Cavia porcellus as a Model for Experimental Infection by Trypanosoma cruzi

Abstract

The guinea pig (Cavia porcellus) is a natural reservoir for Trypanosoma cruzi but has seldom been used as an experimental infection model. We developed a guinea pig infection model for acute and chronic Chagas disease. Seventy-two guinea pigs were inoculated intradermally with 10 4 trypomastigotes of T. cruzi strain Y (experimental group); 18 guinea pigs were used as control group. Eight animals from the experimental group and two from the control group were Chagas disease, caused by the protozoan parasite Trypanosoma cruzi, is one of the most important parasitic infections of the Americas. Despite successful vector control programs in many countries, the disease remains a major public health problem in Latin America, with 8 to 10 million people currently infected, an annual incidence of 65,000 new cases in 15 countries, and 14,000 deaths associated with the infection per year. 1 Transmission to the mammalian host occurs when a T. cruzi-infected triatomine vector deposits infectious trypomastigotes in feces during a blood meal and the parasites invade through the bite wound or intact mucosal membranes. 2 Trypomastigotes can infect virtually any nucleated cell, where they transform to amastigotes and replicate in the host cell cytoplasm. Amastigotes convert back to trypomastigotes inside the cell and then rupture the cell and circulate in the bloodstream. After weeks, the host immune response usually controls the acute infection but does not completely clear the parasite. This results in lifelong chronic infection of the host.The initial weeks after T. cruzi infection are characterized by a high-level parasitemia and symptoms that vary from mild nonspecific febrile illness to life-threatening myocarditis and/or meningoencephalitis. 2 Acute symptoms and detectable parasitemia resolve spontaneously within 2 to 3 months, and the infected individual passes into the chronic phase of the disease. Nearly all individuals in the early period of chronic infection are asymptomatic and are said to have the indeterminate form of the disease. In most individuals, the infection remains silent for life and is detectable only by anti-T. cruzi serologic tests and, in a proportion of individuals, by molecular Supported by NIH training grant in infectious and tropical diseases 5 T35 AI065385 and NIH grant 1R01AI087776-01.

Domestic Pig (Sus scrofa) as an Animal Model for Experimental Trypanosoma cruzi Infection

“…Unfortunately, none of the infected pigs were positive for the presence of DNA in the heart, which is different from other animal models studied such as guinea pig in which the heart was the most frequent tissue with parasite kinetoplast DNA as compared with other organs at three different stages of infection: acute phase (5-55 days), early chronic phase (115-165 days), and chronic phase (365 days). 32 Interestingly, our study found histopathological alterations in the heart without the detection of the parasite. This is similar to a study with chronically infected dogs that demonstrated no detectable parasites in the heart either by microscopic observation or by PCR.…”

“…The number of blood samples positive for PCR decreased during the chronic phase, and similar results were observed in the chronic model of T. cruzi infection in guinea pigs. 32 PCR was also used in the evaluation of different tissues collected at 150 dpi. Unfortunately, none of the infected pigs were positive for the presence of DNA in the heart, which is different from other animal models studied such as guinea pig in which the heart was the most frequent tissue with parasite kinetoplast DNA as compared with other organs at three different stages of infection: acute phase (5-55 days), early chronic phase (115-165 days), and chronic phase (365 days).…”

“…DNA was purified by Proteinase K digestion (Invitrogen, Carlsbad, CA) and was extracted following a standard phenol-chloroform extraction protocol as previously described. 32,33 We used 500 μL of clot or 25 mg of tissue stored in ethanol. The extracted DNA was resuspended in 100 μL of 10 mM TrisHCl and 1 mM ethylenediaminetetraacetic acid.…”

“…35 DNA extraction from urine samples was developed according to published protocols with some modifications. 32 Briefly, 25 μL of concentrated urine samples were incubated with 10 mmol/L TrisHCl, pH 7.6, 10 mmol/L NaCl, and mixed gently for 5 minutes. Sodium dodecyl sulfate (SDS) and Proteinase K (Invitrogen) were added to reach concentrations of 0.25% and 0.50 mg/mL, respectively, and the specimens were incubated for 1 hour at 56°C.…”

“…The kinetics for the production of IgM and IgG to T. cruzi was determined by epimastigote alkaline extract (EAE)-ELISA, as described previously with some modifications. 32 Briefly, ELISA 96-well plates (Immulon 2; Thermo Labsystems, Franklin, MA) were coated with 2 μg/mL EAE antigen in 1 M bicarbonate buffer (pH 9.6) overnight and blocked with 5% skim milk (Nestle) in PBS (pH 7.2) and 0.05% Tween 20 (PBST). The ELISA plates were incubated with pig serum diluted 1:100 (IgG detection) or 1/50 (IgM detection) in 1% dried skim milk/PBST for 1 h at 37°C.…”