test. Serial dilutions of two antimicrobial agents were mixed together so that each row (and column) contained a fixed amount of the first agent and increasing amounts of the second one. The results indicate that the oils had a high inhibitory activity against tested bacteria, except for Pseudomonas aeruginosa. In parallel with the increase of cellular killing, the release of 260nm-absorbing materials from bacterial cells, treated with EOs, increased in response to oil concentration. Out of 80 combinations tested between EOs and antibiotics, 71% showed total synergism, 20% had partial synergistic interaction and 9% showed no effect. Combination with carvacrol, the major constituent of T. maroccanus and T. broussonetii, showed also an interesting synergistic effect in combination with ciprofloxacin. The effect on Gram-positive bacteria was more important than on Gram-negative bacteria. These findings are very promising since the use of these combinations for nosocomial infections treatment is likely to reduce the minimum effective dose of the antibiotics, thus minimizing their possible toxic side effects and treatment cost. However, further investigations are needed to assess the potential for therapeutic application.

Anticandidal activity of the essential oils of Thymus maroccanus and Thymus broussonetii and their synergism with amphotericin B and fluconazol

The discovery of antifungal drugs had eradicated some infections that ravaged the humankind. But their indiscriminate use has led to the development of multidrug resistant pathogens. One strategy employed to overcome these resistance mechanisms is the use of combination of the essential oils (EOs) of medicinal plants and conventional drugs. In this study, we investigated a possible synergistic effect of the EOs of two Moroccan endemic thymes (Thymus broussonetii and T. maroccanus) with amphotericin B (Amp B) and fluconazol against Candida albicans. The fractional inhibitory concentration indices (FICI) of T. maroccanus and T. broussonetii EOs combined with Amp B and fluconazol, calculated from the checkerboard titer assay, were 0.49, 0.27, 0.37 and 0.3, respectively. Also, our results indicate that the synergistic effect of EOs with fluconazol was stronger than the combination with Amp B. All these data highlight that the EOs tested potentiate the antifungal action of Amp B and fluconazol, suggesting a possible utilization of these EOs in addition to antifungal drugs for the treatment of some candidiasis due to C. albicans. The use of these combinations is likely to reduce the minimum effective dose of the drugs, thus minimizing their toxic side effects and the treatment cost.

Essential oils from Moroccan plants as potential chemosensitisers restoring antibiotic activity in resistant Gram-negative bacteria

Bacterial drug resistance is a worrying public health problem. Antibiotic efflux is a major non-specific resistance mechanism used by bacteria, and efflux pumps are involved in the low-level susceptibility of various important Gram-negative pathogens. Use of molecules that can block bacterial pumps is an attractive strategy, but several studies report only partial efficacy owing to limits of these molecules (stability, selectivity, bioavailability, toxicity, etc.). The objective of this study was to search for natural sources of molecules able to inhibit efflux pump systems of resistant Gram-negative bacteria (Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, Salmonella enterica serotype Typhimurium and Pseudomonas aeruginosa). The results indicate that the studied essential oils exhibit interesting activity against the tested bacteria. This activity was significantly enhanced in the presence of an efflux pump inhibitor such as phenylalanine arginyl β-naphthylamide (PAβN). The role of lipopolysaccharide (LPS) structure in the effect of essential oils was also reported in Salmonella LPS deep-rough mutants. In addition, essential oils of Thymus maroccanus and Thymus broussonetii, used at a low concentration (a fraction of the minimum inhibitory concentration), are able to significantly increase chloramphenicol susceptibility of several resistant isolates. These results demonstrate that these essential oils can alter efflux pump activity and may be attractive candidates to develop new drugs for chemosensitising multidrug-resistant strains to clinically used antibiotics.

Resistance to and Accumulation of Heavy Metals by Actinobacteria Isolated from Abandoned Mining Areas

Accumulation of high concentrations of heavy metals in environments can cause many human health risks and serious ecological problems. Nowadays, bioremediation using microorganisms is receiving much attention due to their good performance. The aim of this work is to investigate heavy metals resistance and bioaccumulation potential of actinobacteria strains isolated from some abandoned mining areas. Analysis of mining residues revealed that high concentration of zinc “Zn” was recorded in Sidi Bouatman, Arbar, and Bir Nhass mining residues. The highest concentration of lead “Pb” was found in Sidi Bouatman. Copper “Cu,” cadmium “Cd,” and chromium “Cr” were found with moderate and low concentrations. The resistance of 59 isolated actinobacteria to the five heavy metals was also determined. Using molecular identification 16S rRNA, these 27 isolates were found to belong to Streptomyces and Amycolatopsis genera. The results showed different levels of heavy metal resistance; the minimum inhibitory concentration (MIC) recorded was 0.55 for Pb, 0.15 for Cr, and 0.10 mg·mL−1 for both Zn and Cu. Chemical precipitation assay of heavy metals using hydrogen sulfide technic (H2S) revealed that only 27 isolates have a strong ability to accumulate Pb (up to 600 mg of Pb per g of biomass for Streptomyces sp. BN3).

Essential Oil Composition and Antimicrobial Activity of Wild and Cultivated Moroccan Achillea ageratum L.: a Rare and Threatened Medicinal Species

The essential oils of leaves and flowers of the wild and cultivated Moroccan Achillea ageratum L., a rare and threatened medicinal species, were examined by GC/MS, and their chemical compositions were compared. At least nine components were identified in both wild and cultivated A. ageratum oils, representing more than 95% of the oils. Artemisyl acetate (62.34–78.79%), yomogi alcohol (4.89–12.40%), santolina alcohol (4.86–11.77%), and artemisia alcohol (3.36–7.04%) were the major compounds. Terpene‐alcohol proportion was higher in wild A. ageratum than in cultivated A. ageratum. The antibacterial analysis showed that both oils presented high activity against all the studied Gram‐positive strains in a range of MIC values from 2.55 to 7.02 mg/ml, but they appeared not effective against the tested Gram‐negative ones (MIC values 20.40–41.10 mg/ml). They also exhibited remarkable antifungal activities against Candida species with MIC values ranging from 5.83 to 8.42 mg/ml. From these results, it was concluded that domestication of this threatened medicinal species using clonal propagation did not significantly affect its chemical composition and consequently its antimicrobial properties.

Pril‐ampicillin‐dextrin‐ethanol agar for the isolation and quantification of Aeromonas spp. from polluted environmental waters

Several selective media were evaluated for their suitability for the isolation and quantification of mesophilic Aeromonas species from naturally polluted samples. Satisfactory recoveries were obtained with most of them but only when densities of background microflora were low. When analysed samples were from highly polluted waters, results were inconsistent because they did not give quantitative recovery of mesophilic aeromonads or they did not permit ready differentiation of Aeromonas species from the competitive bacteria. A new medium was developed on the basis of the combination of some positive aspects of several published media, pril‐ampicillin‐dextrin‐ethanol (PADE) agar. The medium employs dextrin (Merck 3006) as a fermentable carbohydrate and pril, ampicillin and ethanol as inhibitory substances. Recovery on PADE agar from suspensions of 15 tested strains of Aeromonas prepared from pure cultures was excellent. The confirmation rate of typical colonies designated Aeromonas spp. isolated from polluted samples exceeded 90%. Recoveries of stressed aeromonad strains on both PADE agar and a non‐selective medium (TSA) did not show any significant difference (P 0.05). PADE agar was more reliable for quantitative recovery of mesophilic aeromonads than the other selective media because of its characteristics: (i) inhibition of the swarming of Proteus, (ii) good reduction of the background, (iii) inhibition of the over growth of Klebsiella spp., (iv) absence of NaCl makes it unfavourable for the growth of halophilic vibrios, (v) combination of two pH indicators permitted a very easy differentiation between Aeromonas colonies and the competitive microflora. The medium can also be used for isolation of aeromonads from various sources by membrane filtration.

Thymus maroccanus essential oil, a membranotropic compound active on Gram-negative bacteria and resistant isolates

The effects of Thymus maroccanus essential oil (EO) on the integrity of the cell membranes and the permeability of the outer membrane (OM) and inner membrane (IM) of Escherichia coli, Enterobacter aerogenes and Salmonella enterica Typhimurium were investigated.

Chemical Composition, in vitro Antioxidant, Antimicrobial and Insecticidal Activities of Essential Oil from Cladanthus arabicus

Abstract The essential oil obtained from the aerial parts of Cladanthus arabicus (L.) Cass was studied for its chemical composition, antioxidant, antimicrobial and insecticidal activities. The essential oil (EO) was analyzed by GC-MS. Sixty seven compounds representing 94.2 % of the oil were identified. The major components of C. arabicus EO were sabinene (31.1 %), β-pinene (16.7 %), myrcene (12.3 %) and α-pinene (5.3 %). Moreover, the antioxidant and antimicrobial activities of the essential oil against six bacteria and four Candida strains were studied. The results showed the existence of a strong antioxidant activity with IC50 of 55.4 ± 0.12 μg/mL and 57.2 ± 0.21 μg/mL, respectively for DPPH and FRAP methods. The EO exhibited great antimicrobial activity especially against Micrococcus luteus (MIC = 0.187 mg/mL) and Candida albicans (MIC = 0.187 mg/mL). LD50 and LD90 values were 43.6 and 74.1 ppm against 1d-old larvae of Aedes aegypti, respectively.

Synergistic antimicrobial and antioxidant activity of saponins-rich extracts from Paronychia argentea and Spergularia marginata.

Aims: The crude saponins extracted from the aerial parts of Paronychia argentea and the roots of Spergularia marginata were tested for their antioxidant, antimicrobial and synergistic effects with antibiotics. Methodology: Antioxidant activity was evaluated using the 2,2-diphenyl-1-picryl-hydrazyl (DPPH) free radical, β-carotene-linoleic acid and reducing power assays. However, the antibacterial activity was assessed by the agar disc diffusion method, whereas the MIC determination and the Original Research Article Malika Ait Sidi Brahim et al.; EJMP, 7(4): 193-204, 2015; Article no.EJMP.2015.082 194 synergistic interaction with antibiotics were evaluated using microdilution method. Results: Saponins-rich extract from Paronychia argentea showed a higher antioxidant activity than that from Spergularia marginata. Using DPPH assay, the IC50 values for saponins-rich extracts from P. argentea and S. marginata were 19.08 and 29.65 μg/ml, respectively. For β-carotene-linoleic acid assay, IC50 values were 98.24 and 614 μg/ml respectively for P. argentea and S. marginata. However, for reducing power assays, the IC50 values for saponins-rich extracts from P. argentea and S. marginata were respectively 27.22 and 61.44 μg/ml. The result of MIC assay showed that both saponins-rich extracts was found to be active against the majority of Candida strains and Gram-positive bacteria. However, crude saponins extracted from S. marginata was more active on microorganisms than that from P. argentea. In fact, the in vitro association of saponin extracts and some commercial antibiotics showed a synergistic effect. For bacteria strains, 30 combinations were studied, 17 (56.66%) combinations had total synergism, 7 (23.33%) had partial synergism, 4 (13.33%) had no effect and 2 (6.66%) had antagonism effect. For Candida strains, 8 combinations of saponins extracts and fluconazol are tested. All of these combinations (100%) exhibited a total synergism with FICi ranging from 0,31 to 0,50. Conclusion: The results founded suggested that further work should be performed on the isolation and identification of the antioxidative and antimicrobial components of these saponins-rich extracts.

Research of Secondary Metabolites with Non-polyenic Antifungal Activity Produced by Actinomycetes Isolated from Different Moroccan Habitats

Fungal diseases are currently a serious public health problem; they have become one of the major causes of death, especially for immunocompromised people around the world.