Development and validation of PCR-based assays for diagnosis of American cutaneous leishmaniasis and identificatio nof the parasite species

Graça, Grazielle Cardoso da; Volpini, Ângela; Romero, Gustavo Adolfo Sierra; Neto, Manoel Paes de Oliveira; Hueb, Márcia; Porrozzi, Renato; Boité, Mariana Côrtes; Cupolillo, Elisa

doi:10.1590/s0074-02762012000500014

Cited by 112 publications

(123 citation statements)

References 57 publications

(66 reference statements)

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“…8). These were applied in Brazil (200), Peru (201), and several Old World countries (202) and on various clinical sample types. The sensitivities of these assays have not been compared systematically to those of other species typing PCRs but were generally found to be 60 to 90% compared to those of higher-copy-number targets, such as rRNA genes or kDNA minicircles, which are used for diagnostic Leishmania genus detection.…”

Section: Gp63mentioning

confidence: 99%

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Species Typing in Dermal Leishmaniasis

2015

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SUMMARY Leishmania is an infectious protozoan parasite related to African and American trypanosomes. All Leishmania species that are pathogenic to humans can cause dermal disease. When one is confronted with cutaneous leishmaniasis, identification of the causative species is relevant in both clinical and epidemiological studies, case management, and control. This review gives an overview of the currently existing and most used assays for species discrimination, with a critical appraisal of the limitations of each technique. The consensus taxonomy for the genus is outlined, including debatable species designations. Finally, a numerical literature analysis is presented that describes which methods are most used in various countries and regions in the world, and for which purposes.

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Section: Gp63mentioning

confidence: 99%

“…They therefore have limited general use. Since they are based on single-copy genes, their sensitivity is lower than that of other methods based on multicopy targets, such as ITS1, hsp70, and kDNA minicircles (200), but they have been applied to clinical specimens (30).…”

Section: Carbohydrate Metabolism Enzymesmentioning

confidence: 99%

Species Typing in Dermal Leishmaniasis

2015

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“…by PCR targeting the ITS1 intergenic region was employed (GRAÇA et al, 2012). Restriction Fragment Length Polymorphism (RFLP) was performed for Leishmania species identification.…”

Section: Molecular Detection Of Leishmania Sppmentioning

confidence: 99%

Hepatozoon canis and Leishmania spp. coinfection in dogs diagnosed with visceral leishmaniasis

Morgado

Cavalcanti

Miranda

et al. 2016

Rev. Bras. Parasitol. Vet.

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This study describes the occurrence of dogs naturally co-infected with Hepatozoon canis and two Leishmania species: L. infantum or L. braziliensis. Four dogs serologically diagnosed with Visceral Leishmaniasis were euthanized. Liver and spleen samples were collected for histopathological analysis and DNA isolation. H. canis meronts were observed in tissues from all four dogs. H. canis infection was confirmed by PCR followed by sequencing of a fragment of 18S rRNA gene. Leishmania detection and typing was confirmed by ITS1' PCR-RFLP and parasite burden was calculated using ssrRNA quantitative qPCR. A DPP -Dual Path platform test was performed. One out (Dog #2) of four animals was asymptomatic. Dogs #1 and #4 were infected by L. infantum and were DPP test positive. Dogs #2 and #3 were infected by L. braziliensis and were DPP test negative. Furthermore, visceral dissemination was observed in Dogs #2 and #3, since L. braziliensis was detected in liver and spleen samples. The visceral dissemination of L. braziliensis associated with systemic signs suggested that this co-infection could influence the parasite burden and disease progression.Keywords: Hepatozoonosis, canine visceral leishmaniasis, Leishmania infantum, Hepatozoon canis, co-infection, Leishmania braziliensis. ResumoO presente estudo descreve a ocorrência de coinfecção com Hepatozoon canis e duas espécies de Leishmania (L. infantum ou L. braziliensis) em cães. Quatro cães sorologicamente diagnosticados com leishmaniose visceral foram eutanasiados. Amostras do baço e fígado foram submetidas à histopatologia e extração de DNA. Merontes de H. canis foram observados nos quatro cães. A infecção por H. canis foi confirmada por PCR e sequenciamento de um fragmento do gene 18S rRNA. A infecção por Leishmania e tipagem foram realizadas por PCR-RFLP do região intergênica ITS1. A carga parasitária foi calculada pela qPCR quantitativa baseada no gene ssrRNA. O teste DPP -Dual Path platform foi realizado. Apenas o Cão #2 era assintomático. Os cães #1 e #4 estavam infectados com L. infantum e foram positivos no DPP. Os cães #2 e #3 estavam infectados com L. braziliensis e foram negativos no DPP. Além disso, visceralização foi observada nos cães #2 e #3, nos quais L. braziliensis foi detectada em amostras de baço e fígado. A visceralização da L. braziliensis associada a sinais clínicos sistêmicos sugerem que esta coinfecção pode ter influenciado na carga parasitária e progressão da doença.Palavras-chave: Hepatozoonose, leishmaniose visceral canina, Leishmania infantum, Hepatozoon canis, coinfecção, Leishmania braziliensis.

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“…are small subunit ribosomal DNA, microsatellite, internal transcribed spacer, mini-exon, and heat shock protein sequences. [97][98][99][100] Due to their sensitivity, these sequences enable distinction among the main species causing TL in South America, when associated with nested PCR, restriction fragment length polymorphism, or sequencing. Although present in lower copy numbers than kinetoplast DNA (kDNA), these targets have high sensitivity to detect parasites as well.…”

Section: Leishmaniasismentioning

confidence: 99%