Fatty Acid Profiles of Leishmania major Derived from Human and Rodent Hosts in Endemic Cutaneous Leishmaniasis Areas of Tunisia and Algeria

Leishmaniasis is a protozoal vector-borne disease that affects both humans and animals. In the Mediterranean Basin, the primary reservoir hosts of Leishmania spp. are mainly rodents and canids. Lipidomic approaches have allowed scientists to establish Leishmania spp. lipid profiles for the identification of cell stage specific biomarkers, drug mechanisms of action, and host immune response. Using an in silico approach of global network interaction between genes involved in fatty acid (FA) synthesis followed by the GC-MS approach, we were able to characterize the fatty acid profiles of L. major derived from human and rodent hosts. Our results revealed that the lipid profile of L. major showed similarities and differences with those already reported for other Leishmania species. Phospholipids are the predominant lipid class. FA composition of rodent parasites was characterized by a lower abundance of the precursor C18:2(n-6). One of the rodent clones, which also expressed the lowest lipid abundance in PL and TAG, was the least sensitive clone to the miltefosine drug and has the lowest infection efficiency. Our findings suggest that the lipid composition variation may explain the response of the parasite toward treatment and their ability to infect their host.

High genome plasticity and frequent genetic exchange in Leishmania tropica isolates from Afghanistan, Iran and Syria

Background The kinetoplastid protozoan Leishmania tropica mainly causes cutaneous leishmaniasis in humans in the Middle East, and relapse or treatment failure after treatment are common in this area. L. tropica’s digenic life cycle includes distinct stages in the vector sandfly and the mammalian host. Sexual reproduction and genetic exchange appear to occur more frequently than in other Leishmania species. Understanding these processes is complicated by chromosome instability during cell division that yields aneuploidy, recombination and heterozygosity. This combination of rare recombination and aneuploid permits may reveal signs of hypothetical parasexual mating, where diploid cells fuse to form a transient tetraploid that undergoes chromosomal recombination and gradual chromosomal loss. Methodology/principal findings The genome-wide SNP diversity from 22 L. tropica isolates showed chromosome-specific runs of patchy heterozygosity and extensive chromosome copy number variation. All these isolates were collected during 2007–2017 in Sweden from patients infected in the Middle East and included isolates from a patient possessing two genetically distinct leishmaniasis infections three years apart with no evidence of re-infection. We found differing ancestries on the same chromosome (chr36) across multiple samples: matching the reference genome with few derived alleles, followed by blocks of heterozygous SNPs, and then by clusters of homozygous SNPs with specific recombination breakpoints at an inferred origin of replication. Other chromosomes had similar marked changes in heterozygosity at strand-switch regions separating polycistronic transcriptional units. Conclusion/significance These large-scale intra- and inter-chromosomal changes in diversity driven by recombination and aneuploidy suggest multiple mechanisms of cell reproduction and diversification in L. tropica, including mitotic, meiotic and parasexual processes. It underpins the need for more genomic surveillance of Leishmania, to detect emerging hybrids that could spread more widely and to better understand the association between genetic variation and treatment outcome. Furthering our understanding of Leishmania genome evolution and ancestry will aid better diagnostics and treatment for cutaneous leishmaniasis caused by L.tropica in the Middle East.

2,3-Dihydroquinazolin-4(1H)-one as a New Class of Anti-Leishmanial Agents: A Combined Experimental and Computational Study

Leishmaniasis is a neglected parasitic disease caused by various Leishmania species. The discovery of new protozoa drugs makes it easier to treat the disease; but, conventional clinical issues like drug resistance, cumulative toxicity, and target selectivity are also getting attention. So, there is always a need for new therapeutics to treat Leishmaniasis. Here, we have reported 2,3-dihydroquinazolin-4(1H)-one derivative as a new class of anti-leishmanial agents. Two derivatives, 3a (6,8-dinitro-2,2-disubstituted-2,3-dihydroquinazolin-4(1H)-ones) and 3b (2-(4-chloro-3-nitro-phenyl)-2-methyl-6,8-dinitro-2,3-dihydro-1H-quinazolin-4-one) were prepared that show promising in silico anti-leishmanial activities. Molecular docking was performed against the Leishmanial key proteins including Pyridoxal Kinase and Trypanothione Reductase. The stability of the ligand-protein complexes was further studied by 100 ns MD simulations and MM/PBSA calculations for both compounds. 3b has been shown to be a better anti-leishmanial candidate. In vitro studies also agree with the in-silico results where IC50 for 3a and 3b was 1.61 and 0.05 µg/mL, respectively.

Long-Term In Vitro Passaging Had a Negligible Effect on Extracellular Vesicles Released by Leishmania amazonensis and Induced Protective Immune Response in BALB/c Mice

Depending on Leishmania species and the presence/absence of virulence factors, Leishmania extracellular vesicles (EVs) can differently stimulate host immune cells. This work is aimed at characterizing and evaluating the protective role of EVs released by Leishmania amazonensis promastigotes under different maintenance conditions. Initially, using a control strain, we standardized 26°C as the best release temperature to obtain EVs with a potential protective role in the experimental leishmaniasis model. Then, long-term (LT-P) promastigotes of L. amazonensis were obtained after long-term in vitro culture (100 in vitro passages). In vivo-derived (IVD-P) promastigotes of L. amazonensis were selected after 3 consecutive experimental infections in BALB/c mice. Those strains developed similar lesion sizes except for IVD-P at 8 weeks post infection. No differences in EV production were detected in both strains. However, the presence of LPG between LT-P and IVD-P EVs was different. Groups of mice immunized with EVs emulsified in the adjuvant and challenged with IVD-P parasites showed decreased lesion size and parasitic load compared with the nonimmunized groups. The immunization regimen with two doses showed high IFN-γ and IgG2a titers in challenged mice with either IVD-P or LT-P EVs. IL-4 and IL-10 were detected in immunized mice, suggesting a mixed Th1/Th2 profile. EVs released by either IVD-P or LT-P induced a partial protective effect in an immunization model. Thus, our results uncover a potential protective role of EVs from L. amazonensis for cutaneous leishmaniasis. Moreover, long-term maintenance under in vitro conditions did not seem to affect EV release and their immunization properties in mice.

Effects of Leishmania Species on Immune Response against Malaria Parasite in Malaria Leishmania Coinfections

Both malaria and leishmania are most widespread protozoon parasitic diseases, certainly in tropical countries of the world. Malaria leishmania coinfection is common in leishmaniasis endemic areas which is mostly endemic to malaria too. Researchers notice that in cases of malaria leishmania coinfection , leishmania species find the some extent the outcome of malaria infection , but also behavior of malaria parasite species play a significant role to figure the consequences of it. While L. donovani protect from severe malaria complications by suppression of major histocompatibility class Ⅱ , so it diminish the clinical severity of malaria but not malaria parasite density due to dysfunction of major histocompatibility class I, which controlled by suppressed one, In another side L. mexicana tends to sequester in macrophages and lead to severe clinical outcomes when it coexisted with malaria parasite at same host. Experimental studies required to know more information about coinfection of different malaria and leishmania species to establish clinical research. Leishmania infection excluded when studies aim to assess the immune response to only malaria parasite, experimental studies required involving different species of malaria and leishmania.

Antileishmanial Drug Discovery and Development: Time to Reset the Model?

Leishmaniasis is a vector-borne parasitic disease caused by Leishmania species. The disease affects humans and animals, particularly dogs, provoking cutaneous, mucocutaneous, or visceral processes depending on the Leishmania sp. and the host immune response. No vaccine for humans is available, and the control relies mainly on chemotherapy. However, currently used drugs are old, some are toxic, and the safer presentations are largely unaffordable by the most severely affected human populations. Moreover, its efficacy has shortcomings, and it has been challenged by the growing reports of resistance and therapeutic failure. This manuscript presents an overview of the currently used drugs, the prevailing model to develop new antileishmanial drugs and its low efficiency, and the impact of deconstruction of the drug pipeline on the high failure rate of potential drugs. To improve the predictive value of preclinical research in the chemotherapy of leishmaniasis, several proposals are presented to circumvent critical hurdles—namely, lack of common goals of collaborative research, particularly in public–private partnership; fragmented efforts; use of inadequate surrogate models, especially for in vivo trials; shortcomings of target product profile (TPP) guides.

Highly diluted natural complex (M2) effects on leishmanias

Introduction: Leishmaniosis is regarded as a serious public health issue either by its magnitude, morbidity or mortality. Depending on Leishmania species disease ranges from cutaneous, which is relatively confined and controlled, until a progressive and fatal visceral disease [4]. In order to complete their lifecycle Leishmania undergo transformations in the vector digestive tract to get to metacyclic infective form [3,5]. Those include secretion of the “promastigote secretory gel” (PSG) that protect parasites from digestive enzymes. PSG acts on survival and colonization of parasites inside the vector, on transmission and infection development in mammals, and also facilitates and increases transmission [1,2]. Drugs used for leishmaniosis treatment show high toxicity and several side effects, leading patients to quit the treatment and consequently generating resistant strains. The search for new therapeutic approaches is considered a strategic research priority by the World Health Organization. Highly diluted natural products show efficacy in modifying immunological response by stimulating the immune system through macrophages activation, then favoring the organism in many pathological conditions. Aim: To assess the direct action of M2 treatment on promastigotes of different Leishmania species. Methods: 3x106 were cultured in the presence of 20% of M2 plus booster 1% doses every 24h for up to 96h at 25ºC in humidified incubator and then submitted to assays for determining mitochondrial activity by MTT (5mg/mL), cell proliferation trough cell counting on Neubauer chamber and rosettes formation (around PSG) by light microscopy. Results of 3 independent experiments were statistically analyzed using t-test. Results: M2 treatment changed mitochondrial metabolic activity in all tested Leishmania strains. L. amazonensis proliferation decreased after 96h treatment and likewise the rosettes formation (total, closed and open ones). Discussion: Our data show that Leishmania cultures treated with M2 present decreased number of rosettes and these rosettes may be slowing the production of PSG, that is typically synthesized by Leishmania during metacyclic infective phase. Rosettes are also a place for fusion between 2 or more leishmanias, a process that involves nuclear and kinetoplast genetic material exchange. Conclusion: M2 acts on L. amazonensis promastigote forms by reducing the total number of rosettes (the same for open and closed rosettes) that are related to infective form of promastigotes, which produce PSG while on rosettes. This result suggests that M2 treatment is capable of decreasing Leishmania infectivity. Although our results are preliminary, these changes open new perspectives for the disease treatment and/or prevention using M2.

Leukocyte Behavior in Mesenteric Microcirculation upon Exper-imental By Leishmania Spp. in BALB/c Mice

Background: We aimed to determine the cellular recruitment (leukocyte rolling and adhesion) by which the Leishmania (Viannia) braziliensis, L. (Leishmania) amazonensis, and L. (Leishmania) major species in the mesenteric microcirculation of BALB/c mice. Methods: Five experimental groups were considered: group 1 (L. braziliensis); group 2 (L. amazonensis); group 3 (L. major); group 4 (control group with PBS); group 5 (negative control group), analyzed 3, 6, 12, and 24 h after parasite inoculation. Results: Infections by the different Leishmania species caused an increase in the number of rolling leukocytes: L. braziliensis a peak at 6 h; L. amazonensis and L. major a peak at 3 h. The Leishmania infections induced leukocyte adhesion: L. major and L. amazonensis showed an increase after 3 and 6 h, respectively. Conclusion: The kinetics of cellular recruitment in Leishmania infections, leading to infection susceptibility or resistance, indicates that distinct mechanisms regulate the initial response to Leishmania infection and determine its course.