Molecular epidemiology of Leishmania infantum on the island of Majorca: a comparison of phenotypic and genotypic tools

Abstract

In the Mediterranean basin, Leishmania infantum is the causative agent of both visceral and cutaneous leishmaniasis, and is an important opportunistic parasite in patients infected with human immunodeficiency virus (HIV). The commonest method used to study the variability of Leishmania spp. is isoenzyme analysis. In addition to this, we employed 3 assays based on the polymerase chain reaction (PCR): random amplified polymorphic deoxyribonucleic acid (RAPD), intergenic region typing (IRT), based on the amplification of ribosomal ribonucleic acid internal transcribed spacers and restriction fragment length polymorphism (PCR-RFLP). We used 54 L. infantum stocks isolated from HIV co-infected patients, 38 isolated from dogs, 3 isolated from immunocompetent patients and 3 isolated from 1826 sand files in the island of Majorca (Spain), a closed ecological niche. Zymodemes MON-1 (70%), MON-24 (11%) and MON-34 (18%) were found among the human isolates, and MON-1 (95%) and MON-108 (5%) among those from dogs. RAPD and IRT could not discriminate among the strains as they all gave the same pattern, even when different zymodemes were examined. In contrast, PCR-RFLP was able to distinguish the strains and, furthermore, a dendrogram (unweighted pair group method with arithmetic average [UPGMA]) was constructed from the genetic distances derived from RFLP data. The Leishmania isolates from HIV-infected subjects formed a single cluster, supporting the existence of an artificial anthroponotic cycle previously proposed by our group, in which syringes have been substituted for sand flies, and in which certain clones have been spread among intravenous drug users. This contrasts with the clusters representing a zoonotic cycle, involving dogs, sand flies and both immunocompetent and immunocompromised humans.

Molecular Epidemiology of Leishmania ( Viannia ) guyanensis in French Guiana

“…Because of the high level of polymorphism in the composition of the ITS locus, several molecular epidemiological studies have been conducted by RFLP analysis in New World taxa (5,10,21,38), but most studies have been performed on Old World Leishmania species (7,19,34,(44)(45)(46)(47). Analyses of the ribosomal ITS regions of different parasites have demonstrated the utility of this marker in detecting genetic diversity and any association of this diversity with several epidemiological aspects (16,33).…”

Use of PCR-Restriction Fragment Length Polymorphism Analysis To Identify the Main New WorldLeishmaniaSpecies and Analyze Their Taxonomic Properties and Polymorphism by Application of the Assay to Clinical Samples

“…Among these methods, restriction fragment length polymorphism (RFLP) analyses of PCR-amplified sequences of multicopy genes have shown promising results (1,12,23,28,30,33,52). Since the work of Cupolillo et al (12), PCR-RFLP methods on the internal transcribed spacers (ITS) of the ribosomal genes have been performed on New World (NW) Leishmania species (4, 10, 19, 39), but these methods have been conducted mostly on Old World Leishmania species (7,17,35,(42)(43)(44)(45). However, up to now, no study has integrated complete analyses of specificity, sensitivity, taxonomic reliability, intraspecies polymorphism, and trial on clinical samples for diagnosis of leishmaniasis in the field for the two subgenera of Leishmania.…”

Aspectos Gerais Da Leishmaniose Visceral

“…Anteriormente ao aparecimento da AIDS no Mediterrâneo, a LV era uma doença que tradicionalmente afetava crianças. Entretanto, a doença passou a acometer principalmente adultos entre 31 e 50 anos de idade, ocorrendo na forma de co-infecção em 70% dos casos (CHICHARRO et al, 2002). Assim, estima-se que ao menos 59.000 pessoas morram anualmente, estando a maioria dos casos na Índia e Sudão (COSTA, 2011).…”

“…Portanto, uma vez infectado, o hospedeiro torna-se reservatório do agente e o cão é o principal reservatório doméstico . Ainda, segundo CHICHARRO et al (2002) a ausência de cura dos cães infectados, atribuem a estes animais um importante papel na cadeia epidemiológica da enfermidade.…”