Experimental Evidence of Biological Interactions among Different Isolates of Trypanosoma cruzi from the Chaco Region

March Experimental Evidence of Biological Interactions among Different Isolates of Trypanosoma cruzi from the Chaco Region Paula G. Ragone 0 1 2 Cecilia Prez Brandn 0 1 2 Mercedes Monje Rumi 0 1 2 Nicols Tomasini 0 1 2 Juan J. Lauthier 0 1 2 Rubn O. Cimino 0 1 2 Alejandro Uncos 0 1 2 Federico Ramos 0 1 2 Anah M. Alberti DAmato 0 1 2 Miguel A. Basombro 0 1 2 Patricio Diosque 0 1 2 0 1 Unidad de Epidemiologia Molecular, Instituto de Patologia Experimental, CONICET (Consejo Nacional de Investigaciones Cientificas y Tecnicas), Universidad Nacional de Salta, Salta-Capital, Argentina, 2 Instituto de Patologia Experimental, CONICET (Consejo Nacional de Investigaciones Cientificas y Tecnicas), Universidad Nacional de Salta , Salta-Capital, Argentina, 3 Catedra de Quimica Biologica , Facultad de Ciencias Naturales, Universidad Nacional de Salta , Salta-Capital , Argentina 1 Funding: This manuscript was supported by funding agency Argentina: Agencia Nacional de Promocion Cientifica y Tecnologica (http://www.agencia.mincyt. gob.ar), through a grant given to Dr. Patricio Diosque (PICT-2012-2174). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript 2 Academic Editor: Herbert B. Tanowitz, Albert Einstein College of Medicine, UNITED STATES Many infectious diseases arise from co-infections or re-infections with more than one genotype of the same pathogen. These mixed infections could alter host fitness, the severity of symptoms, success in pathogen transmission and the epidemiology of the disease. Trypanosoma cruzi, the etiological agent of Chagas disease, exhibits a high biological variability often correlated with its genetic diversity. Here, we developed an experimental approach in order to evaluate biological interaction between three T. cruzi isolates belonging to different Discrete Typing Units (DTUs TcIII, TcV and TcVI). These isolates were obtained from a restricted geographical area in the Chaco Region. Different mixed infections involving combinations of two isolates (TcIII + TcV, TcIII + TcVI and TcV + TcVI) were studied in a mouse model. The parameters evaluated were number of parasites circulating in peripheral blood, histopathology and genetic characterization of each DTU in different tissues by DNA hybridization probes. We found a predominance of TcVI isolate in blood and tissues respect to TcIII and TcV; and a decrease of the inflammatory response in heart when the damage of mice infected with TcVI and TcIII + TcVI mixture were compared. In addition, simultaneous presence of two isolates in the same tissue was not detected. Our results show that biological interactions between isolates with different biological behaviors lead to changes in their biological properties. The occurrence of interactions among different genotypes of T. cruzi observed in our mouse model suggests that these phenomena could also occur in natural cycles in the Chaco Region. - Competing Interests: The authors have declared that no competing interests exist. Advances in molecular typing techniques show that many infectious diseases may arise from co-infections or re-infections with more than one genotype of the same pathogen. In these mixed infections the co-infecting parasites may be interacting among each other within the same host determining host fitness, severity of disease symptoms, parasite transmission successful rate and epidemiology of the disease [1]. Various mechanisms can cause interactions between parasite species or among different genotypes of the same species within an individual host. For example, parasites can infect the same target site within a host and directly interact among each other by interference competition, or indirectly by resources competition or via the host immune system [2]. In general, biological interactions between protozoan parasites have been divided into two main groups: those who involved parasites belonging to the same species and the ones that occur between closely to distantly related different species [3]. In this sense, several research studies have reported mixed infections in Leishmania spp [4,5,6], Plasmodium spp [7,8,9], Trypanosoma brucei and Trypanosoma congolense [10,11,12]. Trypanosoma cruzi is the etiologic agent of Chagas disease, an illness that affects several million people in Latin America and still remains an important public health problem in certain endemic areas of Argentina. This parasite shows a high genetic variability which has been the basis to classify it into six Discrete Typing Units (DTUs), TcI to TcVI [13]. In addition to this genetic diversity, in vitro and in vivo T. cruzi infection models showed a high biological variability among different genotypes of T. cruzi [14,15,16,17,18,19]. Although it is supposed that genetic and biological diversities of the parasite are essential to determine the clinical course of Chagas disease, specific associations between particular clinical manifestations and a determined lineage have not been clearly demonstrated [20]. Furthermore, the host genetics and its ability to establish an immune response to control the infection are very important in the outcome of the disease [21]. The consequences of mixed infections by different T. cruzi DTUs have been studied in animal models using different laboratory strains. It has been demonstrated in vivo that the tissue tropism of one T. cruzi genotype could change in the presence of another genotype of a different DTU [22,23]. Even more, the histopathological damage and the intensity of the inflammatory process resulting of these co-infections also present remarkable variations [24,25]. Other studies involving T. cruzi mixed infections showed that the parasite load in peripheral blood could be altered either increasing or decreasing according to the co-infecting strains [26,27,28]. Even the outcome of specific chemotherapy has been proven to be altered by these events of concomitant infection by T. cruzi [26,29]. In several geographical areas of the Southern Cone of America, the occurrence of natural mixed infections by different genotypes of T. cruzi have been widely reported in humans [20,30,31,32,33], in wild and domestic animals [34,35] and in the vector Triatoma infestans [34,36]. In a previous work we described the different biological properties displayed by three selected isolates obtained from the Chaco region of Argentina and belonging to DTUs TcIII, TcV and TcVI [17]. These isolates have the particularity of circulating sympatrically in a restricted geographical area; therefore, here we describe the biological outcome resulting of in vivo experimental dual-mixed infections with these T. cruzi strains. Our working hypothesis is the existence of biological interactions among different T. cruzi isolates in the vertebrate host. All animal protocols adhered to the National Institutes of Health (NIH) Guide for the care and use of laboratory animals and were app (...truncated)