Parasitemia Curve Research Articles

Heterologous expression in Caenorhabditis elegans as an alternative approach to functional studies in Schistosoma mansoni.

The lack of an accurate diagnosis has been a serious obstacle to the advancement of the anti-Trypanosoma cruzi chemotherapy and long-term infection can result in different health risks to human. PCRs are alternative methods, more sensitive than conventional parasitological techniques, which due to their low sensitivities are considered unsuitable for these purposes. The aim of this study was to investigate a sensitive diagnostic strategy to quantify blood and cardiac tissues parasites based on real-time PCR tools during acute and chronic phases of murine Chagas disease, as well as to monitor the evolution of infection in those mice under specific treatment. In parallel, fresh blood examination, immunological analysis and quantification of cardiac inflammation were also performed to confront and improve real-time PCR data. Similar profiles of parasitemia curves were observed in both quantification techniques during the acute phase of the infection. In contrast, parasites could be quantified only by real-time PCR at 60 and 120 days of infection. In cardiac tissue, real-time PCR detected T. cruzi DNA in 100% of infected mice, and using this tool a significant Pearson correlation between parasite load in peripheral blood and in cardiac tissue during acute and chronic phases was observed. Levels of serum CCL2, CCL5 and nitric oxide were coincident with parasite load but focal and diffuse mononuclear infiltrates was observed, even with significant (p < 0.05) reduction of parasitism after 60 days of infection. Later, this methodology was used to monitor the evolution of infection in animals treated with itraconazole (Itz). Itz-treatment induced a reduction of parasite load in both blood and cardiac muscle at the treatment period, but after the end of chemotherapy an increase of parasitism was detected. Interestingly, inflammatory mediators levels and heart inflammation intensity had similar evolution to the parasite load, in the group of animals treated. Taken together, our data show that real-time PCR strategy used was suitable for studies of murine T. cruzi infection and may prove useful in investigations involving experimental chemotherapy of the disease and the benefits of treatment in relation to parasitism and inflammatory respons.

Hematological alterations during experimental canine infection by Trypanosoma cruzi

To confirm that Beagle dogs are a good experimental model for Chagas disease, we evaluated hematological alterations during the acute and chronic phases in Beagle dogs infected with the Y, Berenice-78 (Be-78) and ABC strains of Trypanosomacruzi, correlating clinical signs with the parasitemia curve. We demonstrate that the acute phase of infection was marked by lethargy and loss of appetite. Simultaneously, we observed anemia, leukocytosis and lymphocytosis. Also,we describe hematological alterations and clinical signs that were positively correlated with the parasitemia during the experimental infection with the three strains of T.cruzi, and demonstrate that experimental infection of Beagle is a trustworthy model for Chagas disease.

Real-time PCR strategy for parasite quantification in blood and tissue samples of experimental Trypanosoma cruzi infection

The lack of an accurate diagnosis has been a serious obstacle to the advancement of the anti-Trypanosoma cruzi chemotherapy and long-term infection can result in different health risks to human. PCRs are alternative methods, more sensitive than conventional parasitological techniques, which due to their low sensitivities are considered unsuitable for these purposes. The aim of this study was to investigate a sensitive diagnostic strategy to quantify blood and cardiac tissues parasites based on real-time PCR tools during acute and chronic phases of murine Chagas disease, as well as to monitor the evolution of infection in those mice under specific treatment. In parallel, fresh blood examination, immunological analysis and quantification of cardiac inflammation were also performed to confront and improve real-time PCR data. Similar profiles of parasitemia curves were observed in both quantification techniques during the acute phase of the infection. In contrast, parasites could be quantified only by real-time PCR at 60 and 120 days of infection. In cardiac tissue, real-time PCR detected T. cruzi DNA in 100% of infected mice, and using this tool a significant Pearson correlation between parasite load in peripheral blood and in cardiac tissue during acute and chronic phases was observed. Levels of serum CCL2, CCL5 and nitric oxide were coincident with parasite load but focal and diffuse mononuclear infiltrates was observed, even with significant (p<0.05) reduction of parasitism after 60 days of infection. Later, this methodology was used to monitor the evolution of infection in animals treated with itraconazole (Itz). Itz-treatment induced a reduction of parasite load in both blood and cardiac muscle at the treatment period, but after the end of chemotherapy an increase of parasitism was detected. Interestingly, inflammatory mediators levels and heart inflammation intensity had similar evolution to the parasite load, in the group of animals treated. Taken together, our data show that real-time PCR strategy used was suitable for studies of murine T. cruzi infection and may prove useful in investigations involving experimental chemotherapy of the disease and the benefits of treatment in relation to parasitism and inflammatory response.

Influence ofTriatoma dimidiatain Modulating the Virulence ofTrypanosoma cruziMexican Strains

The epidemiology of Chagas disease is complex. There are different vectors and reservoirs and different clinical manifestations. In order to assess whether the biological behavior of three strains isolated in southeastern Mexico (H4 isolated from human, Z17 isolated from Didelphis sp., and V isolated from T. dimidiata) could be modified during passage through the vector T. dimidiata, the parasitemia curve, the amount of amastigote nests, and mortality of BALB/c infected with blood trypomastigotes of T. cruzi were evaluated. Strains were maintained in continuous passage from mouse to mouse and in animals infected with metacyclic trypomastigotes. The parasitemia curves were significantly different (P < 0.05) between mice to mice and triatoma to mice groups in strains H4 and Z17, and was also observed fewer amastigote nests in cardiac tissue (P < 0.05 strain H4 with higher number versus all groups and Z17 between mice to mice and triatoma to mice) 45 days after inoculation. It is concluded that T. dimidiata influences in modulating the virulence of strains of T. cruzi in the region. Further studies of the intestinal tract of the insect in search for some protein molecules involved in regulating may clarify the virulence of the parasite.

Morphological, biological and molecular characterization of three strains ofTrypanosoma cruziChagas, 1909 (Kinetoplastida, Trypanosomatidae) isolated fromTriatoma sordida(Stal) 1859 (Hemiptera, Reduviidae) and a domestic cat

SUMMARYA study was conducted of the biological, morphological and molecular characters of 3 strains of Trypanosoma cruzi (SI5, SI8 and SIGR3) isolated from specimens of Triatoma sordida collected in Santo Inácio and a domestic cat. In order to carry out the study, the following parameters were evaluated: pre-patent period, parasitaemia curves, morphology of the parasites, mortality rates, histopathological lesions and molecular typing. The strains presented variable pre-patent periods, low parasitaemia and no animal mortality. The morphological study of trypomastigotes showed a predominance of intermediate-width and short-length forms, as well as low nuclear index. Epimastigotes presented a low nuclear index, intermediate-width forms in strains SI5 and SI8, and large-width forms in SIGR3. A shorter length could be noted in strains SI8 and SIGR3, whereas SI5 displayed an intermediate length. The histopathological study did not detect amastigote nests in tissues. The amplification of the divergent domain of 24Sα rRNA, HSP60 and GPI genes of strains SI5, SI8 and SIGR3 classified the 3 strains into Group II. Biological parameters made it possible to classify the strains isolated in Santo Inácio (BA) into Biodeme III, Zymodeme 1 and Group II of T. cruzi.

Organ tropism during the acute and chronic phases of Trypanosoma cruzi infection in BALB/c mice

The aim of the present study was to investigate the presence of Trypanosoma cruzi in the heart, liver, lung, and kidneys, using hemoculture and PCR analysis, of mice infected with different parasite strains during the acute and chronic phases of infection. Parasitemia curves revealed strain-specific biological behaviors. For the Y and JLP strains, the acute phase of infection started at days six and ten post-infection, parasitemia peaked at days seven and 15 post-infection, the chronic phase started at days nine and 28 post-infection, and animals started dying at days 19 and 120 post-infection, respectively. When the two strains were compared, the JLP strain exhibited reduced and slower replication rates associated with a delayed peak of parasitism and reduced parasite burdens. However, parasites were detected in all studied organs using PCR analysis. The capacity of both strains to infect different organs likely influences disease pathogenesis.

Trypanosoma cruzi infection: do distinct populations cause intestinal motility alteration?

Chagas disease, caused by Trypanosoma cruzi, is an important public health problem in Latin America. Disturbances in gastrointestinal motility are observed in 15-20% of patients at the chronic phase. We previously observed a decrease in intestinal motility in mice infected with Y strain from T. cruzi. Thus, we decided to test if infection with other T. cruzi strains also caused the intestinal disturbance. Male adult Swiss mice were infected intraperitoneally with CL-Brener clone (CL-B), Brazil strain (Br), or Dm28 clone (Dm) of T. cruzi. All infected mice presented a low cumulative mortality (CL-B, 17%; Br, 8%; Dm, 25%) at 35 days post infection (dpi) and their typical parasitemia curves. Br and Dm groups exhibited a maximal reduction of intestinal motility at 35 dpi (176.8 +/- 51.3 and 198.3 +/- 52.6 min, respectively), when compared with non-infected mice (90.2 +/- 19.5 min). However, CL mice presented the peak of delayed intestinal transit at 12 dpi (191.0 +/- 33.3 min), when compared with non-infected mice (105.6 +/- 26.4 min), very close to the 15 dpi for the intense alteration (310.2 +/- 67.4 min) observed with the Y strain. We clearly demonstrate a reduction in intestinal motility in mice infected with different groups of T. cruzi during the acute phase of the infection. Since Br, Dm, and CL strains presented low mortality rates in adult Swiss mice, a prospective study concerning the chronic intestinal alteration is encouraged, particularly for studies of alternative therapies.

Trypanosoma cruzi: Different methods of data analysis to evaluate the genetics–biology relationship

The correlation of genetic and biological diversity in Trypanosoma cruzi was studied. Strains of T. cruzi II, isolated from humans; and of T. cruzi I, isolated from wild-animal reservoirs and from triatomines in the state of Paraná, Brazil, were used. Thirty-six biological parameters measured in vitro and six in vivo, related to growth kinetics and metacyclogenesis, susceptibility to benznidazole, macrophage infection, and experimental infection in mice were evaluated. Data from RAPD and SSR-PCR were used as genetic parameters. Mantel’s test, group analysis, principal components analysis (PCA), and cladistical analyses were applied. With the Mantel’s test, a low correlation was observed when parameters related to growth kinetics and metacyclogenesis in vitro and development of the experimental infection in vivo were included. The group analysis defined two groups that were separated as to whether they produced patent parasitemia in BALB/c mice. In the larger group, strains derived from wild reservoirs were separated from strains derived from triatomines and humans. The PCA identified two groups that differed as to whether they produced a parasitemia curve in mice. The cladistical analysis supported the previous results. This study shows the importance of the parasite–host relationship for the behavior of the strains, and that the combination of methods supports, extends, and clarifies the available information.

Trypanosoma cruzi: Impact of dual-clone infections on parasite biological properties in BALB/c mice

Herein, we have analyzed major biological properties following dual-clone Trypanosoma cruzi infections in BALB/c mice. Eight T. cruzi clonal stocks, two of each principal genotype, including genotype 19 and 20 ( T. cruzi I), hybrid genotype 39 ( T. cruzi) and 32 ( T. cruzi II) were combined into 24 different dual-clone infections. Special attention was given to characterize biological parameters assayed including: prepatent period, patent period, maximum of parasitemia, day of maximum parasitemia, area under the parasitemia curve, infectivity, mortality, and hemoculture positivity. Our findings clearly demonstrated that features resultant of dual-clone infections of T. cruzi clonal stocks did not display either the characteristics of the corresponding monoclonal infections or the theoretical mixture based on the respective monoclonal infections. Significant changes in the expected values were observed in 4.2–79.2% of the mixtures considering the eight biological parameters studied. A lower frequency of significant differences was found for mixtures composed by phylogenetically distant clonal stocks. Altogether, our data support our hypothesis that mixed T. cruzi infections have a great impact on the biological properties of the parasite in the host and re-emphasizes the importance of considering the possible occurrence of natural mixed infections in humans and their consequences on the biological aspects of ongoing Chagas’ disease.

Effects of specific treatment on parasitological and histopathological parameters in mice infected with different Trypanosoma cruzi clonal genotypes.

The goal of this study was to verify the effect of specific treatment on parasitological and histopathological parameters in mice experimentally infected with different Trypanosoma cruzi clonal genotypes. Twenty cloned stocks were selected, representative of the whole phylogenetic diversity of the protozoan and belonging to the clonal genotypes 19 and 20 (T. cruzi I) and 39 and 32 (T. cruzi II). The stocks were inoculated in 40 BALB/c mice divided into four groups: (i) treated with benznidazole, (ii) treated with itraconazole and (iii and iv) untreated control groups (NT) for each drug, respectively. Seven parameters related to parasitaemia curves and histopathological lesions were analysed. Four during the acute phase (AP) and three during both the AP and chronic phase (CP) of infection. Statistical comparison between benznidazole-treated and NT groups for the biological parameters showed significant differences for all genotypes. Benznidazole treatment led to lower patent period, maximum of parasitaemia, day of maximum parasitaemia and area under the parasitaemia curve for all genotypes analysed. Percentage of positive haemoculture during AP and CP was lower for genotypes 19 and 32. Tissue parasitism (TP) and inflammatory process (IP) during AP were lower for genotypes 19 and 32, respectively. In general, itraconazole treatment induced a smaller reduction in these same parameters between treated and NT animals in relation to benznidazole treatment. Our results indicate that phylogenetic divergence among T. cruzi clonal genotypes must be taken in account in chemotherapy and studies dealing with all aspects of the parasite and the disease.

Selenium supplementation at low doses contributes to the decrease in heart damage in experimental Trypanosoma cruzi infection.

Chagasic patients with cardiomyopathy have low levels of selenium (Se), a fundamental trace element. We evaluated the effect of supplementing infected mice with Se (0.25-16 ppm). Supplementation with 0.25 or 1 ppm Se led to parasitaemia and survival curves similar to those of the control group. Mice treated with 4-16 ppm showed a dose-dependent decrease of parasitaemia, significant for the highest concentration. This was probably due to a direct effect on the parasites, which were lysed after in vitro incubation with Se. Survival rates did not change significantly; however, heart damage was reduced in infected mice supplemented with 4 ppm Se, as indicated by a lower cardiac isoform of creatine kinase levels. Our results imply that Se supplementation does not lead to a general protection during infection, but may help protect the heart from inflammatory damage. The effect of Se supplementation in the course of T. cruzi infection depends on the host-parasite pair employed.

Hemoglobin concentration in children in a malaria holoendemic area is determined by cumulated Plasmodium falciparum parasite densities.

In malaria holoendemic areas children are anemic, but the exact influence of falciparum malaria on hemoglobin (Hb) concentration remains largely unsettled. Prospective data were therefore collected in children < 24 months of age during five months in a Tanzanian village. Children with mean asymptomatic parasitemia > or = 400/microl had lower median Hb levels during the study than those with mean density < 400/microl. The difference was 9.7 g/L (95% confidence interval [CI] 2.8-17). In children with one or more clinical malaria episodes, the median Hb was 8.3 g/L (95% CI 0.9-16) lower than those without episode. If early treatment failure was recorded, the immediate effect on Hb was particularly important with a mean drop of 17 g/L. Interestingly, at study-end the Hb concentration represented a function of the area under the parasitemia curve (AUPC) during the previous five months, adjusting for age. In conclusion, stepwise deterioration in median Hb levels was found by asymptomatic parasitemia, clinical malaria episode, and most significantly, treatment failure.

Life cycle of Trypanosoma cruzi (Y strain) in mice.

Since 1958, we have studied experimental Chagas' disease (CD) by subcutaneous inoculation of 1,000 blood forms of Trypanosoma cruzi (Y strain) in Balb/C. mice. Evolution of parasitemia remained constant, beginning on the 5th and 6th day of the disease, increasing progressively, achieving a maximum on about the 30th day. After another month, only a few forms were present, and they disappeared from the circulation after the third month, as determined from direct examination of slides and the use of a Neubauer Counting Chamber. These events coincided with the appearance of amastigote nests in the tissues (especially the cardiac ones), starting the first week, and following the Gauss parasitemia curve, but they were not in parallel until the chronic stage. In 1997, we began to note the following changes: Parasites appeared in the circulation during the first week and disappeared starting on the 7th day, and there was a coincident absence of the amastigote nests in the tissues. A careful study verified that young forms in the evolutionary cycle of T. cruzi (epi + amastigotes) began to appear alongside the trypomastigotes in the circulation on the 5th and 7th post-inoculation day. At the same time, rounded, oval, and spindle shapes were seen circulating through the capillaries and sinusoids of the tissues, principally of the hematopoietic organs. Stasis occurs because the diameter of the circulating parasites is greater than the vessels, and this makes them more visible. Examination of the sternal bone marrow revealed young cells with elongated forms and others truncated in the shape of a "C" occupying the internal surface of the blood cells that had empty central portions (erythrocytes?). We hypothesize that there could be a loss of virulence or mutation of the Y strain of Trypanosoma cruzi.

Influence of the Host Photoperiodicity on the Schizogonic Cycle and the Synchronism of the Rodent Malaria Plasmodium chabaudi chabaudi

This study intended to determine whether LD (Light:Dark) regimens different from the usual 12:12 hours could impair the schizogonic cycle and the synchronism of the rodent malaria parasite Plasmodium chabaudi chabaudi. Five illumination regimens of 12:12 LD, 5:5 LD, 18:18 LD, DD (constant dark) and LL (constant light) were used. Mice were kept in these regimens three months prior to and throughout the experiment. The total and the differential parasitaemia were checked every hour, during more than 24 hours. The parasitaemia data indicated that changes in the LD regimen, except for the LD 18:18, do not affect the length of the developmental cycle of this malaria parasite which remains 24 hours. In both the LL and 18:18 LD regimens, the synchronisation of parasites were impaired, mostly in the LL regimen. Also, we noticed that the schizogony which usually occurs in the dark part of the cycle may happen in the light part too. A circadian rhythm in the frequency of the schizogonic cycle and a time dependent occurrence of ring forms, trophozoites and schizonts were observed. At high parasitaemia, the infection was desynchronised. The total parasitaemia curves displayed a plateau region, followed by a drop at the end of the plateau, and an increase after the schizogony to reach the next plateau level.

Biological Characterization ofTrypanosoma cruziStocks from Chilean Insect Vectors

Solari, A., Wallace, A., Ortiz, S., Venegas, J., and Sanchez, G. Biological characterization ofTrypanosoma cruzistocks from Chilean insect vectors.Experimental Parasitology89,312–322. Fifty-sevenTrypanosoma cruzistocks isolated fromTriatoma infestansandTriatoma spinolaiof the five different geographic endemic areas of Chile were studied by schizodeme and molecular karyotype analysis. Four different genotypes are found in the sylvaticT. spinolaivector and five in theT. infestansdomiciliary vector. Of these genotypes, two common genotypes overlap on both transmission cycles exclusively in the extreme northern endemic areas of Chile. Metacyclic trypomastigotes obtainedin vitroor cell-derived trypomastigotes proved to be infective in γ-irradiated Balb/c mice for the study of the immune response and biological behavior. Of a total of 57T. cruzistocks obtained, 19 of them, representing all the different genotypes found in Chile, were tested on a murine experimental model and then fully studied. Female compared with male animals demonstrated greater resistance to Chagas disease with all theT. cruzistocks tested. The immune response was assessed by lytic antibodies that were studied by thein vitroantibody-dependent complement-mediated lytic assay with the use of bloodstream trypomastigotes as target cells. In one unique parasite genotype the elicited lytic antibodies reacted in a genotype-specific manner, in contrast with lytic antibodies generated by otherT. cruzigenotypes. Parasitemias were high, moderate, and low, with mortality ranges of 6–50%, 0–45%, and 0–10%, respectively. No association was found between specific infective genotypes and virulence or mortality. Independently of theT. cruzistrain studied, each population displayed a characteristic parasitemia curve and prepatent period. A considerable number of the parasite stocks proved to be mixed populations, according to molecular karyotype patterns obtained before and after differentiation and amplification of the parasites. This fact created difficulty in assessing the identity of the genotype really infective to mice.

Chronobiology of Plasmodium chabaudi chabaudi: analysis of hourly recorded total and differential parasitaemia during a schizogonic cycle.

Detailed total and differential parasitaemia curves of asexual Plasmodium chabaudi chabaudi erythrocytic stages were recorded and analysed. Female, inbred, CBA/Ca mice were infected with the virulent IP-PC1 strain after in vitro synchronization of the parasites. Thin blood smears were made on an hourly basis, and the total and differential parasitaemia of ring forms, trophozoites and schizonts were counted after Giemsa staining. These curves reveal information that remains hidden when less detailed curves are examined: the duration and periodicity of the schizogonic cycle, the existence of a plateau, indications of a schizont withdrawal from the peripheral blood, the timing of the rise of the parasitaemia at each schizogony, and the invasion rate of the merozoites. In the perspective of developing a rational and efficient strategy for chronotherapy of malaria, such information should be taken into account.

Trypanosoma cruzi:IL-10, TNF, IFN-γ, and IL-12 Regulate Innate and Acquired Immunity to Infection

Control of the acute phase ofTrypanosoma cruziinfections is critically dependent on cytokine-mediated macrophage activation to intracellular killing. We investigated the roles of IL-10, TNF, IFN-γ, and IL-12 in the control of parasitism by innate and specific immunity. Mice with disrupted IL-10 genes (IL-10 KO) infected with Y strainT. cruzihave lower parasite numbers in the blood and tissues and higher IFN-γ and nitric oxide (NO) production by spleen cells than wild type (WT) mice. Treatment of IL-10 KO and WT mice with recombinant IL-10 resulted in increased parasitemia. Mice with disrupted recombinase-activating genes (RAG/KO) that lack B and T cells provided a model for determining the importance of innate immunity to resistance. RAG/KO and WT mice had similar parasitemia levels until Day 13 of infection, suggestive of effective control of parasitism by the innate immune system during the early phase of infection; from then on parasitemia was higher in RAG/KO. Double RAG/IL-10 KO mice and RAG/KO mice had superimposable parasitemia curves, indicating that in the absence of T and B cells, endogenous IL-10 does not limit the efficacy of the innate immune system. Treatment of infected RAG/KO, IL-10/KO, and WT mice with anti-IFN-γ, anti-TNF, or anti-IL-12 neutralizing mAbs increased parasitemia levels showing the importance of endogenous production of these cytokines in the control of parasitism by innate and specific immune responses. Spleen cells from anti-IL12-treated WT mice had diminished production of IFN-γ and NO, suggesting that early IFN-γ synthesis is most dependent on IL-12 stimulation.

Selection of different strains of Plasmodium falciparum for testing blood-stage vaccines in Aotus nancymai monkeys.

Three strains of Plasmodium falciparum, Vietnam Oak Knoll (FVO), Uganda Palo Alto (Hawaii) (FUP-H) and Uganda Palo Alto (Cayenne) (FUP-C), were examined in 154 Aotus nancymai monkeys as suitable models for testing blood-stage vaccines. The Vietnam Oak Knoll strain had the greatest number of animals with maximum parasite counts > 200,000/microliters. Uniformity of the parasitemia curve increased from passage 4 to passage 6 with an accompanying decrease in the number of days required to reach maximum parasitemia or required treatment. The Uganda Palo Alto (Hawaii) strain was highly infectious, but many animals had extended prepatent periods and extended days to maximum parasitemia. The FUP-H strain would require a greater number of animals per test group to detect partial protection because of the greater number of low-density maximum parasite counts in control animals. The Uganda Palo Alto (Cayenne) strain was poorly adapted to intact A. nancymai. However, five of six splenectomized monkeys inoculated during passage 6 with 10(5) parasites had maximum parasite counts > 200,000/microliters. For the testing of vaccines against primary parasitemia in the A. nancymai model system, the FVO at passage 4 level would appear preferable to passage 6 parasites following a challenge with 10(5) parasites. A similar pattern could be obtained using FUP-H if the challenge was 10(6) parasites. To measure immune memory against recrudescence or rechallenge infection, FUP-C at an early passage in splenectomized A. nancymai would appear to be the appropriate model.

Trypanosoma cruzi and experimental Chagas' disease: characterization of a stock isolated from a patient with associated digestive and cardiac form.

A new Trypanosoma cruzi stock isolated from a patient in the chronic phase of Chagas' disease with the digestive and cardiac form of the disease was characterized by experimental infection in isogenic, susceptible, A/Sn strain mice. Parasitemia curves showed up to 1.7 x 10(6) parasites/ml and no mortality was observed up to 300 days post infection. Specific IgM was found in mice in the acute phase up to 40 days and also in the chronic phase. IgG antibodies were detected in the acute and chronic phase. Histopathology examination demonstrated myotropism to the digestive tract muscle layers and to the heart.

Isolation of Trypanosoma cruzi from blood by histopaque and continuous percoll gradient centrifugations.

Separation of the blood forms of trypanosomes from the blood of infected animals is difficult, especially in the case of Trypanosoma cruzi Y strain. Two procedures to isolate the Y strain blood forms of T. cruzi using polyvinyl pyrrolidone-coated silica (percoll) and histopaque are reported in this study. The recovery rates of parasites were 16 +/- 5 and 68 +/- 16%, respectively. The parasites isolated by these methods presented normal motility and morphology and were infective to albino mice with prepatent periods, parasitemia curves, and polymorphism patterns during the infection that were similar to those of control parasites. In addition, the preservation of surface antigens was confirmed by immunocytochemical studies.