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In human cutaneous leishmaniasis (HCL) caused by Leishmania ( L. ) major , the cutaneous lesions heal spontaneously and induce a Th1-type immunity that confers solid protection against reinfection. The same holds true for the experimental leishmaniasis induced by L. major in C57BL/6 mice where residual parasites persist after spontaneous clinical cure and induce sustainable memory immune responses and resistance to reinfection. Whether residual parasites also persist in scars of cured HCL caused by L. major is still unknown. Cutaneous scars from 53 volunteers with healed HCL caused by L. major were biopsied and the tissue sample homogenates were analyzed for residual parasites by four methods: i) microscope detection of amastigotes, ii) parasite culture by inoculation on biphasic medium, iii) inoculation of tissue exctracts to the footpad of BALB/c mice, an inbred strain highly susceptible to L. major , and iv) amplification of parasite kDNA by a highly sensitive real-time PCR (RT-PCR). Our results show that the scars of healed lesions of HCL caused by L. major do not contain detectable residual parasites, suggesting that this form likely induces a sterile cure at least within the scars. This feature contrasts with other Leishmania species causing chronic, diffuse, or recidivating forms of leishmaniasis where parasites do persist in healed lesions. The possibility that alternative mechanisms to parasite persistence are needed to boost and maintain long-term immunity to L. major , should be taken into consideration in vaccine development against L. major infection., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Sghaier, Benhnini, Guerfali, Attia, Bali, Zaatour, Mkannez, Gharbi, Belhaj-Hamida, Dridi, Ben-Salah, Dellagi and Laouini.)

Zoonotic cutaneous leishmaniasis caused by Leishmania (L.) major parasites affects urban and suburban areas in the center and south of Tunisia where the disease is endemo-epidemic. Several cases were reported in human patients for which infection due to L. major induced lesions with a broad range of severity. However, very little is known about the mechanisms underlying this diversity. Our hypothesis is that parasite genomic variability could, in addition to the host immunological background, contribute to the intra-species clinical variability observed in patients and explain the lesion size differences observed in the experimental model. Based on several epidemiological, in vivo and in vitro experiments, we focused on two clinical isolates showing contrasted severity in patients and BALB/c experimental mice model. We used DNA-seq as a high-throughput technology to facilitate the identification of genetic variants with discriminating potential between both isolates. Our results demonstrate that various levels of heterogeneity could be found between both L. major isolates in terms of chromosome or gene copy number variation (CNV), and that the intra-species divergence could surprisingly be related to single nucleotide polymorphisms (SNPs) and Insertion/Deletion (InDels) events. Interestingly, we particularly focused here on genes affected by both types of variants and correlated them with the observed gene CNV. Whether these differences are sufficient to explain the severity in patients is obviously still open to debate, but we do believe that additional layers of -omic information is needed to complement the genomic screen in order to draw a more complete map of severity determinants., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Components of Pelagia noctiluca (P. noctiluca) venom were evaluated for their anticancer and nitric Oxide (NO) inhibition activities. Three fractions, out of four, obtained by gel filtration on Sephadex G75 of P. noctiluca venom revealed an important selective anti-proliferative activity on several cell lines such as human bladder carcinoma (RT112), human glioblastoma (U87), and human myelogenous leukemia (K562) but not on mitogen-stimulated peripheral blood mononuclear cells. Interestingly, P. noctiluca components showed an important dose-dependent anti-inflammatory activity, through inhibition of NO production via transcriptional regulation of Inducible NO Synthase (iNOS), in IFN-γ/LPS stimulated RAW 264.7 macrophages. These data strongly suggest that P. noctiluca venom could be used as a natural inhibitor of cancer cell lines and a potent anti-inflammatory agent for the treatment of anti-inflammatory diseases., (Copyright © 2016 Elsevier Masson SAS. All rights reserved.)

Synthesized lipophilic tyrosyl ester derivatives with increasing lipophilicity were effective against Leishmania (L.) major and Leishmania infantum species in vitro. These findings prompted us to test in vivo leishmanicidal properties of these molecules and their potential effect on the modulation of immune responses. The experimental BALB/c model of cutaneous leishmaniasis was used in this study. Mice were infected with L. major parasites and treated with three in vitro active tyrosyl esters derivatives. Among these tested tyrosylcaprate (TyC) compounds, only TyC10 exhibited an in vivo anti-leishmanial activity, when injected sub-cutaneously (s.c.). TyC10 treatment of L. major-infected BALB/c mice resulted in a decrease of lesion development and parasite load. TyC10 s.c. treatment of non-infected mice induced an imbalance in interferon γ/interleukin 4 (IFN-γ/IL-4) ratio cytokines towards a Th1 response. Our results indicate that TyC10 s.c. treatment improves lesions' healing and parasite clearance and may act on the cytokine balance towards a Th1 protective response by decreasing IL-4 and increasing IFN-γ transcripts. TyC10 is worthy of further investigation to uncover its mechanism of action that could lead to consider this molecule as a potential drug candidate.

Tunisia is endemic for zoonotic cutaneous leishmaniasis (ZCL), a parasitic disease caused by Leishmania (L.) major. ZCL displays a wide clinical polymorphism, with severe forms present more frequently in emerging foci where naive populations are dominant. In this study, we applied the multi-locus microsatellite typing (MLMT) using ten highly informative and discriminative markers to investigate the genetic structure of 35 Tunisian Leishmania (L.) major isolates collected from patients living in five different foci of Central Tunisia (two old and three emerging foci). Phylogenetic reconstructions based on genetic distances showed that nine of the ten tested loci were homogeneous in all isolates with homozygous alleles, whereas one locus (71AT) had a 58/64-bp bi-allelic profile with an allele linked to emerging foci. Promastigote-stage parasites with the 58-bp allele tend to be more resistant to in vitro complement lysis. These results, which stress the geographical dependence of the genetic micro-heterogeneity, may improve our understanding of the ZCL epidemiology and clinical outcome., (Copyright © 2016 Elsevier B.V. All rights reserved.)

In the present study, the potential effect anti tumor and the chemical composition of different fractions of Gokshur was evaluated. Commonly known as puncture vine, it has been used for a long time in both the Indian and traditional Chinese medicine. It is popularly used as a remedy for fertility disorder in Ayurveda. Samples were collected during June-September 2014 in the Cap Bon (north east of the northern Tunisia). Different organs were separated and extracted by sequential process to compare and investigate their potential anti-tumor effect. For the first time, we report the antiproliferatif effect of leaves n-butannolic fraction against cancer cell lines. The better anti-tumor fraction (94.76±1.52%) has been detected and performed by RP-HPLC has shown a great peak area (5578.21Mau). Novel designed natural derivatives from Gokshur, a cyclotrisiloxane, major compound identified by GC-MS., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Background: Leishmania (L.) are intracellular protozoan parasites able to survive and replicate in the hostile phagolysosomal environment of infected macrophages. They cause leishmaniasis, a heterogeneous group of worldwide-distributed affections, representing a paradigm of neglected diseases that are mainly embedded in impoverished populations. To establish successful infection and ensure their own survival, Leishmania have developed sophisticated strategies to subvert the host macrophage responses. Despite a wealth of gained crucial information, these strategies still remain poorly understood. MicroRNAs (miRNAs), an evolutionarily conserved class of endogenous 22-nucleotide non-coding RNAs, are described to participate in the regulation of almost every cellular process investigated so far. They regulate the expression of target genes both at the levels of mRNA stability and translation; changes in their expression have a profound effect on their target transcripts., Methodology/principal Findings: We report in this study a comprehensive analysis of miRNA expression profiles in L. major-infected human primary macrophages of three healthy donors assessed at different time-points post-infection (three to 24 h). We show that expression of 64 out of 365 analyzed miRNAs was consistently deregulated upon infection with the same trends in all donors. Among these, several are known to be induced by TLR-dependent responses. GO enrichment analysis of experimentally validated miRNA-targeted genes revealed that several pathways and molecular functions were disturbed upon parasite infection. Finally, following parasite infection, miR-210 abundance was enhanced in HIF-1α-dependent manner, though it did not contribute to inhibiting anti-apoptotic pathways through pro-apoptotic caspase-3 regulation., Conclusions/significance: Our data suggest that alteration in miRNA levels likely plays an important role in regulating macrophage functions following L. major infection. These results could contribute to better understanding of the dynamics of gene expression in host cells during leishmaniasis.

In this study, biological activities of methanolic extracts from Artemisia herba-alba, Ruta chalpensis L. and Peganum harmala L. plants, collected in Centre of Tunisia, were investigated. Results showed an important phenolic composition of Artemisia herba-alba (123.95±4.3g GAE/kg of dry mass). The extract of this plant showed, using different antioxidant assays (DPPH, ABTS and AAPH/linoleic acid methods) and an IFN-γ/LPS induced RAW 264.7 murine macrophages' assay, the highest antioxidant (IC50 (DPPH assay) 20.64±0.84mg/L) and anti-inflammatory (72% inhibition at 150mg/L) activities, respectively. Excepting Peganum harmala L. extract, the two other extracts showed a high anticancer activity against several cell lines (human bladder carcinoma RT112, human laryngeal carcinoma Hep2 and human myelogenous leukemia K562), for A. herba-laba IC50=81.59±4.4, 59.05±3.66 and 90.96mg/L respectively, but not on normal peripheral blood mononuclear cells. All these biological activities are well correlated with the phenolic contents of these extracts. These findings demonstrate the remarkable potential of these plants as valuable source of antioxidants with exhibit original and interesting anti-inflammatory and anticancer capacities., (Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.)

In this study, we tested 10 essential oils (EOs) extracted from 10 plants issued from Sned region (Tunisia) to evaluate both their leishmanicidal effects against Leishmania major and L. infantum, and their cytotoxicity against murine macrophage cell line RAW 264.7 (ATCC, TIB-71). The antioxidant activity was also monitored by the DDPH method, while the chemical composition of active EO was assessed by GC-MS analysis. The results showed that the EOs obtained from Thymus hirtus sp. algeriensis (rich on monoterpenoids, especially linalool at 17.62% and camphor at 13.82%) is significantly active against both L. major and L. infantum, whereas Ruta chalepensis EO (rich on 2-undecanone at 84.28%) is only active against L. infantum. Both oil extracts showed low cytotoxicity towards murine macrophages. The characteristic ratios (IC₈₀ Raw264.7 cells/IC₅₀ L. infantum and IC₈₀ Raw264.7 cells/IC₅₀ L. major) were, respectively, 2.7 and 1.57 for T. hirtus sp. algeriensis, and 1.34 and 0.19 for R. chalepensis. However, when measuring the antioxidant effects (DDPH method), the two latter EOs presented a moderate 2,2-diphenyl-2-picrylhydrazyl hydrate scavenging effects compared to EOs from Eucaliptus globulus, Pinus halepensis, Pituranthos tortuosus, Rosmarinus officinalis, Tetraclinis articulata or to BHT.

It is well established that MHC class II restricted-CD4 T cells are dominant during the development of immunity against Leishmania (L) in the C57BL/6-resistant mouse strain. However and in agreement with a number of previous observations indicating that specific CD8 T cells are primed during natural infection or vaccination in humans, a great deal of evidence obtained recently with the susceptible BALB/c murine model of infection by Leishmania major indicates that CD8 T cells participate in both pathogenesis and immunity to cutaneous leishmaniasis. Our goal herein was to identify in silico all parasitic peptides present in the whole L. major predicted proteome, using several public computational systems for the prediction of peptide binding to all MHC (histocompatibility complex-2) molecules in BALB/c and C57BL/6 mice (Syfpeithi, Rankpep, PRED(BALB/c) and Bimas). Peptides that were predicted to bind to different H2 molecules were then analysed for their homology with any of the murine proteins annotated so far, using the BLAST algorithm. Sets of selected peptides for each H2 molecule were defined by different prediction systems and compared to each other. Surprisingly, the results showed that a higher number of L. major peptides were predicted to bind H2 BALB/c molecules and very few or none to bind H2 C57BL/6 molecules. Our finding illustrates how a hybrid immuno-computational approach may be useful for biologists to target an in silico set of selected proteins to define potential candidate antigens for experimental vaccination with greater accuracy as well as a reduced number of T cell antigens.