Effects of Amphotericin B-Conjugated Functionalized Carbon Nanoparticles in the Treatment of Cutaneous Leishmaniasis.

Leishmaniasis is a parasitic disease spread by the bite of an infected sandfly and caused by protozoan parasites of the genus Leishmania. Currently, there is no vaccine available for leishmaniasis in humans, and the existing chemotherapy methods face various clinical challenges. The majority of drugs are limited to a few toxic compounds, with some parasite strains developing resistance. Therefore, the discovery and development of a new anti-leishmanial compound is crucial. One promising strategy involves the use of nanoparticle delivery systems to accelerate the effectiveness of existing treatments. In this study, Amphotericin B (AmB) was incorporated into functionalized carbon nanotube (f-CNT) and evaluated for its efficacy against Leishmania major in vitro and in a BALB/c mice model. The increase in footpad thickness was measured, and real-time PCR was used to quantify the parasite load post-infection. Levels of nitric oxide and cytokines IL-4 and IFN-γ were also determined. We found that f-CNT-AmB significantly reduced the levels of promastigotes and amastigotes of the Leishmania parasite. The nanoparticle showed strong anti-leishmanial activity with an IC50 of 0.00494 ± 0.00095 mg/mL for promastigotes and EC50 of 0.00294 ± 0.00065 mg/mL for amastigotes at 72 h post-infection, without causing harm to mice macrophages. Treatment of infected BALB/c mice with f-CNT-AmB resulted in a significant decrease in cutaneous leishmania (CL) lesion size in the foot pad, as well as reduced Leishmania burden in both lymph nodes and spleen. The levels of nitric oxide and IFN-γ significantly increased in the f-CNT-AmB treated groups. Also, our results showed that the level of IL-4 significantly decreased after f-CNT-AmB treatment in comparison to other groups. In conclusion, our results demonstrate that AmB loaded into f-CNT is significantly more effective than AmB alone in inhibiting parasite propagation and promoting a shift towards a Th1 response.

Investigation of parasite genetic variation and systemic immune responses in patients presenting with different clinical presentations of cutaneous leishmaniasis caused by Leishmania aethiopica.

BACKGROUND: Cutaneous leishmaniasis (CL) is a neglected tropical skin disease, caused by the protozoan parasite Leishmania. In Ethiopia, CL is mainly caused by Leishmania aethiopica and can present in different clinical forms. The aim of this study was to assess whether these different forms are associated with differences in parasite genetic and host systemic immune signatures. METHODS: Here we analysed the whole genome sequence data for 48 clinical parasite isolates and the systemic immune signature from a cohort of CL patients, who were recruited in Nefas Mewcha, Northern Ethiopia, from January 2019 to January 2022. RESULTS: Our results show that parasites from CL cases with different presentations in a single Ethiopian setting are from the same genetic population based on a permutation test of genome-wide similarity. Furthermore, a logistic regression test for genome wide association did not identify any individual genetic variants significantly associated with disease presentation. We also measured plasma chemokine and cytokine levels of 129 CL patients presenting with different forms of CL. None of the chemokine [eotaxin, eotaxin-3, interleukin (IL)-8, interferon (IFN)-γ-induced protein-10 (IP-10), monocyte chemoattractant protein (MCP)-1, MCP-4, macrophage-derived chemokines (MDC), macrophage inflammatory protein (MIP)-1α, MIP-1ß and thymus- and activation-regulated chemokine (TARC)] or cytokine (IFN-γ, IL-1ß, interleukin-2, IL-4, IL-6, IL-10, IL-12p70, IL-13, tumor necrosis factor-α) levels measured were significantly different between the different clinical presentations of CL, as measured by Kruskal-Wallis test. We also compared those with healthy nonendemic controls: our results show a chemokine (IP-10, MCP-1, MCP-4, MDC, MIP-1α, MIP-1ß and TARC) but not a cytokine immune signature in patients with CL as compared to healthy nonendemic controls, as measured by Mann-Whitney test. CONCLUSIONS: The results of our study did not identify a systemic immune signature or parasite genetic factors associated with different clinical presentation of CL.

First case of urban leishmaniasis in the Campeche State, Mexico.

Cutaneous leishmaniasis represents 99% of all reported leishmaniasis cases in Mexico and typically occurs in agricultural or sylvatic areas. Campeche State is endemic for leishmaniasis; however, there are no previous records of urban Leishmania transmission. This report presents a case of cutaneous leishmaniasis in a 75-year-old man residing in an urban area. The patient presented with a three-month-old lesion on the right ear following an initial misdiagnosis of a bacterial infection. Given the suspicion of leishmaniasis, a tissue imprint was collected, revealing the presence of Leishmania amastigotes. Subsequently, amplification and sequencing of the Alanine aminotransferase and Internal transcribed spacer subunit 1 genes confirmed the presence of Leishmania mexicana. The patient was then treated with intralesional meglumine antimoniate. This case is significant as it marks the first confirmed human transmission of L. mexicana in an urban environment in Campeche State, demonstrating the importance of considering this pathology in patients with skin lesions originating from non-endemic areas in Mexico.

Functional plasticity shapes neutrophil response to Leishmania major infection in susceptible and resistant strains of mice.

Neutrophils rapidly infiltrate sites of infection and possess several microbicidal strategies, such as neutrophil extracellular traps release and phagocytosis. Enhanced neutrophil infiltration is associated with higher susceptibility to Leishmania infection, but neutrophil effector response contribution to this phenotype is uncertain. Here, we show that neutrophils from susceptible BALB/c mice (B/c) produce more NETs in response to Leishmania major than those from resistant C57BL/6 mice (B6), which are more phagocytic. The absence of neutrophil elastase contributes to phagocytosis regulation. Microarray analysis shows enrichment of genes involved in NET formation (mpo, pi3kcg, il1b) in B/c, while B6 shows upregulation of genes involved in phagocytosis and cell death (Arhgap12, casp9, mlkl, FasL). scRNA-seq in L. major-infected B6 showed heterogeneity in the pool of intralesional neutrophils, and we identified the N1 subset as the putative subpopulation involved with phagocytosis. In vivo, imaging validates NET formation in infected B/c ears where NETing neutrophils were mainly uninfected cells. NET digestion in vivo augmented parasite lymphatic drainage. Hence, a balance between NET formation and phagocytosis in neutrophils may contribute to the divergent phenotype observed in these mice.

Molecular Characterization of Sterol C4-Methyl Oxidase in Leishmania major.

Sterol biosynthesis requires the oxidative removal of two methyl groups from the C-4 position by sterol C-4-demethylase and one methyl group from the C-14 position by sterol C-14-demethylase. In Leishmania donovani, a CYP5122A1 (Cytochrome P450 family 5122A1) protein was recently identified as the bona fide sterol C-4 methyl oxidase catalyzing the initial steps of C-4-demethylation. Besides CYP5122A1, Leishmania parasites possess orthologs to ERG25 (ergosterol pathway gene 25), the canonical sterol C-4 methyl oxidase in Saccharomyces cerevisiae. To determine the contribution of CYP5122A1 and ERG25 in sterol biosynthesis, we assessed the essentiality of these genes in Leishmania major, which causes cutaneous leishmaniasis. Like in L. donovani, CYP5122A1 in L. major could only be deleted in the presence of a complementing episome. Even with strong negative selection, L. major chromosomal CYP5122A1-null mutants retained the complementing episome in both promastigote and amastigote stages, demonstrating its essentiality. In contrast, the L. major ERG25-null mutants were fully viable and replicative in culture and virulent in mice. Deletion and overexpression of ERG25 did not affect the sterol composition, indicating that ERG25 is not required for C-4-demethylation. These findings suggest that CYP5122A1 is the dominant and possibly only sterol C-4 methyl oxidase in Leishmania, and inhibitors of CYP5122A1 may have strong therapeutic potential against multiple Leishmania species.

Canine leishmaniasis caused by Leishmania Tropica in southeastern Iran: a case series study.

Dogs are the primary reservoirs of Leishmania infantum (L. infantum), but Leishmania tropica (L. tropica) infection is also possible in dogs and can transmitted to humans. The southeast of Iran experiences a high prevalence of canine leishmaniasis (CanL), and veterinarians frequently encounter symptomatic cases. Therefore, from December 2017 to November 2022, the present case series was designed to assess the prevalence of CanL in owned dogs with cutaneous manifestations resembling CanL. A total of 500 owned dogs with dermal lesions from two endemic cities, Zabol and Kerman, were enrolled. Impression smears from skin lesions and popliteal lymph nodes were prepared from all cases, whereas blood samples were gathered from amastigote-positive dogs for serological and molecular surveys. Commercial ELISA was done on sera samples, and two-step nested PCR was used on extracted DNA to amplify variable fragments of the Leishmania species' kinetoplast DNA (kDNA). Leishman bodies were microscopically detected in 7.2% (36/500) of dermal smears. Of the 360 owned dogs from Zabol, 2 have been diagnosed with L. tropica, and 10 have been diagnosed with L. infantum. Out of 140 owned dogs from Kerman, 8 were identified with L. tropica infection, and 16 were infected with L. infantum. Molecular results revealed the presence of 750 full dual-band bases related to the L. tropica species in 10/36 (27.7%) cases, which showed a considerable increase in canine cutaneous leishmaniosis compared with previous studies in southeastern Iran. The noticeable prevalence of L. tropica in owned dogs indicated that the dog's role in cutaneous leishmaniosis should be re-evaluated as a possible animal reservoir in endemic areas.

Epoxy-a-lapachone in nanosystem: a prototype drug for leishmaniasis assessed in the binomial BALB/c - Leishmania (Leishmania) amazonensis.

This perspective presents and supports arguments for a new formulation of epoxy-α-lapachone loaded microemulsion (ELAP-ME), a nanosystem, as a prototype drug for the treatment of leishmaniasis. The benefits of ELAP as a multitarget compound, with properties that affect key physiological pathways of Leishmania spp. are discussed. ELAP-ME demonstrated efficacy in murine infection models, particularly with the binomial BALB/c-Leishmania (Leishmania) amazonensis. Furthermore, it is proposed that the technological maturity of ELAP-ME be classified as Technology Readiness Level 4 (TLR 4) within the context of innovative drugs for American Cutaneous Leishmaniasis (ACL).

Evaluation of Loopamp Leishmania detection kit for the diagnosis of cutaneous leishmaniasis in Ethiopia.

BACKGROUND: Cutaneous leishmaniasis (CL) in Ethiopia and some parts of Kenya is predominantly caused by Leishmania aethiopica. While skin-slit (SS) microscopy is routinely used for CL diagnosis, more sensitive molecular tests are available. The Loopamp™ Leishmania detection kit (Loopamp) is a robust loop-mediated isothermal amplification (LAMP) assay with the potential for implementation in primary healthcare facilities. In this study, we comparatively assessed the diagnostic accuracy of four methods currently used to diagnose CL: Loopamp, kinetoplast DNA (kDNA) PCR, spliced leader RNA (SL-RNA) PCR and SS microscopy. METHODS: A study on 122 stored tape disc samples of suspected CL patients was conducted in Gondar, northwestern Ethiopia. Routine SS microscopy results were obtained from all patients. Total nucleic acids were extracted from the tapes and subjected to PCR testing targeting kDNA and SL-RNA, and Loopamp. Diagnostic accuracy was calculated with SS microscopy as a reference test. The limit of detection (LoD) of Loopamp and kDNA PCR were determined for cultured L. aethiopica and Leishmania donovani. RESULTS: Of the 122 patients, 64 (52.5%) were identified as CL cases based on SS microscopy. Although the PCR tests showed a sensitivity of 95.3% (95% confidence interval [CI] 91.6-99.1), Loopamp only had 48.4% (95% CI 39.6-57.3) sensitivity and 87.9% (95% CI 82.1-93.7) specificity. The LoD of Loopamp for L. donovani was 100-fold lower (20 fg/µl) than that for L. aethiopica (2 pg/µl). CONCLUSIONS: The Loopamp™ Leishmania detection kit is not suitable for the diagnosis of CL in Ethiopia, presumably due to a primer mismatch with the L. aethiopica 18S rRNA target. Further research is needed to develop a simple and sensitive point-of-care test that allows the decentralization of CL diagnosis in Ethiopia.

Type 1 diabetes and parasite infection: An exploratory study in NOD mice.

Microorganisms have long been suspected to influence the outcome of immune-related syndromes, particularly autoimmune diseases. Type 1 diabetes (T1D) results from the autoimmune destruction of the insulin-producing beta cells of pancreatic islets, causing high glycemia levels. Genetics is part of its aetiology, but environmental factors, particularly infectious microorganisms, also play a role. Bacteria, viruses, and parasites influence the outcome of T1D in mice and humans. We used nonobese diabetic (NOD) mice, which spontaneously develop T1D, to investigate the influence of a parasitic infection, leishmaniasis. Leishmania amazonensis is an intracellular eukaryotic parasite that replicates predominantly in macrophages and is responsible for cutaneous leishmaniasis. The implication of Th1 immune responses in T1D and leishmaniasis led us to study this parasite in the NOD mouse model. We previously constructed osteopontin knockout mice with a NOD genetic background and demonstrated that this protein plays a role in the T1D phenotype. In addition, osteopontin (OPN) has been found to play a role in the immune response to various infectious microorganisms and to be implicated in other autoimmune conditions, such as multiple sclerosis in humans and experimental autoimmune encephalomyelitis (EAE) in mice. We present herein data demonstrating the role of OPN in the response to Leishmania in NOD mice and the influence of this parasitic infection on T1D. This exploratory study aimed to investigate the environmental infectious component of the autoimmune response, including Th1 immunity, which is common to both T1D and leishmaniasis.

Diagnostic performance of Leishmania braziliensis and Leishmania peruviana antigens in the immunoblot method for the detection of american tegumentary leishmaniasis. / Rendimiento diagnóstico de los antígenos de Leishmania braziliensis y Leishmania peruviana en el método de inmunoblot para la detección de la leishmaniasis tegumentaria americana.

BACKGROUND: Motivation for the study. To contribute to the immunogenic character of soluble and excretion/secretion antigens of Leishmania braziliensis and Leishmania peruviana with the aim of identifying proteins with diagnostic potential. BACKGROUND: Main findings. The soluble antigen of Leishmania braziliensis has a sensitivity in the detection of ATL of 87.7%, specificity of 100% and a false positive rate of 20% against sera from patients with Chagas disease and 8.3% with mycosis. BACKGROUND: Implications. Immunoblot can improve the resolution capacity in the serological diagnosis of American tegumentary Leishmaniasis, particularly in patients where the length of the disease and the clinical form make difficult the diagnosis by parasitological methods. BACKGROUND: This study aimed to determine the performance of Leishmania braziliensis and Leishmania peruviana antigens in the detection of ATL by using serum samples obtained between 2013 - 2016. The obtained soluble and excretion/secretion antigens were transferred to membrane nitrocellulose by immunoblot assay. The evaluation was carried out against sera confirmed for ATL, at a confidence level of 95%, determining that the soluble antigen of Leishmania braziliensis had a sensitivity of 87.7%, specificity of 100% and area under the curve of 0.95; on the other hand, Leishmania peruviana showed values of 92.3%, 95.7% and 0.94, respectively. According to the results, we recommend that the reported immunogenic regions should be characterized and analyzed in order to continue with the development of recombinant and synthetic proteins, aimed at improving the efficiency of the serological diagnosis of the disease.

BACKGROUND: El presente estudio se planteó determinar el rendimiento de antígenos de Leishmania braziliensis y Leishmania peruviana en la detección de LTA, fue desarrollado a partir de muestras de suero obtenidas entre 2013 - 2016. Los antígenos solubles y de excreción/secreción obtenidos fueron transferidos a membrana de nitrocelulosa mediante un ensayo de inmunotransferencia. Se realizó la evaluación frente a sueros confirmados para LTA, a un nivel de confianza al 95%, logrando determinar que, el antígeno soluble de Leishmania braziliensis presenta una sensibilidad del 87,7%, especificidad del 100% y área bajo la curva de 0,95; mientras que, Leishmania peruviana se encontró valores de 92,3%, 95,7% y 0,94 respectivamente. De acuerdo a los resultados, recomendamos realizar la caracterización y análisis de las regiones inmunogénicas reportadas a fin de continuar con el desarrollo de proteínas recombinantes y sintéticas, orientadas a mejorar la eficiencia del diagnóstico serológico de la enfermedad. BACKGROUND: Motivación para realizar el estudio. Aportar sobre el carácter inmunogénico de los antígenos solubles y de excreción/secreción de Leishmania braziliensis y Leishmania peruviana con la finalidad de identificar proteínas con potencial diagnóstico. BACKGROUND: Principales hallazgos. El antígeno soluble de Leishmania braziliensis presenta una sensibilidad en la detección de LTA del 87,7%, especificidad del 100% y una proporción de falsos positivos de 20% frente a sueros de pacientes con la enfermedad de Chagas y del 8,3% con micosis. BACKGROUND: Implicancias. El Inmunoblot puede mejorar la capacidad resolutiva en el diagnóstico serológico de Leishmaniasis tegumentaria americana, especialmente en pacientes donde el tiempo de la enfermedad y la forma clínica hacen difícil el diagnóstico mediante los métodos parasitológicos.

Salivary antigens rPagSP02 and rPagSP06 are a reliable composite biomarker for evaluating exposure to Phlebotomus argentipes in Sri Lanka.

Phlebotomus argentipes is the established vector of leishmaniasis in the Indian sub-continent. Antibodies to sand fly salivary antigens are biomarkers for vector-host exposure in leishmaniasis-endemic regions. Ph. argentipes transmits Leishmania donovani in Sri Lanka, primarily causing cutaneous leishmaniasis (CL). Our study compared the performance of salivary gland homogenate (SGH) from a lab-reared local strain of Ph. argentipes females to a composite recombinant salivary biomarker (rPagSP02 + rPagSP06) in a CL-endemic population. Sera from 546 healthy individuals, 30 CL patients, and 15 non-endemic individuals were collected. Western blot analysis of Ph. argentipes SGH identified immunogenic bands between 15 kDa and 67 kDa, with bands of predicted molecular weight ∼of 15 kDa (SP02) and ∼28-30 kDa (SP06) as the major antibody targets. Indirect ELISAs using SGH or rPagSP02 + rPagSP06 antigens showed high sensitivity (96.7%) and specificity (100%), detecting comparable seropositivity in endemic populations. rPagSP02 + rPagSP06 exhibited enhanced discriminatory ability, supported by a strong positive correlation (r = 0.869) with SGH. Our findings indicate that the composite rPagSP02 + rPagSP06 salivary biomarker effectively identifies Ph. argentipes exposure in individuals living in Sri Lanka, showing promising potential for use in surveillance. These findings should be further validated to confirm the epidemiological applications in leishmaniasis-endemic regions.

[Special lesion on back of the hand]. / Bijzondere huidafwijking op de handrug.

A 74-year-old man, previously healthy, presented at the Dermatology department with an asymptomatic hand lesion persisting for two months. The general practitioner initially treated the lesion with betamethasone and fusidic acid cream. Examination revealed a 3 cm livid plaque with central crusts. A biopsy confirmed cutaneous leishmaniasis tropica. The patient travelled to Greece, Spain, and Southern France before the lesion occurred, which is consistent with the endemicity of leishmaniasis in the Mediterranean. Treatment involved cryotherapy and antimony injections, resulting in lesion resolution after two sessions. This case underscores the importance of considering travel history and endemic diseases in diagnosing and managing dermatological conditions, especially in regions prone to specific infections.

Cutaneous Leishmaniasis in Dr. Ersin Arslan Training and Research Hospital After Migration and During the Pandemic (2019-2022).

OBJECTIVE: Cutaneous leishmaniasis is a parasitic skin disease transmitted by the bite of sandflies. In our region, which is endemic for this disease, there has been a great migration from a much more endemic region and population movements from our area to Türkiye and abroad. Afterward, a pandemic was experienced. Due to these two extraordinary events and the possible epidemic potential in our region, it is useful to follow-up on the disease. We aimed to contribute to the evaluation of the disease in these processes by analyzing the data of our laboratory in recent years. METHODS: Between January 2019 and December 2022, samples from patients who came to our laboratory with suspected cutaneous leishmaniasis were taken, stained and examined under a microscope. Patients were evaluated in terms of age, gender, nationality, place of residence, lesion site and duration. RESULTS: Out of the 144 examined cases, 64 (44.4%) were positive for cutaneous leishmaniasis. Among these positive cases, 40 (62.5%) were women, 24 (37.5%) were men, and 54 (84.3%) belonged to the 0-9 age group. Of those who tested positive, 54 (84.3%) were Turkish citizens and 23 (35.9%) were Syrian citizens. Fifty-four (84.3%) patients had only single lesion. While the number of applications and positivity rates remained within normal levels in 2019 and 2020, a significant decrease was observed in both from 2021 and 2022. CONCLUSION: Cutaneous leishmaniasis is carried by migration, decreases in large-scale isolations such as pandemics, and its spread can be prevented with correct diagnosis and treatment. Although the number of patients may change over time and place, cutaneous leishmaniasis is a disease that threatens the health of societies and should always be monitored.

Population structure analysis of Phlebotomus papatasi populations using transcriptome microsatellites: possible implications for leishmaniasis control and vaccine development.

BACKGROUND: Phlebotomus papatasi is considered the primary vector of Leishmania major parasites that cause zoonotic cutaneous leishmaniasis (ZCL) in the Middle East and North Africa. Phlebotomus papatasi populations have been studied extensively, revealing the existence of different genetic populations and subpopulations over its large distribution range. Genetic diversity and population structure analysis using transcriptome microsatellite markers is important to uncover the vector distribution dynamics, essential for controlling ZCL in endemic areas. METHODS: In this study, we investigated the level of genetic variation using expressed sequence tag-derived simple sequence repeats (EST-SSRs) among field and colony P. papatasi samples collected from 25 different locations in 11 countries. A total of 302 P. papatasi sand fly individuals were analyzed, including at least 10 flies from each region. RESULTS: The analysis revealed a high-level population structure expressed by five distinct populations A through E, with moderate genetic differentiation among all populations. These genetic differences in expressed genes may enable P. papatasi to adapt to different environmental conditions along its distribution range and likely affect dispersal. CONCLUSIONS: Elucidating the population structuring of P. papatasi is essential to L. major containment efforts in endemic countries. Moreover, the level of genetic variation among these populations may improve our understanding of Leishmania-sand fly interactions and contribute to the efforts of vaccine development based on P. papatasi salivary proteins.

Intralesional gene expression profile of JAK-STAT signaling pathway and associated cytokines in Leishmania tropica-infected patients.

Background: The JAK-STAT signaling pathway is a central cascade of signal transduction for the myriad of cytokines in which dysregulation has been implicated in progression of inflammatory and infectious diseases. However, the involvement of this pathway in human cutaneous leishmaniasis (CL) due to Leishmania (L.) tropica warrants further investigation. Methods: This study sought to investigate differential gene expression of several cytokines and their associated jak-stat genes in the lesions of L. tropica-infected patients byquantitative Real-Time PCR. Further, the expression of five inhibitory immune checkpoint genes was evaluated. Results: Results showed that the gene expression levelsof both Th1 (ifng, il12, il23) and Th2 (il4, il10) types cytokines were increased in the lesion of studied patients. Further, elevated expression levels of il35, il21, il27 and il24 genes were detected in the lesions of CL patients. Notably, the expression of the majority of genes involved in JAK/STAT signaling pathway as well as checkpoint genes including pdl1, ctla4 and their corresponding receptors was increased. Conclusion: Our finding revealed dysregulation of cytokines and related jak-stat genes in the lesion of CL patients. These results highlight the need for further exploration of the functional importance of these genes in the pathogenesis of, and immunity to, CL.

Dogs may carry Leishmania tropica and Leishmania major in their blood circulation: a molecular and hematological study.

BACKGROUND: Dogs may be infected with species of Leishmania parasites that are disseminated through blood circulation and invade the internal organs. In this study, we aim to detect the parasite in the blood of dogs using the PCR technique. The present work was performed from February 2022 to May 2023 in Fars Province, southern Iran, where the disease is endemic. RESULTS: In total, 7(5.1%) out of 135 blood samples, six were identified as Leishmania tropica and one as Leishmania major. We found no trace of Leishmania infantum, which is always known for visceral infection. In addition, no sign of cutaneous lesions or a significant disease was seen in the animals infected with both species. Of 48 dogs with anemia, two were Leishmania positive. The mean value of hematological parameters in the infected dogs was within the normal range except for a significant reduction in the platelet measures (p < 0.05). CONCLUSIONS: Our data revealed that both Leishmania species, tropica and major, may manifest as viscerotropic leishmaniasis. More investigations are needed to understand the conditions under which these species choose the type of infection. Moreover, our data emphasize the role of asymptomatic dogs in carrying these parasites, a crucial factor in spreading the disease.

Entomological aspects of leishmaniasis transmission in the urban area of Teófilo Otoni, a municipality endemic for the disease in the Brazilian state of Minas Gerais.

Brazil is endemic for both visceral (VL) and cutaneous (CL) clinical forms of leishmaniasis, poverty-associated diseases with worldwide distribution. Leishmania parasites are the etiological agents of leishmaniases, which are transmitted to humans through the bites of infected phlebotomine sand flies. From 2018 to 2023, 15 cases of VL and 129 cases of CL were reported in Téofilo Otoni, an important economic center in the Brazilian state of Minas Gerais. Owing to the lack of data on the entomological fauna, the present study aimed to clarify this main aspect of leishmaniasis. From May, 2021 to April, 2023, entomological captures were performed monthly in ten neighborhoods in Teófilo Otoni. The influence of bioclimatic variables on insect populations was evaluated, and natural infection by Leishmania spp. was investigated using molecular methods. A total of 306 specimens of 12 species of phlebotomine sand fly were collected. The majority (91.6%) were proven or putative vectors of leishmaniasis agents. The population of insects tended to increase during the cooler and drier months. Although Leishmania infection was not detected in any of the samples, the presence of vectors provides conditions for the maintenance and expansion of the transmission cycle of leishmaniasis in Teófilo Otoni.

Immunoregulatory effects of soluble antigens of Leishmania sp. in human lymphocytes in vitro.

The clinical manifestations of cutaneous leishmaniasis (CL) depend not only on the infecting species involved, but also on the immune response of the individual. Although not yet well understood in humans, parasite survival and persistence are related to the cytokine profile and T cell proliferation, with the Th1 profile being related to cure, and the Th2 profile to disease progression. Considering the need for studies focused on the species with the highest circulation in the state of Amazonas, this study aimed to analyze the immunoregulation stimulated by soluble antigens (SLAs) of Leishmania (L.) amazonensis and Leishmania (V.) guyanensis in human lymphocytes in vitro, in order to understand the immune response of patients with CL. Lymphoproliferation was evaluated against stimuli of SLAs from L. amazonensis (100 µg/mL), SLAs from L. guyanensis (100 µg/mL) and phytohemagglutinin (10 µg/mL) using a BrdU Cell Proliferation ELISA kit after 72 h of incubation. Quantification of the cytokines IL-1b, IL-6, IL-8, IL-10, IL-12 and TNF was performed using the BD™ cytometric bead array human Th1/Th2/Th17 cytokine kit. Our results demonstrated that soluble antigens from L. amazonensis and L. guyanensis stimulated the lymphoproliferation of PBMCs from patients primo-infected with CL. Among the cytokines dosed, the highest concentrations were of IL-6 and IL-8, thus demonstrating that the soluble antigens evaluated are capable of inducing regulatory mechanisms.

NLRP1-dependent activation of Gasdermin D in neutrophils controls cutaneous leishmaniasis.

Intracellular pathogens that replicate in host myeloid cells have devised ways to inhibit the cell's killing machinery. Pyroptosis is one of the host strategies used to reduce the pathogen replicating niche and thereby control its expansion. The intracellular Leishmania parasites can survive and use neutrophils as a silent entry niche, favoring subsequent parasite dissemination into the host. Here, we show that Leishmania mexicana induces NLRP1- and caspase-1-dependent Gasdermin D (GSDMD)-mediated pyroptosis in neutrophils, a process critical to control the parasite-induced pathology. In the absence of GSDMD, we observe an increased number of infected dermal neutrophils two days post-infection. Using adoptive neutrophil transfer in neutropenic mice, we show that pyroptosis contributes to the regulation of the neutrophil niche early after infection. The critical role of neutrophil pyroptosis and its positive influence on the regulation of the disease outcome was further demonstrated following infection of mice with neutrophil-specific deletion of GSDMD. Thus, our study establishes neutrophil pyroptosis as a critical regulator of leishmaniasis pathology.

Leishmania spp. genetic factors associated with cutaneous leishmaniasis antimony pentavalent drug resistance: a systematic review.

BACKGROUND: Leishmaniasis is a neglected zoonosis caused by parasites of Leishmania spp. The main drug used to treat cutaneous leishmaniasis (CL) is the antimoniate of meglumine. This drug, which has strong adverse and toxic effects, is usually administered intravenously, further complicating the difficult treatment. Factors such as Leishmania gene expression and genomic mutations appear to play a role in the development of drug resistance. OBJECTIVES: This systematic review summarises the results of the literature evaluating parasite genetic markers possibly associated with resistance to pentavalent antimony in CL. METHODS: This study followed PRISMA guidelines and included articles from PubMed, SciELO, and LILACS databases. Inclusion criteria were studies that (i) investigated mutations in the genome and/or changes in gene expression of Leishmania associated with treatment resistance; (ii) used antimony drugs in the therapy of CL; (iii) used naturally resistant strains isolated from patients. The Joanna Briggs Institute Critical Appraisal Checklist was used to assess article quality and risk of bias. FINDINGS: A total of 23 articles were selected, of which 18 investigated gene expression and nine genomic mutations. Of these 23 articles, four examined gene expression and genomic mutations in the same samples. Regarding gene expression, genes from the ABC transporter protein family, AQP1, MRPA, TDR1 and TRYR were most frequently associated with drug resistance. In one of the articles in which mutations were investigated, a mutation was found in HSP70 (T579A) and in three articles mutations were found in AQP1 (A516C, G562A and G700A). A limitation of this review is that in most of the included studies, parasites were isolated from cultured lesion samples and drug resistance was assessed using in vitro drug susceptibility testing. These approaches may not be ideal for accurate genetic evaluation and detection of treatment failure. MAIN CONCLUSIONS: The development of further studies to evaluate the genetic resistance factors of Leishmania spp. is necessary to elucidate the mechanisms of the parasite and improve patient treatment and infection control.