Publications by authors named "Meryem Lemrani"

30 Publications

  • Page 1 of 1

The Modulation of NADPH Oxidase Activity in Human Neutrophils by Moroccan Strains of and Is Not Associated with p47 Phosphorylation.

Microorganisms 2021 May 10;9(5). Epub 2021 May 10.

Molecular Genetics and Immunophysiopathology Research Team, Health and Environment Laboratory, Aïn Chock Faculty of Sciences, UH2C, 20000 Casablanca, Morocco.

Polymorphonuclear neutrophils (PMNs) are the first phagocyte recruited and infected by . They synthetize superoxide anions (O) under the control of the NADPH oxidase complex. In Morocco, and are the main species responsible for cutaneous leishmaniasis (CL). The impact of these parasites on human PMN functions is still unclear. We evaluated the in vitro capacity of primary Moroccan strains of and to modulate PMN O production and p47 phosphorylation status of the NADPH oxidase complex. PMNs were isolated from healthy blood donors, and their infection rate was measured by microscopy. O production was measured by superoxide dismutase-inhibitable reduction of cytochrome C. P47 phosphorylation was analyzed by Western blot using specific antibodies against Ser328 and Ser345 sites. Whereas we did not observe any difference in PMN infectivity rate, our results indicated that only promastigotes inhibited both fMLF- and PMA-mediated O production independently of p47 phosphorylation. soluble antigens (SLAs) from both species significantly inhibited O induced by fMLF or PMA. However, they only decreased PMA-induced p47phox phosphorylation. and modulated differently O production by human PMNs independently of p47 phosphorylation. The inhibition of ROS production by could be a mechanism of its survival within PMNs that might explain the reported chronic pathogenicity of CL.
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http://dx.doi.org/10.3390/microorganisms9051025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8151549PMC
May 2021

Multilocus sequence analysis provides new insight into population structure and genetic diversity of Leishmania tropica in Morocco.

Infect Genet Evol 2021 May 21;93:104932. Epub 2021 May 21.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco. Electronic address:

Cutaneous leishmaniasis (CL) is one of the most neglected tropical diseases, caused by different Leishmania species. Despite its high incidence in Morocco, CL due to Leishmania tropica is poorly understood in terms of its epidemiological status and population structure. In this study, we used multilocus sequence typing (MLST) in order to explore the genetic heterogeneity of L. tropica strains. Samples (N = 48) were collected from CL patients in two localities in Morocco (Foum Jamaa in the Azilal province and Imintanoute in Chichaoua province). PCR-sequencing of 18 strains was carried out for six housekeeping genes (cytb, me, fh, g6pd, pgd and gpi), Genetic diversity indices showed a high population genetic differentiation between and among populations. There was no shared haplotypes between the two localities studied. Our results reveal a considerable degree of differentiation through the relatively high FST value (> 0.4) and remarkable intraspecific polymorphism (S = 29). Imintanoute strains have more polymorphisms (S = 22) than the Foum Jamaa strains despite their small sample size. These results provide crucial background information of epidemiology in Imintanoute which raises questions about animal involvement in L. tropica transmission cycle.
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http://dx.doi.org/10.1016/j.meegid.2021.104932DOI Listing
May 2021

Entomological study in an anthroponotic cutaneous leishmaniasis focus in Morocco: Fauna survey, Leishmania infection screening, molecular characterization and MALDI-TOF MS protein profiling of relevant Phlebotomus species.

Transbound Emerg Dis 2021 Mar 8. Epub 2021 Mar 8.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco.

In Morocco, leishmaniases are a major public health problem due to their genetic diversity and geographical distribution. Cutaneous leishmaniasis (CL) is an infectious disease caused by various species of Leishmania and transmitted typically by bite of phlebotomine sand flies. This study identifies sand fly fauna in Ibaraghen village, province of Azilal, which is a focus of CL, by combination of morphological and molecular methods (sequencing of COI gene, MALDI-TOF MS protein profiling). Nested-kDNA PCR was used to detect and identify Leishmania species within potential vector species. 432 CDC light traps were placed at different heights above ground level at four capture sites during a whole year. Traps at 1.5 m above the ground yielded capture of sand flies almost double compared to above ground level (29.33%), while the collection reached 55.09% when the traps were placed 2.5 m above ground. A total of 2,830 sand flies were collected, 2,213 unfed specimens were morphologically identified, 990 males (44.73%) and 1,223 females (55.26%) of 13 species; ten Phlebotomus species and three Sergentomyia species. Six species were analysed by MALDI-TOF MS protein profiling (4 Phlebotomus and 2 Sergentomiya species), and their identification was confirmed by COI sequencing. 1,375 unfed females were screened for the presence of Leishmania by nested-kDNA PCR in pools, 11/30 pools of P. sergenti showing a single band of 750 bp corresponding to L. tropica. Our results confirm the role of P. sergenti as a proven vector in Azilal focus of cutaneous leishmaniasis; however, the relative abundance of other species known as vectors of Leishmania species emphasizes the risk of introduction of L. infantum and L. major in this province. For the first time in Morocco, a combined approach to identify sand flies by both morphology and molecular methods based on DNA barcoding and MALDI-TOF MS protein profiling was applied.
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http://dx.doi.org/10.1111/tbed.14064DOI Listing
March 2021

Meta-analysis of -308G > A polymorphism in TNFα gene and susceptibility to leishmaniasis.

Cytokine 2021 Apr 24;140:155437. Epub 2021 Jan 24.

Laboratory of Parasitology and Vector-Borne-diseases, Institut Pasteur du Maroc, Casablanca, Morocco.

The clinical spectrum of leishmaniasis depends on several factors, including Leishmania species and immunogenetic factors. Tumor necrosis factor α (TNFα) plays a central role in immunity against intracellular infections. Many studies have reported that TNFα-308G > A polymorphism is associated with susceptibility to intracellular infections and influences TNFα production. Some studies on the implications of TNFα-308G > A polymorphism in the susceptibility to cutaneous leishmaniasis and visceral leishmaniasis showed controversial results. To draw an overall conclusion using accurate data analysis by increasing the number of cases studied, a meta-analysis was performed based on data from the studies included in the analysis. A total of 1264 patients and 2350 controls were enrolled in the meta-analysis. The results showed no significant association between allele G and allele A of -308G > A polymorphism and leishmaniasis by taking the two subgroups separately [OR = 0.99 (0.84-1.18) and OR = 1.19 (0.88-1.59)] or together [OR = 1.04 (0.90-1.20)]. This meta-analysis insinuates the absence of statistical evidence for an association between allele G and allele A of TNFα-308G > A polymorphism and Leishmania infection outcome. This suggests that TNFα, despite its crucial role in the immune response against Leishmania infection, is not the sole determinant factor. Other factors, such as gene-gene and gene-environment interactions, receptors, and signaling pathway efficiency, may influence TNFα function.
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http://dx.doi.org/10.1016/j.cyto.2021.155437DOI Listing
April 2021

The role of sampling by cotton swab in the molecular diagnosis of cutaneous leishmaniasis.

Transbound Emerg Dis 2021 Jul 6;68(4):2287-2294. Epub 2020 Nov 6.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco.

An appreciated sampling technique is essential for achieving optimum results from diagnostic tests of cutaneous leishmaniasis (CL); conventional sampling methods, such as skin biopsy and dermal scrapping, are painful for the patients and require qualified staff and hospital facilities, while swabbing is patient-friendly more comfortable than invasive traditional techniques and can be carried out under field conditions. The aim of this study is to evaluate a non-invasive sampling method (swab) in the cutaneous leishmaniasis diagnosis, compared to microscopic examination. The recruitment of 205 patients was done during 3 years in six regions known as endemic CL foci in the south-east, the centre, the south-west and the north of Morocco. The results showed that molecular detection of the nuclear marker ITS1 on swab materials is to be less sensitive than microscopic examination, and the difference was statistically significant (p-value = .036). Thereafter, 55 patients were randomly selected to compare the results of two molecular techniques (ITS1-PCR and nested KDNA-PCR), performed both on swab and on stained smears used for microscopic examination; ITS1-PCR results from stained smears reached 87% positivity against 65,2% for cotton swab; and it was statistically significant (p-value = .019); on the other hand for the KDNA marker, results from cotton swab reached 93% of positivity against 91% for stained smears; and statistically the difference was not significant (p-value = .5113). One can presume that rubbing over the lesion with cotton swab implies a low parasitic load collection, but the choice of the amplification target greatly influences the results obtained from cotton swab; on top of that swab remains useful in the cases of patients with multiples lesions or the latter are located in sensitive places difficult to reach with an invasive method such as the eyelids, the lips or other mucous areas.
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http://dx.doi.org/10.1111/tbed.13886DOI Listing
July 2021

The different faces of the NLRP3 inflammasome in cutaneous Leishmaniasis: A review.

Cytokine 2020 Aug 15:155248. Epub 2020 Aug 15.

Molecular Genetics and Immunophysiopathology Research Team, Health and Environment Laboratory, Aïn Chock Faculty of Sciences, Hassan II University of Casablanca (UH2C), Morocco. Electronic address:

Cutaneous leishmaniasis (CL) is a vector-borne parasitic disease caused by Protozoa of the genus Leishmania. Clinical manifestations of this disease are the result of a complex interplay of diverse factors, including the genetic background and the immune status of the host. Understanding the impact of these factors on the CL pathology may provide new targets to manage the infection and improve clinical outcome. The NLRP3 inflammasome, an innate immune complex of several cell types, seems to be involved in the CL physiopathology. Current studies of its role show contradictory effects of this complex on the evolution of Leishmania infection in mice and humans. In this review, we discuss the data regarding different roles of the NLRP3 inflammasome in murine and human CL.
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http://dx.doi.org/10.1016/j.cyto.2020.155248DOI Listing
August 2020

Population structure of leishmania major in southeastern morocco.

Acta Trop 2020 Oct 11;210:105587. Epub 2020 Jun 11.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco. Electronic address:

Cutaneous leishmaniasis (CL) is an infectious disease caused by various Leishmania species. It is among the most neglected tropical diseases and has been considered a major health threat over the past decades in the country. Its zoonotic form caused by Leishmania (L) major is the most prevalent in Morocco. This study investigated the population structure of L. major in southeastern Morocco. Samples (n = 67) were collected from patients with CL in five different endemic areas located in three provinces (Ouarzazate, Tinghir, and Zagora). These samples were then sequenced using two nuclear markers: internal transcribed spacer 1 (ITS1) and a fragment of the virulence factor GP63. Next, the sequences were edited and analyzed. Molecular diversity indices showed a high population genetic diversity but an overall low haplotype diversity. Our results suggest small population differentiation, indicating a low geographic structure. Tajima's D and Fu's Fs tests both suggested recent population expansion based on the significant deviations from neutrality in both tests for all populations except Tinghir, which may be due to a small sample size. Based on our findings, the region is experiencing rapid population expansion caused by recent CL outbreaks, and one of them has been recently studied. In addition, analysis of molecular variance and F suggested gene flow between Zagora and both Ouarzazate and Tinghir. Nonetheless, no gene flow was observed between Tinghir and Ouarzazate. To the best of our knowledge, this is the first analysis of the population structure of L. major in Morocco. The results of this study provide crucial background information for epidemiological studies by showing the presence of gene flow between populations and clonal expansion in cases of an outbreak. This will drive authorities to reconsider the implemented control strategies.
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http://dx.doi.org/10.1016/j.actatropica.2020.105587DOI Listing
October 2020

Possible introduction of to urban areas determined by epidemiological and clinical profiles of patients with cutaneous leishmaniasis in Casablanca (Morocco).

Parasite Epidemiol Control 2020 May 3;9:e00129. Epub 2020 Jan 3.

Immunopathology of Infectious and Systemic Diseases (Lab. Cellular and Molecular Pathology), Faculty of Medicine and Pharmacy, University Hassan II of Casablanca, Morocco.

Leishmaniases are a group of infectious diseases caused by protozoan parasites and are transmitted by the bites of infected phlebotomine sandflies. The heterogeneity of these diseases is influenced by both parasitic properties and host factors. Cutaneous leishmaniasis (CL) is a major public health problem in Morocco, where the geographical expansion of CL (particularly CL caused by ), the heterogeneous appearance of lesions and the difficulty in diagnosing CL contribute to late diagnosis of CL and delayed treatment of patients. Therefore, the main objective of this study was to describe the epidemiological and clinical profiles of patients with CL diagnosed in Casablanca (Morocco), which is a non-endemic area for CL. A cross-sectional study was conducted between 2010 and 2016, during which epidemiological and clinical data were collected from patients that met the inclusion criteria through an information sheet. Then, samples were obtained from each patient for parasitological and molecular diagnosis, and only patients with positive polymerase chain reaction and genotyping results were included in the study. Overall, 106 cases of CL were genotyped, of which 61 (57.5%) were caused by L. , 38 (35.9%) by L. and 7 (6.6%) by L. . While all age groups were affected, CL cases wherein L. was the causative agent were most frequently diagnosed in children aged 0-9 years ( = 0.005), whereas those caused by L. were more frequently diagnosed in elderly patients ( = 0.004). Multivariate logistic regression analysis showed that two clinical variables were significantly associated with CL caused by L. : lesion size ( = 0.002) and occurrence of lesion on the face ( = 0.005). Furthermore, the results of our survey highlighted the association of infection when travelling to endemic areas. The high number of endemic foci where patients with CL were infected with L. illustrated the tendency of this form to spread and generate epidemics, exposing young people to a greater degree to the disease. The epidemic status of CL caused by L. in Morocco and the increased movement of the population from rural to urban areas indicate a possible introduction of this species to urban areas.
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http://dx.doi.org/10.1016/j.parepi.2019.e00129DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7171526PMC
May 2020

Epidemiological features of a recent zoonotic cutaneous leishmaniasis outbreak in Zagora province, southern Morocco.

PLoS Negl Trop Dis 2019 04 9;13(4):e0007321. Epub 2019 Apr 9.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco.

Background: Leishmania major is an endemic vector-borne disease in Morocco that causes zoonotic cutaneous leishmaniasis (ZCL), especially in arid pre-Saharan regions where its unique vector and reservoir are Phlebotomus papatasi and Meriones shawi, respectively, and may cause epidemics. In late 2017, the Zagora province, an endemic focus for ZCL in southern Morocco, had CL outbreak. The main objective of our investigation was to analyze the epidemiological features of this latest ZCL outbreak.

Methodology/principal Findings: We analyzed epidemiological features of this latest ZCL outbreak. The Regional Delegation of Health, Zagora, recorded 4,402 CL patients between October 2017 and end of March 2018. Our findings showed that 24 municipalities were affected and majority (55.1%) of infected cases belonged to the Tinzouline rural municipality. Majority of patients were females (57.2%). While all age group patients were affected, those aged <10 years were the most affected (42.1%). During this outbreak over 5 days in December 2017, we conducted a survey in Tinzouline and recruited and sampled 114 CL patients to confirm CL diagnosis by parasitological (direct examination and culture) and molecular (ITS1-PCR) methods and identify the etiological agent of infection using ITS1-PCR-RFLP and sequencing. We completed a detailed questionnaire including clinical and epidemiological data for each patient and found 72.8% of patients presenting multiple lesions (≥2), with an average number of lesions of 5.16 ± 0.5. Lesions were more prevalent in the upper limbs, with the most common type being the ulcerocrusted lesion (60.5%). We detected no associations between lesion type and patients' sex or age.

Conclusions/significance: Among 114 clinically diagnosed CL patients, we confirmed 90.35% and identified L. major as the species responsible for this outbreak. Self-medication using various products caused superinfection and inflammation of lesions and complicated the diagnosis and treatment. Thus, ZCL remains a major public health problem in the Zagora province, and commitment of all stakeholders is urgently required to implement a sustainable regional control program.
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http://dx.doi.org/10.1371/journal.pntd.0007321DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6474635PMC
April 2019

Genome Dynamics during Environmental Adaptation Reveal Strain-Specific Differences in Gene Copy Number Variation, Karyotype Instability, and Telomeric Amplification.

mBio 2018 11 6;9(6). Epub 2018 Nov 6.

Unité de Parasitologiemoléculaire et Signalisation, Institut Pasteur, Paris, France

Protozoan parasites of the genus adapt to environmental change through chromosome and gene copy number variations. Only little is known about external or intrinsic factors that govern genomic adaptation. Here, by conducting longitudinal genome analyses of 10 new clinical isolates, we uncovered important differences in gene copy number among genetically highly related strains and revealed gain and loss of gene copies as potential drivers of long-term environmental adaptation in the field. In contrast, chromosome rather than gene amplification was associated with short-term environmental adaptation to culture. Karyotypic solutions were highly reproducible but unique for a given strain, suggesting that chromosome amplification is under positive selection and dependent on species- and strain-specific intrinsic factors. We revealed a progressive increase in read depth towards the chromosome ends for various isolates, which may represent a nonclassical mechanism of telomere maintenance that can preserve integrity of chromosome ends during selection for fast growth. Together our data draw a complex picture of genomic adaptation in the field and in culture, which is driven by a combination of intrinsic genetic factors that generate strain-specific phenotypic variations, which are under environmental selection and allow for fitness gain. Protozoan parasites of the genus cause severe human and veterinary diseases worldwide, termed leishmaniases. A hallmark of biology is its capacity to adapt to a variety of unpredictable fluctuations inside its human host, notably pharmacological interventions, thus, causing drug resistance. Here we investigated mechanisms of environmental adaptation using a comparative genomics approach by sequencing 10 new clinical isolates of the , , and complexes that were sampled across eight distinct geographical regions. Our data provide new evidence that parasites adapt to environmental change in the field and in culture through a combination of chromosome and gene amplification that likely causes phenotypic variation and drives parasite fitness gains in response to environmental constraints. This novel form of gene expression regulation through genomic change compensates for the absence of classical transcriptional control in these early-branching eukaryotes and opens new venues for biomarker discovery.
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http://dx.doi.org/10.1128/mBio.01399-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6222132PMC
November 2018

Genetic diversity of Leishmania tropica in Morocco: does the dominance of one haplotype signify its fitness in both predominantly anthropophilic Phlebotomus sergenti and human beings?

Transbound Emerg Dis 2019 Jan 22;66(1):373-380. Epub 2018 Oct 22.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco.

In Morocco, cutaneous leishmaniasis (CL) caused by Leishmania tropica is endemic to locations where the predominantly anthropophilic blood-feeding Phlebotomus sergenti and humans co-perpetuate. The objective of this study was to explore whether the range of epidemiological features of CLcould be linked to the range of L. tropica genetic heterogeneity and to further explore the relationships between the genetic diversity of L. tropica in both P. sergenti and humans. L. tropica DNAwas extracted from dermal scarping smears of 125 CLpatients. Genetic polymorphisms were analyzed by sequencing the internal transcribed spacer (ITS) 1 and 5.8S rDNAgene. Nucleotide diversity (π), haplotype diversity (Hd) and Tajima's D test for neutrality, as well as overall and pairwise FSTvalues, were calculated using Arlequin ver 3.5 software. Out of the 125 amplified DNAsequences, 93 were completely sequenced and 13 L. tropica haplotypes were identified, which confirmed the significant genetic heterogeneity of L. tropica in Morocco. The most common haplotype included 74 out of 93 sequences; this haplotype is not only widely represented but was also detected in P. sergenti, which is known to be the most abundant species in the studied foci. Considering the negative value calculated using Tajima's D index, we briefly discussed the hypothesis that the L. tropica common haplotype propagation could be a sign of its fitness in P. sergenti and human hosts. Furthermore, analysis of molecular variance (AMOVA) shows significant correlations between intraspecific variants of L. tropica and patients' geographic origins. The long-term goals of the present pilot study are to further explore the relationships between the genetic diversity of L. tropica in human and P. sergenti populations.
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http://dx.doi.org/10.1111/tbed.13031DOI Listing
January 2019

Genetic polymorphism in Leishmania infantum isolates from human and animals determined by nagt PCR-RFLP.

Infect Dis Poverty 2018 Jun 14;7(1):54. Epub 2018 Jun 14.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco.

Background: Leishmania infantum is the causative agent of human visceral leishmaniasis (VL) and sporadic human cutaneous leishmaniasis (CL) in the Mediterranean region. The genetic variation of the Leishmania parasites may result in different phenotypes that can be associated with the geographical distribution and diversity of the clinical manifestations. The main objective of this study was to explore the genetic polymorphism in L. infantum isolates from human and animal hosts in different regions of Morocco.

Methods: The intraspecific genetic variability of 40 Moroccan L. infantum MON-1 strains isolated from patients with VL (n = 31) and CL (n = 2) and from dogs (n = 7) was evaluated by PCR-RFLP of nagt, a single-copy gene encoding N-acetylglucosamine-1-phosphate transferase. For a more complete analysis of L. infantum polymorphism, we included the restriction patterns of nagt from 17 strains available in the literature and patterns determined by in-silico digestion of three sequences from the GenBank database.

Results: Moroccan L. infantum strains presented a certain level of genetic diversity and six distinct nagt-RFLP genotypes were identified. Three of the six genotypes were exclusively identified in the Moroccan population of L. infantum: variant M1 (15%), variant M2 (7.5%), and variant M3 (2.5%). The most common genotype (65%), variant 2 (2.5%), and variant 4 (7.5%), were previously described in several countries with endemic leishmaniasis. Phylogenetic analysis segregated our L. infantum population into two distinct clusters, whereas variant M2 was clearly distinguished from both cluster I and cluster II. This distribution highlights the degree of genetic variability among the Moroccan L. infantum population.

Conclusion: The nagt PCR-RFLP method presented here showed an important genetic heterogeneity among Moroccan L. infantum strains isolated from human and canine reservoirs with 6 genotypes identified. Three of the six Moroccan nagt genotypes, have not been previously described and support the particular genetic diversity of the Moroccan L. infantum population reported in other studies.
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http://dx.doi.org/10.1186/s40249-018-0439-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6001066PMC
June 2018

G119S ace-1 mutation conferring insecticide resistance detected in the Culex pipiens complex in Morocco.

Pest Manag Sci 2019 Jan 21;75(1):286-291. Epub 2018 Aug 21.

Institut Pasteur du Maroc, Service de Parasitologie et des Maladies Vectorielles, Casablanca, Morocco.

Background: Arboviruses are controlled through insecticide control of their mosquito vector. However, inconsiderate use of insecticides often results in the selection of resistance in treated populations, so that monitoring is required to optimize their usage. Here, Culex pipiens (West Nile and Rift Valley Fever virus vector) specimens were collected from four Moroccan cities. Levels of susceptibility to the organophosphate (OP) insecticide malathion were assessed using World Health Organization (WHO)-recommended bioassays. Individual mosquitoes were tested for the presence of the G119S mutation in the ace-1 gene, the main OP-target resistance mutation.

Results: Bioassays showed that mosquitoes from Mohammedia were significantly more resistant to malathion than those from Marrakech. Analyzing the ace-1 genotypes in dead and surviving individuals suggested that other resistance mechanisms may be present in Mohammedia. The ace-1 resistance allele frequencies were relatively moderate (< 0.4). Their analyses in three Moroccan cities (Tangier, Casablanca and Marrakech) however showed disparities between two coexisting Cx. pipiens forms and revealed that the G119S mutation tends to be more frequent in urban than in rural collection sites.

Conclusion: These findings provide a reference assessment of OP resistance in Morocco and should help the health authorities to develop informed and sustainable vector control programs. © 2018 Society of Chemical Industry.
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http://dx.doi.org/10.1002/ps.5114DOI Listing
January 2019

First Molecular Identification and Phylogeny of Moroccan Anopheles sergentii (Diptera: Culicidae) Based on Second Internal Transcribed Spencer (ITS2) and Cytochrome c Oxidase I (COI) Sequences.

Vector Borne Zoonotic Dis 2018 09 5;18(9):479-484. Epub 2018 Jun 5.

1 Laboratory of Vector-Borne-Diseases, Institut Pasteur du Maroc , Casablanca, Morocco .

Anopheles sergentii known as the "oasis vector" or the "desert malaria vector" is considered the main vector of malaria in the southern parts of Morocco. Its presence in Morocco is confirmed for the first time through sequencing of mitochondrial DNA (mDNA) cytochrome c oxidase subunit I (COI) barcodes and nuclear ribosomal DNA (rDNA) second internal transcribed spacer (ITS2) sequences and direct comparison with specimens of A. sergentii of other countries. The DNA barcodes (n = 39) obtained from A. sergentii collected in 2015 and 2016 showed more diversity with 10 haplotypes, compared with 3 haplotypes obtained from ITS2 sequences (n = 59). Moreover, the comparison using the ITS2 sequences showed closer evolutionary relationship between the Moroccan and Egyptian strains than the Iranian strain. Nevertheless, genetic differences due to geographical segregation were also observed. This study provides the first report on the sequence of rDNA-ITS2 and mtDNA COI, which could be used to better understand the biodiversity of A. sergentii.
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http://dx.doi.org/10.1089/vbz.2018.2269DOI Listing
September 2018

Molecular identification of Leishmania infection in the most relevant sand fly species and in patient skin samples from a cutaneous leishmaniasis focus, in Morocco.

PLoS Negl Trop Dis 2018 03 2;12(3):e0006315. Epub 2018 Mar 2.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco.

Background: Cutaneous leishmaniasis (CL) is an infectious disease caused by various species of Leishmania and transmitted by several species of sand flies. CL is among the most neglected tropical diseases, and it has represented a major health threat over the past 20 years in Morocco. The main objectives of this study were to identify relevant sand fly species and detect Leishmania infection in the most prevalent species and patient skin samples in Taza, a focus of CL in North-eastern Morocco.

Methodology And Finding: A total of 3672 sand flies were collected by CDC miniature light traps. Morphological identification permitted the identification of 13 species, namely 10 Phlebotomus species and 3 Sergentomyia species. P. longicuspis was the most abundant species, comprising 64.08% of the total collected sand flies, followed by P. sergenti (20.1%) and P. perniciosus (8.45%). Using nested-kDNA PCR, seven pools of P. sergenti were positive to Leishmania tropica DNA, whereas 23 pools of P. longicuspis and 4 pools of P. perniciosus tested positive for Leishmania infantum DNA. The rates of P. longicuspis and P. perniciosus Leishmania infection were 2.51% (23/915) and 7.27% (4/55), respectively, whereas the infection prevalence of P. sergenti was 3.24%. We also extracted DNA from lesion smears of 12 patients suspected of CL, among them nine patients were positive with enzymatic digestion of ITS1 by HaeIII revealing two profiles. The most abundant profile, present in eight patients, was identical to L. infantum, whereas L. tropica was found in one patient. The results of RFLP were confirmed by sequencing of the ITS1 DNA region.

Conclusion: This is the first molecular detection of L. tropica and L. infantum in P. sergenti and P. longicuspis, respectively, in this CL focus. Infection of P. perniciosus by L. infantum was identified for the first time in Morocco. This study also underlined the predominance of L. infantum and its vector in this region, in which L. tropica has been considered the causative agent of CL for more than 20 years.
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http://dx.doi.org/10.1371/journal.pntd.0006315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5851645PMC
March 2018

New epidemiological pattern of cutaneous leishmaniasis in two pre-Saharan arid provinces, southern Morocco.

Acta Trop 2017 Sep 17;173:11-16. Epub 2017 May 17.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco. Electronic address:

Three Leishmania species are responsible of cutaneous leishmaniasis (CL) in Morocco. Zoonotic CL due to Leishmania major and Leishmania infantum, the first is known as established in the eastern arid regions, whereas the latter evolves sporadically, especially in the North. While Leishmania tropica, classically considered anthroponotic, is endemic in the semi-arid regions and is largely distributed throughout the country. The aim of this study was to identify the Leishmania species causing CL in two Provinces in arid pre-Saharan region known as zoonotic CL foci, and to contribute an update to the national data concerning the distribution of Leishmania species in both regions. The recruitment of patients was done in six localities in Ouarzazate and Zagoura provinces in 2015 and 2016. Out of 81 samples collected, 66 were positive (81%) by ITS1-PCR amplification of Leishmania DNA extracted from stained smears. The highest rate of Leishmania infection was registered in children aged 9 years or less (71,2%). The ITS1-PCR- RFLP analysis revealed the predominance of L. major infecting 52 patients (79%), followed by L. tropica in 12 patients (18%) and L. infantum in 2 patients who had no history of travel outside the studied area (3%). The sequencing of the ITS1 of both L. infantum, showed 100% similarities with L. infantum strains isolated from dogs and visceral leishmaniasis patients from the south and north of Morocco. The coexistence of the 3 Leishmania species in the same focus, and the difficult distinction of infections associated to the different Leishmania species based only on clinical lesions' aspects complicate the diagnosis and then the national control strategy, as well as the therapeutic management. The epidemiological pattern of CL in the studied areas appears to have changed during the last decades, from a predominant zoonotic CL caused by L. major to a polymorphic disease that can be due to any of the 3 Leishmania species. The expansion of L. infantum and L. tropica in southern parts of Morocco, calls for in depth epidemiological investigations for a better understanding of the CL situations in Southern parts of the country and for an assessment of the climate impact and environment changes on the leishmaniasis transmission system.
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http://dx.doi.org/10.1016/j.actatropica.2017.05.016DOI Listing
September 2017

Intraspecific genetic variability in a population of Moroccan Leishmania infantum revealed by PCR-RFLP of kDNA minicircles.

Acta Trop 2017 May 14;169:142-149. Epub 2017 Feb 14.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco. Electronic address:

In Morocco, Leishmania infantum is the main etiologic agent of human and canine visceral leishmaniasis (VL). This species has been proven to be an opportunistic agent in HIV+ patients and is also responsible of sporadic cutaneous leishmaniasis (CL).This work aims to evaluate the genetic variability of Moroccan L. infantum strains based on PCR-RFLP analysis of the kinetoplastid DNA (kDNA) minicircles. A total of 75 DNA samples extracted from positive Giemsa-stained smears (n=32) and from L. infantum cultures (n=43) was studied. The samples have been taken from VL patients infected (n=7) or not (n=56) by HIV, patients with CL (n=2) and finally from infected dogs (n=10). An hypervariable region of kDNA was amplified using the primers MC1 and MC2; the PCR products were digested separately by a panel of nine restriction enzymes. The presence or absence of restriction fragments was scored in a binary matrix and the SplitsTree4 software was used for the construction of a Neighbor-Net network. Moroccan L. infantum population showed an important level of variability with the identification of 6 genotypes. For each genotype a PCR product was sequenced, confirming the presence of all the expected restriction sites. The predominant profile was the genotype B. A new genotype, named Q was detected for the first time, whereas the four other genotypes (G, K, N and O) were reported sporadically in the Mediterranean basin. The Neighbor-Net network segregates our L. infantum population into 3 clusters: Cluster I includes genotype B, cluster II grouping the genotypes O, Q and G and finally the cluster III contains the genotype N. The kDNA-PCR-RFLP assay is suitable for use directly on biological samples; it reveals an important degree of genetic variability among L. infantum strains even those belonging to the same zymodeme what is of great epidemiological interest.
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http://dx.doi.org/10.1016/j.actatropica.2017.02.016DOI Listing
May 2017

The TLR2 and TLR4 gene polymorphisms in Moroccan visceral leishmaniasis patients.

Acta Trop 2016 Jun 2;158:77-82. Epub 2016 Mar 2.

Laboratory of Parasitology and Vector-Borne-Diseases, Institut Pasteur du Maroc, Casablanca, Morocco. Electronic address:

Visceral leishmaniasis (VL) is endemic in the Mediterranean basin and leads to the most severe form of Leishmania infection, lethal if left untreated. However, most infections are sub-clinical or asymptomatic, reflecting the influence of host genetic background on disease outcome. This study aimed to investigate possible association of TLR4 Asp299Gly, TLR4 Thr399Ile and TLR2 Arg753Gln polymorphisms with VL in Moroccan children. We enrolled 119 children with VL caused by Leishmania infantum as well as 138 unrelated children, 95 asymptomatic subjects and 43 healthy individuals who had no evidence of present or past infection. Polymorphisms were genotyped by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) and amplification refractory mutation system assay (ARMS-PCR). Results showed significant differences in genotype Thr399Ile and recessive model frequencies between VL and delayed-type hypersensitivity (DTH+) groups (p=0.018, OR=0.414CI 0.195-0.880; p=0.029, OR=0.448CI 0.214-0.938], respectively) by having the amino-acid threonine polymorphism as a reference in the VL group. Concerning the Asp299Gly there were a significant associations when comparing VL vs DTH+ (Asp299Gly genotype p=0.002, OR=0.326CI 0.158-0.671, allele frequencies p=0.033, OR=0.396CI 0.164-0.959, recessive model p=0.002, OR=0.343CI 0.172-0.681) and DTH+ vs DTH- groups (Asp299Gly genotype p=2.160E-4, OR=3.065CI 1.672-5.618, Gly299Gly genotype p=0.047, OR=0.368CI 0.299-0.452, allele frequencies p=1.406E-7, OR=29.571CI 3.907-223.8, recessive model p=4.370E-14, OR=36.965CI 8.629-158.3), by having the aspartic acid polymorphism as a reference these results suggest that the allele A (savage) confer protection against the clinical manifestations but not against the infection. Furthermore, there was a significant association regarding the Arg753Gln genotype (p=0.002, OR=0.326CI 0.158-0.671), allele frequencies (p=0.033, OR=0.396CI 0.164-0.959) and when applying a recessive model (p=0.002, OR=0.343CI 0.172-0.681) in the VL vs DTH+ groups. The same results was observed when comparing DTH+ vs DTH- groups (p=4.136E-6, OR=0.211CI 0.104-0.428), allele frequencies (p=0.008, OR=0.327CI 0.137-0.779) and recessive model (p=1.748E-5, OR=0.244CI 0.124-0.480). The results provide evidence that allele C in Thr399Ile and allele G in Arg753Gln polymorphisms may lead to protection against the clinical disease. Our data provide insights into the possible role of TLR2 and TLR4 variations in VL susceptibility.
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http://dx.doi.org/10.1016/j.actatropica.2016.02.020DOI Listing
June 2016

Toscana virus isolated from sandflies, Morocco.

Parasit Vectors 2015 Apr 3;8:205. Epub 2015 Apr 3.

Laboratoire de Parasitologie et Maladies Vectorielles, Institut Pasteur du Maroc, Casablanca, Morocco.

To investigate the transmission of phleboviruses, a total of 7,057 sandflies were collected in well-known foci of cutaneous leishmaniasis and were identified to species level according to morphological characters.Collected sandflies were tested by Nested PCR for the presence of Phleboviruses and subsequently by viral isolation on Vero cells. The corresponding products were sequenced. Toscana virus was isolated, for the first time, from 5 pools of sandflies.Hence, Toscana virus should be considered a potential risk that threatens public health and clinicians should be aware of the role of Toscana virus in cases of meningitis and encephalitis in Morocco.
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http://dx.doi.org/10.1186/s13071-015-0826-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4415314PMC
April 2015

Phlebotomus sergenti in a cutaneous leishmaniasis focus in Azilal province (High Atlas, Morocco): molecular detection and genotyping of Leishmania tropica, and feeding behavior.

PLoS Negl Trop Dis 2015 Mar 31;9(3):e0003687. Epub 2015 Mar 31.

Laboratoire de Parasitologie et Maladies Vectorielles, Institut Pasteur du Maroc, Casablanca, Morocco.

Background: Phlebotomus (Paraphlebotomus) sergenti is at least one of the confirmed vectors for the transmission of cutaneous leishmaniasis caused by Leishmania tropica and distributed widely in Morocco. This form of leishmaniasis is considered largely as anthroponotic, although dogs were found infected with Leishmania tropica, suggestive of zoonosis in some rural areas.

Methodology And Findings: This survey aimed at (i) studying the presence of Leishmania in field caught Phlebotomus sergenti, (ii) investigating genetic diversity within Leishmania tropica and (iii) identifying the host-blood feeding preferences of Phlebotomus sergenti. A total of 4,407 sand flies were collected in three rural areas of Azilal province, using CDC miniature light traps. Samples collected were found to consist of 13 species: Phlebotomus spp. and 3 Sergentomyia spp. The most abundant species was Phlebotomus sergenti, accounting for 45.75 % of the total. 965 female Phlebotomus sergenti were screened for the presence of Leishmania by ITS1-PCR-RFLP, giving a positive rate of 5.7% (55/965), all being identified as Leishmania tropica. Nucleotide heterogeneity of PCR-amplified ITS1-5.8S rRNA gene-ITS2 was noted. Analyses of 31 sequences obtained segregated them into 16 haplotypes, of which 7 contain superimposed peaks at certain nucleotide positions, suggestive of heterozygosity. Phlebotomus sergenti collected were found to feed on a large variety of vertebrate hosts, as determined by Cytochrome b sequencing of the DNA from the blood meals of 64 engorged females.

Conclusion: Our findings supported the notion that Phlebotomus sergenti is the primary vector of Leishmania tropica in this focus, and that the latter is genetically very heterogeneous. Furthermore, our results might be suggestive of a certain level of heterozygosity in Leishmania tropica population. This finding, as well as the feeding of the vectors on different animals are of interest for further investigation.
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http://dx.doi.org/10.1371/journal.pntd.0003687DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4380298PMC
March 2015

SLC11A1 polymorphisms and susceptibility to visceral leishmaniasis in Moroccan patients.

Acta Trop 2014 Dec 20;140:130-6. Epub 2014 Aug 20.

Laboratoire de parasitologie et maladies vectorielles, Institut Pasteur du Maroc, Casablanca, Morocco. Electronic address:

Human visceral leishmaniasis is endemic in the Mediterranean basin. Since most infections are sub-clinical or asymptomatic, host genetics can provide concrete evidence in determining disease outcome. SLC11A1/NRAMP1 is a candidate gene that may be related to host susceptibility versus resistance to intracellular pathogens. This study aimed to determine possible association of SLC11A1 polymorphisms with visceral leishmaniasis among Moroccan children. A total of 106 children who developed visceral leishmaniasis due to Leishmania infantum were enrolled in this study. The control group was composed of 137 unrelated children, 97 asymptomatic subjects (DTH+) and 42 healthy individuals (DTH) who had no evidence of present or past infection. Four polymorphisms were studied by PCR-RFLP and sequencing: (GT)n microsatellite in the 5' exon 1; silent substitutions 469+14G/C in intron 4; amino acid substitution D543N in exon 15 and 823C/T polymorphism in exon 8. Thereafter, the frequencies of genotypes, alleles and haplotypes were estimated. Two polymorphisms were each significantly associated in the genotypes with visceral leishmaniasis: 823C/T in exon 8 and D543N in exon 15 when comparing visceral leishmaniasis and DTH+ groups. The results of haplotype frequencies suggested an evidence of association with resistance to visceral leishmaniasis for the "286GTG" and "288GCA" haplotypes, whereas, the "286GCG" haplotype appears to increase the risk to visceral leishmaniasis susceptibility.Our data provide insights into the possible role of SLC11A1 variation in visceral leishmaniasis susceptibility. These results must be regarded as preliminary but suggestive that further study with larger populations is worthwhile.
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http://dx.doi.org/10.1016/j.actatropica.2014.08.013DOI Listing
December 2014

Molecular detection and identification of Leishmania infection in naturally infected sand flies in a focus of cutaneous leishmaniasis in northern Morocco.

Parasit Vectors 2014 Jul 2;7:305. Epub 2014 Jul 2.

Laboratoire de Parasitologie et Maladies Vectorielles, Institut Pasteur du Maroc, 1 Place Louis Pasteur, 20360 Casablanca, Morocco.

Background: Cutaneous leishmaniasis is an infectious disease caused by various species of the flagellate protozoan Leishmania. During the past 20 years, cutaneous leishmaniasis has emerged as a major public health threat in Morocco. The main objective of this study was to study the occurrence of Leishmania infection in vectors and to identify sand fly blood meal sources in an endemic locality of cutaneous leishmaniasis within Sefrou province, where the vectors of leishmaniasis were still unknown.

Methods: 2650 sand flies were collected using CDC miniature light traps and identified morphologically. The identified sand flies were tested for Leishmania infection by nested PCR. The source of blood meal of 10 freshly engorged females: 6 Phlebotomus longicuspis and 4 Phlebotomus sergenti, was determined using the Cyt b sequence.

Results: The collected sand flies consisted of 10 species, seven of which belonged to the genus Phlebotomus and three to the genus Sergentomyia. The most abundant species was P. longicuspis, accounting for 72% of the total sand flies collected. In females of three P. longicuspis and four P. sergenti, Leishmania infantum and Leishmania tropica DNA was detected, respectively.The source of blood meal of engorged females showed that all sand flies tested fed on humans.

Conclusions: We report for the first time the natural infection of P. longicuspis with L. infantum in Morocco. The high frequency of this species in this region, in addition to its anthropophilic character make P. longicuspis the putative vector of L. infantum in this cutaneous leishmaniasis focus where L. tropica is confirmed as the causative agent of the disease and P. sergenti as its vector. The presence of L. infantum, and its presumed vector in this area, makes this a site of high risk of visceral leishmaniasis, mostly because of the proximity of a focus of human and canine visceral leishmaniasis.
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http://dx.doi.org/10.1186/1756-3305-7-305DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4094396PMC
July 2014

Phlebotomus sergenti a common vector of Leishmania tropica and Toscana virus in Morocco.

J Vector Borne Dis 2014 Jun;51(2):86-90

Laboratoire de parasitologie et de maladies vectorielles, Institut Pasteur du Maroc, Casablanca, Morocco.

Background & Objectives: An entomological study using CDC miniature light-traps was performed in El Hanchane locality, where cutaneous leishmaniasis (CL) was emerging during the summer of 2011. The aim of this study is to identify the vectors of Leishmania and of phleboviruses.

Methods: In the field, a total of 643 sandfly specimens were collected, identified by morphological keys and categorized by sex and species. A total of nine distinct species were morphologically identified where seven belonged to the Phlebotomus genus and two species to the Sergentomyia genus. Phlebotomus sergenti was the most abundant species (76%). Phleboviruses were detected by nested RT-PCR using 30 pooled sandflies while P. sergenti females were tested individually for infections of Leishmania species.

Results: By using ITS1-PCR-RFLP approach, Leishmania tropica DNA was detected in 10 females, caught in this emerging focus, and provide additional evidence in favour of the role of P. sergenti as vector of L. tropica in Morocco. Real-time PCR screening for phlebovirus RNA, using an assay targeting the polymerase gene, showed positive result in one pool of male P. sergenti.

Interpretation & Conclusion: In this study, P. sergenti were infected by L. tropica and Toscana virus. To our knowledge, actually this is the first time that Toscana virus has been detected in P. sergenti.
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June 2014

Moroccan Leishmania infantum: genetic diversity and population structure as revealed by multi-locus microsatellite typing.

PLoS One 2013 17;8(10):e77778. Epub 2013 Oct 17.

Faculty of Pharmacy, Alquds University, Jerusalem, Palestine ; Institute of Microbiology and Hygiene, Charité University Medicine, Berlin, Germany.

Leishmania infantum causes Visceral and cutaneous leishmaniasis in northern Morocco. It predominantly affects children under 5 years with incidence of 150 cases/year. Genetic variability and population structure have been investigated for 33 strains isolated from infected dogs and humans in Morocco. A multilocus microsatellite typing (MLMT) approach was used in which a MLMtype based on size variation in 14 independent microsatellite markers was compiled for each strain. MLMT profiles of 10 Tunisian, 10 Algerian and 21 European strains which belonged to zymodeme MON-1 and non-MON-1 according to multilocus enzyme electrophoresis (MLEE) were included for comparison. A Bayesian model-based approach and phylogenetic analysis inferred two L.infantum sub-populations; Sub-population A consists of 13 Moroccan strains grouped with all European strains of MON-1 type; and sub-population B consists of 15 Moroccan strains grouped with the Tunisian and Algerian MON-1 strains. Theses sub-populations were significantly different from each other and from the Tunisian, Algerian and European non MON-1 strains which constructed one separate population. The presence of these two sub-populations co-existing in Moroccan endemics suggests multiple introduction of L. infantum from/to Morocco; (1) Introduction from/to the neighboring North African countries, (2) Introduction from/to the Europe. These scenarios are supported by the presence of sub-population B and sub-population A respectively. Gene flow was noticed between sub-populations A and B. Five strains showed mixed A/B genotypes indicating possible recombination between the two populations. MLMT has proven to be a powerful tool for eco-epidemiological and population genetic investigations of Leishmania.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0077778PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3798341PMC
June 2014

Detection and molecular typing of Leishmania tropica from Phlebotomus sergenti and lesions of cutaneous leishmaniasis in an emerging focus of Morocco.

Parasit Vectors 2013 Jul 26;6:217. Epub 2013 Jul 26.

Laboratoire de Parasitologie et Maladies Vectorielles, Institut Pasteur du Maroc, 1 Place Louis Pasteur, Casablanca, Morocco.

Background: Cutaneous leishmaniasis is an infectious disease caused by flagellate protozoa of the genus Leishmania. In Morocco, anthroponotic cutaneous leishmaniasis due to Leishmania tropica is considered as a public health problem, but its epidemiology has not been fully elucidated. The main objective of this study was to detect Leishmania infection in the vector, Phlebotomus sergenti and in human skin samples, in the El Hanchane locality, an emerging focus of cutaneous leishmaniasis in central Morocco.

Methods: A total of 643 sand flies were collected using CDC miniature light traps and identified morphologically. Leishmania species were characterized by ITS1 PCR-RFLP and ITS1-5.8S rRNA gene nested-PCR of samples from 123 females of Phlebotomus sergenti and 7 cutaneous leishmaniasis patients.

Results: The sand flies collected consisted of 9 species, 7 of which belonged to the genus Phlebotomus and two to the genus Sergentomyia. Phlebotomus sergenti was the most predominant (76.67%).By ITS1 PCR-RFLP Leishmania tropica was found in three Phlebotomus sergenti females and four patients (4/7). Using nested PCR Leishmania tropica was identified in the same three Phlebotomus sergenti females and all the 7 patients. The sequencing of the nested PCR products recognized 7 haplotypes, of which 6 have never been described.

Conclusions: This is the first molecular detection and identification of Leishmania tropica in human skin samples and Phlebotomus sergenti in support of its vector status in El Hanchane. The finding of seven Leishmania tropica haplotypes underscores heterogeneity of this species at a high level in Morocco.
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http://dx.doi.org/10.1186/1756-3305-6-217DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3751070PMC
July 2013

Epidemiologic study and molecular detection of Leishmania and sand fly species responsible of cutaneous leishmaniasis in Foum Jamâa (Azilal, Atlas of Morocco).

Acta Trop 2013 Jul 21;127(1):1-5. Epub 2013 Mar 21.

Laboratory of Management and Valorization of Naturals Resources, FST, Sultan Moulay, Slimane University, M'GHILA Route de Fes, B.P. 523, Beni Mellal 23000, Morocco.

The region of Foum Jamâa (province of Azilal) has become endemic for cutaneous leishmaniasis (CL) since 2006. The objective of this study was to investigate molecular identification of the etiological agent of CL in this region; we also carried out an entomological survey of Phlebotomine sand flies (Diptera: Psychodidae) in this focus to study the sand fly fauna, species composition, and the monthly prevalence of sand flies during 1 year. In the period between 2009 and 2010, skin scrapings spotted on glass slides were collected from 119 patients, aged from 9 months to 70 years old, who came from 43 localities distributed in 3 sectors in Foum Jamâa (FJ). The ITS1 PCR-RFLP was used to identify the Leishmania parasite responsible for the recent cases of CL in FJ. Our results revealed that the disease is caused by L. tropica. No significant association was observed between gender and the rate of CL in presenting patients, while the highest rate of positive lesions was found in the age group of 9 years old or under (86.67%). In this study, we found also that L. tropica infection mostly caused single lesions (67.90%) that were located in the face (96.30%). Morphological identification was performed on a total of 1152 sand flies (23% females and 77% males) collected by sticky paper traps. 57% of the total collected flies were identified as Phlebotomus (Paraphlebotomus) sergenti (Parrot).
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http://dx.doi.org/10.1016/j.actatropica.2013.03.005DOI Listing
July 2013

A variant in the promoter of MBL2 is associated with protection against visceral leishmaniasis in Morocco.

Infect Genet Evol 2013 Jan 18;13:162-7. Epub 2012 Sep 18.

Laboratoire des recherches et d'étude sur les Leishmanioses, Institut Pasteur du Maroc, Casablanca, Morocco.

Progressive visceral leishmaniasis (VL) is fatal if not treated; yet, most infections with the causative agents are asymptomatic. We hypothesized that genetic factors contribute to this variable response to infection. The mannose-binding lectin 2 gene (MBL2) is a candidate that merits examination in the context of VL because it enhances infection with intracellular pathogens. Four functional MBL2 polymorphisms at codons 52, 54, 57 and in the promoter at the -221 position (X/Y) are known to be associated with the outcome of several diseases. The aim of the present study was to investigate whether these functional variants were associated with VL in Moroccan children. Here, we genotyped polymorphisms by sequencing and PCR-RFLP in 112 individuals with VL, 97 asymptomatic subjects and 42 healthy individuals who had no evidence of present or past infection. Regression analysis showed no significant association between polymorphisms in exon 1 genotypes and outcome of infection with Leishmania infantum. However, the genotype XY in -221 conferred a protective role against VL in our study population with a significant difference (OR=0.291; CI [0.158-0.538]; p=0.0006). Subjects with YY genotypes in -221 had a higher risk to developing VL. We concluded that MBL2 polymorphism at the -221 promoter region plays a protective role in L. infantum infection.
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http://dx.doi.org/10.1016/j.meegid.2012.09.002DOI Listing
January 2013

PCR detection of Leishmania in skin biopsies.

J Infect Dev Ctries 2009 Sep 15;3(2):115-22. Epub 2009 Sep 15.

Laboratoire des recherche et d'étude sur les Leishmanioses, Institut Pasteur du Maroc, Casablanca, Morocco.

Introduction: Cutaneous leishmaniasis (CL) is an endemic disease and one of the major health problems in Morocco. In 2006, the recorded total number of cases of CL was 3361, occurring predominantly in the rural population. A new and more sensitive diagnostic technique than current methods used is needed in this setting. The aim of this study was to assess the efficacy of polymerase chain reaction (PCR) to detect leishmanial parasites in skin biopsies of patients from different areas of endemicity in Morocco.

Methodology: Biopsies from 26 patients with cutaneous ulcers suggestive of leishmaniasis were analysed by PCR using primers from the small subunit ribosomal gene. The ability of PCR to detect Leishmania was compared with smear-stained and in vitro culture.

Results: PCR exhibited superior sensitivity (84,6%) compared with direct microscopy smear (69,2%) and in vitro culture (69,2%). Our PCR assay also showed good specificity (100%).

Conclusions: PCR should be considered a valuable, sensitive, and faster diagnostic tool in the diagnosis of cutaneous leishmaniasis, especially for those patients with negative parasitologic examination.
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http://dx.doi.org/10.3855/jidc.511DOI Listing
September 2009

Antileishmanial and antibacterial activity of a new pyrazole derivative designated 4-[2-(1-(ethylamino)-2-methyl- propyl)phenyl]-3-(4-methyphenyl)-1-phenylpyrazole.

Arch Pharm (Weinheim) 2006 Jun;339(6):291-8

Laboratoire de Microbiologie et Biologie Moléculaire, Département de Biologie, Faculté des Sciences Ben M'sik-Casablanca, Université Hassan Il-Mohammedia, Casablanca, Morocco.

Here, we report for the first time the synthesis and the antileishmanial activity of a new pyrazole derivative, namely 4-[2-(1-(ethylamino)-2-methylpropyl)phenyl]-3-(4-methyphenyl)-1-phenylpyrazole). Micromolar concentrations of this compound were found to inhibit the in vitro multiplication of Leishmania tropica, Leishmania major, and Leishmania infantum, three species causing different forms of leishmaniasis. Furthermore, the 50% inhibitory concentration (IC50) values for the compound are only slightly higher than those of amphotericin B, one of the most active antileishmanial agents used as a satisfactory substitute in cases not responding to pentostam. The IC50 values after 48 h for L. tropica, L. major, and L. infantum promastigote growth were 0.48 microg/mL, 0.63 microg/mL and 0.40 microg/mL, respectively for the compound, while they were 0.23 microg/mL, 0.29 microg/mL and 0.24 microg/mL, respectively for amphotericin B. We also tested this compound for its antibacterial activity against several bacteria. The strongest antibacterial activity was observed against Entrococcus feacalis and Staphylococcus aureus with a minimal inhibitory concentration (MIC) of 60 microg/mL.
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http://dx.doi.org/10.1002/ardp.200500266DOI Listing
June 2006

Antileishmanial activity of a new 8-hydroxyquinoline derivative designed 7-[5'-(3'-phenylisoxazolino)methyl]-8-hydroxyquinoline: preliminary study.

Farmaco 2004 Mar;59(3):195-9

Département de Biologie, Laboratoire de Microbiologie et Biologie Moléculaire, Faculté des Sciences Ben M'sik-Casablanca, Université Hassan II-Mohammedia, Sidi Othmane, Casablanca B.P. 7955, Morocco.

7-[5'-(3'-phenylisoxazolino)methyl]-8-hydroxyquinoline a new synthetic 8-hydroxyquinoline derivative, was found, for the first time, to inhibit the multiplication in vitro of Leishmania tropica, Leishmania major and Leishmania infantum at micromolar concentrations. For each test we calculated a 50% inhibitory concentration (IC50) and the IC50 values found after 48 h are: 0.4 microgram/ml for L. tropica, 0.88 microgram/ml for L. major and 0.62 microgram/ml for L. infantum. As positive control, amphotericin B, a standard antileishmanial drug was used.
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http://dx.doi.org/10.1016/j.farmac.2003.11.001DOI Listing
March 2004