Expresión transcripcional de receptores tipo Toll y citoquinas proinflamatorias en macrófagos infectados con Leishmania braziliensis nativa / Transcriptional expression of Toll-like receptors and proinflammatory cytokines in macrophages infected with native Leishmania braziliensis

RESUMEN Introducción. El receptor de tipo Toll (TLR) que interactúe con el promastigote de Leishmania spp. determina la vía de activación celular. Objetivo. Identificar la expresión transcripcional de TLR-3, TLR-4, TLR-9, IL-12 y TNF-α en macrófagos infectados con una cepa nativa de L. braziliensis (Lbn). Métodos. La identificación de Lbn se hizo empleando qPCR para secuencias del DNA del cinetoplasto. Los macrófagos peritoneales de ratones fueron infectados con promastigotes y se midieron la producción de óxido nítrico (ON). Se cuantificaron los niveles transcripcionales para TLRs y citoquinas empleando qRT-PCR. Resultados. Lbn presentó 96% de homología con L. braziliensis. En los infectados con promastigotes se observó elevada producción de ON a las 2 h; significativa expresión transcripcional especialmente de TLR-3 y TLR-9 que se correspondió con la expresión para citoquinas. Conclusión. Lbn activó fuertemente a los macrófagos mediante los TLRs endosomales lo cual puede ser aplicado en el diseño de agonistas para tratar la enfermedad.

ABSTRACT Introduction. The Toll-like receptor (TLR) interacting with the promastigote of Leishmania spp. determines the cellular activation pathway. Objective. To determine the transcriptional expression of TLR-3, TLR-4, TLR-9, IL-12 and TNF-α in macrophages infected with a native strain of L. braziliensis (Lbn). Materials and Methods. Identification of Lbn was performed by qPCR for kinetoplast DNA sequences. Mouse peritoneal macrophages were infected with promastigotes (MI) and nitric oxide (NO) production was measured; transcript levels for TLRs and cytokines were quantified by qRT-PCR. Results. Lbn showed 96% homology to L. braziliensis. High ON production was observed in IMs at 2 h; significant transcriptional expression especially of TLR-3 and TLR-9, which corresponded with expression for cytokines. Conclusions. Lbn strongly activated macrophages via endosomal TLRs, which can be applied in the design of agonists to treat the disease.

The absence of eosinophils is associated with early metastatic lesions in Leishmania amazonensis-infected mice

BACKGROUND Eosinophils are granulocytes that rapidly increase frequency in the bloodstream during helminthic infections and allergic responses. They are found in tissue infected by Leishmania during early disease, but their role during infection is not entirely understood. OBJECTIVES We aim to compare the disease due to Leishmania amazonensis in BALB/c and Δdbl-GATA1 mice, which lack eosinophils. METHODS BALB/c and Δdbl-GATA1 mice infected with L. amazonensis were observed for several weeks. The parasite load and dissemination pattern were assessed. FINDINGS The Δdbl-GATA1 mice developed an anticipated dissemination of L. amazonensis and a worsening disease. No differences were found in the lesion development or the parasite load in the footpad among Δdbl-GATA1 mice and BALB/c eight weeks after infection. However, nine weeks after infection, massive growth of metastatic lesions appeared in several parts of the skin in Δdbl-GATA1 mice, weeks earlier than BALB/c. We observed increased parasites in the bloodstream, probably an essential dissemination route. Thirteen weeks after infection, metastatic lesions were found in all Δdbl-GATA1 mice. MAIN CONCLUSION These results suggest a protective role of eosinophils in delaying the disease caused by L. amazonensis, although several limitations of this mice strain must be considered.

FVB/NJ strain as a mouse model for cutaneous leishmaniasis by Leishmania (L.) amazonensis

BACKGROUND Leishmaniases encompass a spectrum of neglected diseases caused by parasites of the genus Leishmania, grouped in two forms: tegumentary and visceral leishmaniasis. OBJECTIVES In this study, we propose Friend Virus B NIH Jackson (FVB/NJ) mouse strain as a new experimental model of infection with Leishmania (Leishmania) amazonensis, the second most prevalent agent of tegumentary leishmaniasis in Brazil. METHODS AND FINDINGS We performed in vitro infections of FVB/NJ macrophages and compared them with BALB/c macrophages, showing that BALB/c cells have higher infection percentages and a higher number of amastigotes/cell. Phagocytosis assays indicated that BALB/c and FVB/NJ macrophages have similar capacity to uptake parasites after 5 min incubations. We also investigated promastigotes' resistance to sera from FVB/NJ and BALB/c and observed no difference between the two sera, even though FVB/NJ has a deficiency in complement components. Finally, we subcutaneously infected FVB/NJ and BALB/c mice with 2 × 106 parasites expressing luciferase. Analysis of lesion development for 12 weeks showed that FVB/NJ and BALB/c mice have similar lesion profiles and parasite burdens. MAIN CONCLUSIONS This work characterises for the first time the FVB/NJ mouse as a new model for tegumentary leishmaniasis caused by Leishmania (L.) amazonensis.

GSK-3 kinase a putative therapeutic target in trypanosomatid parasites

ABSTRACT Trypanosomatids are an important group of parasites that predominate in tropical and subtropical areas of the planet, which cause diseases that are classified as forgotten and neglected by the world health organization. In this group of parasites, we find Trypanosoma cruzi, Trypanosoma brucei, Trypanosoma brucei rhodesiense and Leishmania spp, for which there is no vaccine available, and its control has focused mainly on pharmacological treatment. Due to the poverty situation where these diseases are found and the biological complexity of these parasites, there are multiple variables to control, including the diversity of species, the complexity of their life cycles, drug resistance, cytotoxicity, the limited use in pregnant women, the high costs of treatment and the little-known pharmacological mechanisms of action, among others. It is therefore necessary to find new strategies and approaches for the treatment of these parasitic diseases. Among these new approaches is the rational search for new targets based on the allosteric inhibition of protein kinases, which have been little studied in trypanosomatids. Among these kinases, we find Glycogen Synthase Kinase-3 (GSK-3), a kinase of great pharmacological interest, which is under intense basic and clinical research by pharmaceutical companies for the treatment of cancer. This kinase, highly studied in the PI3K/AKT/mTOR pathway signaling in humans, has an orthologous gene in these parasites (GSK-3 s), which has proven to be essential for them in response to different challenges; Therefore, it is notable to increase research in this kinase in order to achieve a broad structural and functional characterization in the different species of trypanosomatids.

Leishmania (Sauroleishmania) tarentolae versus pathogenic species: comparative evaluation of protease activity, glycoconjugates, resistance to complement and metabolome composition

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.

Estudo epidemiológico descritivo dos casos notificados de Leishmaniose visceral no estado de Goiás no período de 2011 a 2020 / Descriptive epidemiological study of notified cases of visceral Leishmaniasis in the state of Goiás in the period from 2011 to 2020 / Estudio epidemiológico descriptivo de los casos notificados de Leishmaniasis visceral en el estado de Goiás en el período de 2011 a 2020

A leishmaniose visceral, conhecida também como calazar, trata-se da forma mais grave das leishmanioses por ser uma enfermidade sistêmica e com um grande espectro clínico e um caráter irregular. Desse modo, objetivou-se avaliar a situação epidemiológica da leishmaniose visceral no Estado de Goiás, bem como o caráter dessa doença e o perfil mais atingido. Para isso, foi realizada uma pesquisa retrospectiva compreendendo o período de 2012 a 2021, com os casos notificados de leishmaniose visceral no Estado de Goiás. A busca pelos dados ocorreu por meio dos SINAN/DATASUS do Ministério da Saúde. Os dados encontrados foram tabulados e analisados por meio do programa Excel da Microsoft®. No período avaliado, foram notificados 388 casos de LV em Goiás, distribuídos em 47 municípios goianos. 66,23% das ocorrências pertenciam ao sexo masculino; 25,51% eram adultos com idade entre 20- 39 anos; 74,48% eram moradores da zona urbana; além disso, 9,27% do total de casos notificados em Goiás evoluíram para óbito. Há ainda uma a taxa de coinfecção LV/HIV de 12,11% do total de casos notificados. Constatou-se, portanto, que algumas localidades possuem registros consequentes de LV, levando a existência de áreas endêmicas no estado. Existe um perfil de acometimento em Goiás, no período de análise, direcionado para o sexo masculino, de cor parda, jovens adultos, com baixa escolaridade, vivendo nas cidades e grandes centros urbanos. Destaca-se o elevado quantitativo de dados identificados como ignorado/branco, que podem interferir significativmente em análises epidemiológicas.

Visceral leishmaniasis, also known as kala-azar, is the most serious form of leishmaniasis because it is a systemic disease with a wide clinical spectrum and an irregular character. Thus, the objective was to evaluate the epidemiological situation of visceral leishmaniasis in the State of Goiás, as well as the character of this disease and the most affected profile. For this, a retrospective survey was carried out covering the period from 2012 to 2021, with reported cases of visceral leishmaniasis in the State of Goiás. The search for data took place through the SINAN/DATASUS of the Ministry of Health. The data found were tabulated and analyzed using the Microsoft® Excel program. In the period evaluated, 388 cases of VL were reported in Goiás, distributed in 47 munic- ipalities in Goiás. 66.23% of the occurrences were male; 25.51% were adults aged be- tween 20-39 years; 74.48% were residents of the urban area; in addition, 9.27% of the total cases reported in Goiás evolved to death. There is also a LV/HIV co-infection rate of 12.11% of the total reported cases. It was found, therefore, that some localities have consequent records of VL, leading to the existence of endemic areas in the state. There is a profile of involvement directed towards males, of mixed race, young adults, with low education, living in cities and large urban centers. The high quantity of data identified as ignored/white stands out, which can significantly interfere in epidemiological analyses. KEYWORDS: Epidemiology; Leishmania; Notification.

La leishmaniasis visceral, también conocida como kala-azar, es la forma más grave de leishmaniasis por tratarse de una enfermedad sistémica de amplio espectro clínico y carácter irregular. Así, el objetivo fue evaluar la situación epidemiológica de la leishmaniasis visceral en el Estado de Goiás, así como el carácter de esta enfermedad y el perfil más afectado. Para ello, se realizó una encuesta retrospectiva que abarcó el período de 2012 a 2021, con los casos notificados de leishmaniasis visceral en el Estado de Goiás. La búsqueda de datos se realizó a través del SINAN/DATASUS del Ministerio de Salud. Los datos encontrados fueron tabulados y analizados utilizando el programa Microsoft® Excel. En el período evaluado, fueron notificados 388 casos de LV en Goiás, distribuidos en 47 municipios de Goiás. 66,23% de las ocurrencias fueron del sexo masculino; 25,51% eran adultos con edad entre 20-39 años; 74,48% eran residentes del área urbana; además, 9,27% del total de casos notificados en Goiás evolucionaron a muerte. Existe también una tasa de coinfección LV/VIH de 12,11% del total de casos notificados. Se constató, por lo tanto, que algunas localidades tienen registros consecuentes de LV, llevando a la existencia de áreas endémicas en el estado. Existe un perfil de envolvimiento dirigido a hombres, mestizos, adultos jóvenes, con baja escolaridad, residentes en ciudades y grandes centros urbanos. Se destaca la elevada cantidad de datos identificados como ignorados/blancos, lo que puede interferir significativamente en los análisis epidemiológicos.

Epidemiological aspects of American and visceral cutaneous leishmaniasis in the city of Cametá, Pará, Amazon / Aspectos epidemiológicos da leishmaniose tegumentar americana e visceral na cidade de Cametá, Pará, Amazônia / Aspectos epidemiológicos de la leishmaniasis cutánea americana y visceral en la ciudad de Cametá, Pará, Amazonas

Background and objectives: leishmaniases are anthropozoonosis considered a major public health problem in tropical regions and endemic in some areas of constant expansion. This study aimed to assess the main epidemiological aspects of American tegumentary leishmaniasis (ATL) and visceral leishmaniasis (VL) in the municipality of Cametá, in the state of Pará, from 2007 to 2017. Methods: this is a descriptive-exploratory analysis, of time series, with data collected in the Department of Epidemiological Surveillance of the Department of Sanitary Surveillance of Cametá. Statistical calculations were performed, and, for the coefficient of incidence of ATL and VL, the standard formula was used to obtain the indicator. Results: a total of 94 and 294 cases of ATL and VL were reported, with the highest incidence rate in 2008. The disease affected all established age groups, with high frequency in children under ten years of age for VL (n=174), and between 20 and 30 years of age, for ATL (n=71). The disease was more prevalent in males (ATL (89.4%) and VL (58.2%)), because men are more related to economic activities. Conclusion: considering the high number of rural cases, it is noteworthy that reporting in urban areas is also worrisome, in addition to the livelihood of local families, because it has made them vulnerable to the disease. Furthermore, there is concern about the possible expansion and change in the pattern of ATL in the municipality. The Municipal Department as well as the epidemiological surveillance must pay attention to promote investments and campaigns to combat and treat this important disease.(AU)

Justificativa e objetivos: as leishmanioses são antropozoonoses consideradas um grande problema para a saúde pública em regiões tropicais e endêmicas em algumas áreas de constante expansão. Este estudo teve como objetivo avaliar os principais aspectos epidemiológicos da leishmaniose tegumentar americana (LTA) e leishmaniose visceral (LV) no município de Cametá, no estado do Pará, no período de 2007 a 2017. Métodos: trata-se de uma análise descritiva-exploratória, de série temporal, com dados coletados no Departamento de Vigilância Epidemiológica da Secretaria de Vigilância Sanitária de Cametá. Realizaram-se os cálculos estatísticos, e, para o coeficiente de incidência de LTA e LV, utilizou-se a fórmula padrão para a obtenção do indicador. Resultados: foram notificados 94 e 294 casos de LTA e LV, com maior taxa de incidência em 2008. A doença atingiu todas as faixas etárias estabelecidas, com alta frequência nos menores de dez anos para LV (n=174), e, entre 20 e 30 anos de idade, para LTA (n=71). A doença foi mais prevalente no sexo masculino (LTA (89,4%) e LV (58,2%)), em virtude dos homens estarem mais relacionados com as atividades econômicas. Conclusão: em vista do alto número de casos rurais, ressalta-se que a notificação em área urbana também é preocupante, além dos meios de subsistência das famílias locais, pois vem tornando-as vulneráveis para o adoecimento. Ademais, há a preocupação com a possível expansão e mudança no padrão da LTA no município. A Secretaria Municipal, bem como de vigilância epidemiológica, deve atentar-se a promover investimentos e campanhas de combate e tratamento deste importante agravo.(AU)

Justificación y objetivos: las leishmaniasis son antropozoonosis consideradas un importante problema de salud pública en las regiones tropicales y endémicas en algunas zonas de constante expansión. Este estudio tuvo como objetivo evaluar los principales aspectos epidemiológicos de la leishmaniasis tegumentaria americana (LTA) y la leishmaniasis visceral (LV) en el municipio de Cametá, en el estado de Pará, de 2007 a 2017. Métodos: se trata de un análisis descriptivo-exploratorio, de serie temporal, con datos recolectados en el Departamento de Vigilancia Epidemiológica de la Secretaría de Vigilancia Sanitaria de Cametá. Se realizaron cálculos estadísticos y, para el coeficiente de incidencia de LCA y LV, se utilizó la fórmula estándar para obtener el indicador. Resultados: se reportaron 94 y 294 casos de LTA y LV, con la mayor tasa de incidencia en 2008. La enfermedad afectó a todos los grupos de edad establecidos, con alta frecuencia en menores de diez años para LV (n=174), y entre 20 y 30 años. años de edad, para LTA (n=71). La enfermedad fue más prevalente en el sexo masculino (LTA (89,4%) y VL (58,2%)), debido a que los hombres están más relacionados con actividades económicas. Conclusión: dado el alto número de casos rurales, cabe señalar que la notificación en las zonas urbanas también es motivo de preocupación, además de los medios de subsistencia de las familias locales, ya que las ha vuelto vulnerables a la enfermedad. Además, existe preocupación por la posible expansión y cambio en el patrón de LTA en el municipio. La Secretaría Municipal, así como la de vigilancia epidemiológica, debe prestar atención a promover inversiones y campañas para combatir y tratar este importante problema.(AU)

Conocimientos, actitudes y prácticas sobre Leishmaniasis visceral en tres comunas de Neiva, Huila

Introducción: La leishmaniasis visceral es una enfermedad zoonótica, transmitida por vectores del género Lutzomyia, de distribución en 98 países, incluyendo Colombia y que puede ocasionar un cuadro clínico grave, que en ausencia de tratamiento puede ser fatal. Objetivo: Determinar los conocimientos, actitudes y prácticas (CAP) adquiridas sobre vectores, reservorios y características de la Leishmaniasis visceral en las comunas 8, 9 y 10 de Neiva (Huila) en 2019. Materiales y métodos: Se realizó un estudio descriptivo de corte transversal en el período de junio a diciembre de 2019, en el que se muestrearon tres comunas (8, 9 y 10), en un total de 30 barrios de la ciudad de Neiva-Huila. Se aplicaron 385 encuestas tipo CAP, con un margen de error de 5 % y una confiabilidad de 95 %. Se incluyeron personas adultas de 18 años o más (267 mujeres y 117 hombres), que voluntariamente desearon participar, con previo consentimiento informado. Resultados: Según la información analizada, se determinó que 77 % de los encuestados no conoce la leishmaniasis visceral, 52 % no la identifican como una enfermedad zoonótica, 82 % no conocen el agente causal y 44 % no tienen claridad sobre los síntomas que se presentan en humanos. Conclusiones: Aunque se han realizado campañas preventivas en las comunas afectadas, el nivel de conocimiento sobre la enfermedad, el vector, el reservorio y las prácticas preventivas específicas para contrarrestar la leishmaniasis visceral son poco conocidos en la población de estudio, a pesar de que las prácticas y actitudes identificadas son positivas.

Introduction: Visceral leishmaniasis is a zoonotic disease transmitted by vectors of the Lutzomyia genus, distributed in 98 countries, including Colombia, which can cause a serious clinical picture that, in the absence of treatment, can be fatal. Objective: To determine the knowledge, attitudes, and practices (KAP) acquired on vectors and reservoirs, as well as the characteristics of visceral Leishmaniasis in communes 8, 9 and 10 of the city of Neiva in 2019. Material and Methods: A descriptive cross-sectional study was carried out from June to December 2019. Three communes (8, 9 y 10) were sampled from a total of 30 neighborhoods of the city of Neiva - Huila. In addition, 385 CAP-type surveys were applied, with a margin of error of 5% and a reliability of 95%. People aged 18 years or older (267 women and 117 men) who voluntarily wished to participate, with prior informed consent, were included. Results: According to the analyzed information, it was determined that the majority of the surveyed population did not know about visceral leishmaniasis (77%), did not identify it as a zoonotic disease (52%), nor did they know who the causative agent is (82%); furthermore, they were not clear about the symptoms that occur in humans, nor the treatment used to manage them. Conclusions: Although preventive campaigns have been carried out in the affected communes, the level of knowledge about the disease, the vector, the reservoir, and the specific preventive practices to counteract visceral leishmaniasis are little known in the study population, despite the fact that the identified practices and attitudes are positive.

Expression profile in Leishmania major exposed to Staphylococcus aureus and group A beta-hemolytic Streptococcus

Background & objectives: The interaction of Leishmania spp. with microbiota inside the midgut vector has significant output in pathogenesis. This study aimed to identify the profile of Leishmania major gene expression of LACK, gp63, and hsp70 after exposure to Staphylococcus aureus and group A beta-hemolytic Streptococci (GABHS). Methods: Leishmania major (MRHO/IR/75/ER) promastigotes were exposed with S. aureus, with GABHS, and with both GABHS and S. aureus at 25°C for 72 h. The gene expression analysis of Lmgp63, Lmhsp70, and LmLACK was assessed using SYBR Green real-time PCR by ??Ct. All experiments were repeated in triplicate. Statistical analysis was done using two-way ANOVA. A P-value less than 0.05 was considered significant. Results: Lmgp63 was expressed in the group exposed to GABHS with 1.75-fold lower than the control group (p=0.000). The LmLACK had expression in both groups exposed with GABHS and GABHS with S. aureus with 2.8 and 1.33-fold more than the control group, respectively (p=0.000). The Lmhsp70 gene expression was reported in the group exposed with GABHS with relative quantification of 5.7-fold more than the control group. Interpretation & conclusion: This study showed that the important genes encoding LACK, gp63, and hsp70 changed their expression after exposure to the S. aureus and GABHS.

Uso de anfotericina B liposomal en leishmaniasis cutánea diseminada causada por Leishmania braziliensis en un paciente pediátrico con síndrome de Down / Use of liposomal anfotericin B in diseminated cutaneous leishmaniasis caused by Leishmania braziliensis in a pediatric patient with Down syndrome

Se presenta un caso de leishmaniasis selvática cutánea diseminada con manifestación extensa en una paciente pediátrica con síndrome de Down. El caso se confirmó a través de estudios parasitológicos e inmunológicos, mientras que la identificación se realizó mediante la técnica de reacción en cadena de la polimerasa-polimorfismos de longitud de fragmentos de restricción (PCR-RFLP, por sus siglas en inglés), determinándose la especie como Leishmania (Viannia) braziliensis. La manifestación clínica agresiva y prolongada con poca respuesta a estibogluconato y anfotericina desoxicolato pueden deberse al déficit inmunológico que se presenta como parte del síndrome de Down. La paciente eventualmente recibió tratamiento con anfotericina B liposomal y al término de la terapia, mostró mejoría clínica de las lesiones. El presente reporte ilustra los desafíos tanto de diagnóstico como tratamiento de leishmaniasis cutánea en pacientes pediátricos inmunosuprimidos, especialmente en un entorno de difícil acceso social, económico y geográfico, a los servicios de salud. Se recomienda considerar a la leishmaniasis en el diagnóstico diferencial cuando se atienda ulceras crónicas dermatológicas atípicas; así como tener en cuenta el uso de anfotericina liposomal en pacientes inmunocomprometidos.

We present a case of disseminated cutaneous leishmaniasis with extensive manifestation in a pediatric patient with Down syndrome. The case was confirmed by parasitological and immunological tests. The species was identified as Leishmania (Viannia) braziliensis by polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLP). The immune deficit that occurs as part of Down syndrome may have been the reason for the aggressive and prolonged clinical manifestations as well as the poor response to stibogluconate and deoxycholate amphotericin. The patient was treated with liposomal amphotericin B and at the end of therapy, showed clinical improvement of the lesions. This report highlights the challenges of the diagnosis and treatment of cutaneous leishmaniasis in immunosuppressed pediatric patients, especially under difficult social, economic and geographic conditions. Leishmaniasis should be considered as a differential diagnosis when treating atypical chronic dermatologic ulcers; the use of liposomal amphotericin in immunocompromised patients should also be considered in these cases.

The first imported case of visceral leishmaniasis in Shenzhen City / 中国血吸虫病防治杂志

A patient with fever, chills, and pancytopenia as major clinical manifestations was presented. To investigate the cause, the patient’s peripheral blood was collected for pathogen screening using metagenomic next - generation sequencing (mNGS). The DNA sequence of Leishmania donovani was detected, and Leishmania amastigotes were found in bone marrow smears using microscopy. The case was therefore definitively diagnosed as visceral leishmaniasis, and was cured and discharged from hospital following treatment with liposomal amphotericin B for 14 days. This is the first imported case of visceral leishmaniasis since the founding of Shenzhen City in 1979.

Correlation between different clinical manifestations and changes of blood indexes in canine leishmaniasis / 中华地方病学杂志

Objective:To study the main blood parameters of Leishmania-infected dogs with different clinical symptoms in the high incidence area of visceral leishmaniasis in Gansu Province, and to provide scientific basis for early diagnosis and monitoring of canine leishmaniasis. Methods:In May 2019, Wenxian County of Longnan City and Diebu County of Gannan Tibetan Autonomous Prefecture, the historical endemic areas of visceral leishmaniasis in Gansu Province, were selected as the investigation sites, and 40 dogs with positive Leishmania DNA were investigated. According to their clinical symptoms, they were divided into three groups: asymptomatic ( n = 17), minimally symptomatic ( n = 12) and symptomatic ( n = 11). At the same time, 10 healthy dogs were selected as controls. Major blood indexes were detected by an automatic animal blood five-classification cell analyzer. Results:Compared with the control group, the number of red blood cells, hemoglobin content, hematocrit, and the number of lymphocytes, monocytes and eosinophils in the asymptomatic, minimally symptomatic and symptomatic groups were decreased ( P < 0.05), while the number of white blood cells and neutrophils in minimally symptomatic and symptomatic groups were decreased ( P < 0.05). The above blood indexes were lower in the symptomatic group than those in the asymptomatic group. The number of red blood cells (control group vs symptomatic group) decreased from (6.82 ± 0.05) × 10 12/L to (4.19 ± 0.08) × 10 12/L, hemoglobin content decreased from (15.84 ± 0.74) g/L to (9.65 ± 0.18) g/L, hematocrit decreased from (46.41 ± 0.95)% to (27.86 ± 0.42)%, the number of white blood cells decreased from (14.90 ± 0.22) × 10 9/L to (10.23 ± 0.24) × 10 9/L, the number of neutrophils decreased from (7.25 ± 0.18) × 10 9/L to (6.18 ± 0.11) × 10 9/L, the number of lymphocytes decreased from (2.80 ± 0.07) × 10 9/L to (1.64 ± 0.06) × 10 9/L, the number of monocytes decreased from (1.13 ± 0.01) × 10 9/L to (0.53 ± 0.02) × 10 9/L, and the number of eosinophils decreased from (1.70 ± 0.06) × 10 9/L to (0.71 ± 0.03) × 10 9/L. Conclusion:Different clinical symptoms of Leishmania-infected dogs have different blood test results.

Antileishmanial effects, cellular mechanisms, and cytotoxicity of Elettaria cardamomum essential oil against Leishmania major infection

@#Leishmaniasis is an infectious disease with various clinical manifestations. We studied the therapeutic effects of Elettaria cardamomum essential oil (ECEO) against Leishmania major infection. In vitro effects of ECEO against L. major were examined by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and macrophage assays. Nitric oxide (NO) production, infection inhibition in macrophages, and the apoptotic activity of ECEO in treated parasites were also measured. By calculating the 50% cytotoxic concentrations (CC50), we studied the cytotoxicity effects of ECEO on human macrophage cells (THP-1). The efficacy of ECEO for improving cutaneous leishmaniasis (CL) lesions in mice (BALB/c) was determined by evaluating the size of lesions and the number of amastigotes before and after four weeks of treatment. The effects of ECEO on liver and kidney function in the tested mice were also evaluated. ECEO dose-dependently (p<0.001) inhibited the viability and the mean number of promastigotes and amastigote forms of L. tropica. Four weeks of treatment with ECEO at the doses of 2.5 and 5 mg/kg/ day significantly (p<0.001) improved the CL lesions and reduced the number of parasites in the infected mice. ECEO significantly increased NO production, apoptosis induction, and infection rate in parasites. The CC50 value for ECEO and MA was 303.4 µg/mL and 835.2 µg/mL, respectively. In the mice receiving ECEO at the doses of 2.5 and 5 mg/kg/day for 28 days, no significant change was reported between the serum level of liver enzymes and kidney factors when compared with the control group. ECEO displayed promising efficacy in parasite reduction in vitro and in the animal model. ECEO can thus be used as an alternative medicine to treat CL.

Desenvolvimento e avaliação de uma proteína quimérica recombinante para o diagnóstico e prognóstico das leishmanioses tegumentar e visceral / Development and evaluation of a recombinant chimeric protein for the diagnosis and prognosis of tegumentary and visceral leishmaniasis

As leishmanioses são doenças tropicais negligenciadas com alta endemicidade e que afetam milhares de pessoas no mundo. Sua infecção é causada por parasitos protozoários do gênero Leishmania. A diversidade biológica entre as espécies é quem permite determinar as manifestações clínicas, sendo elas na forma de leishmaniose visceral (LV) ou leishmaniose tegumentar (LT). Dentre estas manifestações, a LV é considerada a mais grave, devido sua alta letalidade e grande emergência em indivíduos com a infecção provocada pelo vírus da imunodeficiência humana (HIV). Atualmente, as medidas de controle e prevenção adotadas pela Organização Mundial da Saúde (OMS), baseiam-se em uma combinação de estratégias de intervenção contra a infecção, uma vez que o diagnóstico eficaz e precoce é indispensável para que se possa intervir com o tratamento adequado, diminuindo índices de mortalidade e a evolução de complicações clínicas. Entretanto, os testes sorológicos utilizados apresentam sensibilidade e especificidade prejudicadas em pacientes com leishmanioses e/ou coinfectados LV/HIV, devido a baixos ní-veis de anticorpos antileishmanial ou pela presença de doenças que causem reação cruzada, levando a resultados falso-positivos. A sensibilidade torna-se também variável em pacientes tratados, uma vez que a sorologia pode manter-se positiva por meses ou anos após o fim do tratamento e cura da doença. Buscando resolver tal problemática, a identificação de novos antígenos, por meio de análises de bioinformática associadas à imunoproteômica, tem permitido a detecção de novas proteínas com potencial aplicação diagnóstica. Em estudos anteriores, as proteínas hipotéticas LiHyT, LiHyD, LiHyV e LiHyP foram encontradas em espécies de Leishmania spp, e avaliadas em suas versões recombinantes por meio de ensaios de ELISA, obtendo resultados satisfatórios para a detecção da LV humana e canina. Com base nessas informações, o presente trabalho teve como objetivo desenvolver uma proteína quimera recombinante base-ada na predição de epítopos lineares específicos de células B das quatro proteínas antigênicas de L. infantum citadas e avaliar o potencial diagnóstico, assim como dos peptídeos individuais que a constituíram, frente à leishmaniose humana, bem como com a coinfecção com HIV, além de testar os antígenos como marcadores prognóstico após o tratamento da LV e LT. As sequências de aminoácidos das proteínas foram avaliadas e oito epítopos de células B foram preditos e utilizados na construção de uma nova proteína quimérica. A proteína foi expressa, purificada e avaliada como antígeno recombinante em ELISA para o diagnóstico de LV, LT, coinfecção LV/HIV e prognóstico em amostras de pacientes tratados de LV e LT. Os epítopos de células B usados na construção da quimera foram sintetizados e também testados em ELISA frente às mesmas amostras, assim como um extrato antigênico solúvel de Leishmania braziliensis (SLA). Os resultados mostraram que a proteína quimera apresentou sensibilidade e especificidade de 100% para diagnosticar a LV, LT e LV/HIV, enquanto os peptídeos sintéticos apresentaram sensibilidade variando entre eles de 9,1% a 90,9% para amostras de LT e 76,8% a 99,2% para amostras de LV e LV/HIV, já os valores de especificidade atingiram 98,3% a 99,1% para LT e 67,1% a 95,7% para LV e LV/HIV. O SLA apresentou sensibilidade e especificidade de 18,2% e 98,3% para LT, e 56,8% a 69,5% para amostras de LV e LV/HIV, respectivamente. Uma avaliação prognóstica preliminar mostrou ainda que os anticorpos anti-quimera diminuíram em níveis significativos, quando comparada a reatividade sorológica antes e seis meses após o tratamento, sugerindo um possível papel prognóstico da quimera para as leishmanioses. O presente estudo, mostrou-se eficaz na construção e avaliação de novos candidatos, que demonstram ter um bom desempenho na detecção diagnóstica e prognóstica para as leishmanioses e dos casos de coinfecção LV/HIV.

Leishmaniasis are neglected tropical diseases with high endemicity that affect thousands of people in the world. Infection is caused by protozoan parasites of the genus Leishmania. The biological diversity between species is what allows determining the clinical manifestations, either in the form of visceral leishmaniasis (VL) or tegumentary leishmaniasis (TL). Among these clinical manifestations, VL is considered the most serious, due to its high lethality and great emergence in individuals with infection caused by the human immunodeficiency virus (HIV). Currently, the control and prevention measures adopted by the World Health Organiza-tion (WHO) are based on a combination of intervention strategies against the infection, since an effective and early diagnosis is essential to intervene with the appropriate treatment, decrea-sing mortality rates and evolution of clinical complications. However, the serological tests used show impaired sensitivity and specificity in patients with leishmaniasis and/or coinfected with VL/HIV, due to low levels of anti-leishmanial antibodies or the presence of diseases that cause cross-reaction, leading to false-positive results. The sensitivity also becomes variable in treated patients, since the serology can remain positive for months or years after the end of the trea-tment and cure of the disease. Seeking to solve this problem, the identification of new antigens, through bioinformatic analysis associated with immunoproteomic, has allowed the detection of new proteins with potential diagnostic application. In previous studies, the hypothetical proteins LiHyT, LiHyD, LiHyV and LiHyP were found in species of Leishmania spp, and evaluated in their recombinant versions through ELISA assays, and satisfactory results were obtained for the detection of human and canine VL. Based on this information, the present work aimed to develop a recombinant chimera protein through on the prediction of specific linear epitopes of B cells derived from these four antigenic proteins of L. infantum and to evaluate its diagnostic potential, as well as the individual peptides that constitute it, against human leishmaniasis, as well as co-infection with HIV, in addition to testing them as possible prognostic markers of patients after VL and TL treatment. The amino acid sequences of the proteins were evaluated and eight B cell epitopes were predicted and used in the construction of a new chimeric protein. The protein was expressed, purified and evaluated as a recombinant antigen in ELISA for the diagnosis of VL, TL, VL/HIV co-infection and prognosis in samples from patients treated for VL and TL. The B cell epitopes used in the construction of the chimera were synthesized and also tested in ELISA against the same samples, as well as a soluble Leishmania braziliensis antigenic extract (SLA). The results showed that the chimera protein apresented sensitivity and specificity of 100% for diagnosing VL, TL and VL/HIV, while the synthetic peptides showed sensitivity ranging from 9.1% to 90.9% for TL samples and 76.8 % to 99.2% for VL and VL/HIV samples, while the specificity values reached from 98.3% to 99.1% for TL and 67.1% to 95.7% for VL and VL/HIV. The SLA showed sensitivity and specificity of 18.2% and 98.3% for TL, and 56.8% to 69.5% for VL and VL/HIV samples, respectively. A preliminary prog-nostic evaluation also showed that anti-chimera antibodies significantly decreased when com-pared to serological reactivity before and six months after treatment, suggesting a possible prognostic role of the antigen for leishmaniasis. The present study proved to be effective in the construction and evaluation of new candidates, who demonstrate good performance in diagnos-tic and prognostic detection for leishmaniasis and VL/HIV co-infection.

Genetic variability of Leishmania (Leishmania) infantum causing human visceral leishmaniasis in the Southeastern Brazil

ABSTRACT Leishmania infantum is a protozoan that causes visceral leishmaniasis (VL) in the Americas and some regions of Europe. The disease is mainly characterized by hepatosplenomegaly and fever, and can be fatal. Factors related to the host and parasite can contribute to the transmission of Leishmania and the clinical outcome. The intraspecific genetic variability of L. infantum strains may be one of these factors. In this study, we evaluated the genetic variability of L. infantum obtained from bone marrow smear slides from patients in the Sao Paulo State, Brazil. For this, the minicircle of the kDNA hypervariable region was used as target by Sanger sequencing. By analyzing the similarity of the nucleotides and the maximum likelihood tree (Fasttree), we observed a high similarity (98%) among samples. Moreover, we identified four different profiles of L. infantum. In conclusion, L. infantum strains from Sao Paulo State, Brazil, showed low diversity measured by minicircle of the kDNA hypervariable region.

Chemical composition, in vitro larvicidal and antileishmanial activities of the essential oil from Citrus reticulata Blanco fruit peel / Composição química, atividades larvicida e leishmanicida in vitro do óleo essencial da casca do fruto de Citrus reticulata Blanco

Abstract Numerous studies have investigated the chemical composition and biological activities of essential oils from different Citrus species fruit peel, leaves and flowers. This paper aims to investigate the chemical composition, larvicidal and antileishmanial activities of essential oil from Citrus reticulata fruit peel (CR-EO). CR-EO was obtained by hydrodistillation in a Clevenger-type apparatus and its chemical composition was analyzed by GC-MS and GC-FID. Limonene (85.7%), ɣ-terpinene (6.7%) and myrcene (2.1%) were identified as its major components. CR-EO showed high activity against promastigote forms of Leishmania amazonensis (IC50 = 8.23 µg/mL). CR-EO also exhibited high larvicidal activity against third instar Aedes aegypti larvae at a lethal concentration (LC50 = 58.35 µg/mL) and 100% mortality at 150 µg/mL. This study suggests, for the first time, the potential use of CR-EO against this important mosquito-borne viral disease caused by the genus Aedes.

Resumo Numerosos estudos têm investigado a composição química e as atividades biológicas de óleos essenciais extraídos de cascas dos frutos, folhas e flores de diferentes espécies de Citrus. Este trabalho tem como objetivo investigar a composição química e as atividades larvicida e leishmanicida in vitro do óleo essencial das cascas dos frutos de Citrus reticulata (CR-EO). CR-EO foi obtido pela técnica de extração em aparelho Clevenger e sua composição química foi determinada por CG-EM e CG-DIC. Limoneno (85,7%), ɣ-terpineno (6,7%) and mirceno (2,1%) foram identificados como os constituintes majoritários. CR-EO mostrou alta atividade contra as formas promastigota de Leishmania amazonensis (CI50 = 8,23 µg/mL). CR-EO também exibiu alta atividade larvicida contra as larvas do terceiro estágio do Aedes aegypti com concentração letal (CL50 = 58,35 µg/mL) e mortalidade de 100% em 150 µg/mL. Este estudo sugere, pela primeira vez, o uso potencial de CR-EO contra esta importante doença viral transmitida por mosquitos do gênero Aedes.

Chemical composition, in vitro larvicidal and antileishmanial activities of the essential oil from Citrus reticulata Blanco fruit peel / Composição química, atividades larvicida e leishmanicida in vitro do óleo essencial da casca do fruto de Citrus reticulata Blanco

Numerous studies have investigated the chemical composition and biological activities of essential oils from different Citrus species fruit peel, leaves and flowers. This paper aims to investigate the chemical composition, larvicidal and antileishmanial activities of essential oil from Citrus reticulata fruit peel (CR-EO). CR-EO was obtained by hydrodistillation in a Clevenger-type apparatus and its chemical composition was analyzed by GC-MS and GC-FID. Limonene (85.7%), ɣ-terpinene (6.7%) and myrcene (2.1%) were identified as its major components. CR-EO showed high activity against promastigote forms of Leishmania amazonensis (IC50 = 8.23 µg/mL). CR-EO also exhibited high larvicidal activity against third instar Aedes aegypti larvae at a lethal concentration (LC50 = 58.35 µg/mL) and 100% mortality at 150 µg/mL. This study suggests, for the first time, the potential use of CR-EO against this important mosquito-borne viral disease caused by the genus Aedes.

Numerosos estudos têm investigado a composição química e as atividades biológicas de óleos essenciais extraídos de cascas dos frutos, folhas e flores de diferentes espécies de Citrus. Este trabalho tem como objetivo investigar a composição química e as atividades larvicida e leishmanicida in vitro do óleo essencial das cascas dos frutos de Citrus reticulata (CR-EO). CR-EO foi obtido pela técnica de extração em aparelho Clevenger e sua composição química foi determinada por CG-EM e CG-DIC. Limoneno (85,7%), ɣ-terpineno (6,7%) and mirceno (2,1%) foram identificados como os constituintes majoritários. CR-EO mostrou alta atividade contra as formas promastigota de Leishmania amazonensis (CI50 = 8,23 µg/mL). CR-EO também exibiu alta atividade larvicida contra as larvas do terceiro estágio do Aedes aegypti com concentração letal (CL50 = 58,35 µg/mL) e mortalidade de 100% em 150 µg/mL. Este estudo sugere, pela primeira vez, o uso potencial de CR-EO contra esta importante doença viral transmitida por mosquitos do gênero Aedes.

Prevalence of cutaneous leishmaniasis in the largest populated city Karachi, Pakistan / Prevalência de leishmaniose cutânea na maior cidade populosa de Karachi, Paquistão

The study was undertaken from September 2018 to April 2019 to determine the prevalence of cutaneous leishmaniasis in human beings living in six districts of Karachi. Suspected persons were screened for the disease and positive cases were identified on the basis of skin lesions and blood samples. Samples were observed by mounting their smear. A total of 207 subjects of different ages and sex groups were investigated, however, only 192 (92%) of the suspected cases were found to have the disease 64% of cases were male which were significantly high (p<0.05), than female 36%. The lesion was more frequently detected among the youth ages of 21-30 years (31%) as compared to other groups. In both sexes, legs were found to be more infected (25% male + 20% female) followed by arms (20% male + 0% female) and face (15% male +11% female). The mixed body parts had shown the lowest infections such as (4% in males + 5%) in females. In conclusion, the highest and lowest leishmaniasis infections were observed in District West (23% male + 9% female) followed by District East (15% male + 7% female), District Malir (11% male+ 4% female), District Central (7% male + 5% female), District Korangi (4% male + 7% female) and District South (4% male + 4% female) respectively.

O estudo foi realizado de setembro de 2018 a abril de 2019 para determinar a prevalência de leishmaniose tegumentar em seres humanos que vivem em seis distritos de Karachi. Pessoas suspeitas foram rastreadas para a doença e os casos positivos foram identificados com base em lesões de pele e amostras de sangue. As amostras foram observadas montando seu esfregaço. Um total de 207 indivíduos de diferentes idades e grupos sexuais foi investigado, no entanto apenas 192 (92%) dos casos suspeitos foram encontrados para ter a doença; 64% dos casos eram do sexo masculino, que foram significativamente elevados (p < 0,05), e do sexo feminino 36%. A lesão foi detectada com maior frequência entre os jovens de 21 a 30 anos (31%) em comparação com os outros grupos. Em ambos os sexos, as pernas estavam mais infectadas (25% homens + 20% mulheres), seguidas pelos braços (20% homens + 0% mulheres) e rosto (15% homens + 11% mulheres). As partes mistas do corpo mostraram as infecções mais baixas (4% homens + 5% mulheres). Em conclusão, as infecções de leishmaniose mais altas e mais baixas foram observadas no Distrito Oeste (23% homens + 9% mulheres) seguido pelo Distrito Leste (15% homens + 7% mulheres), Distrito Malir (11% homens + 4% mulheres), Distrito Central (7% homens + 5% mulheres), Distrito Korangi (4% homens + 7% mulheres) e Distrito Sul (4% homens + 4% mulheres), respectivamente.

Chemical composition, in vitro larvicidal and antileishmanial activities of the essential oil from Citrus reticulata Blanco fruit peel

Abstract Numerous studies have investigated the chemical composition and biological activities of essential oils from different Citrus species fruit peel, leaves and flowers. This paper aims to investigate the chemical composition, larvicidal and antileishmanial activities of essential oil from Citrus reticulata fruit peel (CR-EO). CR-EO was obtained by hydrodistillation in a Clevenger-type apparatus and its chemical composition was analyzed by GC-MS and GC-FID. Limonene (85.7%), -terpinene (6.7%) and myrcene (2.1%) were identified as its major components. CR-EO showed high activity against promastigote forms of Leishmania amazonensis (IC50 = 8.23 µg/mL). CR-EO also exhibited high larvicidal activity against third instar Aedes aegypti larvae at a lethal concentration (LC50 = 58.35 µg/mL) and 100% mortality at 150 µg/mL. This study suggests, for the first time, the potential use of CR-EO against this important mosquito-borne viral disease caused by the genus Aedes.

Resumo Numerosos estudos têm investigado a composição química e as atividades biológicas de óleos essenciais extraídos de cascas dos frutos, folhas e flores de diferentes espécies de Citrus. Este trabalho tem como objetivo investigar a composição química e as atividades larvicida e leishmanicida in vitro do óleo essencial das cascas dos frutos de Citrus reticulata (CR-EO). CR-EO foi obtido pela técnica de extração em aparelho Clevenger e sua composição química foi determinada por CG-EM e CG-DIC. Limoneno (85,7%), -terpineno (6,7%) and mirceno (2,1%) foram identificados como os constituintes majoritários. CR-EO mostrou alta atividade contra as formas promastigota de Leishmania amazonensis (CI50 = 8,23 µg/mL). CR-EO também exibiu alta atividade larvicida contra as larvas do terceiro estágio do Aedes aegypti com concentração letal (CL50 = 58,35 µg/mL) e mortalidade de 100% em 150 µg/mL. Este estudo sugere, pela primeira vez, o uso potencial de CR-EO contra esta importante doença viral transmitida por mosquitos do gênero Aedes.

Prevalence of cutaneous leishmaniasis in the largest populated city Karachi, Pakistan

Abstract. The study was undertaken from September 2018 to April 2019 to determine the prevalence of cutaneous leishmaniasis in human beings living in six districts of Karachi. Suspected persons were screened for the disease and positive cases were identified on the basis of skin lesions and blood samples. Samples were observed by mounting their smear. A total of 207 subjects of different ages and sex groups were investigated, however, only 192 (92%) of the suspected cases were found to have the disease 64% of cases were male which were significantly high (p 0.05), than female 36%. The lesion was more frequently detected among the youth ages of 21-30 years (31%) as compared to other groups. In both sexes, legs were found to be more infected (25% male + 20% female) followed by arms (20% male + 0% female) and face (15% male +11% female). The mixed body parts had shown the lowest infections such as (4% in males + 5%) in females. In conclusion, the highest and lowest leishmaniasis infections were observed in District West (23% male + 9% female) followed by District East (15% male + 7% female), District Malir (11% male+ 4% female), District Central (7% male + 5% female), District Korangi (4% male + 7% female) and District South (4% male + 4% female) respectively.

Resumo O estudo foi realizado de setembro de 2018 a abril de 2019 para determinar a prevalência de leishmaniose tegumentar em seres humanos que vivem em seis distritos de Karachi. Pessoas suspeitas foram rastreadas para a doença e os casos positivos foram identificados com base em lesões de pele e amostras de sangue. As amostras foram observadas montando seu esfregaço. Um total de 207 indivíduos de diferentes idades e grupos sexuais foi investigado, no entanto apenas 192 (92%) dos casos suspeitos foram encontrados para ter a doença; 64% dos casos eram do sexo masculino, que foram significativamente elevados (p 0,05), e do sexo feminino 36%. A lesão foi detectada com maior frequência entre os jovens de 21 a 30 anos (31%) em comparação com os outros grupos. Em ambos os sexos, as pernas estavam mais infectadas (25% homens + 20% mulheres), seguidas pelos braços (20% homens + 0% mulheres) e rosto (15% homens + 11% mulheres). As partes mistas do corpo mostraram as infecções mais baixas (4% homens + 5% mulheres). Em conclusão, as infecções de leishmaniose mais altas e mais baixas foram observadas no Distrito Oeste (23% homens + 9% mulheres) seguido pelo Distrito Leste (15% homens + 7% mulheres), Distrito Malir (11% homens + 4% mulheres), Distrito Central (7% homens + 5% mulheres), Distrito Korangi (4% homens + 7% mulheres) e Distrito Sul (4% homens + 4% mulheres), respectivamente.