Biological diversity of T. cruzi

In the vertebrate host

T. cruzi displays great polymorphism or biological diversity in several aspects, which may be correlated with the morphology of this parasite in the bloodstream. Since the early studies of Chagas disease discovery,1 blood polymorphism has been interpreted by some authors as indicative of sexual dimorphism of the parasite.71 Although the clonal theory of T. cruzi structure and evolution is accepted,72 the hypothesis of T. cruzi sexuality remains an open question for some authors.73

Some evidence suggests that the morphological variations of T. cruzi are related to the parasites’ physiological variations. Studying different T. cruzi populations, Brener74 and Brener and Chiari75 verified that in some strains the slender forms (Fig. 16.7A) are present throughout the course of infection, whereas in others the slender parasites are present only in the first days of infection and are gradually replaced by the broad forms (Fig. 16.7B) that become predominant in the later days of the acute infection. Therefore, when blood trypomastigotes are inoculated via the intravenous route in normal mice, the slender forms quickly disappear from the circulation to reproduce within the cell. Moreover, the broad forms remain in the circulation without interacting and infecting the host cells.76 Additionally, it has been demonstrated that naturally surviving animals previously infected with T. cruzi and intravenously inoculated with slender trypomastigotes, are rapidly destroyed, while the broad ones are resistant and remain in the circulation for a long time. Afterward, Howells and Chiari77 demonstrated that broad trypomastigotes present greater ability to infect the triatomine vectors than the slender parasites, which degenerate without carrying out metacyclogenesis. Interestingly, blood trypomasti- gotes are antigenically different from trypomastigotes obtained in cellular culture.75 Moreover, blood trypomastigotes have a different ability to infect vertebrate and invertebrate hosts.79,80

This dual behavior in strains with predominantly slender (Fig. 16.7A) or intermediate (Fig. 16.7D) versus large or stout trypomastigotes (Figs. 16.7B and 16.7C) led8 to purposing the term “polar strains” for the Y and CL strains. Striking differences are observed in the parasitemia curves induced by the two strains in experimental hosts such as mice, dogs, and rabbits. The Y strain infects mice more efficiently and higher parasitemia is observed early in the course of infection followed by early mortality, whereas in animals infected with the CL strain, infection, increase of parasitemia and mortality are observed later. In addition, the Y strain exhibits tropism for cells from the mononuclear phagocytic system as demonstrated by the peculiar parasitism of macrophages from the spleen, liver, and bone marrow; in contrast, the CL strain induces a negligible parasitism in these cells. As both strains infect muscle cells, the concept of macrophagotropic and nonmacrophago- tropic strains has been suggested to characterize these distinct tropisms. In vitro experiments with mouse peritoneal resident macrophages confirmed the in vivo findings. In addition, blood forms of the Y strain (but not the CL strain) collected

Figure 16.7 Morphological variation of blood trypomastigotes in Trypanosoma cruzi. (A) slender; (B) broad; (C) stout; and (D) intermediate.⁷⁴

from infected mice in the acute phase are readily lysed by complement via membrane-bound specific immunoglobulins. The resistance of the CL strain to the complement-mediated lyses strongly suggests that broad blood forms are equipped with evasion mechanisms that are lacking in the Y population.

The observation that different tissue tropism (cardiotropism or myotropism) of T. cruzi is verified in individuals infected with parasites with distinct genetic characteristics⁸¹ led Macedo et al.⁵ to propose the “clonal histiotropic model theory” for this parasite since the correlation between T. cruzi genetic and tissue tropism could be associated with the different clinical manifestations of the disease (cardiopathy, megaesophagus, and megacolon), alone or in combination.

These variations in T. cruzi morphology are correlated with several other aspects of its infection in the murine model such as parasitemia curves, tissue tropism, mortality and resistance or susceptibility to treatment. These associations led Andrade⁸² to propose the subdivision of T. cruzi populations into types I, II, and III. Type I includes the Y and Peruvian strains, which present predominance of slender trypo- mastigotes, rapid multiplication in mice, a higher and early parasitemia peak (Fig. 16.8A), higher and early mortality, and particularly tropism for macrophagic cells during the acute phase of infection, and are susceptible or partially resistant to benznidazole and nifurtimox. Type II includes the strains typical of “Reconcavo baiano,” Bahia state, Brazil, which presents a predominance of broad trypomasti- gotes but with slender forms at the beginning of infection, slow multiplication in mice (Fig. 16.8B), irregular peaks of parasitemia between 12 and 20 days, null mortality in the acute phase, myocardial tropism, and is partially resistant or resistant to treatment. Type III includes, e.g., the Colombian strain, which presents a predominance of broad trypomastigotes throughout infection, slow multiplication in mice, higher peaks of parasitemia between 20 and 30 days of infection (Fig. 16.8C), lower rates of mortality occurring slowly throughout acute infection, tropism for the skeletal musculature, and resistance to treatment. Types I, II, and III correspond to Biodemes I, II, and III, respectively,⁸³ since these associations were observed with the isoenzymatic profile of the parasites. Several other authors observed similar associations between the parasite genetics and its biological properties when comparatively studying clonal stocks in acellular culture,⁸⁴ cellular culture,⁸⁵ and mice⁸⁶ of the three principal T. cruzi genotypes,⁸⁷ such as principal genotypes 19 and 20, or T. cruzi I, principal genotype 32 or T. cruzi II, and principal genotype 39 or T. cruzi hybrid genotype, currently called T. cruzi I, T. cruzi II, and T. cruzi V groups, respectively, according to the recent genetic T. cruzi classification.² They are equivalent to Biodemes I, II, and III,⁸³ respectively. These authors demonstrated higher values for the growth and metacyclogenesis variables in acellular culture,⁸⁴ infectivity and differentiation in trypomastigotes in cellular culture,⁸⁵ virulence, and parasitemia in mice⁸⁶ for the T. cruzi I group in relation to T. cruzi II group, differently from the published data about TcI T. cruzi strains isolated from the Occidental Amazonia in Brazil⁸⁸; whereas T. cruzi V presented intermediate values for these same variables. With the same group of clonal stocks, Revollo et al.⁸⁵ and Toledo et al.⁸⁹ also demonstrated that T. cruzi I was more resistant to benznidazole and nifurtimox (in vitro) and benznidazole and itraconazole (in mice), whereas T. cruzi II was the most susceptible, and the hybrid stocks (T. cruzi V) presented intermediate results. Moreover, all stocks belonging to the principal genotype 20 were 100% resistant to treatment with all the compounds assayed in vitro and in vivo. These results may explain the regional differences observed in human chemotherapy, which differs from region to region, and explains why it is easier to cure patients from Argentina and Chile, where T. cruzi II is predominant, than in the north of the Americas, where T. cruzi I is more widespread.^90,91

Figure 16.8 Parasitemia curves in mice infected with Y (A), Be-78 (B) and Colombiana (C) strains of Trypanosoma cruzi corresponding to Types I, II, and III, respectively, according to the Andrade (1974)⁷⁹ classification.

The genetic polymorphism in T. cruzi is also related to trypomastigote small surface antigen (TSSA) sequences, a highly antigenic surface glycosylphosphatidyl inositol (GPI)-anchored mucin-like protein. Thus, Di Noia et al.⁹² described the first immunological marker in T. cruzi that discriminated the two T. cruzi lineage groups (T. cruzi I and T. cruzi II).