Prevalence and Distribution of Leishmania RNA Virus 1 in Leishmania Parasites from French Guiana.

In South America, the presence of the Leishmania RNA virus type 1 (LRV1) was described in Leishmania guyanensis and Leishmania braziliensis strains. The aim of this study was to determine the prevalence distribution of LRV1 in Leishmania isolates in French Guiana given that, in this French overseas department, most Leishmania infections are due to these parasite species. The presence of the virus was observed in 74% of Leishmania spp. isolates, with a highest presence in the internal areas of the country.

Presence of Leishmania RNA Virus 1 in Leishmania guyanensis Increases the Risk of First-Line Treatment Failure and Symptomatic Relapse.

Treatment failure and symptomatic relapse are major concerns in American tegumentary leishmaniasis (TL). Such complications are seen frequently in Leishmania guyanensis infections, in which patients respond variously to first-line antileishmanials and are more prone to develop chronic cutaneous leishmaniasis. The factors underlying this pathology, however, are unknown. Recently, we reported that a double-stranded RNA virus, Leishmania RNA virus 1 (LRV1), nested within L. guyanensis parasites is able to exacerbate experimental murine leishmaniasis by inducing a hyperinflammatory response. This report investigates the prevalence of LRV1 in human L. guyanensis infection and its effect on treatment efficacy, as well as its correlation to symptomatic relapses after the completion of first-line treatment. In our cohort of 75 patients with a diagnosis of primary localized American TL, the prevalence of LRV1-positive L. guyanensis infection was elevated to 58%. All patients infected with LRV1-negative L. guyanensis were cured after 1 dose (22 of 31 [71%]) or 2 doses (31 of 31 [100%]) of pentamidine. In contrast, 12 of 44 LRV1-positive patients (27%) presented with persistent infection and symptomatic relapse that required extended therapy and the use of second-line drugs. Finally, LRV1 presence was associated with a significant increase in levels of intra-lesional inflammatory markers. In conclusion, LRV1 status in L. guyanensis infection is significantly predictive (P = .0009) of first-line treatment failure and symptomatic relapse and has the potential to guide therapeutic choices in American TL.

Severe Cutaneous Leishmaniasis in a Human Immunodeficiency Virus Patient Coinfected with Leishmania braziliensis and Its Endosymbiotic Virus.

Leishmaniaparasites cause a broad range of disease, with cutaneous afflictions being, by far, the most prevalent. Variations in disease severity and symptomatic spectrum are mostly associated to parasite species. One risk factor for the severity and emergence of leishmaniasis is immunosuppression, usually arising by coinfection of the patient with human immunodeficiency virus (HIV). Interestingly, several species ofLeishmaniahave been shown to bear an endogenous cytoplasmic dsRNA virus (LRV) of theTotiviridaefamily, and recently we correlated the presence of LRV1 withinLeishmaniaparasites to an exacerbation murine leishmaniasis and with an elevated frequency of drug treatment failures in humans. This raises the possibility of further exacerbation of leishmaniasis in the presence of both viruses, and here we report a case of cutaneous leishmaniasis caused byLeishmania braziliensisbearing LRV1 with aggressive pathogenesis in an HIV patient. LRV1 was isolated and partially sequenced from skin and nasal lesions. Genetic identity of both sequences reinforced the assumption that nasal parasites originate from primary skin lesions. Surprisingly, combined antiretroviral therapy did not impact the devolution ofLeishmaniainfection. TheLeishmaniainfection was successfully treated through administration of liposomal amphotericin B.

Implication of different domains of the Leishmania major metacaspase in cell death and autophagy.

Metacaspases (MCAs) are cysteine peptidases expressed in plants, fungi and protozoa, with a caspase-like histidine-cysteine catalytic dyad, but differing from caspases, for example, in their substrate specificity. The role of MCAs is subject to debate: roles in cell cycle control, in cell death or even in cell survival have been suggested. In this study, using a Leishmania major MCA-deficient strain, we showed that L. major MCA (LmjMCA) not only had a role similar to caspases in cell death but also in autophagy and this through different domains. Upon cell death induction by miltefosine or H2O2, LmjMCA is processed, releasing the catalytic domain, which activated substrates via its catalytic dyad His/Cys and a proline-rich C-terminal domain. The C-terminal domain interacted with proteins, notably proteins involved in stress regulation, such as the MAP kinase LmaMPK7 or programmed cell death like the calpain-like cysteine peptidase. We also showed a new role of LmjMCA in autophagy, acting on or upstream of ATG8, involving Lmjmca gene overexpression and interaction of the C-terminal domain of LmjMCA with itself and other proteins. These results allowed us to propose two models, showing the role of LmjMCA in the cell death and also in the autophagy pathway, implicating different protein domains.

Proteases de Leishmania: novos alvos para o desenvolvimento racional de fármacos

Leishmania causes tegumental and visceral diseases called leishmaniasis. Disease control is possible interrupting the transmission cycle, but HIV co-infection, chemotheraphy toxicity and lack of a vaccine are paramount difficulties. So, is necessary to study new Leishmania molecules and investigate the possibility to develop rational drugs using these molecules as targets. Leishmania express many peptidases during their life, and cysteine are the most abundant protease and many inhibitors were developed but failed to kill parasites. On the other hand, inhibitors of serine proteases killed promastigotes, indicating the possibility of these enzymes to be important targets in the development of anti-Leishmania drugs.

Síntese, caracterização e estudo da atividade inibitória de novas dialquilfosforilarilidrazonas sobre o crescimento de tripanossomatídeos

A new series of dialkylphosphorylhydrazones was synthesized through the condensation of aromatic aldehydes with different phosphorylhydrazines. All synthesized compounds were characterized by IR, ¹H-NMR, 13C-NMR and 31P-NMR spectroscopies. The in vitro investigation of the activity of these compounds against Leishmania amazonensis promastigotes and epimastigotes of T. cruzi, showed an efficient inhibition of proliferation, at non toxic concentrations to mammalian cells. The results have shown some derivatives as potential antiparasitic agents against trypanosomatids.

Estudo químico e atividades antiproliferativa, tripanocida e leishmanicida de Maxillaria picta

The chemical study of the orchid Maxillaria picta resulted in the isolation of the bioactive stilbenes phoyunbene B and phoyunbene C, in addition to four phenolic acids, one xanthone, steroidal compounds and two triterpenes. Crude extract, fractions, subfractions and the isolated xanthone were evaluated for anticancer activity against human tumor cell lines and against evolutionary forms of T. cruzi and L. amazonensis. The structures of the compounds were determined by GC-MS, and ¹H NMR, 13C NMR spectral methods as well as bidimensional techniques.

An enzyme-linked immunosorbent assay (ELISA) for the detection of IgM antibodies against Leishmania chagasi in dogs

Visceral leishmaniasis is an emergent zoonosis with an increasing number of new cases in Brazil where the domestic dog is an important parasite reservoir in the infectious cycle of Leishmania chagasi. An enzyme-linked immunosorbent assay (ELISA), based upon the use of a total soluble antigenic preparation of L. chagasi, was adapted for the detection of IgM antibodies in the serum of infected dogs. Optimal dilutions of the antigen, using positive and negative reference sera, were determined by checkboard titrations. The specificity and sensitivity of the ELISA were 100 %. A total of 110 serum samples were taken from dogs in Belo Horizonte, Minas Gerais, Brazil, and examined for anti-L. chagasi IgM antibody by ELISA and indirect fluorescent antibody test (IFAT). About 25% (n=27) of all the dogs tested were found serologically positive for L. chagasi by IFAT, while 89.09% (n=98) were seropositive by ELISA. The results obtained by ELISA and IFAT were significantly different (P<0.01). The combined use of ELISA and IFAT is recommended in order to enable veterinary services to more efficiently detect canine visceral leishmaniasis.

Análise histomorfométrica da matriz extracelular do linfonodo poplíteo de cães naturalmente infectados por Leishmania (L.) chagasi

Nas Américas, a leishmaniose visceral canina é causada por Leishmania (Leishmania) chagasi, um protozoário intracelular obrigatório do sistema fagocítico mononuclear; as principais alterações histológicas associadas a essa doença ocorrem nos em órgãos linfóides. Apesar de o cão ser considerado o principal mantenedor e disseminador da leishmaniose no ambiente urbano, são escassos estudos dos aspectos histopatológicos e histomorfométricos, em cães naturalmente infectados com L. chagasi, que investiguem a interação entre o parasito e a matriz extracelular. Este estudo visou caracterizar e quantificar as alterações dos componentes celulares e da matriz extracelular (colágenos I e III) do linfonodo poplíteo de 22 cães com infecção natural por L. chagasi detectada através da reação de imunofluorescência indireta (RIF) e compará-las com as alterações encontradas no linfonodo poplíteo de 10 cães não-infectados, negativos na RIF e clinicamente saudáveis. Fragmentos dos linfonodos foram seccionados longitudinalmente, processados rotineiramente para exame histológico e corados por hematoxilina-eosina. Cortes adicionais do mesmo linfonodo incluídos em glicol metacrilato foram corados pelo azul de toluidina para histomorfometria. Linfonodos de cães infectados apresentaram linfadenopatia generalizada, aumento do tamanho e do número dos folículos linfóides, hipertrofia da cápsula e hiperplasia linfóide significativa. Nos linfonodos de cães do grupo infectado, a análise quantitativa de fibras colágenas mostrou significativo predomínio do colágeno I sobre o colágeno III. Esses resultados demonstram que cães infectados por L. chagasi apresentam degradação dos constituintes da matriz extracelular e conseqüente destruição do arcabouço linfóide, alterando a morfologia do órgão.

Presença de formas amastigotas de Leishmania chagasi e perfil leucocitário no aparelho reprodutivo de cães

A leishmaniose visceral (LV) é uma zoonose causada pelo protozoário Leishmania (Leishmania) chagasi. A leishmaniose visceral canina (LVC) é a doença de maior relevância zoonótica. Usualmente, a infecção ocorre entre um hospedeiro invertebrado para um hospedeiro vertebrado, entretanto, a transmissão na ausência do vetor já é conhecida. O objetivo principal deste estudo foi identificar a presença de formas amastigotas, quantificar as células leucocitárias, estimar o risco relativo da presença de formas amastigotas no aparelho reprodutivo de cães sorologicamente positivos com e sem sinais clínicos. Para isso, foram utilizados cães sem raça definida, sexualmente maduros e testados sorologicamente para LVC (com sinais clínicos, n=25; sem sinais clínicos, n=25), que após eutanásia, tiveram fragmentos de testículo, epidídimo (cabeça, corpo e cauda) e glândula prostática (selecionados ao acaso) impressos em lâminas. Um grupo de 20 cãs sorologicamente negativos e sem sinais clínicos foi usado como controle. Amostras do baço foram incluídas como controle parasitológico positivo. O percentual de linfócitos foi superior (P<0,05) no corpo e cauda do epidídimo, assim como no testículo. Macrófagos foram superiores (P<0,05) apenas nas regiões do corpo e cauda epididimais. A presença de amastigotas correlacionou-se entre as distintas regiões do aparelho reprodutivo. Nos sintomáticos variaram entre 0,50 a 0,80 e entre 0,79 a 0,95 nos assintomáticos. A presença de amastigotas no testículo dos cães sintomáticos foi 6,5 vezes superior aos cães assintomáticos. Os resultados obtidos demonstram o potencial epidemiológico da transmissão venérea da doença, principalmente em áreas onde os programas de controle da LVC não consideram esta forma de transmissão, que pode ser importante em populações caninas não esterilizadas.