PCR-RFLP of ribosomal internal transcribed spacers highlights inter and intra-species variation among Leishmania strains native to La Paz, Bolivia.

Human leishmaniasis is highly endemic in Bolivia and shows a growing incidence. This report reveals the genetic variability of 35 isolates mainly belonging to Leishmania braziliensis and Leishmania amazonensis species. Among them, 31 were from human patients with different clinical presentations, 3 strains from Lutzomya nuneztovari anglesi (the proven vector of L. amazonensis) and 1 strain of a mammal (Conepatus chinga). The isolates were analyzed by isoenzyme electrophoresis (MLEE) and PCR-RFLP of ITS rRNA genes, a genetic marker highly polymorphic and better adapted to sub-structuring of populations. MLEE and RFLP-ITS were in agreement to discriminate the species, 12 belong to L. (V.) braziliensis, 21 to L. (L.) amazonensis, 1 to Leishmania (V.) lainsoni and 1 to Leishmania (L.) chagasi. Among L. (V.) braziliensis the RFLP-ITS only highlights variability. Ten isolates from either cutaneous or mucocutaneous clinical forms, were grouped together (bootstrap value of 99.8%) apart from two others, one from a mammal (C. chinga), the other from a patient with a cutaneous form. Among L. (L.) amazonensis both markers detect variability but no significant sub-division was identified including isolates from different clinical forms. Moreover, the high frequency of several isolates from cutaneous forms occurred during an outbreak, with putative hybrid character (multiloci heterozygous patterns depicted by MLEE) could be linked to better fitness of these parasites. However, in the absence of observation of hypothetical parents, their hybrid status remains a question.

Polymerase chain reaction detection and serologic follow-up after treatment with benznidazole in Bolivian children infected with a natural mixture of Trypanosoma cruzi I and II.

Thirty-five Bolivian children (5-10 years of age) seropositive for infection with T. cruzi underwent specific chemotherapy with benznidazole. Before treatment, 57.1% had a positive parasitologic diagnosis. Some patients presented an early conversion by polymerase chain reaction of blood samples, while others were still positive four and seven months after the end of the treatment, which indicated an absence of parasite clearance. Strain typing showed that most patients were infected by a mixture of clones I and II of T. cruzi. Serologic conversion in conventional tests and antibodies to shed acute-phase antigen were observed in two and four patients, respectively. For the other patients, the average rate of antibody decay was half the initial rate. The parasitologic and serologic data indicated that chemotherapy acts throughout the course of infection in a long-lasting process in which the decrease of specific antibody production is related to the reduction of the live parasite load.

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.

Detección e identificación de complejos de leishmania a traves de la reacción en cadena de la polimerasa (PCR) e hibridación con sondas especificas de complejo / Valores hematologicos en recien nacidos sanos habitantes de altura: 3600 msnm

Pregutna de investigación. ¿Los iniciadores L1 y L2 serán útiles en la detección e identificación de complejos de Leishmania, logrando una alta sensibilidad y escificidade de la Reacción en Cadena de la Polimerasa? Objetivo. Evaluar los iniciadores L1 y L2 para la identifcación de complejos de Leishmania a trvés de técnicas moleculares: Reacción en Cadena de la Polimerasa (PCR) e hibridación con osndas específicas de ADN de Kinetoplasto. Diseño. Básico experimental. Métodos.El presente estudio se realizó en 27 cepas de referencia caracterizadas previamente por electroforesis de isoenzimas. para la Reacción en Cadena de la Polimerasa se utilizo iniciadores L1 y L2, diseñados a partir de la secuenciación realizado por 1. Estos se alinean en la parte consevada y amplifican la arte variable de los minicírculos de ADN de kinetoplasto. Las sondas fuern elaboradas a partir de losproductos de PCR, seleccionado bandas mayores para tres complejos: brasiliensis (MHOM/BO90/CG); mexicana (MNYC/BZ/6"/M379); y denovani (MHOM/BR/74/PP75). La tecnica de hibridación fue realizada bajo condiciones de alta astringencia que nos da la seguridad de una alta homología entre las sondas y el ADN reconocido por ellas. Resultados. Se ha obtenido un perfil polimórfico para cada una de las cepas en estudio, con una alta selsibilidad de la técnica de PCR, amplificando varios Kinetoplastidae además de Leishmania, entre ellos Trypanisoma cruzi, Trypanisoma rangeli y Tripanisoma brucei. Las sondas presentan una alta especificidad logrando así racionalizar el amplio polimorfismo obtenido por PCR. Las sondas son una herramienta uútil para la detección e identificación directa de complejos de Leishmania. Conclusiones. Los idicadores L1 y L2 dan un perfil polimófico para cada cepa, presentando una sensibilidad muy alta, sin embargo no son específicos lde Leishmania, amplifican otros Kinetoplastidae. Este alto polimorfismo se racionaliza con la utilización de las osndas construidas a partir del polimorfismo obtenido, las que son especificas de complejo.

Leishmaniasis visceral subclínica en 123 individuos de un cantón de la provincia Caranavi-La Paz / Sub-clinical infections by visceral leishmaniasis in Caranavi district: study in 123 individuals

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 individuos clínicamente sanos y un individuo con infección positiva a Leishmania cutánea, Se demostró la circulación de Leishmania sp. en 32,3% de sujetos estudiados. El 14.4% 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.

In 1999, in the "Hospital del Niño". a new case of visceral leishmaniasis was identified in a 2 years old child from Taipiplaya in the Caranavi district. For this reason, a visceral leishmaniasis evaluation using serological and molecular tests was realized on 122 healthy people and also on one leishmania cutanea infected person. Leishmania spp was present in 32,3 % of the studied people and 14,4 % had an anti-rk39 antibody, attesting the existence of Leishmania chagasi responsible for visceral leishmaniasis. The possibility of mixed infections with other Leishmania species as well as mixed infections Leishmania spp and Trypanosama cruzi, responsible for chagas disease should not be discarded.

Is Rhodnius robustus (Hemiptera: Reduviidae) responsible for Chagas disease transmission in Western Venezuela?

We present evidence for the putative role of Rhodnius robustus as extradomestic vector of Chagas disease in Western Venezuela. First, we assessed the validity of this triatomine species by genetic characterization in relation with some other species of the prolixus group. Random amplified polymorphic DNA data showed a clear separation between this species and R. prolixus and indicated a probable genetic heterogeneity within R. robustus. Faeces and gut contents were microscopically examined in 54 of 137 R. robustus collected in palm trees. According to this morphological examination, 18% were positive for Trypanosoma cruzi, 11% harboured T. rangeli and 11% showed mixed infection. Five of the seven samples examined gave a polymerase chain reaction major band of 270 bp specific of T. cruzi. The hybridization probes showed that R. robustus may transmit clones 20 and 39 (or genetically related ones) in Venezuela. Such a transmission might occur when, in absence of domestic R. prolixus and attracted by artificial light, R. robustus enters houses and feeds on humans, or when people are bitten outdoors. The lack of bugs inside houses could mean that the insects leave houses after feeding, or die without reproducing there.

Sylvatic population of Triatoma infestan from the bolivian Chaco: from field collection to characterization

A sylvatic Triatoma infestans DM (dark morph) population detected in the Bolivian Chaco was characterized and compared with various domestic ones. The degree of differentiation of DM was clearly within the T. infestans intra-specific level. Nevertheless marked chromatic and morphometric differences as well as differences in antennal pattern, chromosome banding and randomly amplified polymorphic DNA support the hypothesis of a distinct population. Continuous exchange of insects between wild and domestic habitats seems unlikely in the Chaco