Coinfecções por Leishmania infantum, Neospora caninum e Toxoplasma gondii em cães necropsiados da região central do Rio Grande do Sul, Brasil / Coinfections by Leishmania infantum, Neospora caninum and Toxoplasma gondii in necropsied dogs from the central region of Rio Grande do Sul, Brazil

O objetivo do presente trabalho foi determinar a presença de anticorpos para Leishmania infantum, Neospora caninum e Toxoplasma gondii, por meio da reação de imunofluorescência indireta (RIFI), em cães (n=78) provenientes da região central do Rio Grande do Sul, necropsiados no Hospital Veterinário da Universidade Federal de Santa Maria (UFSM), bem como avaliar os dados epidemiológicos, sazonais e anátomo-histopatológicos. Do total de animais avaliados, 67,9% (53/78) apresentaram soropositividade para ao menos um agente. A ocorrência de anticorpos para L. infantum, N. caninum e T. gondii foi de 33,3 (26/78), 37,1 (29/78) e 43,5% (34/78), respectivamente. Detectaram-se monoinfecções em 9,4% (5/53) para L. infantum, 18,8% (10/53) para N. caninum e 20,7% (11/53) para T. gondii. As coinfecções foram observadas em 27/53 (50,9%) dos animais. As infecções ocorreram independentemente de idade, sexo, procedência ou raça (P>0,05). Não se verificaram lesões anatomo-histopatológicas relacionadas aos agentes pesquisados, caracterizando-os como animais assintomáticos. Os resultados confirmaram a exposição de cães a esses protozoários na região central do RS e, em especial, demonstraram a circulação do agente causador da leishmaniose em uma área considerada indene para a enfermidade.(AU)

The present paper is aimed to determine the presence of antibodies for Leishmania infantum, Neospora caninum and Toxoplasma gondii by indirect immunofluorescence (IIF) in dogs (n=78) from the central region in the state of Rio Grande do Sul necropsied in the Veterinary Hospital from Universidade Federal de Santa Maria (UFSM). The data was evaluated regarding epidemiological, anatomic, and histopathologic findings. Of the total animals evaluated, 67.9% (53/78) showed seropositivity for at least one agent. The occurrence of antibodies to L. infantum, N. caninum and T. gondii was 33.3% (26/78) 37.1% (29/78) and 43.5% (34/78), respectively. The mono infections were detected in 9.4% (5/53) of L. infantum, 18.8% (10/53) for N. caninum and 20.7% (11/53) T. gondii. The coinfections occurred in 50.9% (27/53) of animals. There were not anatomical and histopathological lesions regarding these surveyed agents, characterizing them as subclinical animals. The results confirmed the exposition of dogs to these protozoa in the central region of the RS, highlighting the circulation of the causer agent of leishmaniasis in an area considered harmless for the disease.(AU)

Evaluation of the presence of Leishmania infantum in bone stored in glycerol or sterilized by autoclave for cortical bone grafting / Avaliação da presença de Leishmania infantum em ossos armazenados em glycerol ou esterilizados em autoclave para enxertia óssea cortical

This study was performed to evaluate the effects of autoclaving and storage in 85% glycerol on cortical bone from dogs infected with Leishmania sp. We used 42 cadavers with leishmaniasis. The dogs were evaluated for the presence of Leishmania sp. in culture of bone marrow and by culturing cortical bone. From the infected animals, we harvested 42 diaphysis of the right femur for culture of cortical bone before and after autoclaving or storage in glycerol. There was no significant difference in growth of the parasite in culture of bone marrow or cortical bone. There was no growth of Leishmania sp. in culture of samples after autoclaving or storage in glycerol. Both treatments were effective in preventing the growth of the parasite in vitro, so it was considered viable for grafting.(AU)

Este estudo teve por objetivo avaliar os efeitos da esterilização em autoclave e do armazenamento em glicerol a 85% no osso cortical de cães infectados por Leishmania sp. Foram utilizados 42 cadáveres de cães com leishmaniose. Os cães foram avaliados para a presença de Leishmania sp. em cultura de medula óssea e de osso cortical. Foram coletadas 42 diáfises do fêmur direito para cultura do osso cortical antes e após a esterilização e o armazenamento em glicerol. Não houve diferença significativa no crescimento do parasito em cultura utilizando-se medula óssea ou osso cortical. Não houve crescimento de Leishmania sp. em cultura de amostras de osso cortical após a autoclavagem ou o armazenamento em glicerol. Ambos os tratamentos foram eficazes na prevenção do crescimento do parasita in vitro, de modo que foram considerados viáveis para enxerto.(AU)

Estudo da coinfecção Leishmania infantum e Ehrlichia canis em cães numa área endêmica para leishmaniose visceral canina / Study co infection Leishmania infantum and Ehrlichia canis in dogs in an endemic area for canine visceral leishmaniasis

A Erliquiose Monocítica Canina (EMC) e a Leishmaniose Visceral Canina (LVC) são duas doenças, transmitidas por vetores, com ampla distribuição mundial. Os agentes causadores dessas doenças são Ehrlichia canis e Leishmania infantum, respectivamente. Enquanto a EMC é transmitida por carrapatos, principalmente Rhipicephalus sanguineus, a Leishmania é inoculada no hospedeiro através do inseto-vetor da subfamilia Phlebotominae. A maioria dos sinais físicos são comuns às duas enfermidades, o que dificulta o diagnóstico clínico e o tratamento, especialmente em áreas endêmicas. Poucos são os estudos relacionados à coinfecção da erliquiose e leishmaniose caninas no Brasil. No Piauí, particularmente em Teresina, ainda não se tem dados concretos sobre a prevalência dessas afecções.Os sinais físicos variam com a severidade da infecção, a resposta imune do hospedeiro e a presença de coinfecção. Assim, buscamos avaliar cães, independente dos sinais físicos, atendidos no hospital universitário e clínicas particulares da cidade de Teresina, Piauí quanto a frequência de erliquiose e leishmaniose caninas no período de março de 2012 a setembro de 2014...

The Canine Monocytic Ehrlichiosis (CME) and Canine Visceral Leishmaniasis (CVL) are the two vector-borne diseases with worldwide distribution. The causative agents of these diseases are Ehrlichia canis and Leishmania infantum, respectively. While CME is transmitted by ticks, especially Rhipicephalus sanguineus, Leishmania is inoculated into the host through the insect vector of the subfamily Phlebotominae. Most clinical signs are common to both conditions, which hinders the clinical diagnosis and treatment, especially in endemic areas. There are few studies related to the coinfection of canine ehrlichiosis and leishmaniasis in Brazil. In Piaui, particularly in Teresina, there is not yet concrete data on the prevalence of these diseases. The clinical signs vary according to the severity of the infection, to the host immune response and to the presence of coinfection. Thus, we sought to evaluate dogs, regardless of clinical signs, seen at university hospital and private clinics in the city of Teresina, Piauí, for the frequency of canine ehrlichiosis and leishmaniasis from March 2012 to September 2014...

Genes that encodes NAGT, MIF1 and MIF2 are not virulence factors for kala-azar caused by Leishmania infantum

Introduction Kala-azar is a disease resulting from infection by Leishmania donovani and Leishmania infantum. Most patients with the disease exhibit prolonged fever, wasting, anemia and hepatosplenomegaly without complications. However, some patients develop severe disease with hemorrhagic manifestations, bacterial infections, jaundice, and edema dyspnea, among other symptoms, followed by death. Among the parasite molecules that might influence the disease severity are the macrophage migration inhibitory factor-like proteins (MIF1 and MIF2) and N-acetylglucosamine-1-phosphotransferase (NAGT), which act in the first step of protein N-glycosylation. This study aimed to determine whether MIF1, MIF2 and NAGT are virulence factors for severe kala-azar. Methods To determine the parasite genotype in kala-azar patients from Northeastern Brazil, we sequenced the NAGT genes of L. infantum from 68 patients as well as the MIF1 and MIF2 genes from 76 different subjects with diverse clinical manifestations. After polymerase chain reaction (PCR), the fragments were sequenced, followed by polymorphism identification. Results The nucleotide sequencing of the 144 amplicons revealed the absence of genetic variability of the NAGT, MIF1 and MIF2 genes between the isolates. The conservation of these genes suggests that the clinical variability of kala-azar does not depend upon these genes. Additionally, this conservation suggests that these genes may be critical for parasite survival. Conclusions NAGT, MIF1 and MIF2 do not alter the severity of kala-azar. NAGT, MIF1 and MIF2 are highly conserved among different isolates of identical species and exhibit potential for use in phylogenetic inferences or molecular diagnosis. .

Papel do fator de ativação de plaquetas na infecção de macrófagos por leishmania infantum e identificação de uma Paf-Acetilhidrolase no parasita / Role of platelet-activating factor in macrophage infection by Leishmania infantum and identification of the parasite PAF acetylhydrolase

A leishmaniose visceral é causada pelo parasita Leishmania infantum. A infecção ocorre quando flebótomos infectados se alimentam na derme do hospedeiro vertebrado, inoculando o parasita. A infecção produz uma resposta com diversas moléculas inflamatórias, como os mediadores lipídicos. O fator de ativação de plaquetas (PAF) é um potente mediador lipídico derivado de um lisofosfolipídio. PAF participa da fisiologia normal da célula e possui um perfil pró-inflamatório. A participação de mediadores lipídicos, como eicosanóides e PAF, já foi identificada na imunopatogênese das leishmanioses. PAF gerado pelo hospedeiro tem efeito leishmanicida e de controle da infecção por L. amazonensis. PAF-acetilhidrolases (PAF-AH) são fosfolipases A2 que hidrolisam PAF e foi demonstrado que PAF-AH podem ser um fator de virulência devido a essa habilidade. O objetivo desse estudo foi avaliar o papel do PAF e de uma PAF-AH na infecção de macrófagos por L. infantum. Foi observado que PAF 1μM, quando adicionado durante e após a infecção, foi capaz de diminuir 50% da infecção após 72 horas, bem como a viabilidade dos parasitas dentro dos macrófagos num mecanismos independente do seu receptor PAFR e da produção de óxido nítrico. PAF 10μM interrompeu o crescimento de promastigotas de L. infantum em cultura axênica. Uma PAFAH, com elevada identidade e semelhança com PLA2/PAF-AH de outros tripanossomatídeos, foi identificada no genoma de L. infantum. A clonagem e expressão recombinante produziu uma proteína de cerca de 69kDa, com atividade PAF-AH. Frações celulares do parasita, enriquecidas com estruturas de membrana também apresentaram atividade PAF-AH. Os resultados indicam que PAF é capaz de diminuir a infecção de macrófagos por L. infantum e que o parasita possui uma PAF-AH funcional possivelmente envolvida com sua virulência.

Visceral leishmaniasis is caused by Leishmania infantum parasites. Infection occurs when infected sandflies feed on vertebrate host skin delivering the parasite which survive, multiply and spread on the parasitophorous vacuoles of macrophages. The inflammatory response during the infection leads to the production of diverse bioactive molecules, as lipid mediators. The platelet activating factor (PAF) is a lipid mediator derived from a lysophospholipid. PAF has a role in normal cellular physiology, acting as proinflamatory molecule. The participation of some lipid mediators, as eicosanoids and PAF has been identified in leishmaniasis. PAF produced by the host is able to kill the parasite and control the infection by L. amazonensis. PAF-acetylhydrolases (PAF-AH) are phospholipases A2 (PLA2) that hydrolyse PAF, and possibly involved in pathogen virulence. The aim of this study was to evaluate the role of PAF on macrophages infection by L. infantum and identify a PAF-AH expressed by the parasite. PAF 1μM, added during and after the infection, was able to reduce approximately 50% of infection, as well as, the viability of parasites inside macrophages. Apparently this reduction occurs by an classical PAF receptor and nitric oxide production independent mechanism. PAF 10μM inhibited L. infantum promastigotes growing in axenic culture. A PAF-AH with high identity to PLA2/PAF-AH of others trypanosomatids was identified in L. infantum genome. The cloning and recombinant expression produced a 69kDa protein with PAF-AH activity. Cellular fractions from parasites, with membrane structures also presented PAF-AH activity. The results suggest that PAF is able to decrease machophage infection by L. infantum witch has a functional PAFAH possibly related to its virulence.

Visceral Leishmaniasis without Fever in an 11-Month-Old Infant: a Rare Clinical Feature of Kala-azar

Visceral leishmaniasis or kala-azar is an endemic parasitic disease in some parts of the world which is characterized by fever, splenomegaly, and pancytopenia in most of the cases. Herein we report an 11 month-old male infant with diagnosis of kala-azar who presented with pallor, hepatosplenomegaly, failure to gain weight, and no history of fever. Surprisingly, fever started after beginning of meglumine antimoniate treatment in this patient. As far as we are aware of, this is a rare presentation of visceral leishmaniasis. Therefore, clinicians especially in endemic areas are highly recommended to include kala-azar among differential diagnosis of unexplained anemia without fever to prevent misdiagnosis of this potentially fatal, but treatable condition.

Susceptibility of peritoneal macrophage from different species of neotropical primates to Ex vivo Leishmania (L.) infantum chagasi-infection / Susceptibilidade do macrófago peritoneal de diferentes espécies de primatas neotropicais para a infecção ex vivo por Leishmania (L.) infantum chagasi

This study examined the susceptibility of peritoneal macrophage (PM) from the Neotropical primates: Callithrix jacchus, Callithrix penicillata, Saimiri sciureus, Aotus azarae infulatus and Callimico goeldii to ex vivo Leishmania (L.) infantum chagasi-infection, the etiological agent of American visceral leishmaniasis (AVL), as a screening assay for evaluating the potential of these non-human primates as experimental models for studying AVL. The PM-susceptibility to infection was accessed by the PM-infection index (PMI) at 24, 72 h and by the mean of these rates (FPMI), as well as by the TNF-α, IL-12 (Capture ELISA) and Nitric oxide (NO) responses (Griess method). At 24h, the PMI of A. azarae infulatus (128) was higher than those of C. penicillata (83), C. goeldii (78), S. sciureus (77) and C. jacchus (55). At 72h, there was a significant PMI decrease in four monkeys: A. azarae infulatus (128/37), C. penicillata (83/38), S. sciureus (77/38) and C. jacchus (55/12), with exception of C. goeldii (78/54). The FPMI of A. azarae infulatus (82.5) and C. goeldii (66) were higher than C. jacchus (33.5), but not higher than those of C. penicillata (60.5) and S. sciureus (57.5). The TNF-a response was more regular in those four primates which decreased their PMI at 24/72 h: C. jacchus (145/122 pg/mL), C. penicillata (154/130 pg/mL), S. sciureus (164/104 pg/mL) and A. azarae infulatus (154/104 pg/mL), with exception of C. goeldii (38/83 pg/mL). The IL-12 response was mainly prominent in A. infulatus and C. goeldii which presented the highest FPMI and, the NO response was higher in C. goeldii, mainly at 72 h. These findings strongly suggest that these New World primates have developed a resistant innate immune response mechanism capable of controlling the macrophage intracellular growth of L. (L.) i. chagasi-infection, which do not encourage their use as animal model for studying AVL.

Este estudo examinou a susceptibilidade do macrófago peritoneal (PM) dos primatas neotropicais: Callithrix jacchus, Callithrix penicillata, Saimiri sciureus, Aotus azarae infulatus e Callimico goeldii para a infecção ex vivo por Leishmania (L.) infantum chagasi, o agente etiológico da leishmaniose visceral americana (LVA), como método de triagem para avaliar o potencial desses primatas como modelo de estudo da LVA. A susceptibilidade do PM para a infecção foi investigada através do índice de infecção do PM (PMI) a intervalos de 24, 72 horas e, ainda, pela média dessas taxas (FPMI), assim como, pelas respostas do TNF-α, IL-2 (ELISA de captura) e óxido nítrico (NO) (método de Griess). Às 24hs da infecção experimental, o PMI do primata A. azarae infulatus (128) foi maior que aqueles de C. penicillata (83), C. goeldii (78), S. sciureus (77) e C. jacchus (55). Às 72hs, houve uma redução significativa do PMI de quatro primatas: A. azarae infulatus (128/37), C. penicillata (83/38), S. sciureus (77/38) e C. jacchus (55/12), com exceção de C. goeldii (78/54). O FPMI dos primatas A. azarae infulatus (82.5) e C. goeldii (66) foi maior que do primata C. jacchus (33.5), porém, não foi maior que dos primatas C. penicillata (60.5) e S. sciureus (57.5). A resposta do TNF-α foi mais regular nos quatro primatas que reduziram o PMI no intervalo de 24-72hs: C. jacchus (145/122 pg/µL), C. penicillata (154/130 pg/µL), S. sciureus (164/104 pg/µL) e A. azarae infulatus (154/104 pg/µL), com exceção de C. goeldii (38/83 pg/µL). A resposta de IL-12 foi, principalmente, marcante nos primatas A. azarae infulatus e C. goeldii, os quais apresentaram as maiores taxas do FPMI, e a resposta do NO foi maior no primata C. goeldii, em especial no intervalo de 72hs. Estes achados sugerem, fortemente, que estes primatas neotropicais parecem ter desenvolvido mecanismos resistentes de resposta imune inata capaz de controlar o crescimento intracelular da infecção por L. (L.) i. chagasi no macrófago, o que não encoraja o uso destes primatas como modelo de estudo da LVA.

Susceptibility of Cebus apella monkey (Primates: Cebidae) to experimental Leishmania (L.) infantum chagasi-infection / Susceptibilidade do macaco Cebus apella (Primata: Cebidae) à infecção experimental por Leishmania (L.) infantum chagasi

In Amazonian Brazil, the Cebus apella monkey (Primates: Cebidae) has been associated with the enzootic cycle of Leishmania (V.) shawi, a dermotropic parasite causing American cutaneous leishmaniasis (ACL). It has also been successfully used as animal model for studying cutaneous leishmaniasis. In this work, there has been investigated its susceptibility to experimental Leishmania (L.) infantum chagasi-infection, the etiologic agent of American visceral leishmaniasis (AVL). There were used ten C. apella specimens, eight adult and two young, four males and six females, all born and raised in captivity. Two experimental infection protocols were performed: i) six monkeys were inoculated, intra-dermal via (ID), into the base of the tail with 2 x 10(6) promastigotes forms from the stationary phase culture medium; ii) other four monkeys were inoculated with 3 x 10(7) amastigotes forms from the visceral infection of infected hamsters by two different via: a) two by intravenous via (IV) and, b) other two by intra-peritoneal via (IP). The parameters of infection evaluation included: a) clinical: physical exam of abdomen, weigh and body temperature; b) parasitological: needle aspiration of the bone-marrow for searching of amastigotes (Giemsa-stained smears) and promastigotes forms (culture medium); c) immunological: Indirect fluorescence antibody test (IFAT) and, Delayed-type hypersensitivity (DTH). In the six monkeys ID inoculated (promastigotes forms) all parameters of infection evaluation were negative during the 12 months period of follow-up. Among the four monkeys inoculated with amastigotes forms, two IV inoculated showed the parasite in the bone-marrow from the first toward to the sixth month p.i. and following that they cleared the infection, whereas the other two IP inoculated were totally negative. These four monkeys showed specific IgG-antibody response since the third month p.i. (IP: 1/80 and IV: 1/320 IgG) toward to the 12th month (IP: 1/160 and IV: 1/5120). The DTH-conversion occurred in only one IV inoculated monkey with a strong (30 mm) skin reaction. Considering these results, we do not encourage the use of C. apella monkey as animal model for studying the AVL.

Na Amazônia Brasileira, o macaco Cebus apella (Primata: Cebidae) tem sido associado com o ciclo enzoótico da Leishmania (V.) shawi, um parasito dermotrópico causador da Leishmaniose Tegumentar Americana (LTA). Ele tem sido também empregado com sucesso como modelo experimental para estudo da leishmaniose tegumentar. Neste trabalho, foi investigada sua susceptibilidade à infecção experimental por Leishmania (L.) infantum chagasi, o agente etiológico da Leishmaniose Visceral Americana (LVA). Foram usados dez espécimes de C. apella oito adultos e dois jovens, quatro machos e seis fêmeas, todos nascidos e criados em cativeiro. Dois protocolos de infecção experimental foram feitos: i) seis macacos foram inoculados por via intradérmica (ID), na base da cauda com 2x10(6) formas promastigotas em fase estacionária de crescimento; ii) outros quatro macacos foram inoculados com 3x10(7) formas amastigotas de infecção visceral de hamsteres por duas vias diferentes: a) dois por via intravenosa (IV) e, b) outros dois pela via intraperitoneal (IP). A avaliação da infecção incluiu parâmetros: clínico: exame físico do abdômen, peso e temperatura corporal; b) parasitológico: aspirado de medula óssea por agulha para procura de amastigotas (esfregaço corado por Giemsa) e formas promastigotas (meio de cultura); c) imunológico: Reação de Imunofluorescência Indireta (RIFI) e, resposta de hipersensibilidade tardia (DTH). Nos seis macacos inoculados ID (formas promastigotas) todos os parâmetros de avaliação da infecção foram negativos durante o período de 12 meses. Entre os quatro macacos inoculados com formas amastigotas, dois inoculados IV mostraram parasitos na medula óssea do primeiro ao sexto mês p.i. e em seguida houve a resolução da infecção, no entanto os outros dois inoculados IP foram totalmente negativos. Esses quatro macacos apresentaram resposta específica de anticorpo IgG desde o terceiro mês p.i. (IP: 1/80 e IV: 1/320) até o décimo segundo mês (IP: 1/160 e IV: 1/5120). A conversão DTH ocorreu em apenas um macaco inoculado IV com uma forte reação na pele (30 mm). Considerando esses resultados, nós não recomendamos o uso do macaco C. apella como modelo animal para estudo da LVA.

Detecção de anticorpos anti-Leishmania infantum syn chagasi em cães de São José do Rio Preto, São Paulo / Anti-Leishmania infantum syn chagasi antibodies in dogs from São José do Rio Preto, São paulo

A leishmaniose visceral (LV) é uma zoonose que vem se expandido por todo o território paulista desde 1998, quando foi identificado o primeiro caso canino autóctone, no município de Araçatuba. O objetivo do presente estudo foi determinar a ocorrência de anticorpos anti-Leishmania infantum syn chagasi em amostras de soro de 584 cães de São José do Rio Preto, São Paulo, área não endêmica para a doença. Cinco cães (0,86%) foram soro reagentes pela técnica de ELISA e um (0,17%) por imunocromatografia. A reação de imunofluorescência indireta, realizada em 138 animais que possuíam densidades ópticas acima ou próximas ao ponto de corte do ELISA evidenciou dois cães (1,45%) com títulos acima de 1:40. Apenas um animal foi sororeagente nas três técnicas sorológicas. Apesar deste cão não apresentar histórico de deslocamento para áreas endêmicas, havia sido adquirido em região com casos caninos e humanos de LV. Os resultados obtidos no presente estudo fazem supor que não existiam casos autóctones de LV canina na população estudada.

Visceral leishmaniasis (VL) has been a widespread zoonosis in São Paulo since 1998, when the first autochthonous canine case was identified in Araçatuba. The aim of this study was to determine the occurrence of anti–Leishmania infantum syn chagasi antibodies in serum samples of 584 dogs from São José do Rio Preto, São Paulo, a non endemic area for the disease. Five dogs (0.86%) seroconverted by ELISA and one (0.17%) by immunochromatography. The indirect immunofluorescent reaction, carried out in 138 animals whose optical densities were above or close to ELISA’s cutt-off point, evidenced two dogs (1.45%) with titers above 1:40. Only one dog was serum-reactive on the three techniques. Although there was not a history of displacing this animal to endemic areas, the dog had been acquired in a region with canine and human cases of VL. These results suggests that there were no autochthonous cases of canine VL in this population.

Leishmania chagasi y Trypanosoma cruzi: Conducta trófica en cultivos axénicos puros y mixtos / Leishmania chagasi and Trypanosoma cruzi: Trophic behavior in pure and mixed axenic cultures

El mecanismo adaptativo entre Leishmania y Trypanosoma ante la competencia por los recursos nutricionales en ambientes compartidos ha sido pobremente investigado. En particular, el estudio de la conducta trófica entre Leishmania chagasi y Trypanosoma cruzi podría contribuir al entendimiento de situaciones de gran relevancia médica en humanos, como son las infecciones mixtas por ambos tripanosomatídeos. En este trabajo se establecieron cultivos axénicos puros de Leishmania chagasi y Trypanosoma cruzi, así como cultivos axénicos mixtos (L. chagasi + T. cruzi) en médio BHI. Se registraron los cambios de dinámica poblacional (organismos por mililitro), la evolución de la estructura de las poblaciones, las variaciones de las concentraciones de colesterol, glucosa y proteínas totales, así como los cambios de pH en el medio de cultivo. El manejo cuantitativo de los conjuntos numéricos generados experimentalmente se abordó con técnicas univariadas (Análisis de la Varianza) y multivariadas (Análisis Discriminante Múltiple). Los resultados demuestran diferencias estadísticas significativas entre las medias de los parámetros considerados y prueban que el comportamiento “in vitro” investigado en el cultivo mixto L. chagasi – T. cruzi difiere taxativamente del estudiado en los cultivos puros de L. chagasi y T. Cruzi

The adaptive mechanisms betweenLeishmania and Trypanosoma facing the competencefor the nutritional resources in shared environmentshave been poorly investigated. Particularly, thestudy of the trophic behavior between Leishmaniachagasi and Trypanosoma cruzi could contribute tothe understanding of relevant medical situations, asmixed human infections. In this work pure axeniccultures of Leishmania chagasi and Trypanosomacruzi, as well as mixed cultures (L. chagasi + T.cruzi) were established in BHI medium. Changes ofpopulation dynamics (organisms/mL), the evolutionof the population’s structure, the variations of theconcentration of cholesterol, glucose and total proteins,as well as the changes of the medium’s pH wereregistered. The generated numerical sets were tackledwith univariate (Analysis of Variance) and multivariate(Multiple Discriminant Analysis) techniques. Theresults demonstrate significant statistical differencesbetween the media of the considered parameters andprove that the “in vitro” behavior investigated in theL. chagasi – T. cruzi mixed culture precisely differsfrom the L. chagasi and T. cruzi pure cultures

Prevalencia de la infección por Leishmania spp. en un área periurbana de Altagracia de Orituco, estado Guárico, Venezuela / Prevalence of infection with Leishmania spp. in a suburban area of Altagracia de Orituco, Guarico State, Venezuela

Leishmania infantum es el agente etiológico de la leishmaniasis visceral (LV). Sin embargo, tanto en el Viejo Mundo como en el Nuevo Mundo(América Central), este protozoario ha sido involucrado en casos de leishmaniasis cutánea atípica (LCA). Evidencias clínicas, parasitológicas, moleculares y entomológicas han demostrado que una situación similar está ocurriendo en un área periurbana de Altagracia de Orituco (Guárico, Venezuela). Con la finalidad de contribuir al entendimiento de este nuevo escenario epidemiológico, se realizó un estudio transversal en el cual se determinó la reactividad a la prueba de leishmanina [Leishmanin skin test (LST)] y la seroprevalencia a antígenos crudos de Leishmania spp. y al antígeno específico rK39 mediante la prueba de ELISA en una muestra de ocho casos de LCA detectados en el periodo 1997-2000, en sus cohabitantes (n=15) y en una muestra de 233 de la comunidad igualmente expuestas a riesgo. La dermoprevalencia resultó ser de 31,6% (67/212). El 28,8% (67/233) resultó positivo a la prueba de ELISA usando promastigotas de L. infantum (=L. chagasi) y 13,3% (31/233) poseían anticuerpos anti-L. braziliensis. Así mismo, la prueba de ELISA usando el antígeno rK39 resultó positiva en uno de los ocho casos estudiados y en dos de sus cohabitantes; así como también en 13,7% (32/233) de los individuos residentes en la comunidad. Los resultados obtenidos indican la circulación de especies del subgénero Leishmania y del subgénero Viannia en el área en estudio y que la presencia de anticuerpos anti-Leishmania en general se encontró asociada a la edad y al tiempo de residencia en la zona

Leishmania infantum is the etiological agent of the visceral leishmaniasis (VL). However, in the OldWorld as well in the New World (Central America), thisprotozo an has also been involved in cases of atypicalcutaneous leishmaniasis (ACL). Clinical, molecularand entomological evidences have demonstrated thata similar situation is happening in a peri-urban areaof Altagracia de Orituco (Guárico state, Venezuela). With the purpose of contributing to the understandingof such an epidemiological scenario, a cross-sectionalstudy was carryed out using the leishmanin skin test(LST) and screening for the seroprevalence to crudeantigens of Leishmania spp. and rK39 by means ofthe ELISA test in cases of ACL reported in the period1997-2000 (n=8), among their co-inhabitants (n=15) and in a randomly selected sample of 233 people. Thedermoprevalence was 31.6% (67/212); 28.8% (67/233)were positive to the ELISA test using promastigotes of L. infantum (= L. chagasi) and 13.3% (31/233) hadantibodies anti-L. braziliensis. In addition, the test of ELISA using the antigen rK39 was positive in oneout of the eight cases studied and in two out of theirco-inhabitants; as well as in 13.7% (32/233) of theinhabitants of the community. The results obtained inthis epidemiological study indicate the circulation of species of the sub-genus Leishmania as well as of thesubgenus Viannia. The presence of antibodies anti-Leishmania in general was associated to the age andthe time of residence in the zone

[Comparison between serology and PCR methods for the diagnosis of visceral Leishmaniasis]

Canine visceral leishmaniasis [CVL] caused by Leishmania Infantum is endemic in most Mediterranean basin and its seroprevalence ranges from 10 to 37%. Diagnosis of Infection is very important especially in asymptomatic dogs for control of human leishmaniasis for control of human visceral leishmaniasis. This study was aimed to compare three methods for detection of canine visceral leishmaniasis. In this research process study, 71 dogs were selected from 4 endemic villages in Meshkin-Shahr district. Peripheral blood samples were tested by serologic [DAT and Dipstick rK39] and molecular [PCR] methods. Skin samples were tested by molecular [PCR] methods. Twelve samples of PCR products were sequenced that all of them were identified as Leishmania infantum and 2 nucleotide sequence data submitted to the GenBank database. From 71 dogs that were studied, 21.1% were symptomatic and others were asymptomatic[78.9%]. 17 dogs [23.9%] had >/= 1:320 titer of antibody by direct agglutination test [DAT]. Twenty two dogs[31%] were positive by Dipstick rK39 test, 21 dogs [29.6%] were positive by PCR on skin samples, 31 dogs [43.7%] were positive in blood PCR and 38 dogs [53.5%] were positive by skin/blood PCR. The highest correlation was between DAT and Dipstick test [76%].According to the results of this study, we can diagnose infection in symptomatic and asymptomatic dogs by DAT as a suitable method and PCR is suitable to follow parasite DNA in skin and other tissues of dogs

Leishmaniasis visceral subclínicaen 123 inpiduos de un cantón de la provincia Caranavi-La Paz: study in 123 inpiduals / Sub-clinical infections by visceral leishmaniasis in Caranavi district

En 1999, en el Hospital del Niño se registró un nuevo caso de Leishmaniasis visceral sobre un niño de dos años proveniente del cantón de Taipiplaya (Provincia Caranavi). Por este motivo se realiza en este cantón una evaluación transversal de la leishmaniasis visceral mediante pruebas serológicas y moleculares involucrando a 122 inpiduos clínicamente sanos y un individuo con infección positiva a Leishmania cutánea, Se demostró la circulación de Leishmania sp. en 32,3 por ciento de sujetos estudiados. El 14,4 por ciento de la población examinada presentó anticuerpos anti-rk39, demostrándose la circulación de Leishmania chagasi responsable de la leishmaniasis visceral. No podemos descartar la posibilidad de la existencia de coinfecciones mixtas inter-especie de Leishmania como también de coinfecciones mixtas por Leishmania sp y Trypanosoma cruzi, responsable de la enfermedad de Chagas.

Experimental infection of canine peritoneal macrophages with visceral and dermotropic Leishmania strains

A study was carried out using macrophages cultured from the peritoneal exudate of dogs infected in vitro with three species of Leishmania: L. (L.) chagasi, L. (Viannia) braziliensis and L. (L.) amazonensis with the aim of investigating the growth kinetics and infectivity of these species in the host cell. Results were expressed as the percentage of macrophages infected measured at 24 hr intervals over six days in RPMI - 1640 culture medium at a temperature of 34-35oC. The findings open the possibility of using canine peritoneal cells as a model for the screenning of leishmanicide drugs and to study the pathogenesis of these species.

histopathological picture of the liver hamsters experimentally infected with leishmania d. infantum on top of schistosoma mansoni infection

The histopathological picture of the liver of Syrian golden hamsters experimentally infected with Leishmania d. infantum on top of pre- infection with S. mansoni was studied. The results showed that the liver was markedly affected by the concomitant infection than infection with either parasite alone. The superadded Leishmania infection caused the early appearance of cell necrosis and fatty changes. The schistosomal granulomas were rounded in shape and well circumscribed. However, they were smaller in size and less in number when compared with schistosomal control. Also, there was a comparative decrease in the number of their cellular constitutions, but without a change in the cell types. The schistosomal granulomas were surrounded with a rim of amastigotes laden macrophages. On the other hand, the leishmanial granulomas were irregular in shape, but comparatively increased in size and in number on the behalf of the schistosomal granulomas. Their cellular constitutions comparatively increased. So, it was concluded that the L. d. infantum infection on the top of S. mansoni infection suppressed the schistosomal pathological picture in the liver of hamsters

Renal changes in golden hamsters experimentally infected with leishmania d. infantum on top of schistosoma mansoni infection

This work aimed to study the concomitant infection of L. d. infantum on top of S. mansoni infection in Syrian golden hamsters, the model animal for both parasites. The results showed that the renal changes were more or less the same as infection with either parasite alone. Both parasites produce mesangial cell proliferation and mesangial matrix increase. Some glomeruli showed complete replacement of the capillary tuft with structureless homogenous material. In one S. mansoni positive control hamster, Schistosoma eggs were seen within granulomas and atrophic glomeruli. However, the leishmanial infection, more or less suppressed the schistosomal infection. The leishmanial infection itself became more prominent and the renal lesion appeared earlier and was more obvious

Histopathological studies of Syrian golden hamsters experimentally infected with Leishmania d infantum

In this paper, Syrian Golden hamsters were chosen as a model animal for histopathological studies of L. d. infantum. 24 male hamsters were experimentally infected with L. D. Infantum. Every week, 2 hamsters were sacrificed and parts from the liver, heart, lung, spleen, small intestine and kidney were paraffin sectioned and stained with hematoxylin and eosin or with Giemsa stain. The leishmanial granuloma was the main histopathological change and the liver was the main organ affected. Leishmanial granulomas were common in the liver, but few in lung and small intestine. The hepatic granulomas increased in surface areas and in number in the late stage of the infection [<6 weeks]. They were formed mainly of histiocytes, less lymphocytes and very few eosinophils with necrosis inside and bridging in between granulomas, mainly occurred in the late stage of infection. Neither fibrosis nor peripheral delimitation was seen. Kupffer cell hyperplasia was seen throughout the experimental period [12 weeks]. The spleen showed expansion of the red pulp and atrophy of the white pulp. The kidney showed mesangial cell proliferation and the mesangial matrix increased accompanied with amyloid deposition. All the tissue sections were photographed and the results were discussed