Molecular diagnosis of trypanosomatids in Didelphis marsupialis from Los Montes de María: a first report of Trypanosoma rangeli from Colombian Caribbean region.

Didelphis marsupialis is a primary reservoir of Trypanosoma cruzi, etiologic agent of American Trypanosomiasis-AT or Chagas Disease-CD, in America. Some findings of Trypanosoma rangeli have been recorded in this mammal, in sympatry with T. cruzi. In Los Montes de María, Bolívar, Colombian Caribbean, triatomine insects and potential parasite host has been registered, but little is known about the relationship between these parasites and D. marsupialis. We investigated the natural trypanosomatids infection rate in D. marsupialis, applying a parasitological and molecular diagnosis. Twenty D. marsupialis was investigated between 2018 and 2019 using 21 Tomahawk® traps placed on the sylvatic/domestic corridors. Blood was drawn by cardiopuncture after sedation. An aliquot of blood samples was cultured in Novy, Nicolle, McNeal/Roswell Park Memorial Institute medium at 24 °C/60 days for the detection of motile trypomastigotes. Parasite DNA was obtained by salting out methods from positive blood cultures. Trypanosomatids diagnosis was done by Polymerase Chain Reaction-sequencing of V7V8 region of 18S ribosomal RNA (18S-rRNA) gene. Amplicons were sequenced, and consensus sequences were aligned with reference sequences from GenBank. Four isolates corresponded to T. rangeli (20%) and one to T. cruzi (5%). The natural infection of D. marsupialis by T. rangeli and T. cruzi constitutes the first record of these parasites in didelphids in Los Montes de María and the first record of T. rangeli in this marsupial, in the Colombian Caribbean.

Surveillance of phlebotomine fauna and Didelphis marsupialis (Didelphimorphia: Didelphidae) infection in an area highly endemic for visceral leishmaniasis in Colombia. / Vigilancia de la fauna flebotomínea e infección de Didelphis marsupialis (Didelphimorphia: Didelphidae) en un área de alta endemia de leishmaniasis visceral en Colombia

INTRODUCTION: The study of the interaction between the parasite, the vector and the mammalian hosts, including man, allows to understand the behavior of the leishmaniases. OBJECTIVE: To determine the presence of Lutzomyia species and to detect the Leishmania infection in Didelphis marsupialis in an endemic area for visceral leishmaniasis. MATERIALS AND METHODS: Phlebotomine fauna and individuals of D. marsupialis were collected with CDC and Tomahawk™ traps, respectively. The species of Lutzomyia were identified using the Young and Duncan key (1994). Ear and tail biopsies and blood samples from D. marsupialis were taken to identify the Leishmania species by amplifying a fragment of the gene associated with the 70 kD heat shock protein. RESULTS: Seven Lutzomyia species were identified: Lu. evansi, Lu. gomezi, Lu. panamensis, Lu. dubitans, Lu. cayennensis cayennensis, Lu. rangeliana and Lu. trinidadensis. The first three species have epidemiological importance in Colombia because of their implications in the transmission of the Leishmania parasite. Sixty-five tissue samples from 19 D. marsupialis individuals were negative for Leishmania spp. CONCLUSIONS: The presence of the Lutzomyia species that have been identified as vectors for Leishmania inside and around houses in the village of El Bledo, in El Carmen de Bolívar represents a risk of infection. Furthermore, the presence of Lu. panamensis is reported for first time in El Carmen de Bolívar in Colombia. Although the lack of detection of Leishmania spp. in D. marsupialis samples may suggest that D. marsupialis does not play an important role in the transmission cycle of Leishmania in this region, it is necessary to carry out further longitudinal studies to confirm this hypothesis.

Introducción. El estudio de la interacción entre el parásito, el vector y los huéspedes mamíferos, incluido el hombre, permite entender el comportamiento de la leishmaniasis. Objetivo. Determinar la presencia de especies del género Lutzomyia y detectar la infección por Leishmania spp. en Didelphis marsupialis en un área endémica de leishmaniasis visceral. Materiales y métodos. Se recolectaron flebotomíneos y D. marsupialis con trampas CDC y Tomahawk™, respectivamente. Las especies de Lutzomyia se identificaron usando la clave de Young y Duncan, 1994. Se tomaron biopsias de oreja, cola y muestras de sangre de D. marsupialis para diagnosticar Leishmania spp. mediante la amplificación de un fragmento del gen de la proteína de choque térmico de 70 kD. Resultados. Se identificaron siete especies de Lutzomyia: Lu. evansi, Lu. gomezi, Lu. panamensis, Lu. dubitans, Lu. cayennensis cayennensis, Lu. rangeliana y Lu. trinidadensis. Las tres primeras especies son reconocidas como vectores en el país por estar implicadas en la transmisión de Leishmania spp. En total, 65 muestras de tejidos de oreja, cola y de sangre provenientes de 19 D. marsupialis fueron negativas para Leishmania spp. en la PCR-HSP70. Conclusiones. La presencia de flebotomíneos con importancia epidemiológica en la zona evaluada representa un riesgo de transmisión. Asimismo, Lu. panamensis es reportada por primera vez en El Bledo (Carmen de Bolívar). La ausencia de Leishmania spp. en D. marsupialis podría sugerir que esta especie no tiene un papel importante en el ciclo de transmisión de Leishmania en la vereda El Bledo, por lo que es necesario profundizar en estudios longitudinales para corroborar esta hipótesis.

Surveillance of phlebotomine fauna and Didelphis marsupialis (Didelphimorphia: Didelphidae) infection in an area highly endemic for visceral leishmaniasis in Colombia / Vigilancia de la fauna flebotomínea e infección de Didelphis marsupialis (Didelphimorphia: Didelphidae) en un área de alta endemia de leishmaniasis visceral en Colombia

Abstract Introduction: The study of the interaction between the parasite, the vector and the mammalian hosts, including man, allows to understand the behavior of the leishmaniases. Objective: To determine the presence of Lutzomyia species and to detect the Leishmania infection in Didelphis marsupialis in an endemic area for visceral leishmaniasis. Materials and methods: Phlebotomine fauna and individuals of D. marsupialis were collected with CDC and Tomahawk™ traps, respectively. The species of Lutzomyia were identified using the Young and Duncan key (1994). Ear and tail biopsies and blood samples from D. marsupialis were taken to identify the Leishmania species by amplifying a fragment of the gene associated with the 70 kD heat shock protein. Results: Seven Lutzomyia species were identified: Lu. evansi, Lu. gomezi, Lu. panamensis, Lu. dubitans, Lu. cayennensis cayennensis, Lu. rangeliana and Lu. trinidadensis. The first three species have epidemiological importance in Colombia because of their implications in the transmission of the Leishmania parasite. Sixty-five tissue samples from 19 D. marsupialis individuals were negative for Leishmania spp. Conclusions: The presence of the Lutzomyia species that have been identified as vectors for Leishmania inside and around houses in the village of El Bledo, in El Carmen de Bolívar represents a risk of infection. Furthermore, the presence of Lu. panamensis is reported for first time in El Carmen de Bolívar in Colombia. Although the lack of detection of Leishmania spp. in D. marsupialis samples may suggest that D. marsupialis does not play an important role in the transmission cycle of Leishmania in this region, it is necessary to carry out further longitudinal studies to confirm this hypothesis.

Resumen Introducción. El estudio de la interacción entre el parásito, el vector y los huéspedes mamíferos, incluido el hombre, permite entender el comportamiento de la leishmaniasis. Objetivo. Determinar la presencia de especies del género Lutzomyia y detectar la infección por Leishmania spp. en Didelphis marsupialis en un área endémica de leishmaniasis visceral. Materiales y métodos. Se recolectaron flebotomíneos y D. marsupialis con trampas CDC y Tomahawk™, respectivamente. Las especies de Lutzomyia se identificaron usando la clave de Young y Duncan, 1994. Se tomaron biopsias de oreja, cola y muestras de sangre de D. marsupialis para diagnosticar Leishmania spp. mediante la amplificación de un fragmento del gen de la proteína de choque térmico de 70 kD. Resultados. Se identificaron siete especies de Lutzomyia: Lu. evansi, Lu. gomezi, Lu. panamensis, Lu. dubitans, Lu. cayennensis cayennensis, Lu. rangeliana y Lu. trinidadensis. Las tres primeras especies son reconocidas como vectores en el país por estar implicadas en la transmisión de Leishmania spp. En total, 65 muestras de tejidos de oreja, cola y de sangre provenientes de 19 D. marsupialis fueron negativas para Leishmania spp. en la PCR-HSP70. Conclusiones. La presencia de flebotomíneos con importancia epidemiológica en la zona evaluada representa un riesgo de transmisión. Asimismo, Lu. panamensis es reportada por primera vez en El Bledo (Carmen de Bolívar). La ausencia de Leishmania spp. en D. marsupialis podría sugerir que esta especie no tiene un papel importante en el ciclo de transmisión de Leishmania en la vereda El Bledo, por lo que es necesario profundizar en estudios longitudinales para corroborar esta hipótesis.