RESUMEN
We studied the prevalence of infection by trypanosomatid parasites in Didelphis marsupialis and its relationship with morphological/age aspects in a rural area of El Carmen de Bolivar, Colombia. Five visits were made to the Vereda El Alférez; each of which lasted three consecutive nights. During these visits, Tomahawk® traps were installed in the peridomestic and wild ecotopes of the Vereda El Alférez. Body measurements, sex and age were determined from the collected animals. Blood was extracted by cardiopuncture, after sedation, in order to obtain total deoxyribonucleic acid (DNA) and amplify the conserved region of the kinetoplast minicircle DNA (kDNA) of parasitic trypanosomatids. The association between morphological parameters of didelphids and their frequency of infection by parasitic trypanosomatids was determined by binomial regression. Thirty D. marsupialis specimens (60.0% females and 40.0% males/66.7% adults and 33.3% juveniles) were collected. Molecular diagnosis revealed a frequency of trypanosomatid parasite infection of 46.7%. Stage (p=0.024) was a determinant for infection. We discuss the role of D. marsupialis as a potential reservoir of parasitic trypanosomatids in the Vereda El Alférez.
Se evaluó la prevalencia de infección por parásitos tripanosomátidos en Didelphis marsupialis y su relación con los aspectos morfológicos/etarios en una zona rural de El Carmen de Bolívar, Colombia. En cinco visitas (2018-2019) se instalaron trampas Tomahawk® en los ecótopos peridoméstico y silvestre en la Vereda El Alférez, durante tres noches consecutivas/visita. A los animales recolectados, se les determinaron medidas corporales, sexo y edad; y se les extrajo sangre por cardiopuntura, previa sedación, para extracción del ácido desoxirribonucleico (ADN) total y amplificación de la región conservada del ADN de minicírculos de kinetoplasto (ADNk) de parásitos tripanosomátidos. La asociación entre parámetros morfológicos de los didélfidos y su frecuencia de infección por parásitos tripanosomátidos fue determinada mediante una regresión binomial. Se recolectaron 30 individuos de D. marsupialis (60,0% hembras y 40,0% machos/66,7% adultos y 33,3% juveniles). El diagnóstico molecular reveló una frecuencia de infección por parásitos tripanosomátidos del 46,7%. El estadio (p=0,024) fue determinante para la infección. Se discute el papel de D. marsupialis como potencial reservorio de parásitos tripanosomátidos en la zona evaluada.
Asunto(s)
Didelphis , Parásitos , Animales , Masculino , Femenino , Didelphis/parasitología , Reservorios de Enfermedades , Colombia/epidemiología , ADNRESUMEN
Se evaluó la prevalencia de infección por parásitos tripanosomátidos en Didelphis marsupialis y su relación con los aspectos morfológicos/etarios en una zona rural de El Carmen de Bolívar, Colombia. En cinco visitas (2018-2019) se instalaron trampas Tomahawk® en los ecótopos peridoméstico y silvestre en la Vereda El Alférez, durante tres noches consecutivas/visita. A los animales recolectados, se les determinaron medidas corporales, sexo y edad; y se les extrajo sangre por cardiopuntura, previa sedación, para extracción del ácido desoxirribonucleico (ADN) total y amplificación de la región conservada del ADN de minicírculos de kinetoplasto (ADNk) de parásitos tripanosomátidos. La asociación entre parámetros morfológicos de los didélfidos y su frecuencia de infección por parásitos tripanosomátidos fue determinada mediante una regresión binomial. Se recolectaron 30 individuos de D. marsupialis (60,0% hembras y 40,0% machos/66,7% adultos y 33,3% juveniles). El diagnóstico molecular reveló una frecuencia de infección por parásitos tripanosomátidos del 46,7%. El estadio (p=0,024) fue determinante para la infección. Se discute el papel de D. marsupialis como potencial reservorio de parásitos tripanosomátidos en la zona evaluada.
We studied the prevalence of infection by trypanosomatid parasites in Didelphis marsupialis and its relationship with morphological/age aspects in a rural area of El Carmen de Bolivar, Colombia. Five visits were made to the Vereda El Alférez; each of which lasted three consecutive nights. During these visits, Tomahawk® traps were installed in the peridomestic and wild ecotopes of the Vereda El Alférez. Body measurements, sex and age were determined from the collected animals. Blood was extracted by cardiopuncture, after sedation, in order to obtain total deoxyribonucleic acid (DNA) and amplify the conserved region of the kinetoplast minicircle DNA (kDNA) of parasitic trypanosomatids. The association between morphological parameters of didelphids and their frequency of infection by parasitic trypanosomatids was determined by binomial regression. Thirty D. marsupialis specimens (60.0% females and 40.0% males/66.7% adults and 33.3% juveniles) were collected. Molecular diagnosis revealed a frequency of trypanosomatid parasite infection of 46.7%. Stage (p=0.024) was a determinant for infection. We discuss the role of D. marsupialis as a potential reservoir of parasitic trypanosomatids in the Vereda El Alférez.
Asunto(s)
Animales , Zoonosis Bacterianas , Marsupiales , LeishmaniasisRESUMEN
For the molecular diagnosis of Chagas disease by real-time PCR (polymerase chain reaction), optimization of diagnostic accuracy is desirable. The detection limit of real-time PCR assays for the diagnosis of Trypanosoma cruzi in human serum is affected by various influences including the choice of the nucleic acid extraction assay. In this study, three nucleic acid extraction assays were compared regarding their influence on the sensitivity of a T. cruzi-specific real-time PCR with 62 reference sera containing T. cruzi target DNA (deoxyribonucleotide acid). More than 95% of the positive sera were correctly identified after all three nucleic acid extraction strategies with a detection rate ranging from 96.8% (60/62) for the worst assay to 100% (62/62) for the best one. A matched pairs analysis for the comparison of the cycle threshold (Ct) values obtained with the 59 reference samples with positive real-time PCR results after all three nucleic acid extraction schemes indicated differences in a range of about 3 Ct steps. Summarized, all three compared nucleic acid extraction schemes were basically suitable for T. cruzi-specific PCR from serum with some minor differences. However, in the case of low quantities of circulating parasite DNA in the serum of a patient with Chagas disease, even minor effects can make a difference in the individual diagnosis.
RESUMEN
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.
