Molecular diagnosis of cutaneous leishmaniasis and identification of the causative Leishmania species in Morocco by using three PCR-based assays

Background: The diagnosis of cutaneous leishmaniasis (CL) might be difficult, in particular in endemic areas where different species of Leishmania can cause lesions of very similar appearance and where other skin diseases with similar clinical symptoms occur. Even today, the parasitological diagnosis of CL remains the gold standard and it is based on the direct identification of amastigotes in microscopy smears and/or culture of promastigotes from infected tissues. Although these techniques are highly specific, they are not sensitive enough. The objective of this study is to contribute to improving the diagnosis of CL and the identification of Leishmania species in Morocco by comparing three PCR-based assays applied directly on dermal samples. Methods: A total of 58 patients presenting with cutaneous lesions suggestive of CL were sampled for parasitological diagnosis by direct examination (DE), culture in NNN medium, two kinetoplast DNA (kDNA) PCRs (Lmj4/Uni21 and 13A/13B primers) and one rRNA gene internal transcribed spacer 1 (ITS1) PCR (LITSR/L5.8S primers). The techniques were statistically analyzed and compared. Results: According to our consensus positive, 44 out of 58 samples were true positives. The 13A/13B-PCR and ITS1-PCR showed the highest sensitivities (100%). Parasite microscopy and culture detected 43% and 29% of the true positives, respectively, while culture and microscopy together improved sensitivity to 52%. PCRs 13A/13B and ITS1 were associated to four and one false positives, respectively, while the other assays were 100% specific. Furthermore, the ITS1-PCR-RFLP assay clearly identified the Leishmania species for all the true positives (44/44), whereas Lmj4/Uni21-PCR identified 35/44 samples. The comparison between the Leishmania molecular characterizations and the expected species according to the national data from the Ministry of Health indicate 7 discrepant results. Conclusions: The PCR-based assays tested on our samples increased the speed and sensitivity of the diagnosis of CL compared to the conventional techniques. Furthermore, we showed that we can not base the species identification on the national data from the Ministry of Health. Finally, we suggest the use of PCR-ITS1-RFLP for diagnosis and simultaneous identification of the species in the Moroccan epidemiological context, but also in similar areas of the Mediterranean Basin.This study was supported by the University of Granada, Spain (CICODE program), the Faculty of Medicine and Pharmacy of Casablanca (Morocco), and the PHC Volubilis program

Psoriasis and staphylococcus aureus skin colonization in Moroccan patients

Psoriatic lesions are rarely complicated by recurrent infections. The aim of our study is to determine skin colonisation and nasal carriage of Staphylococcus aureus in patients with psoriasis and in healthy  persons. Patients and methods: a comparative study that include 33 patients with psoriasis and 33  healthy persons.Samples were taken from lesional and non lesional psoriatic skin and from healthy skin of control group. For S. aureus nasal carriage, we used sterile cotton tipped swabs. Out of165 samples (66 skin samples and 33 nasal swabs), 26 S. Aureus strains were isolated in 26 persons, 57.69% in the  control group and 42.3% in the psoriasisgroup. S. aureus skin colonization was found in one case (3%) inlesional psoriatic skin vs 9 cases (27.3%) in control skin OR=0.08 IC 95% (0.01-0.70) p=0.02 and in 12,1% in non lesional soriatic skin vs 27, 3% in control skin (p =0,13). This colonization was less important in lesional psoriatic skin (3%) than in non lesional psoriatic skin (12.1%) p= 0.20. Nasal screening identified (7/33) 21, 21% S. aureus carriers in psoriasis group and in control group. Our results are in consensus withliterature findings. They have confirmed the importance of antimicrobial peptides in Innateimmunity of human skin. These peptides are normally produced bykeratinocytes in response to inflammatory stimuli such as psoriasis. Their high  expression in psoriasis skin reduces the risk of skin infection and skin colonization with S. Aureus.Key words: Antimicrobial peptides, innate immunity, nasal carriage, psoriasis, skin colonization, staphylococcus aureu

Concomitant visceral and localized cutaneous leishmaniasis in two Moroccan infants

Background: Leishmaniases are vector-borne diseases caused by the protozoa of the Leishmania genus. The clinical spectrum of these diseases extends from benign dermal lesions to visceral forms. In the Mediterranean region, zoonotic visceral leishmaniasis (ZVL) is caused by L. infantum. If untreated within two years, the disease usually leads to death. In Morocco, ZVL is endemic in the north, with a hundred cases notified each year, mostly in children aged below five years. Here, we report on two clinical observations in infants presenting unusual concomitant VL and cutaneous leishmaniasis (CL) in Morocco. Case presentation: In this case study, we report on two infants aged nine and 12 months old. They both have a history of febrile splenomegaly, anemia, and pallor of mucous membranes. Visceral leishmaniasis was confirmed by parasitological diagnosis (positive bone marrow smear and screening of anti-L. infantum antibodies). However, the clinical examination also showed cutaneous lesions that suggested the presence of CL. This was reinforced by the patients having a history of living or traveling to endemic foci. Thus, direct examination, culture, and PCR-RFLP (ITS1- Hae 3) were carried out on the patients’ dermal exudates. In one of the infants, CL was associated with L. infantum, while in the other it was associated with L. tropica. The infants were treated as according to the recommendations of the Ministry of Health. Both patients were cured in two months; defervescence, reduction of splenomegaly, and healing of cutaneous lesions were all observed. Conclusions: These singular patients illustrate the clinical polymorphism of CL and the necessity of updating the differential diagnosis of leukemia-like syndromes, including VL, in children living in or travelling to known endemic areas. These observations suggest a change in the Mediterranean VL phenotype that may be associated with CL

HLA-C Genotyping Reveals Haplotype C*07 as a Potential Biomarker of Late Psoriasis Onset in Moroccan Patients

Psoriasis still has an unknown etiology. Genetic predisposition shows the association between HLA-Cw6 allele and psoriasis. Although biotherapies have been proven effective in psoriasis treatment, methotrexate (MTX) is still used as a first-line systemic therapy due to its efficacy/affordability, but the differential response to MTX is mostly related to interindividual genetic variability and remains an issue. Our study aimed to analyze HLA-C allele frequencies in a sample of Moroccan psoriatic patients and assess the therapeutic response to MTX. Whole blood of 54 Moroccan psoriatic patients was collected and DNA was extracted. Patients’ HLA-C locus was genotyped by PCR-SSO. Results were analyzed with Luminex xMAP Technology and Match-it DNA Evolution 3.4. HLA-C typing results of 77 sex- and age-matched unrelated non-psoriatic healthy subjects were included. We observed no difference in the allelic distribution of HLA-C between patients and healthy controls, suggesting that none of the HLA-C alleles were significantly associated with psoriasis. Moreover, the HLA-C*07 allele was associated with a late age at disease onset (>30 years old) (p = 0.007). No statistically significant association was found between HLA-C allele expression and response to MTX, despite a higher frequency of HLA-C*06 in responders compared to non-responders. Thus, HLA-C*07 could be a biomarker of late psoriasis onset in the Moroccan population

Additional file 1: of Concomitant visceral and localized cutaneous leishmaniasis in two Moroccan infants

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