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1.
Sci Rep ; 14(1): 12981, 2024 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-38839916

RESUMO

Micro RNAs (miRNAs, miRs) and relevant networks might exert crucial functions during differential host cell infection by the different Leishmania species. Thus, a bioinformatic analysis of microarray datasets was developed to identify pivotal shared biomarkers and miRNA-based regulatory networks for Leishmaniasis. A transcriptomic analysis by employing a comprehensive set of gene expression profiling microarrays was conducted to identify the key genes and miRNAs relevant for Leishmania spp. infections. Accordingly, the gene expression profiles of healthy human controls were compared with those of individuals infected with Leishmania mexicana, L. major, L. donovani, and L. braziliensis. The enrichment analysis for datasets was conducted by utilizing EnrichR database, and Protein-Protein Interaction (PPI) network to identify the hub genes. The prognostic value of hub genes was assessed by using receiver operating characteristic (ROC) curves. Finally, the miRNAs that interact with the hub genes were identified using miRTarBase, miRWalk, TargetScan, and miRNet. Differentially expressed genes were identified between the groups compared in this study. These genes were significantly enriched in inflammatory responses, cytokine-mediated signaling pathways and granulocyte and neutrophil chemotaxis responses. The identification of hub genes of recruited datasets suggested that TNF, SOCS3, JUN, TNFAIP3, and CXCL9 may serve as potential infection biomarkers and could deserve value as prognostic biomarkers for leishmaniasis. Additionally, inferred data from miRWalk revealed a significant degree of interaction of a number of miRNAs (hsa-miR-8085, hsa-miR-4673, hsa-miR-4743-3p, hsa-miR-892c-3p, hsa-miR-4644, hsa-miR-671-5p, hsa-miR-7106-5p, hsa-miR-4267, hsa-miR-5196-5p, and hsa-miR-4252) with the majority of the hub genes, suggesting such miRNAs play a crucial role afterwards parasite infection. The hub genes and hub miRNAs identified in this study could be potentially suggested as therapeutic targets or biomarkers for the management of leishmaniasis.


Assuntos
Biomarcadores , Biologia Computacional , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Leishmaniose , MicroRNAs , Mapas de Interação de Proteínas , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Leishmaniose/genética , Leishmaniose/parasitologia , Biologia Computacional/métodos , Biomarcadores/metabolismo , Perfilação da Expressão Gênica/métodos , Mapas de Interação de Proteínas/genética , Transcriptoma , Leishmania/genética
2.
BMC Infect Dis ; 24(1): 551, 2024 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-38824500

RESUMO

BACKGROUND: Leishmaniasis, an illness caused by protozoa, accounts for a substantial number of human fatalities globally, thereby emerging as one of the most fatal parasitic diseases. The conventional methods employed for detecting the Leishmania parasite through microscopy are not only time-consuming but also susceptible to errors. Therefore, the main objective of this study is to develop a model based on deep learning, a subfield of artificial intelligence, that could facilitate automated diagnosis of leishmaniasis. METHODS: In this research, we introduce LeishFuNet, a deep learning framework designed for detecting Leishmania parasites in microscopic images. To enhance the performance of our model through same-domain transfer learning, we initially train four distinct models: VGG19, ResNet50, MobileNetV2, and DenseNet 169 on a dataset related to another infectious disease, COVID-19. These trained models are then utilized as new pre-trained models and fine-tuned on a set of 292 self-collected high-resolution microscopic images, consisting of 138 positive cases and 154 negative cases. The final prediction is generated through the fusion of information analyzed by these pre-trained models. Grad-CAM, an explainable artificial intelligence technique, is implemented to demonstrate the model's interpretability. RESULTS: The final results of utilizing our model for detecting amastigotes in microscopic images are as follows: accuracy of 98.95 1.4%, specificity of 98 2.67%, sensitivity of 100%, precision of 97.91 2.77%, F1-score of 98.92 1.43%, and Area Under Receiver Operating Characteristic Curve of 99 1.33. CONCLUSION: The newly devised system is precise, swift, user-friendly, and economical, thus indicating the potential of deep learning as a substitute for the prevailing leishmanial diagnostic techniques.


Assuntos
Aprendizado Profundo , Leishmania , Leishmaniose , Microscopia , Telemedicina , Humanos , Leishmaniose/parasitologia , Leishmaniose/diagnóstico , Leishmania/isolamento & purificação , Microscopia/métodos , COVID-19 , SARS-CoV-2/isolamento & purificação
3.
J Cell Biol ; 223(9)2024 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-38829962

RESUMO

Two sets of motor proteins underpin motile cilia/flagella function. The axoneme-associated inner and outer dynein arms drive sliding of adjacent axoneme microtubule doublets to periodically bend the flagellum for beating, while intraflagellar transport (IFT) kinesins and dyneins carry IFT trains bidirectionally along the axoneme. Despite assembling motile cilia and flagella, IFT train speeds have only previously been quantified in immobilized flagella-mechanical immobilization or genetic paralysis. This has limited investigation of the interaction between IFT and flagellar beating. Here, in uniflagellate Leishmania parasites, we use high-frequency, dual-color fluorescence microscopy to visualize IFT train movement in beating flagella. We discovered that adhesion of flagella to a microscope slide is detrimental, reducing IFT train speed and increasing train stalling. In flagella free to move, IFT train speed is not strongly dependent on flagella beat type; however, permanent disruption of flagella beating by deletion of genes necessary for formation or regulation of beating showed an inverse correlation of beat frequency and IFT train speed.


Assuntos
Flagelos , Leishmania , Microtúbulos , Axonema/metabolismo , Axonema/genética , Transporte Biológico , Cílios/metabolismo , Cílios/genética , Dineínas/metabolismo , Dineínas/genética , Flagelos/metabolismo , Flagelos/genética , Cinesinas/metabolismo , Cinesinas/genética , Leishmania/citologia , Leishmania/genética , Leishmania/metabolismo , Proteínas de Protozoários/metabolismo , Proteínas de Protozoários/genética , Microtúbulos/metabolismo
4.
Cell Rep ; 43(5): 114203, 2024 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-38722744

RESUMO

Leishmania is the causative agent of cutaneous and visceral diseases affecting millions of individuals worldwide. Pseudouridine (Ψ), the most abundant modification on rRNA, changes during the parasite life cycle. Alterations in the level of a specific Ψ in helix 69 (H69) affected ribosome function. To decipher the molecular mechanism of this phenotype, we determine the structure of ribosomes lacking the single Ψ and its parental strain at ∼2.4-3 Å resolution using cryo-EM. Our findings demonstrate the significance of a single Ψ on H69 to its structure and the importance for its interactions with helix 44 and specific tRNAs. Our study suggests that rRNA modification affects translation of mRNAs carrying codon bias due to selective accommodation of tRNAs by the ribosome. Based on the high-resolution structures, we propose a mechanism explaining how the ribosome selects specific tRNAs.


Assuntos
Pseudouridina , RNA de Transferência , Ribossomos , Pseudouridina/metabolismo , Ribossomos/metabolismo , RNA de Transferência/metabolismo , RNA de Transferência/genética , Leishmania/metabolismo , Leishmania/genética , Microscopia Crioeletrônica , RNA Ribossômico/metabolismo , RNA Ribossômico/química , RNA Ribossômico/genética , Conformação de Ácido Nucleico , Modelos Moleculares
5.
PLoS Negl Trop Dis ; 18(5): e0012175, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38768213

RESUMO

In Brazil, Leishmania amazonensis is the etiological agent of cutaneous and diffuse cutaneous leishmaniasis. The state of Maranhão in the Northeast of Brazil is prevalent for these clinical forms of the disease and also has high rates of HIV infection. Here, we characterized the drug susceptibility of a L. amazonensis clinical isolate from a 46-year-old man with diffuse cutaneous leishmaniasis coinfected with HIV from this endemic area. This patient underwent several therapeutic regimens with meglumine antimoniate, liposomal amphotericin B, and pentamidine, without success. In vitro susceptibility assays against promastigotes and intracellular amastigotes demonstrated that this isolate had low susceptibility to amphotericin B, when compared with the reference strain of this species that is considered susceptible to antileishmanial drugs. Additionally, we investigated whether the low in vitro susceptibility would affect the in vivo response to amphotericin B treatment. The drug was effective in reducing the lesion size and parasite burden in mice infected with the reference strain, whereas those infected with the clinical isolate and a resistant line (generated experimentally by stepwise selection) were refractory to amphotericin B treatment. To evaluate whether the isolate was intrinsically resistant to amphotericin B in animals, infected mice were treated with other drugs that had not been used in the treatment of the patient (miltefosine, paromomycin, and a combination of both). Our findings demonstrated that all drug schemes were able to reduce lesion size and parasite burden in animals infected with the clinical isolate, confirming the amphotericin B-resistance phenotype. These findings indicate that the treatment failure observed in the patient may be associated with amphotericin B resistance, and demonstrate the potential emergence of amphotericin B-resistant L. amazonensis isolates in an area of Brazil endemic for cutaneous leishmaniasis.


Assuntos
Anfotericina B , Antiprotozoários , Resistência a Medicamentos , Anfotericina B/farmacologia , Anfotericina B/uso terapêutico , Animais , Brasil , Pessoa de Meia-Idade , Antiprotozoários/farmacologia , Antiprotozoários/uso terapêutico , Humanos , Masculino , Camundongos , Leishmania/efeitos dos fármacos , Leishmania/isolamento & purificação , Leishmania/classificação , Leishmania mexicana/efeitos dos fármacos , Leishmania mexicana/isolamento & purificação , Leishmaniose Cutânea/tratamento farmacológico , Leishmaniose Cutânea/parasitologia , Infecções por HIV/complicações , Infecções por HIV/tratamento farmacológico , Testes de Sensibilidade Parasitária , Camundongos Endogâmicos BALB C , Leishmaniose Tegumentar Difusa/parasitologia , Leishmaniose Tegumentar Difusa/tratamento farmacológico
6.
Microb Pathog ; 192: 106721, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38815776

RESUMO

Ticks play an important role in the transmission of parasitic diseases, especially pathogenic protozoa in canine hosts, and it is very important to determine the role and extent of their infection with these pathogens in order to determine important control strategies. This study assessed the molecular prevalence of three protozoan pathogens including Hepatozoon canis, Leishmania spp. and Babesia spp., in ticks using PCR. A total 300 stray dogs were investigated and 691 ticks (171 male, 377 female and 143 nymph) were detected directly from 45 infested dogs. Species, stage of growth, and gender were determined for each tick. DNA extracted from 224 ticks (26 male, 165 female and 33 nymph). The molecular presence of three protozoan pathogens including Hepatozoon spp. (18S rRNA gene), Leishmania infantum (kinetoplastid minicircle DNA) and Babesia spp. (ssrRNA gene) were investigated using PCR method. One species of ticks, Rhipicephalus sanguineus was identified. Two of the target pathogens, Hepatozoon spp. (7/83; 8.43 %) and Babesia spp. (1/83; 1.2 %), were detected by PCR method. Sequence analysis of the ssrRNA gene of detected Babesia spp. showed a close relationship to the deposited strains of Babesia vulpis in the gene bank. To the best of our knowledge, this is the first study to undertake a phylogenetic analysis of H. canis and Babesia spp. in stray dogs in Alborz province, Iran and the first report about molecular detection of Babesia vulpis from tick infesting dogs in Iran. According to the above results, it seems necessary to implement tick control programs in dogs.


Assuntos
Babesia , Doenças do Cão , Filogenia , RNA Ribossômico 18S , Animais , Cães , Irã (Geográfico)/epidemiologia , Doenças do Cão/parasitologia , Doenças do Cão/epidemiologia , Babesia/genética , Babesia/isolamento & purificação , Babesia/classificação , Feminino , Masculino , RNA Ribossômico 18S/genética , DNA de Protozoário/genética , Reação em Cadeia da Polimerase , Rhipicephalus sanguineus/parasitologia , Carrapatos/parasitologia , Eucoccidiida/genética , Eucoccidiida/isolamento & purificação , Eucoccidiida/classificação , Leishmania infantum/genética , Leishmania infantum/isolamento & purificação , Leishmania infantum/classificação , Infestações por Carrapato/veterinária , Infestações por Carrapato/epidemiologia , Infestações por Carrapato/parasitologia , Leishmania/genética , Leishmania/classificação , Leishmania/isolamento & purificação
7.
Mol Biochem Parasitol ; 259: 111629, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38750697

RESUMO

Leishmaniases comprise a group of infectious parasitic diseases caused by various species of Leishmania and are considered a significant public health problem worldwide. Only a few medications, including miltefosine, amphotericin B, and meglumine antimonate, are used in current therapy. These medications are associated with severe side effects, low efficacy, high cost, and the need for hospital support. Additionally, there have been occurrences of drug resistance. Additionally, only a limited number of drugs, such as meglumine antimonate, amphotericin B, and miltefosine, are available, all of which are associated with severe side effects. In this context, the need for new effective drugs with fewer adverse effects is evident. Therefore, this study investigated the anti-Leishmania activity of a dichloromethane fraction (DCMF) extracted from Arrabidaea brachypoda roots. This fraction inhibited the viability of L. infantum, L. braziliensis, and L. Mexicana promastigotes, with IC50 values of 10.13, 11.44, and 11.16 µg/mL, respectively, and against L. infantum amastigotes (IC50 = 4.81 µg/mL). Moreover, the DCMF exhibited moderate cytotoxicity (CC50 = 25.15) towards RAW264.7 macrophages, with a selectivity index (SI) of 5.2. Notably, the DCMF caused damage to the macrophage genome only at 40 µg/mL, which is greater than the IC50 found for all Leishmania species. The results suggest that DCMF demonstrates similar antileishmanial effectiveness to isolated brachydin B, without causing genotoxic effects on mammalian cells. This finding is crucial because the isolation of the compounds relies on several steps and is very costly while obtaining the DCMF fraction is a simple and cost-effective process. Furthermore, In addition, the potential mechanisms of action of brachydins were also investigated. The computational analysis indicates that brachydin compounds bind to the Triosephosphate isomerase (TIM) enzyme via two main mechanisms: destabilizing the interface between the homodimers and interacting with catalytic residues situated at the site of binding. Based on all the results, DCMF exhibits promise as a therapeutic agent for leishmaniasis due to its significantly reduced toxicity in comparison to the adverse effects associated with current reference treatments.


Assuntos
Antiprotozoários , Bignoniaceae , Flavonoides , Leishmania , Simulação de Acoplamento Molecular , Extratos Vegetais , Bignoniaceae/química , Antiprotozoários/farmacologia , Antiprotozoários/química , Antiprotozoários/isolamento & purificação , Flavonoides/farmacologia , Flavonoides/química , Animais , Leishmania/efeitos dos fármacos , Leishmania/genética , Extratos Vegetais/farmacologia , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Camundongos , Concentração Inibidora 50 , Macrófagos/efeitos dos fármacos , Macrófagos/parasitologia , Células RAW 264.7
8.
Acta Trop ; 256: 107265, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38772434

RESUMO

In this present study, carried out between November 2020 and July 2023 at Londrina's University Hospital, patients with active lesions of cutaneous leishmaniasis (CL) were analyzed regarding pain perception and anatomopathological aspects of the ulcers. Pain was assessed using a numerical rating scale (NRS) to compare five patients diagnosed with CL with four control patients diagnosed with vascular skin ulcers. Histopathological evaluations were used to investigate the nociceptor neuron-Leishmania interface. Patients with CL ulcers reported less pain compared to patients with vascular ulcers (2.60 ± 2.30 and 7.25 ± 0.95, respectively, p = 0.0072). Histopathology evidenced Leishmania spp. amastigote forms nearby sensory nerve fibers in profound dermis. Schwann cells marker (S100 protein) was detected, and caspase-3 activation was not evidenced in the in the nerve fibers of CL patients' samples, suggesting absence of apoptotic activity in nerve endings. Additionally, samples taken from the active edge of the lesion were negative for bacilli acid-alcohol resistant (BAAR), which excludes concomitant leprosy, in which painless lesions are also observed. Thus, the present data unveil for the first time anatomopathological and microbiological details of painless ulcers in CL patients, which has important clinical implications for a better understanding on the intriguing painless clinical characteristic of CL.


Assuntos
Apoptose , Leishmania , Leishmaniose Cutânea , Úlcera Cutânea , Humanos , Masculino , Feminino , Leishmaniose Cutânea/patologia , Leishmaniose Cutânea/parasitologia , Adulto , Pessoa de Meia-Idade , Úlcera Cutânea/parasitologia , Úlcera Cutânea/patologia , Células Receptoras Sensoriais/patologia , Neurônios/patologia , Idoso , Pele/parasitologia , Pele/patologia , Pele/inervação
9.
PLoS Negl Trop Dis ; 18(5): e0011897, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38739677

RESUMO

Leishmania, the dixenous trypanosomatid parasites, are the causative agents of leishmaniasis currently divided into four subgenera: Leishmania, Viannia, Sauroleishmania, and the recently described Mundinia, consisting of six species distributed sporadically all over the world infecting humans and/or animals. These parasites infect various mammalian species and also cause serious human diseases, but their reservoirs are unknown. Thus, adequate laboratory models are needed to enable proper research of Mundinia parasites. In this complex study, we compared experimental infections of five Mundinia species (L. enriettii, L. macropodum, L. chancei, L. orientalis, and four strains of L. martiniquensis) in three rodent species: BALB/c mouse, Chinese hamster (Cricetulus griseus) and steppe lemming (Lagurus lagurus). Culture-derived parasites were inoculated intradermally into the ear pinnae and progress of infection was monitored for 20 weeks, when the tissues and organs of animals were screened for the presence and quantity of Leishmania. Xenodiagnoses with Phlebotomus duboscqi were performed at weeks 5, 10, 15 and 20 post-infection to test the infectiousness of the animals throughout the experiment. BALB/c mice showed no signs of infection and were not infectious to sand flies, while Chinese hamsters and steppe lemmings proved susceptible to all five species of Mundinia tested, showing a wide spectrum of disease signs ranging from asymptomatic to visceral. Mundinia induced significantly higher infection rates in steppe lemmings compared to Chinese hamsters, and consequently steppe lemmings were more infectious to sand flies: In all groups tested, they were infectious from the 5th to the 20th week post infection. In conclusion, we identified two rodent species, Chinese hamster (Cricetulus griseus) and steppe lemming (Lagurus lagurus), as candidates for laboratory models for Mundinia allowing detailed studies of these enigmatic parasites. Furthermore, the long-term survival of all Mundinia species in steppe lemmings and their infectiousness to vectors support the hypothesis that some rodents have the potential to serve as reservoir hosts for Mundinia.


Assuntos
Arvicolinae , Modelos Animais de Doenças , Leishmania , Leishmaniose , Camundongos Endogâmicos BALB C , Animais , Leishmania/classificação , Leishmaniose/parasitologia , Camundongos , Cricetinae , Arvicolinae/parasitologia , Cricetulus , Feminino
10.
Chem Biol Drug Des ; 103(5): e14535, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38772877

RESUMO

Despite efforts, available alternatives for the treatment of leishmaniasis are still scarce. In this work we tested a class of 15 quinolinylhydrazone analogues and presented data that support the use of the most active compound in cutaneous leishmaniasis caused by Leishmania amazonensis. In general, the compounds showed activity at low concentrations for both parasitic forms (5.33-37.04 µM to promastigotes, and 14.31-61.98 µM to amastigotes). In addition, the best compound (MHZ15) is highly selective for the parasite. Biochemical studies indicate that the treatment of promastigotes with MHZ15 leads the loss of mitochondrial potential and increase in ROS levels as the primary effects, which triggers accumulation of lipid droplets, loss of plasma membrane integrity and apoptosis hallmarks, including DNA fragmentation and phosphatidylserine exposure. These effects were similar in the intracellular form of the parasite. However, in this parasitic form there is no change in plasma membrane integrity in the observed treatment time, which can be attributed to metabolic differences and the resilience of the amastigote. Also, ultrastructural changes such as vacuolization suggesting autophagy were observed. The in vivo effectiveness of MHZ15 in the experimental model of cutaneous leishmaniasis was carried out in mice of the BALB/c strain infected with L. amazonensis. The treatment by intralesional route showed that MHZ15 acted with great efficiency with significantly reduction in the parasite load in the injured paws and draining lymph nodes, without clinical signs of distress or compromise of animal welfare. In vivo toxicity was also evaluated and null alterations in the levels of hepatic enzymes aspartate aminotransferase, and alanine aminotransferase was observed. The data presented herein demonstrates that MHZ15 exhibits a range of favorable characteristics conducive to the development of an antileishmanial agent.


Assuntos
Apoptose , Hidrazonas , Leishmaniose Cutânea , Camundongos Endogâmicos BALB C , Mitocôndrias , Animais , Apoptose/efeitos dos fármacos , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Hidrazonas/farmacologia , Hidrazonas/química , Leishmaniose Cutânea/tratamento farmacológico , Leishmaniose Cutânea/parasitologia , Antiprotozoários/farmacologia , Antiprotozoários/química , Antiprotozoários/uso terapêutico , Leishmania/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Feminino , Leishmania mexicana/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos
11.
Sci Rep ; 14(1): 11575, 2024 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-38773273

RESUMO

Leishmaniasis is a disease caused by a protozoan of the genus Leishmania, affecting millions of people, mainly in tropical countries, due to poor social conditions and low economic development. First-line chemotherapeutic agents involve highly toxic pentavalent antimonials, while treatment failure is mainly due to the emergence of drug-resistant strains. Leishmania arginase (ARG) enzyme is vital in pathogenicity and contributes to a higher infection rate, thus representing a potential drug target. This study helps in designing ARG inhibitors for the treatment of leishmaniasis. Py-CoMFA (3D-QSAR) models were constructed using 34 inhibitors from different chemical classes against ARG from L. (L.) amazonensis (LaARG). The 3D-QSAR predictions showed an excellent correlation between experimental and calculated pIC50 values. The molecular docking study identified the favorable hydrophobicity contribution of phenyl and cyclohexyl groups as substituents in the enzyme allosteric site. Molecular dynamics simulations of selected protein-ligand complexes were conducted to understand derivatives' interaction modes and affinity in both active and allosteric sites. Two cinnamide compounds, 7g and 7k, were identified, with similar structures to the reference 4h allosteric site inhibitor. These compounds can guide the development of more effective arginase inhibitors as potential antileishmanial drugs.


Assuntos
Arginase , Inibidores Enzimáticos , Leishmania , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Arginase/antagonistas & inibidores , Arginase/química , Arginase/metabolismo , Leishmania/enzimologia , Leishmania/efeitos dos fármacos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Relação Quantitativa Estrutura-Atividade , Proteínas de Protozoários/antagonistas & inibidores , Proteínas de Protozoários/química , Proteínas de Protozoários/metabolismo , Sítio Alostérico , Antiprotozoários/farmacologia , Antiprotozoários/química , Domínio Catalítico
12.
Mem Inst Oswaldo Cruz ; 119: e230243, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38775551

RESUMO

BACKGROUND: Leishmania tarentolae is a non-pathogenic species found in lizards representing an important model for Leishmania biology. However, several aspects of this Sauroleishmania remain unknown to explain its low level of virulence. OBJECTIVES: We reported several aspects of L. tarentolae biology including glycoconjugates, proteolytic activities and metabolome composition in comparison to pathogenic species (Leishmania amazonensis, Leishmania braziliensis, Leishmania infantum and Leishmania major). METHODS: Parasites were cultured for extraction and purification of lipophosphoglycan (LPG), immunofluorescence probing with anti-gp63 and resistance against complement. Parasite extracts were also tested for proteases activity and metabolome composition. FINDINGS: Leishmania tarentolae does not express LPG on its surface. It expresses gp63 at lower levels compared to pathogenic species and, is highly sensitive to complement-mediated lysis. This species also lacks intracellular/extracellular activities of proteolytic enzymes. It has metabolic differences with pathogenic species, exhibiting a lower abundance of metabolites including ABC transporters, biosynthesis of unsaturated fatty acids and steroids, TCA cycle, glycine/serine/threonine metabolism, glyoxylate/dicarboxylate metabolism and pentose-phosphate pathways. MAIN CONCLUSIONS: The non-pathogenic phenotype of L. tarentolae is associated with alterations in several biochemical and molecular features. This reinforces the need of comparative studies between pathogenic and non-pathogenic species to elucidate the molecular mechanisms of virulence during host-parasite interactions.


Assuntos
Glicoconjugados , Leishmania , Metaboloma , Peptídeo Hidrolases , Leishmania/enzimologia , Peptídeo Hidrolases/metabolismo , Animais , Glicoesfingolipídeos/metabolismo , Proteínas do Sistema Complemento
13.
Acta Trop ; 255: 107238, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38710262

RESUMO

Lutzomyia longipalpis is the primary vector of Leishmania infantum in the Americas and a permissive vector for Leishmania amazonensis. Previous studies showed that Leishmania infantum-infected hosts can release different volatile organic compounds (VOCs) compared with uninfected hosts, presenting a higher attractiveness to vectors. In this study, we aimed to evaluate a possible effect of L. amazonensis infection of golden hamsters in three parameters: attractiveness to Lu. longipalpis females; blood volume ingested by sand fly females; and VOCs released by the animals.. Attractiveness was measured indirectly by the number of Lu. longipalpis females that blood fed in each L. amazonensis-infected and uninfected animal. For VOCs extraction, solid phase micro extraction fibers were used, which were analyzed by gas chromatography-mass spectrometry. Behavioral trials did not show any effect of L. amazonensis infection on the attraction of sand flies nor difference on blood meal rates of Lu. longipalpis fed in both goups of hamsters. Additionally, there was no difference between the VOCs profiles of L. amazonensis-infected or uninfected hamsters.


Assuntos
Insetos Vetores , Mesocricetus , Psychodidae , Compostos Orgânicos Voláteis , Animais , Psychodidae/parasitologia , Psychodidae/fisiologia , Compostos Orgânicos Voláteis/análise , Feminino , Cricetinae , Insetos Vetores/parasitologia , Insetos Vetores/fisiologia , Leishmania mexicana , Comportamento Alimentar , Cromatografia Gasosa-Espectrometria de Massas , Leishmania/fisiologia
14.
PLoS One ; 19(5): e0303686, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38781128

RESUMO

The intracellular protozoan parasite Leishmania causes leishmaniasis in humans, leading to serious illness and death in tropical and subtropical areas worldwide. Unfortunately, due to the unavailability of approved vaccines for humans and the limited efficacy of available drugs, leishmaniasis is on the rise. A comprehensive understanding of host-pathogen interactions at the molecular level could pave the way to counter leishmaniasis. There is growing evidence that several intracellular pathogens target RNA interference (RNAi) pathways in host cells to facilitate their persistence. The core elements of the RNAi system are complexes of Argonaute (Ago) proteins with small non-coding RNAs, also known as RNA-induced silencing complexes (RISCs). Recently, we have shown that Leishmania modulates Ago1 protein of host macrophages for its survival. In this study, we biochemically characterize the Ago proteins' interactome in Leishmania-infected macrophages compared to non-infected cells. For this, a quantitative proteomic approach using stable isotope labelling by amino acids in cell culture (SILAC) was employed, followed by purification of host Ago-complexes using a short TNRC6 protein-derived peptide fused to glutathione S-transferase beads as an affinity matrix. Proteomic-based detailed biochemical analysis revealed Leishmania modulated host macrophage RISC composition during infection. This analysis identified 51 Ago-interacting proteins with a broad range of biological activities. Strikingly, Leishmania proteins were detected as part of host Ago-containing complexes in infected cells. Our results present the first report of comprehensive quantitative proteomics of Ago-containing complexes isolated from Leishmania-infected macrophages and suggest targeting the effector complex of host RNAi machinery. Additionally, these results expand knowledge of RISC in the context of host-pathogen interactions in parasitology in general.


Assuntos
Proteínas Argonautas , Macrófagos , Proteínas Argonautas/metabolismo , Proteínas Argonautas/genética , Humanos , Macrófagos/parasitologia , Macrófagos/metabolismo , Proteômica/métodos , Leishmania/metabolismo , Interferência de RNA , Leishmaniose/parasitologia , Leishmaniose/metabolismo
15.
PLoS One ; 19(5): e0302567, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38781235

RESUMO

This study investigated the sand fly fauna of the municipality Iguatama, in the Midwest Region of Minas Gerais state, Brazil, including Leishmania infection rates and blood meal sources. Sand flies were collected during four periods over the course of a single year, encompassing both dry and rainy seasons, using CDC light traps placed in peridomiciles where dogs were seropositive for visceral leishmaniasis (VL). A total of 762 sand fly specimens, representing 12 species across seven genera, were collected. Lutzomyia longipalpis was the most abundant species, comprising 57.6% of the collected specimens, followed by Nyssomyia neivai (19.6%) and Nyssomyia whitmani (10.5%). Species richness and diversity varied among collection periods, with the highest diversity observed in January 2019. Molecular analysis detected Leishmania DNA in 12.5% of the sand fly specimens, with Le. infantum being the predominant species. Blood meal analysis revealed feeding on multiple vertebrate species, including humans, rats, dogs, and chickens. The presence of Leishmania DNA in sand flies, and the identification of human blood meals, highlight the potential role of these species in VL transmission. These findings underscore the importance of continued surveillance and control measures to prevent the spread of VL and reduce transmission risk in the region.


Assuntos
Insetos Vetores , Leishmania , Psychodidae , Animais , Brasil/epidemiologia , Psychodidae/parasitologia , Leishmania/isolamento & purificação , Leishmania/genética , Cães , Humanos , Insetos Vetores/parasitologia , Leishmaniose Visceral/transmissão , Leishmaniose Visceral/epidemiologia , Leishmaniose Visceral/parasitologia , Leishmaniose Visceral/veterinária , Ratos , Galinhas/parasitologia , Comportamento Alimentar , Biodiversidade
16.
An Acad Bras Cienc ; 96(2): e20230375, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38747836

RESUMO

In pursuit of potential agents to treat Chagas disease and leishmaniasis, we report the design, synthesis, and identification novel naphthoquinone hydrazide-based molecular hybrids. The compounds were subjected to in vitro trypanocide and leishmanicidal activities. N'-(1,4-Dioxo-1,4-dihydronaphthalen-2-yl)-3,5-dimethoxybenzohydrazide (13) showed the best performance against Trypanosoma cruzi (IC50 1.83 µM) and Leishmania amazonensis (IC50 9.65 µM). 4-Bromo-N'-(1,4-dioxo-1,4-dihydronaphthalen-2-yl)benzohydrazide (16) exhibited leishmanicidal activity (IC50 12.16 µM). Regarding trypanocide activity, compound 13 was low cytotoxic to LLC-MK2 cells (SI = 95.28). Furthermore, through molecular modeling studies, the cysteine proteases cruzain, rhodesain and CPB2.8 were identified as the potential biological targets.


Assuntos
Desenho de Fármacos , Hidrazinas , Leishmania , Naftoquinonas , Tripanossomicidas , Trypanosoma cruzi , Naftoquinonas/farmacologia , Naftoquinonas/química , Naftoquinonas/síntese química , Trypanosoma cruzi/efeitos dos fármacos , Tripanossomicidas/farmacologia , Tripanossomicidas/síntese química , Tripanossomicidas/química , Leishmania/efeitos dos fármacos , Hidrazinas/química , Hidrazinas/farmacologia , Animais , Antiprotozoários/farmacologia , Antiprotozoários/síntese química , Antiprotozoários/química , Testes de Sensibilidade Parasitária , Concentração Inibidora 50 , Relação Estrutura-Atividade , Cisteína Endopeptidases
17.
Drug Dev Res ; 85(3): e22194, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38704828

RESUMO

The aim the present study was to investigate the impact of novel pentavalent organobismuth and organoantimony complexes on membrane integrity and their interaction with DNA, activity against Sb(III)-sensitive and -resistant Leishmania strains and toxicity in mammalian peritoneal macrophages. Ph3M(L)2 type complexes were synthesized, where M = Sb(V) or Bi(V) and L = deprotonated 3-(dimethylamino)benzoic acid or 2-acetylbenzoic acid. Both organobismuth(V) and organoantimony(V) complexes exhibited efficacy at micromolar concentrations against Leishmania amazonensis and L. infantum but only the later ones demonstrated biocompatibility. Ph3Sb(L1)2 and Ph3Bi(L1)2 demonstrated distinct susceptibility profiles compared to inorganic Sb(III)-resistant strains of MRPA-overexpressing L. amazonensis and AQP1-mutated L. guyanensis. These complexes were able to permeate the cell membrane and interact with the Leishmania DNA, suggesting that this effect may contribute to the parasite growth inhibition via apoptosis. Taken altogether, our data substantiate the notion of a distinct mechanism of uptake pathway and action in Leishmania for these organometallic complexes, distinguishing them from the conventional inorganic antimonial drugs.


Assuntos
Antimônio , Antiprotozoários , Membrana Celular , Resistência a Medicamentos , Compostos Organometálicos , Antimônio/farmacologia , Antimônio/química , Animais , Compostos Organometálicos/farmacologia , Camundongos , Membrana Celular/efeitos dos fármacos , Antiprotozoários/farmacologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/parasitologia , Leishmania/efeitos dos fármacos , DNA de Protozoário , Leishmania infantum/efeitos dos fármacos , Leishmania infantum/genética , Camundongos Endogâmicos BALB C
18.
Medicine (Baltimore) ; 103(18): e38039, 2024 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-38701291

RESUMO

As a result of increasing drug resistance, crossover resistance development, prolonged therapy, and the absence of different agents with innovative methods for implementation, the efficacy of recent antileishmanial medications is severely declining. So, it is vital to look for other medications from botanical remedies that have antileishmanial activity. The latex of Euphorbia abyssinica (E abyssinica) and the leaves of Clematis simensis fresen (C simensis) were macerated in methanol (80%). In vitro antileishmanial activity of the preparation was tried on promastigotes of Leishmania aethiopica (L aethiopica) and Leishmania donovani (L donovani) using resazurin assay, and fluorescence intensity was measured. One percent of dimethyl sulfoxide (DMSO) and media as negative control and amphotericin B as positive control were used. Additionally, hemolytic & phytochemical tests of the preparation were done. The mean and standard errors of each extract were evaluated and interpreted for statistical significance using one-way analysis of variance. From sigmoidal dose-response curves of % inhibition, half maximal inhibitory concentration (IC50) values were determined by GraphPad Prism and Microsoft Excel; outcomes were presented as mean ±â€…standard error of mean of triplicate trials. P < .05 was statistical significance. The phytochemical screening of C simensis and E abyssinica confirmed the existence of steroids, phenols, tannins, saponins, alkaloids, terpenoids, flavonoids and glycosides. C simensis possesses antileishmanial activity with IC50 outcomes of 46.12 ±â€…0.03 and 8.18 ±â€…0.10 µg/mL on the promastigotes of L aethiopica and L donovani, respectively. However, E abyssinica showed stronger activity with IC50 outcomes of 16.07 ±â€…0.05 µg/mL and 4.82 ±â€…0.07 µg/mL on L aethiopica and L donovani, respectively. C simensis and E abyssinica have a less hemolytic effect on human red blood cells at low concentrations. The outcomes from this investigation demonstrated that the preparation of C simensis and E abyssinica indicated significant antileishmanial activity. Therefore, further in vivo assessment of antileishmanial, cytotoxicity activity and quantitative identification of secondary metabolites are highly recommended.


Assuntos
Antiprotozoários , Euphorbia , Látex , Extratos Vegetais , Folhas de Planta , Extratos Vegetais/farmacologia , Euphorbia/química , Látex/farmacologia , Látex/química , Antiprotozoários/farmacologia , Folhas de Planta/química , Humanos , Leishmania donovani/efeitos dos fármacos , Concentração Inibidora 50 , Leishmania/efeitos dos fármacos , Metanol , Solventes , Hemólise/efeitos dos fármacos
19.
Molecules ; 29(10)2024 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-38792079

RESUMO

Infectious diseases caused by trypanosomatids, including African trypanosomiasis (sleeping sickness), Chagas disease, and different forms of leishmaniasis, are Neglected Tropical Diseases affecting millions of people worldwide, mainly in vulnerable territories of tropical and subtropical areas. In general, current treatments against these diseases are old-fashioned, showing adverse effects and loss of efficacy due to misuse or overuse, thus leading to the emergence of resistance. For these reasons, searching for new antitrypanosomatid drugs has become an urgent necessity, and different metabolic pathways have been studied as potential drug targets against these parasites. Considering that trypanosomatids possess a unique redox pathway based on the trypanothione molecule absent in the mammalian host, the key enzymes involved in trypanothione metabolism, trypanothione reductase and trypanothione synthetase, have been studied in detail as druggable targets. In this review, we summarize some of the recent findings on the molecules inhibiting these two essential enzymes for Trypanosoma and Leishmania viability.


Assuntos
Amida Sintases , Glutationa , NADH NADPH Oxirredutases , Trypanosoma , NADH NADPH Oxirredutases/metabolismo , NADH NADPH Oxirredutases/antagonistas & inibidores , Humanos , Amida Sintases/metabolismo , Amida Sintases/antagonistas & inibidores , Trypanosoma/efeitos dos fármacos , Trypanosoma/metabolismo , Glutationa/metabolismo , Glutationa/análogos & derivados , Animais , Espermidina/análogos & derivados , Espermidina/metabolismo , Leishmania/efeitos dos fármacos , Leishmania/metabolismo , Tripanossomicidas/farmacologia , Tripanossomicidas/uso terapêutico , Leishmaniose/tratamento farmacológico , Leishmaniose/metabolismo , Leishmaniose/parasitologia , Trypanosomatina/metabolismo , Trypanosomatina/efeitos dos fármacos , Proteínas de Protozoários/metabolismo , Proteínas de Protozoários/antagonistas & inibidores , Doença de Chagas/tratamento farmacológico , Doença de Chagas/parasitologia , Doença de Chagas/metabolismo
20.
Front Immunol ; 15: 1298275, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38707903

RESUMO

Background: Innate immune responses against infectious agents can act as triggers of inflammatory diseases. On the other hand, various pathogens have developed mechanisms for the evasion of the immune response, based on an inhibition of innate immunity and inflammatory responses. Inflammatory diseases could thus be controlled through the administration of pathogens or pathogen-derived molecules, capable of interfering with the mechanisms at the basis of inflammation. In this framework, the NLRP3 inflammasome is an important component in innate antimicrobial responses and a major player in the inflammatory disease. Parasites of the genus Leishmania are master manipulators of innate immune mechanisms, and different species have been shown to inhibit inflammasome formation. However, the exploitation of pathogenic Leishmania species as blockers of NLRP3-based inflammatory diseases poses safety concerns. Methods: To circumvent safety issues associated with pathogenic parasites, we focused on Leishmania tarentolae, a species of Leishmania that is not infectious to humans. Because NLRP3 typically develops in macrophages, in response to the detection and engulfment microorganisms, we performed our experiments on a monocyte-macrophage cell line (THP-1), either wild type or knockout for ASC, a key component of NLRP3 formation, with determination of cytokines and other markers of inflammation. Results: L. tarentolae was shown to possess the capability of dampening the formation of NLRP3 inflammasome and the consequent expression of pro-inflammatory molecules, with minor differences compared to effects of pathogenic Leishmania species. Conclusion: The non-pathogenic L. tarentolae appears a promising pro-biotic microbe with anti-inflammatory properties or a source of immune modulating cellular fractions or molecules, capable of interfering with the formation of the NLRP3 inflammasome.


Assuntos
Inflamassomos , Inflamação , Leishmania , Proteína 3 que Contém Domínio de Pirina da Família NLR , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Humanos , Inflamassomos/metabolismo , Inflamassomos/imunologia , Leishmania/imunologia , Inflamação/imunologia , Células THP-1 , Macrófagos/imunologia , Macrófagos/metabolismo , Macrófagos/parasitologia , Imunidade Inata , Citocinas/metabolismo
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