Molecular diagnosis and typing of Trypanosoma cruzi populations and lineages in cerebral Chagas disease in a patient with AIDS

Trypanosoma cruzi DNA was amplified from an intracranial biopsy and peripheral blood of an HIV patient with encephalitis; this episode was indicative of AIDS and congenital Chagas disease. The analysis of a microsatellite locus revealed a multiclonal parasite population at the brain lesion with a more complex minicircle signature than that profiled in blood using restriction fragment length polymorphism (RFLP)-PCR and low stringency single primer (LSSP) PCR. Interestingly, different sublineages of T. cruzi II were detected in blood and brain by means of spliced-leader and 24s ribosomal-DNA amplifications. Quantitative-competitive PCR monitored the decrease of parasitic load during treatment and secondary prophylaxis with benznidazole. The synergy between parasiticidal plus antiretroviral treatments probably allowed the patient a longer survival than usually achieved in similar episodes. This is the first case report demonstrating a differential distribution of natural parasite populations and sublineages in Chagas disease reactivation, showing the proliferation of cerebral variants not detectable in peripheral blood.Fil: Burgos, Juan Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Bergher, Sandra B.. Gobierno de la Ciudad de Buenos Aires. Hospital "Ignacio Pirovano"; ArgentinaFil: Freitas, Jorge M.. Universidade Federal de Minas Gerais; BrasilFil: Bisio, Margarita María Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Altcheh, Jaime Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez"; ArgentinaFil: Teijeiro, Ricardo. Gobierno de la Ciudad de Buenos Aires. Hospital "Ignacio Pirovano"; ArgentinaFil: Begher, Sandra B.. Gobierno de la Ciudad de Buenos Aires. Hospital "Ignacio Pirovano"; ArgentinaFil: Freilij, Hector León. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez"; ArgentinaFil: Deccarlini, Florencia. Gobierno de la Ciudad de Buenos Aires. Hospital "Ignacio Pirovano"; ArgentinaFil: Levalle, Jorge. Gobierno de la Ciudad de Buenos Aires. Hospital "Ignacio Pirovano"; ArgentinaFil: Lopez Alcoba, Horacio. Gobierno de la Ciudad de Buenos Aires. Hospital "Ignacio Pirovano"; ArgentinaFil: Burgos, Juan Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Fisiología, Biología Molecular y Celular; ArgentinaFil: Levin, Mariano Jorge. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Duffy, Tomás. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Macedo, Andrea M.. Universidade Federal de Minas Gerais; BrasilFil: Schijman, Alejandro Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentin

Randomized, direct comparison study of Saccharomyces boulardii CNCM I-745 versus multi-strained Bacillus clausii probiotics for the treatment of pediatric acute gastroenteritis

Background: The choice of an appropriate probiotic for pediatric acute gastroenteritis (PAGE) can be confusing. Our aim was to compare the efficacy and safety of 2 probiotics (Saccharomyces boulardii CNCM I-745 vs a 4-strain mixture of Bacillus clausii O/C, SIN, N/R, T) for the treatment of PAGE. Methods: A 2-arm parallel, randomized trial recruited children (6 months to 5 years old) with mild-moderate acute diarrhea, from 8 centers in Argentina. A total of 317 children were enrolled and blindly randomized to 5 days of either S boulardii CNCM I-745 (n = 159) or a 4-strain mixture of B clausii (n = 158), then followed for 7 days post-probiotic treatment. A stool sample was collected at inclusion for pathogen identification. The primary outcome was duration of diarrhea defined as the time from enrollment to the last loose stool followed by the first 24-hour period with stool consistency improvement. Secondary outcomes included frequency of loose stools/day, severity of diarrhea, number reporting no diarrhea at Day 6, time-to-first formed stool, recurrence of diarrhea by study end (Day 12) and safety outcomes. Results: Three hundred twelve (98%) children completed the study. S boulardii CNCM I-745 showed a significant reduction (P =.04) in the mean duration of diarrhea (64.6 hours, 95% confidence interval [CI] 56.5-72.8) compared to those given B clausii (78.0 hours, 95% CI 69.9-86.1). Both probiotics showed improvement in secondary outcomes and were well-tolerated. Conclusion: In this study, S boulardii CNCM I-745 demonstrated better efficacy than B clausii mix for reducing the duration of pediatric acute diarrhea.Fil: Altcheh, Jaime Marcelo. Gobierno de la Ciudad de Buenos Aires. Instituto Multidisciplinario de Investigaciones en Patologías Pediátricas. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto Multidisciplinario de Investigaciones en Patologías Pediátricas; ArgentinaFil: Carosella, Mabel V.. No especifíca;Fil: Ceballos, Ana. Instituto Médico Río Cuarto; ArgentinaFil: D'Andrea, Ulises. Instituto Médico Río Cuarto; ArgentinaFil: Jofre, Sandra M.. No especifíca;Fil: Marotta, Carolina. No especifíca;Fil: Mugeri, Domingo. No especifíca;Fil: Sabbaj, Liliana. No especifíca;Fil: Soto, Adriana. No especifíca;Fil: Josse, Constant. No especifíca;Fil: Montestruc, Francois. No especifíca;Fil: McFarland, Lynne V.. No especifíca

Benznidazole Therapy Modulates Interferon-γ and M2 Muscarinic Receptor Autoantibody Responses in Trypanosoma cruzi-Infected Children

OBJECTIVE: The presence of autoantibodies with adrenergic and cholinergic activity, capable of triggering neurotransmitter receptor-mediated effects, has been associated with pathogenesis in T. cruzi-infected hosts. The goal of this study was to investigate the production of anti-M2 muscarinic receptor autoantibodies (Anti-M2R AAbs) as well as the IFN-γ profile in children at the early stage of Chagas disease, and to examine whether trypanocidal chemotherapy with benzonidazole (BZ) could modify both response patterns. METHODS: This study comprised 30 T. cruzi-infected children (mean age: 13.8 years) and 19 uninfected controls (mean age: 12.7 years). Infected patients were treated with BZ and followed-up. Blood samples collected at diagnosis-T0, end of treatment-T1, and six months later-T2 were analysed by ELISA for detection of Anti-M2R AAbs and circulating levels of IFN-γ. RESULTS: At T0, anti-M2R AAbs were demonstrated in 56.7% of T. cruzi-infected patients, whereas uninfected controls were 100% negative. The average age of Anti-M2R AAbs(+) patients was higher than that from negative population. Infected children also displayed significantly stronger serum IFN-γ responses than controls. Upon BZ treatment, a significant linear decreasing trend in Anti-M2R AAb reactivity was recorded throughout the follow-up, with 29.7-88.1% decrease at T2. IFN-γ circulating levels also declined by T2. CONCLUSION: Anti-M2R AAbs and IFN-γ raise early during chagasic infection in children and are downmodulated by BZ therapy. These findings reinforce the usefulness of early BZ treatment not only to eliminate the parasite but also to reduce potentially pathogenic immune responses

Accurate Real-Time PCR Strategy for Monitoring Bloodstream Parasitic Loads in Chagas Disease Patients

Infection with the parasite Trypanosoma cruzi (T. cruzi), causing American trypanosomiasis or Chagas disease, remains a major public health concern in 21 endemic countries of America, with an estimated prevalence of 8 million infected people. Chagas disease shows a variable clinical course, ranging from asymptomatic to chronic stages with low parasitaemias, whose severest form is heart disease. Diagnosis at the asymptomatic and chronic stages is based on serological detection of anti-T. cruzi antibodies, because conventional parasitological methods lack sensitivity. Current chemotherapies are more effective in recent infections than in the chronic adult population. The criterion of cure relies on serological conversion to negative, which may occur only years after treatment, requiring long-term follow-up. In this context, we aimed to develop a real-time PCR assay targeted to repetitive sequences of T. cruzi for sensitive quantitation of parasitic load in peripheral blood of infected patients. It was applied to monitor treatment response of infected children, allowing rapid evaluation of drug efficacy as well as detection of treatment failure. It was also used for early diagnosis of chagasic reactivation in end-stage heart disease patients who received immunosuppressive drugs after cardiac transplantation. This laboratory strategy may constitute a novel parasitological tool for prompt and sensitive evaluation of anti-parasitic treatment of Chagas disease

Kinetic analysis of the humoral immune response against 3 Toxoplasma gondii-recombinant proteins in infants with suspected congenital toxoplasmosis

Fil: Altcheh, Jaime. Hospital de Niños Dr. Ricardo Gutiérrez. Parasitología; Argentina.Fil: Diaz, Natalia S. Hospital de Niños Dr. Ricardo Gutiérrez. Parasitología; Argentina.Fil: Pepe, Carolina M. Hospital de Niños Dr. Ricardo Gutiérrez. Parasitología; Argentina.Fil: Martin, Valentina. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Nigro, Monica. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Freilij, Héctor. Hospital de Niños Dr. Ricardo Gutiérrez. Parasitología; Argentina.Fil: Angel, Sergio O. CONICET-UNSAM. Laboratorio de Parasitología Molecular, Chascomús; Argentina.The reactivity values of Toxoplasma gondii ROP2, GRA4, and GRA7 recombinant antigens (rAgs) were analyzed by immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA) in 23 congenitally infected (I) and 36 noninfected (NI) infants. The reactivity values observed from the serum samples of I versus NI infants for each recombinant protein were 91% versus 67% for rROP2 (P = .05), 86% versus 29% for rGRA4 (P < .001), and 56% versus 11% for rGRA7 (P = .003). The follow-up showed that serum samples from NI infants became negative for specific IgG at 5.8 months (95% confidence interval [CI], 4.9-6.7) using a commercial assay; meanwhile, by specific recombinant protein ELISA, the samples became negative at 3.7 months with rROP2 (95% CI, 2.8-4.6), at 1.3 months with rGRA4 (95% CI, 0.8-1.8), and at 0.9 months with rGRA7 (95% CI, 0.5-1.3). Kinetic analysis also showed that serum samples from group I presented different IgG-profiles among rAgs. The rROP2 IgG profile was similar to that of the commercial assay, whereas rGRA4 and rGRA7 profiles showed a gradual decrease along the period of the study. The potential of the utility of rAgs to develop a diagnostic system that discriminates congenitally I infants from NI is discussed

An α-Gal antigenic surrogate as a biomarker of treatment evaluation in Trypanosoma cruzi-infected children. A retrospective cohort study.

BackgroundProper evaluation of therapeutic responses in Chagas disease is hampered by the prolonged persistence of antibodies to Trypanosoma cruzi measured by conventional serological tests and by the lack of sensitivity of parasitological tests. Previous studies indicated that tGPI-mucins, an α-Gal (α-d-Galp(1→3)-β-d-Galp(1→4)-d-GlcNAc)-rich fraction obtained from T. cruzi trypomastigotes surface coat, elicit a strong and protective antibody response in infected individuals, which disappears soon after successful treatment. The cost and technical difficulties associated with tGPI-mucins preparation, however, preclude its routine implementation in clinical settings.Methods/principle findingsWe herein developed a neoglycoprotein consisting of a BSA scaffold decorated with several units of a synthetic α-Gal antigenic surrogate (α-d-Galp(1→3)-β-d-Galp(1→4)-β-d-Glcp). Serological responses to this reagent, termed NGP-Tri, were monitored by means of an in-house enzyme-linked immunosorbent assay (α-Gal-ELISA) in a cohort of 82 T. cruzi-infected and Benznidazole- or Nifurtimox-treated children (3 days to 16 years-old). This cohort was split into 3 groups based on the age of patients at the time of treatment initiation: Group 1 comprised 24 babies (3 days to 5 months-old; median = 26 days-old), Group 2 comprised 31 children (7 months to 3 years-old; median = 1.0-year-old) and Group 3 comprised 26 patients (3 to 16 years-old; median = 8.4 years-old). A second, control cohort (Group 4) included 39 non-infected infants (3 days to 5 months-old; median = 31 days-old) born to T. cruzi-infected mothers. Despite its suboptimal seroprevalence (58.4%), α-Gal-ELISA yielded shorter median time values of negativization (23 months [IC 95% 7 to 36 months] vs 60 months [IC 95% 15 to 83 months]; p = 0.0016) and higher rate of patient negative seroconversion (89.2% vs 43.2%, p Conclusions/significanceThe tools evaluated here provide the cornerstone for the development of an efficacious, reliable, and straightforward post-therapeutic marker for pediatric Chagas disease

Benznidazole Treatment of Chagasic Encephalitis in Pregnant Woman with AIDS

We report a case of chagasic meningoencephalitis reactivation in a pregnant woman co-infected with Trypanosoma cruzi and HIV that was successfully managed with benznidazole and highly active antiretroviral therapy. Early diagnosis enabled rapid specific treatment that improved the health of the patient and her baby

Congenital syphilis in Argentina: Experience in a pediatric hospital.

In spite of being preventable, Congenital syphilis (CS) is still an important, and growing health problem worldwide. Fetal infection can be particularly aggressive, but newborns can be asymptomatic at birth and, if left untreated, develop systemic compromise afterwards with poor prognosis. We analyzed 61 CS diagnosis cases between 1987-2019 presenting at the Buenos Aires Children' Hospital. The distribution of cases showed a bimodal curve, with a peak in 1992-1993 and in 2014-2017. Median age at diagnosis was 2 months (IQ 1-6 months). The main clinical findings were: bone alterations (59%); hepatosplenomegaly (54.1%); anemia (62.8%); skin lesions (42.6%) and renal compromise (33.3%). Cerebrospinal fluid (CSF) was abnormal in 5 patients, normal in 45 and was not available for 11 patients. Remarkably, spinal lumbar puncture did not modify therapeutic decisions in any case. Between mothers, only 46% have been tested for syphilis during pregnancy and 60.5% patients had non-treponemal titers equal to or less than fourfold the maternal titer. Intravenous penicillin G was prescribed for all except one patient, who received ceftriaxone with good therapeutic response. During follow-up, 1.6% infants died, 6.5% had persistent kidney disorders and 1.6% showed bone sequelae damage. RPR titers decreased after treatment, reaching negative seroconversion in 43% subjects at a median of 26.4 months. Low adherence to follow up was observed due to inherent vulnerable and low-income population characteristics in our cohort. Our results highlight a rising tendency in cases referred for CS in our population with high morbidity related to delayed diagnosis. A good therapeutic response was observed. CS requires a greater effort from the health system to adequately screen for this disease during pregnancy, and to detect cases earlier, to provide an adequate diagnosis and treatment

Efficacy of voriconazole in a murine model of acute Trypanosoma cruzi infection

OBJECTIVES: Antifungal triazole derivatives have been studied as possible alternatives for the treatment of Chagas' disease. Voriconazole has demonstrated in vitro activity against Trypanosoma cruzi, but its efficacy in vivo has not yet been tested. We aimed to determine the effect of voriconazole in a murine model of acute T. cruzi infection. METHODS: Treatment efficacy was evaluated by comparing parasitaemia, mortality and organ involvement (by histological examination) of infected mice. RESULTS: Treatment with voriconazole significantly lowered parasitaemia and mortality compared with controls, reduced the percentage of mice with amastigote nests in heart and skeletal muscle and moderately decreased myocardial inflammation. CONCLUSIONS: Our findings support the potential of voriconazole for the treatment of acute Chagas' disease and motivate future animal studies using varying doses and treatment schemes. Further evaluation of voriconazole for clinical use in human Chagas' patients is warranted.Fil: Gulin, Julián Ernesto Nicolás. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Servicio de Parasitología y Chagas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Eagleson, M. A.. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Servicio de Parasitología y Chagas; Argentina. Harvard University; Estados UnidosFil: Postan, Miriam. Dirección Nacional de Instituto de Investigación. Administración Nacional de Laboratorio e Instituto de Salud. Instituto Nacional de Parasitología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Cutrullis, Romina Andrea. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Servicio de Parasitología y Chagas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Freilij, Hector León. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Servicio de Parasitología y Chagas; ArgentinaFil: García Bournissen, Facundo. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Servicio de Parasitología y Chagas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Petray, Patricia Beatriz. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Servicio de Parasitología y Chagas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Altcheh, Jaime Marcelo. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Servicio de Parasitología y Chagas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin