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Am. J. Trop. Med. Hyg., 75(3), 2006, pp. 497-501
Copyright © 2006 by The American Society of Tropical Medicine and Hygiene

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Right arrow Chagas Disease

POLYMERASE CHAIN REACTION DETECTION AND SEROLOGIC FOLLOW-UP AFTER TREATMENT WITH BENZNIDAZOLE IN BOLIVIAN CHILDREN INFECTED WITH A NATURAL MIXTURE OF TRYPANOSOMA CRUZI I AND II

MARIA FLORES-CHAVEZ, MARIE-FRANCE BOSSENO, BRIGITTE BASTRENTA, JOSE-LOUIS ALCAZAR DALENZ, MIREILLE HONTEBEYRIE, SUSANA REVOLLO, AND SIMONE FRÉDÉRIQUE BRENIÈRE*
Facultad de Ciencias Farmacéuticas y Bioquímicas, Instituto Seladis, Universidad Mayor de San Andrés, La Paz, Bolivia; Unité de Recherche 008 Pathogénie des Trypanosomatidés, Institut de Recherche pour le Développement, Montpellier, France; Institut Pasteur, Paris, France


ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Thirty-five Bolivian children (5–10 years of age) seropositive for infection with T. cruzi underwent specific chemotherapy with benznidazole. Before treatment, 57.1% had a positive parasitologic diagnosis. Some patients presented an early conversion by polymerase chain reaction of blood samples, while others were still positive four and seven months after the end of the treatment, which indicated an absence of parasite clearance. Strain typing showed that most patients were infected by a mixture of clones I and II of T. cruzi. Serologic conversion in conventional tests and antibodies to shed acute-phase antigen were observed in two and four patients, respectively. For the other patients, the average rate of antibody decay was half the initial rate. The parasitologic and serologic data indicated that chemotherapy acts throughout the course of infection in a long-lasting process in which the decrease of specific antibody production is related to the reduction of the live parasite load.


INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Chagas disease, which affects 17 million people throughout Latin America, is caused by the protozoan Trypanosoma cruzi and is transmitted by hematophagous insects of the family Reduviidae.1 After development of an acute disease phase characterized by intense parasitism and generally transient symptoms such as fever and myocarditis lesions, an asymptomatic indeterminate phase develops with subpatent parasitemia and positive serologic findings. Although most infected patients remain in this phase for life, 20–30% will develop clinical manifestations involving degenerative cardiac or digestive tract lesions with inflammatory foci characteristic of T. cruzi infection.

Two main hypotheses have been formulated to explain the pathogenesis of Chagas disease: 1) T. cruzi induces an autoimmune response that is directed mainly against host tissues and is independent of the persistence of the parasite and/or 2) persistence of the parasite in particular tissues or organs will result in a chronic inflammatory process leading to tissue destruction.2–4 Both hypotheses are supported by cumulative data, but the finding supporting the second hypothesis is that treatment with specific anti-chagasic drugs is beneficial when applied to children as well as to patients with chronic disease with cardiomyopathy.5–7 However, the general assumption until now is that treatment is particularly successful in the early stages of the infection (acute and congenital), but less effective when applied later during the indeterminate chronic phase.1

Two drugs have been used for the treatment of Chagas disease: nifurtimox (a nitrofurane derivative) and benznidazole (a nitroimidazole), with various degrees of sensitivity depending on the parasite strain. Clonal populations of T. cruzi show considerable biologic and genetic variability, and comparative studies have suggested that differences in drug susceptibility are associated with particular genotypes.8–10 Moreover, secondary effects limit the use of these drugs. Thus, there is an urgent need for new and better compounds to increase the number of cured patients, particularly those in the chronic phase.1 However, it is difficult to accurately determine the effect of treatment in the absence of a joint approach that includes both physicians and biologists. The first criterion for cure should be parasitologic clearance. The second one should be seroconversion, the only parameter accessible by routine tests. Nevertheless, the persistence of positive serologic findings, which are generally observed after treatment of chronic cases, represents a major difficulty in accurate evaluation of therapeutic efficacy.

Recently, specific antibodies to T. cruzi were separated into two categories: 1) those associated with acute ongoing infection and 2) those persisting at high levels throughout the life of chagasic patients.11,12 Parasitologic clearance could be monitored by the polymerase chain reaction (PCR), which is the most sensitive parasitologic test, or by measures of antibody decay associated with live parasites. The latter should be clinically beneficial by reducing the probability of developing chronic disease, and could be a criterion for long-term follow-up studies.

We report the results of a field trial focused on evaluating the success of chemotherapy in Bolivian children infected by a natural mixture of two clones belonging to T. cruzi I and II lineages (clones 20 and 39, respectively). Treatment with benznidazole and follow-up were carried out by parasitologic (xenodiagnosis and PCR-hybridization studies) and serologic (conventional and antibodies to shed acute-phase antigen [SAPA]) tests.


MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Study population and benznidazole treatment. This study was conducted at the Buenas Nuevas School and the Harry Williams Hospital in the Huayrakhasa sector of the Cocha-bamba city in Bolivia during 1997 and 1998. The protocol of the study was reviewed and approved by the Institutional Ethical Committee of the Colegio Médico de Bolivia. The parents of the children were informed in detail about the protocol and a consent form was signed by each of them. Thirty-five school children (age range = 5–10 years) were recruited from 40 of 930 children examined who were positive by two independent conventional serologic tests (enzyme-linked immunosorbent assay [ELISA] and immunofluorescence test). Before treatment, any clinical manifestation of acute Chagas disease and other pathology was observed in the selected patients. Toxic effects were assessed before and during follow-up by hemogram, including red blood, hematocrit, hemoglobin (mean cell volume, mean corpuscular hemoglobin, and mean corpuscular hemoglobin concentration), neutrophil, monocyte, lymphocyte, eosinophil, basophil, and platelet counts, and erythrocyte sedimentation rate. Benznidazole was given at a dose of 8 mg per kg of body weight per day for 60 days. The drugs were administrated daily by health workers at the school and to each house on weekends. Prior to treatment, parasites were detected in blood by microscopically observation of four hematocrit tubes at their interphase (hematocrit test).13 Xenodiagnosis was carried out before the treatment and 12 months post-treatment using 2 boxes of 20 third-instar nymphs of Triatoma infestans each for 30 minutes. The feces of the bugs were examined individually two and three months later and collected for further analysis by PCR.

Patient follow-up after treatment. The outcome of treatment was evaluated immediately after treatment and at 6, 9, and 14 months after treatment by an ELISA, detection of specific antibodies against SAPA recombinant protein, and a PCR on blood samples.14,15 Identification of clones 20 and 39 was done by hybridization of PCR products with specific kinetoplast DNA probes as previously described.16

Screening of houses for vectors. Dwellings of the children were examined for triatomines before the study and all positive and adjacent dwellings were sprayed both inside and outside with a deltamethrin-active substance (K-Othrine; Hoechst Schering Agrevo S.A., Berlin, Germany) at a concentration of 1.5 grams/5 liters/100 m2. Dwellings were subsequently sprayed monthly to eliminate transmission during the study.


RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Parasitologic status before treatment. Different parasitologic tests were conducted for 35 seropositive children selected for treatment with benznidazole. The results are summarized in Table 1Go. Only three patients had positive hematocrit test results, while one was also positive by xenodiagnosis and a PCR on blood samples, and the two others were negative by xenodiagnosis but positive on a PCR for blood samples. Conventional xenodiagnosis identified disease in 7 of the 35 patients studied, while PCR and xenodiagnosis confirmed these cases and identified three additional patients. The PCR on blood samples was positive in 37.1% of the patients and appeared to be the most sensitive parasitologic test, although the agreement between xenodiagnosis and the PCR on blood samples was only 51.4% for parasite detection. Twenty-one (57.1%) patients had at least one positive parasitologic test result.


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TABLE 1
Results of parasitologic diagnoses in 35 seropositive Bolivian children before treatment*
 
Post-treatment parasitologic follow-up and detection of clones 20 and 39. All patients completed treatment although some follow-up data were missing, mainly for 12 months after the end of the treatment. Toxic side effects were not observed during treatment. Table 2Go shows the individual results for the parasitologic tests and strain detection before and after treatment. All 22 patients studied tested 12 months after the end of treatment were negative for by conventional xenodiagnosis, PCR and xenodiagnosis, and PCR. Four different patterns were observed. The first pattern (group 1) included 8 patients with positive parasitologic results before treatment and early conversion of PCR results from 4 to 12 months after treatment. The second pattern (group 2) included 12 patients with conversion of PCR results approximately one year post-treatment. The third pattern (group 3) included 15 patients who were parasitologically negative before treatment, of which but 6 were subsequently positive by the PCR. The fourth pattern (group 4) included nine patients who had consistently negative results.


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TABLE 2
Detection of Trypanosoma cruzi clones 20 and 39 by hybridization with kinetoplast DNA-specific probes in 35 chagasic children before and after benznidazole treatment*
 
The two major clones 20 and 39 were identified by hybridization of xenodiagnosis-PCR products and PCR products with specific kinetoplast DNA probes. Before treatment, parasite clones were identified in 20 patients. Similar frequencies of clones 20 (70%) and 39 (65%) were observed, including 45% of mixed infections (positive hybridization with both probes) (Table 2Go). Parasite clones were detected in 18 patients immediately after treatment and at 4, 7, and 12 months after the end of treatment, of which 12 had a positive PCR result before treatment and 6 others had a negative PCR result. Mixed infections were identified in 83.3% of the samples.

Antibodies to T. cruzi and IgG antibodies to SAPA post-treatment. The evolution of specific antibodies against T. cruzi and SAPA recombinant protein after treatment is shown in Table 3Go and Figure 1Go. Before treatment, all 35 patients had IgG antibodies to T. cruzi IgG, but two converted to negative results (patients HUA 243 and HUA 419). One patient showed early conversion 4 months after the end of treatment (samples at 7 and 12 months were not available). The second patient had antibody 7 months after the end of treatment but was negative at 12 months. For the other 33 patients, average levels of antibodies to T. cruzi decreased by 21% immediately after treatment and by 48.6% at 12 months after the end of treatment. Figure 1aGo shows that most (75%) of the patients showed a decrease in antibody level (> 50%) at 12 months. Antibodies to SAPA were detected in 31 patients (88.6%), while 4 showed no antibody response to SAPA before treatment and at 12 after the end of treatment (patients HUA 842, HUA 284, HUA 35, and HUA 489). Among the patients positive for antibodies to SAPA before treatment, 29% were negative after treatment (3 patients immediately after treatment, 5 patients 4 months later, 1 patient 7 months later, and 1 patients 12 months later), while 71% remained positive with decreasing average antibody levels from immediately after treatment to 12 months later, reaching 50% of the initial level at 12 months (Figure 1bGo).


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TABLE 3
Proportion of patients with reactive IgG antibodies against T. cruzi and Sapa recombinant protein before and after treatment*
 

Figure 1
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  		    FIGURE 1. Kinetics of rates of production of (A), antibodies to Trypanosoma cruzi and (B), IgG antibodies to shed acute-phase antigen (SAPA) detected by enzyme-linked immunosorbent assay before and after treatment with benznidazole. Percentages of patients with different levels of antibodies compared with the initial rate before treatment are shown at different times after treatment. 1 = at the end of the treatment (2 months); 2 = 4 months after the end of the treatment; 3 = 7 months after the end of the treatment; 4 = 12 months after the end of the treatment.

 
Comparison of levels of antibodies to T. cruzi and SAPA in patient groups with different parasitologic patterns showed that before treatment positive patients (groups 1 and 2) had no significantly different levels of either antibody type. Only patients in group 4 had a significantly lower level of antibodies either against T. cruzi or SAPA. In this group, although the parasitologic test results were negative at 12 months after the end of treatment, the serologic results were positive.


DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
Although it is not known whether Chagas disease has an autoimmune or parasitic disease pathology, available knowledge indicates specific treatment for this disease.17 Although the benefit of treatment is limited in patients with a long-lasting chronic infection, it is more promising for short-term chronic infection and indeterminate forms of the disease6 Nevertheless, one of the principal challenges concerning Chagas disease control is the absence of reliable criteria for the evaluation of chemotherapeutic efficacy. Two categories of tests concerning parasitologic and serologic evaluations have been proposed. Whatever the pathology, parasite clearance is difficult to ascertain because conventional parasitologic tests are relatively insensitive. Nevertheless, the PCR is currently the most sensitive technique for parasite detection and in the work presented in this report, it was used together with conventional xenodiagnosis before and after treatment.18–21

In this study, PCR-based parasite diagnosis was positive in only 37.1% of the seropositive children. This result differed from our previous report of a higher sensitivity of the PCR-based diagnosis (82%) in a rural population (Mizque village) of Bolivian children living in the same region,16 and indicates that patients living in a rural area may develop higher parasitemias. In both areas, clones 20 and 39 were the principal ones detected in the T. infestans vector, but the seroprevalence was of 4.6% in the urban population and 10-fold higher in the rural area (44%).22 In areas highly endemic for this disease, individuals are exposed continuously to the vector and multiple infections likely occur simultaneously, leading to increased infestation and parasitemia. Experimental infections in dogs showed that partial immunity was acquired after the first infection and that re-infections resulted in an increase of circulating parasites.23 A similar phenomenon could occur in humans in areas highly endemic for this disease. Although PCR results correlate with those obtained for parasitemia, they appear to depend on parasite genotypes, and the results presented here suggest that PCR results should not be the only parameter considered because multiple co-infections probably influence the course of parasitemia, including the persistence of blood parasites and their density.15,24,25

One year post-treatment, the PCR parasite detection test showed negative results for all previously positive patients, but four groups were observed. The first group was composed of eight patients with a positive PCR result before treatment and no detectable parasitemia at the end of the treatment and up to one year of follow-up. These patients were considered cured. The second group was composed of patients who still had parasites detected at the end of treatment and during subsequent follow-up, but after one year, the PCR and xenodiagnosis results were negative. Persistence of the parasite in this group may contribute to induction of a strong specific immune response that eliminates the parasites by one year post-treatment. The third group was composed of 15 patients who were PCR negative before treatment, of whom six converted to a PCR-positive result after treatment and reconverted to a PCR-negative result one year later (group 3). These patients may show an effect of benznidazole on activation of intracellular amastigotes, in which their differentiation is subsequently controlled by the immune system. The increase in detectable mixed infections after treatment may favor this hypothesis. The fourth group was composed of patients who did not have any positive parasitologic test results. This showed that the infection was naturally controlled in these patients. This population should be monitored for possible auto-reactive clinical symptoms caused by antibodies induced by parasite antigens, but continuously stimulated by cross-reactive self antigens.

A diagnosis of T. cruzi infection is generally made by serologic detection of specific IgG against parasite proteins. The persistence of specific antibodies in individuals infected with T. cruzi is a common component of the immune response, while positive serologic results that become negative is indicative of parasite clearance, signifying cure of the infection. Some cases of spontaneous human cure have been reported but these are generally considered to be exceptional.26 After successful treatment of cases with chronic disease, the decrease in antibodies detected by conventional serologic techniques is generally a very slow process, whereas antibodies that have a parasite-specific lytic activity gradually become negative after treatment.27 The low level of seroconversion after only one year post-treatment is consistent with previous data. Nevertheless, the decrease in antibodies to SAPA is more rapid and may provide a better criterion of cure.28–31

In the present study, most patients were parasite negative one year post-treatment, but retained positive serologic results for T. cruzi. The mechanisms underlying this persistence of specific antibodies are not understood. One hypothesis is that small numbers of parasites may be sequestered in body tissues and are able to sustain a persistent immune response.4 Alternatively, an autoimmune response may be elicited during the acute phase of infection that leads to cross-reactive autoantibodies,2 which may persist by permanent stimulation of self antigens. There are no particularly strong arguments favoring either hypothesis and they are not mutually exclusive. Proteomic studies and double-dimension electrophoresis might identify some cross-reactive antigens recognized by the sera of chronically infected individuals. Recent studies have shown that the presence of PCR-detectable parasite DNA in human samples was associated with pathology.32,33 However, no good cure criteria are yet available in spite of several attempts to define a relevant parasite molecule, such as the antigenic counterpart of the lytic antibodies.11

Received February 25, 2005. Accepted for publication October 31, 2005.

Acknowledgments: We thank Daniel Sanchez (Fundación Campomar, Buenos Aires, Argnetina) for providing SAPA recombinant protein, the personnel of the Harry Williams Hospital in Cocha-bamba for medical assistance during the treatment of the children, and Shirley Longacre (Laboratory of Vaccinologie Parasitaire, Institut Pasteur, Paris, France) for help with the English revision of the manuscript.

Financial support: This research was supported by the World Health Organization Special Program for Research and Training in Tropical Disease (ID 940856) and the Institut de Recherche pour le Développement, France.

* Address correspondence to Simone Frédérique Brenière, Unite de Recherche 008 Pathogénie des Trypanosomatidés, Institut de Recherche pour le Développement, 911 Avenue Agropolis, BP 5045, 34032 Montpellier Cedex 1, France. E-mail: breniere{at}mpl.ird.fr Back

Authors’ addresses: Maria Flores-Chavez, Jose-Louis Alcazar Dalenz, and Susana Revollo, Facultad de Ciencias Farmacéuticas y Bioquímicas, Instituto Seladis, Universidad Mayor de San Andrés, Saavedra No. 2224, La Paz, Bolivia. Marie-France Bosseno, Brigitte Bastrenta, and Simone Frédérique Brenière, Unité de Recherche 008 Pathogénie des Trypanosomatidés, Institut de Recherche pour le Développement, 911 Avenue Agropolis, BP 5045, 34032 Montpellier Cedex 1, France, Telephone: 33-4-67-41-62-98, Fax: 33-4-67-41-63-30, E-mail: breniere{at}mpl.ird.fr. Mireille Hontebeyrie, Institut Pasteur, 28, Rue du Dr. Roux, Paris 15, France, E-mail: mhj{at}pasteur.fr.


REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. WHO, 2002. Control of Chagas Disease. Second report of the WHO expert committee. Geneva: World Health Organization.
  2. Kierszenbaum F, 1999. Chagas’ disease and the autoimmunity hypothesis. Clin Microbiol Rev 12: 210–223.[Abstract/Free Full Text]
  3. Kalil J, Cunha-Neto E, 1996. Autoimmunity in Chagas disease cardiomyopathy: fulfilling the criteria at last? Parasitol Today 12: 396–398.[ISI][Medline]
  4. Tarleton RL, Zhang L, 1999. Chagas disease etiology: autoimmunity or parasite persistence? Parasitol Today 15: 94–99.[ISI][Medline]
  5. Andrade AL, Zicker F, de Oliveira RM, Almeida Silva S, Luquetti A, Travassos LR, Almeida IC, de Andrade SS, de Andrade JG, Martelli CM, 1996. Randomised trial of efficacy of benznidazole in treatment of early Trypanosoma cruzi infection. Lancet 348: 1407–1413.[ISI][Medline]
  6. Sosa Estani S, Segura EL, Ruiz AM, Velazquez E, Porcel BM, Yampotis C, 1998. Efficacy of chemotherapy with benznidazole in children in the indeterminate phase of Chagas’ disease. Am J Trop Med Hyg 59: 526–529.[Abstract]
  7. Viotti R, Vigliano C, Armenti H, Segura E, 1994. Treatment of chronic Chagas’ disease with benznidazole: clinical and serologic evolution of patients with long-term follow-up. Am Heart J 127: 151–162.[ISI][Medline]
  8. Carneiro M, Romanha AJ, Chiari E, 1991. Biological characterization of Trypanosoma cruzi strains from different zymodemes and schizodemes. Mem Inst Oswaldo Cruz 86: 387–393.[ISI][Medline]
  9. Andrade SG, Magalhaes JB, 1996. Biodemes and zymodemes of Trypanosoma cruzi strains: correlations with clinical data and experimental pathology. Rev Soc Bras Med Trop 30: 27–35.[Medline]
  10. Revollo S, Oury B, Laurent JP, Barnabe C, Quesney V, Carriere V, Noel S, Tibayrenc M, 1998. Trypanosoma cruzi: impact of clonal evolution of the parasite on its biological and medical properties. Exp Parasitol 89: 30–39.[ISI][Medline]
  11. Krautz GM, Kissinger JC, Krettli AU, 2000. The targets of the lytic antibody response against Trypanosoma cruzi. Parasitol Today 16: 31–34.[ISI][Medline]
  12. Altcheh J, Corral R, Biancardi MA, Freilij H, 2003. Anti-F2/3 antibodies as cure marker in children with congenital Trypanosoma cruzi infection. Medicina (B Aires) 63: 37–40.
  13. La Fuente C, Saucedo E, Urjel R, 1984. The use of microhaematocrit tubes for the rapid diagnosis of Chagas disease and malaria. Trans R Soc Trop Med Hyg 78: 278–279.[ISI][Medline]
  14. Brenière SF, Yaksic N, Telleria J, Bosseno MF, Noireau F, Wincker P, Sanchez D, 1997. Immune response to Trypanosoma cruzi shed acute phase antigen in children from an endemic area for Chagas’ disease in Bolivia. Mem Inst Oswaldo Cruz 92: 503–507.[ISI][Medline]
  15. Wincker P, Bosseno MF, Britto C, Yaksic N, Cardoso MA, Morel CM, Brenière SF, 1994. High correlation between Chagas’ disease serology and PCR-based detection of Trypanosoma cruzi kinetoplast DNA in Bolivian children living in an endemic area. FEMS Microbiol Lett 124: 419–423.[ISI][Medline]
  16. Brenière SF, Bosseno MF, Telleria J, Bastrenta B, Yacsik N, Noireau F, Alcazar JL, Barnabe C, Wincker P, Tibayrenc M, 1998. Different behavior of two Trypanosoma cruzi major clones: transmission and circulation in young Bolivian patients. Exp Parasitol 89: 285–295.[ISI][Medline]
  17. Urbina JA, 2001. Specific treatment of Chagas disease: current status and new developments. Curr Opin Infect Dis 14: 733–741.[ISI][Medline]
  18. Galvao LM, Chiari E, Macedo AM, Luquetti AO, Silva SA, Andrade AL, 2003. PCR assay for monitoring Trypanosoma cruzi parasitemia in childhood after specific chemotherapy. J Clin Microbiol 41: 5066–5070.[Abstract/Free Full Text]
  19. Britto C, Cardoso A, Silveira C, Macedo V, Fernandes O, 1999. Polymerase chain reaction (PCR) as a laboratory tool for the evaluation of the parasitological cure in Chagas disease after specific treatment. Medicina (B Aires) 59 (Suppl 2): 176–178.
  20. Zulantay I, Honores P, Solari A, Apt W, Ortiz S, Osuna A, Rojas A, Lopez B, Sanchez G, 2004. Use of polymerase chain reaction (PCR) and hybridization assays to detect Trypanosoma cruzi in chronic chagasic patients treated with itraconazole or allopurinol. Diagn Microbiol Infect Dis 48: 253–257.[ISI][Medline]
  21. Meira WS, Galvao LM, Gontijo ED, Machado-Coelho GL, Norris KA, Chiari E, 2004. Use of the Trypanosoma cruzi recombinant complement regulatory protein to evaluate therapeutic efficacy following treatment of chronic chagasic patients. J Clin Microbiol 42: 707–712.[Abstract/Free Full Text]
  22. Brenière SF, Bosseno MF, Noireau F, Yacsik N, Liegard P, Aznar C, Hontebeyrie M, 2002. Integrate study of a Bolivian population infected by Trypanosoma cruzi, the agent of Chagas disease. Mem Inst Oswaldo Cruz 97: 289–295.[ISI][Medline]
  23. Machado EM, Fernandes AJ, Murta SM, Vitor RW, Camilo DJ Jr, Pinheiro SW, Lopes ER, Adad SJ, Romanha AJ, Pinto Dias JC, 2001. A study of experimental reinfection by Trypanosoma cruzi in dogs. Am J Trop Med Hyg 65: 958–965.[Abstract]
  24. Veloso VM, Carneiro M, Toledo MJ, Lana M, Chiari E, Tafuri WL, Bahia MT, 2001. Variation in susceptibility to benznidazole in isolates derived from Trypanosoma cruzi parental strains. Mem Inst Oswaldo Cruz 96: 105–111.[ISI][Medline]
  25. Zingales B, Souto RP, Mangia RH, Lisboa CV, Campbell DA, Coura JR, Jansen A, Fernandes O, 1998. Molecular epidemiology of American trypanosomiasis in Brazil based on dimorphisms of rRNA and mini-exon gene sequences. Int J Parasitol 28: 105–112.[ISI][Medline]
  26. Francolino SS, Antunes AF, Talice R, Rosa R, Selanikio J, de Rezende JM, Romanha AJ, Dias JC, 2003. New evidence of spontaneous cure in human Chagas’ disease. Rev Soc Bras Med Trop 36: 103–107.[Medline]
  27. Galvao LM, Nunes RM, Cancado JR, Brener Z, Krettli AU, 1993. Lytic antibody titre as a means of assessing cure after treatment of Chagas disease: a 10-year follow-up study. Trans R Soc Trop Med Hyg 87: 220–223.[ISI][Medline]
  28. Martins-Filho OA, Eloi-Santos SM, Carvalho AT, Oliveira RC, Rassi A, Luquetti AO, Rassi GG, Brener Z, 2002. Double-blind study to evaluate flow cytometry analysis of anti-live trypomastigote antibodies for monitoring treatment efficacy in cases of human Chagas’ disease. Clin Diagn Lab Immunol 9: 1107–1113.
  29. Pereira-Chioccola VL, Fragata-Filho AA, Levy AM, Rodrigues MM, Schenkman S, 2003. Enzyme-linked immunoassay using recombinant trans-sialidase of Trypanosoma cruzi can be employed for monitoring of patients with Chagas’ disease after drug treatment. Clin Diagn Lab Immunol 10: 826–830.
  30. Norris KA, Galvao LM, Schrimpf JE, Cancado JR, Krettli AU, 1994. Humoral immune response to the Trypanosoma cruzi complement regulatory protein as an indicator of parasitologic clearance in human Chagas’ disease. Infect Immun 62: 4072–4074.[Abstract/Free Full Text]
  31. Krautz GM, Galvao LM, Cancado JR, Guevara-Espinoza A, Ouaissi A, Krettli AU, 1995. Use of a 24-kilodalton Trypanosoma cruzi recombinant protein to monitor cure of human Chagas’ disease. J Clin Microbiol 33: 2086–2090.[Abstract]
  32. Schijman AG, Vigliano CA, Viotti RJ, Burgos JM, Brandariz S, Lococo BE, Leze MI, Armenti HA, Levin MJ, 2004. Trypanosoma cruzi DNA in cardiac lesions of Argentinean patients with end-stage chronic chagas heart disease. Am J Trop Med Hyg 70: 210–220.[Abstract/Free Full Text]
  33. Elias FE, Vigliano CA, Laguens RP, Levin MJ, Berek C, 2003. Analysis of the presence of Trypanosoma cruzi in the heart tissue of three patients with chronic Chagas’ heart disease. Am J Trop Med Hyg 68: 242–247.[Abstract/Free Full Text]




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