Infección experimental de vectores de la enfermedad de Chagas (Reduviidae: Triatominae) y de un modelo murino, con cepas de Trypanosoma rangeli (Kinetoplastida: Trypanosomatidae) genéticamente diferentes.

Magíster Scientae en Enfermedades TropicalesSe compararon dos cepas de T. rangeli, de Colombia y de Costa Rica, genotipicamente diferentes (KP1+ y KP1-), las cuales presentaron similitudes y diferencias en cuanto a diversos parámetros observados tanto in vivo como in vitro. Ninguna de ellas logro crecer en medio líquido de Warren, aunque crecieron bien si este se adiciona de suero fetal bovino al 10 o 20 %. Las cepas presentaron diferencias en cuanto al tiempo para alcanzar los picos máximos de crecimiento que siempre fueron mayores para la cepa KP1-. Ninguna de las dos cepas logro infectar células Vero en cultivo de tejidos. En cuanto a la infección comparativa de insectos triatóminos, inoculados por vía hemocelómica, la cepa KP1- logró completar el ciclo en su vector natural R. pallescens, mientras que la cepa KP1+ logró lo mismo tanto en R. prolixus como en R. pallescens, lo cual no se reporta en la literatura. Ambas cepas infectaron únicamente la hemolinfa de T. lecticularia y de T. bruneri ya que no consiguieron completar el ciclo en las glándulas salivales. En las especie T. dimidiata y T. infestans, las infecciones hemolinfáticas, con la única cepa que logró infectarlas (KP1+), fueron muy bajas y transitorias, mientras que P. geniculatus fue totalmente refractario a la infección con ambas cepas. Los cultivos de las dos cepas recién recuperadas de ratones infectados, fueron capaces de infectar nuevos ratones de las razas Balb/c y Swiss, mientras que las mismas cepas mantenidas en el laboratorio en medios de cultivo por mas de 2 años, perdieron su capacidad de infectar ratones, aún usando inóculos altos. No se notó una diferencia significativa en cuanto a la susceptibilidad de ambas razas de ratones para cada una de las cepas. En un estudio morfométrico comparativo de tripomastigotos sanguíneos, se comprobó que la cepa KP1+ aventajó significativamente en longitud y en el índice PN/NA a la cepa KP1-, lo cual sugiere que la morfometría podría ser un criterio de diferenciación entre cepas fenotípicamente diferentes. Estos resultados permiten afirmar que las cepas de T. rangeli de genotipos diferentes presentan diferencias importantes en cuanto al comportamiento biológico y morfológico tanto en condiciones in vitro como in vivo. Palabras claves Trypanosoma rangeli, Insectos triatóminos, Interacción parásito-vector, Cepas KP1- y KP1+, Genotipos, Morfometría x xi ABSTRACT Two genotypically different T. rangeli isolates, (KP1+ and KP1-), from Colombia and Costa Rica, were compared under different in vitro and in vivo parameters, presenting similarities and differences in some aspects. None of them was able to grow in Warren’s liquid medium except when 10 to 20 % fetal calf serum was added to the medium. A difference in time for reaching the peak of the growth curve was noticed for the two isolates and KP1- strain exhibit higher figure at the peak. Also, none of them was able to grow in tissue culture inside Vero cells. In relation to their ability to infected triatomine insects by hemocelomic injection of the parasite, the KP1- strain was able to complete the salivary gland cycle in R. pallescens, whereas the KP1+ strain completed the cycle in both R. prolixus and R. pallescens, something not reported in the literature. Both strains infected the hemolymph of T. lecticularia and of T. bruneri but were unable to complete the cycle in the salivary glands. In T. dimidiata and T. infestans the infection with KP1+, the only strain that infected these bugs, were low and transient, whereas P. geniculatus was completely refractory to the infection by both isolates. The two strains, after reisolation in cultures, were able to infect two breed of mice (Balb/c and Swiss) whereas, even with high inocula, the original strains, maintained in culture for more than 2 years, were unable to infect mice. Both breeds behaved in this respect similarly. Morphometric studies of blood trypomatigotes showed that those from the KP1+ strain were significantly longer and with larger PN/NA index than those from strain KP1-. These results suggest that both genotypes could be differentiated by their size in blood. The results allow us to believe that T. rangeli of different genotypes exhibits some differences in relation to biological and morphological characteristics, both in vitro and in vivo.Universidad Nacional, Costa Rica.Escuela de Medicina Veterinari

Intracellular passage triggers a molecular response in that increases its infectiousness

Brucella abortus is a facultative extracellular-intracellular pathogen that encounters a diversity of environments within the host cell. We report that bacteria extracted from infected cells at late stages (48 h post-infection) of the intracellular life cycle significantly increase their ability to multiply in new target cells. This increase depends on early interaction with the cell surface since bacteria become more adherent and penetrate more efficiently as compared to in vitro grown bacteria. At this late stage of infection, the bacterium locates within an autophagosome-like compartment, facing starving and acidic conditions. At this point, the two-component system BvrR/BvrS becomes activated, and the expression of the transcriptional regulator VjbR and the type IV secretion system VirB increased. Using bafilomycin to inhibit BvrR/BvrS activation and specific inhibitors for VjbR and VirB we showed that the BvrR/BvrS and VjbR systems correlate with the increased interaction with new host cells while the VirB system does not. Bacteria released from infected cells under natural conditions displayed the same phenotype as intracellular bacteria. We propose a model in which the B. abortus BvrR/BvrS system senses the transition from its replicative niche at the endoplasmic reticulum to the autophagosome-like exit compartment. This activation leads to the expression of VirB participating in the release of the bacterium from the cells and an increase in VjbR expression that results in a more efficient interaction with new host cells.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Centro de Investigación en Enfermedades Tropicales (CIET