Abderrahim Sadak

Differential targeting of dense granule proteins in the parasitophorous vacuole of Toxoplasma gondii

The biosynthesis and fate of 4 different dense granule proteins of Toxoplasma gondii were studied with 3 monoclonal antibodies raised against tachyzoites and 1 polyclonal antibody raised against a recombinant protein. These proteins have the following molecular weights: 27 kDa (GRA 1), 28 kDa (GRA 2), 30 kDa (GRA 3) and 40 kDa (GRA 4). All four proteins were found in dense granules by immunoelectron microscopy; in T. gondii-infected cells, they were found in the vacuolar network but, in addition, GRA 3 was also detected on the parasitophorous vacuole membrane. Therefore, dense granule contents undergo differential targeting when exocytosed in the parasitophorous vacuole. Metabolic labelling and immunoprecipitation showed that GRA 2 and GRA 3 were processed from lower molecular weight precursors, and that GRA 2 and GRA 4 incorporated [3H] glucosamine and are thus likely to be glycosylated.

National Serologic Survey of Haematobium Schistosomiasis in Morocco: Evidence for Elimination

The Moroccan Health Ministry launched a Process of Eliminating Schistosomiasis in 1994. During 2005-2009, the epidemiologic status showed a clear interruption of disease transmission at the national level; only a few residual cases were recorded. Our present study is the first systematic serologic survey to evaluate the transmission status in remaining disease-endemic foci. A study population of 2,382 children born after the date of the last autochthonous cases were selected from provinces with histories of high schistosomiasis transmission (Tata, Chtouka Ait Baha, Errachidia, El Kelaa Des Sraghna, and Beni Mellal). To identify the presence of disease, specific antibodies directed against Schistosoma haematobium adult worm microsomal antigens were detected by using an enzyme-linked immunoelectrotransfer blot assay. The results showed an absence of antibodies in all serum samples. Consequently, our findings confirm either a low transmission status or an interruption of schistosomiasis transmission within the last disease endemic foci.

Analysis of MIF, FCGR2A and FCGR3A gene polymorphisms with susceptibility to pulmonary tuberculosis in Moroccan population

In order to investigate the influence of functional polymorphisms of macrophage migration inhibitory factor (MIF), Fcg receptors CD16A (FCGR3A) and CD32A (FCGR2A) genes on susceptibility to pulmonary tuberculosis (PTB) in the Moroccan population, we analyzed 123 patients with PTB and 154 healthy controls. The genotyping for MIF-173 (G/C) (rs755622), FCGR2A-131H/R (rs1801274) and FCGR3A-158V/F (rs396991) was carried out using TaqMan SNP Genotyping Assay method. We found a statistically significant increase of the MIF -173CC homozygote genotype and MIF -173*C allele frequencies in PTB patients compared with healthy controls (17.07%versus 5.84%, P = 0.003; and 35.37%versus 26.30%, P = 0.02; respectively). In contrast, no association was observed between FCGR2A-131H/R and FCGR3A-158V/F polymorphisms and tuberculosis disease. Our finding suggests that MIF -173*C variant may play an important role in the development of active tuberculosis.

Immunolocalization and characterization of the low molecular weight antigen (4-5 kDa) of Toxoplasma gondii that elicits an early IgM response upon primary infection

A striking feature of toxoplasmic seroconversion is the prominent and early IgM response to a low molecular weight antigen of 4-5 kDa. Two different monoclonal antibodies directed against the 4-5 kDa antigen have been generated and used to characterize this molecule. Using these monoclonal antibodies, we could demonstrate the surface localization of the low M(r) antigen by immunofluorescence and immuno-electron microscopy assays. By immunoblotting, we observed that one of the monoclonal antibodies was unable to recognize the 4-5 kDa antigen in tachyzoites propagated in cell culture, indicating an epitope variability between Toxoplasma gondii tachyzoites grown in vivo and in vitro. We discuss the implications of this latter finding in the design of diagnostic reagents.

DNA barcodes confirm the presence of a single member of the Anopheles maculipennis group in Morocco and Algeria: An. sicaulti is conspecific with An. labranchiae

Anopheles labranchiae Falleroni is the only member of the Maculipennis Group known to occur in northern Africa; however, confusion exists as to the taxonomic status of its junior synonym, An. sicaulti Roubaud (type locality: near Rabat, Morocco). Based on morphological and behavioural distinctions, it has been suggested that Moroccan populations have been isolated from other North African populations by the Atlas Mountains, and that Moroccan populations may represent An. sicaulti, originally described as a variety of An. maculipennis Meigen. DNA barcodes (658bp of the mitochondrial COI gene) obtained from 89 An. maculipennis s.l. collected in Morocco (n=79) and Algeria (n=10) in 2007 and 2008 were used to determine if Moroccan populations are genetically isolated from those east of the Atlas Mountains (Algeria), and whether there is molecular evidence to support the presence of more than one member of the Maculipennis Group in the region. No evidence for speciation was found between Moroccan and Algerian populations, or within populations in northern Morocco. Moreover shared COI haplotypes between Algeria and Morocco indicate ongoing gene flow between populations in these countries, suggesting that the Atlas Mountains are not a boundary to gene flow in An. labranchiae. The synonymy of An. sicaulti with An. labranchiae is confirmed. That An. labranchiae comprises the same species in these North African countries is important for malaria control.

Screening of Antioxidant, Antibacterial and Antileishmanial Activities of Salvia officinalis L. Extracts from Morocco

Aims: The aim of this study was to evaluate in vitro antioxidant and antimicrobial activities of organic extracts from Salvia officinalis L. (Lamiaceae) collected in the province of Ouezzane. Study Design: Evaluation of in vitro antimicrobial and antioxidant activities of extracts and determination phenolic contents. Place and Duration of Study: After plant collection from the Province of Ouezzane, further work was carried out in Parasitology Laboratory of the National Institute of health and Laboratory of Original Research Article Et-Touys et al.; BMRJ, 16(5): 1-10, 2016; Article no.BMRJ.28307 2 Biochemistry-Immunology, Faculty of Science, Mohammed V University of Rabat, Morocco from Novembre 2015 to Mai 2016. Methodology: The antioxidant activity was evaluated using DPPH scavanging assay. The antibacterial activity was tested against three reference strains (Staphylococcus aureus, Escherichia coli and Listeria monocytogenes serovar) by the diffusion method and the minimum inhibitory concentration (MIC) by microtitration assay. The antiparasitic activity was tested against Leishmania major using MTT (3(4.5-dimethylthiazol2yl) -2.5-diphenyltetrazolium bromide) assay. The levels of polyphenols and flavonoids extracts were estimated by colorimetric assay. Results: The methanol extract has shown a significant ability to trap the radical DPPH (IC50=65.655 μg/ml) compared to n-hexane and ethanol extracts. This value is higher than that of ascorbic acid (13.198 μg/ml) and Trolox (22.484 μg/ml) used as standards. The three extracts tests revealed the inhibitory power of three bacterial strains with a significant difference in the diameters of inhibition. The largest area was registered by the hexane and methanol extracts against S. aureus (22±8 mm), while the weakest area was 11±0.22 mm expressed by the ethanol extract against E. coli and the methanol extract against L. monocytogenes. The antileishmanial activity was moderate with a value of cytotoxicity (IC50) above 1 mg / ml. The extracts showed high concentrations of polyphenols and flavonoids, while biological activities were not very high when correlated with these levels. Conclusion: These results will be completed by the determination of the active component and the extracts will be tested on other biological systems namely antifungal and antitumor activities.

Schistosoma haematobium detection in snails by DraI PCR and Sh110/Sm-Sl PCR: further evidence of the interruption of schistosomiasis transmission in Morocco

BackgroundThis is the first study in Morocco to estimate snail infection rates at the last historic transmission sites of schistosomiasis, known to be free from new infection among humans since 2004. Screening of large numbers of snails for infection is one way to confirm that Schistosoma haematobium transmission has stopped and does not resurge.MethodsA total of 2703 Bulinus truncatus snails were collected from 24 snail habitats in five provinces of Morocco: Errachidia, El Kelaa des Sraghna, Tata, Beni Mellal, and Chtouka Ait Baha. All visible snails were collected with a scoop net or by hand. We used waders and gloves as simple precautions. Snails were morphologically identified according to Moroccan Health Ministry guide of schistosomiasis (1982).All snails were analyzed in pools by molecular tool, using primers from the newly identified repeated DNA sequence, termed DraI, in the S. haematobium group. To distinguish S. bovis and S. haematobium, the snails were analyzed by Sh110/Sm-Sl PCR that was specific of S. haematobium.ResultsThe results showed that snails from Errachidia, Chtouka Ait Baha, sector of Agoujgal in Tata and sector of Mbarkiya in El kelaa des Sraghna were negative for DraI PCR; but, snails from remaining snail habitats of El Kelaa des Sraghna, Tata and Beni Mellal were positive. This led to suggest the presence of circulating schistosome species (S. haematobium, S. bovis or others) within these positive snail habitats. Subsequently, confirmation with S. haematobium species specific molecular assay, Sh110/Sm-Sl PCR, showed that none of the collected snails were infected by S. haematobium in all historic endemic areas.ConclusionThe absence of S. haematobium infection in snails supports the argument of S. haematobium transmission interruption in Morocco.

In vitro Antileishmanial Activity of Extracts from Endemic Moroccan Medicinal Plant Salvia verbenaca (L.) Briq. ssp verbenaca Maire (S. clandestina Batt. non L)

Aims: The aim of this study was to evaluate in vitro the antileishmanial activities of organic extracts (methanol, n-hexane and dichloromethane extract) from Salvia clandestina (Lamiaceae) used in Moroccan medicinal plant. Study Design: Evaluation of in vitro antileishmanial activity of extracts and determination phenolic contents. Original Research Article Et-Touys et al.; EJMP, 16(1): 1-8, 2016; Article no.EJMP.27891 2 Place and Duration of Study: After plant collection from the region of Rabat-Morocco, further work was carried out in Parasitology Laboratory of the National Institute of health and Laboratory of Biochemistry-Immunology, Faculty of Science, Mohammed V University of Rabat, Morocco from February 2015 to march 2016. Methodology: The plant was extracted using organic solvents and using Soxhlet. The antileishmanial activity of extracts was tested against three leishmanial strains, Leishmania major, Leishmania tropica and Leishmania infantum in their promatigotes form, using MTT assay. The total phenolic content was assessed by the Folin-Ciocalteau assay and total flavonoid content was assessed by aluminium chloride (AlCl3) colorimetric assay. Results: The MTT based colorimetric assay showed reduced promastigotes viability on the all strains tested. The best growth inhibition was observed with n-hexane and dichloromethane extracts of Salvia clandestina (IC50≤ 155.43 μg/ml) compared to N-methyl glucamine antimoniate (Glucantime®) (IC50>1000μg/ml) used as control, after 72 h of treatment. Phenolic content of S. clandestina extracts ranged between 107.52±3.12 and 74.41±4.96 mg GAE/g extract, and the flavonoid content ranged between 24.64±3.65 and 16.31±3.69 mg QE/g extract. Conclusion: The current investigation reveals that S. clandestina extracts possess activity against three Leishmania species. S. clandestina need further investigation so that the pure bioactive antileishmanial compounds should be isolated with cost effective, promising results and less side effects.

Epidemiological characteristics of visceral leishmaniasis in Morocco (1990–2014): an update

Leishmaniases are parasitic diseases frequent in the Mediterranean Basin. Visceral leishmaniasis (VL) is a notifiable parasitic disease that increased in incidence in Morocco over the past few years and has recently emerged in several new foci, causing a public health problem in Morocco. The aim of this study is to describe the spatio-temporal distribution of VL in Morocco between 1990 and 2014 period in order to highlight important features and trends of VL and its epidemiology and to assess whether the activity of the unit reflects the situation of the disease at the national level and whether it could constitute an indicator of public health relevance. Two thousand four hundred and twenty one cases were reported in Morocco between 1990 and 2014 with an average annual reported incidence rate of 0.4 cases per 100.000 inhabitants. Before 1996 the average annual incidence of VL was 50 cases on average. After this date the number of cases increased and then remained stable with around 100-150 cases per year. Children whose age varies between 1 and 4 years old are the most affected with 1327 (74%) of total cases; nevertheless the adult starts to be affected by the disease. In 2000, 65% of positive cases of VL are concentrated at both northern regions: Taza-Al Hoceima- Taounate with 45% of cases, Tanger- Tetouan mainly represented by Chefchaoun with 20% of cases. The Fez-Boulemane region located in the center recorded 12% of cases. Throughout the years the map VL distribution has been progressively changed and spatial spread of the disease to the center is noted in 2007. 2014 has been marked by an even greater extension of the disease to the center and south of Morocco. Nationally in 2014, 34 of 75 provinces and prefectures are affected compared to 2000, when 22 out of 82 provinces and prefectures were affected. Leishmania infantum was identified the causative agent based on species- specific PCR-Lei70 assay. VL remains a sporadically endemic parasitic disease in Morocco with a progressive extension of its range of distribution. Such a situation would relate to the geographical succession of Phlebotomine sand fly vectors, the difficulty of actions against the canine population reservoirs of L. infantum and unfavorable socio-economic factors.

Evaluation of interleukin-10 levels in the plasma of patients with various stages of tuberculosis

Mycobacterium tuberculosis (Mtb) infection remains one of the world’s major causes of illness and mortality. A clear understanding of the host defense against Mtb is imperatively needed forthe control of this epidemic. When tuberculosis (TB) infection occurs, a variety of pro and anti-inflammatory cytokines play a vital role in the pathogenesis of this disease. Interleukin-10 (IL-10) is one of the most important anti-inflammatory cytokines reported to suppress the protective immune response against tuberculosis. Aim The aim of the present study was to evaluate the association of plasma IL-10 levels with various disease stages of TB and the possible effects of treatment on these levels. Materials and methods A group of 30 patients with active pulmonary TB and a control group of 21 healthy individuals were enrolled in this study. The levels of IL-10 were measured before, during, and after treatment using commercially available enzyme-linked immune-sorbent assay (ELISA). Data were analyzed using GraphPad Prism version 5.0. Results The results showed that the levels of IL-10 had significant differences between the TB and control groups (p<0.05). The patients with abnormal chest X-Ray findings had higher IL-10 levels when compared to patients with normal X-Rays (p=0.03). A subgroup of 18 patients were followed during the treatment and the mean plasma concentration of IL-10 in patients before therapy was higher than in patients at 3 months of therapy and in patients after 6 months of therapy (p=0.01). However, the IL-10 level remained significantly higher in patients at the end of treatment compared with controls. These findings could be used in follow-up as clinical biomarker of the success of tuberculosis therapy.