Papers by Alejandro Schijman
PLOS Neglected Tropical Diseases
Background Chagas disease or American trypanosomiasis, a neglected tropical disease, is a persist... more Background Chagas disease or American trypanosomiasis, a neglected tropical disease, is a persistent Public Health problem in Latin America and other, non-endemic, countries. Point-of-care (POC) sensitive methods are still needed to improve and extend early diagnosis in acute infections such as congenital Chagas disease. The objective of this study was to analytically evaluate in the lab the performance of a qualitative POC molecular test (Loop-mediated isothermal amplification (LAMP), Eiken, Japan) for rapid diagnosis of congenital Chagas disease employing FTA cards or Whatman 903 filter paper as solid supports for small-scale volumes of human blood. Methodology/principal findings We used human blood samples artificially infected with cultured T. cruzi strains to assess the analytical performance of the test in comparison with liquid blood anticoagulated with heparin. The DNA extraction process was evaluated using the ultrarapid purification system PURE manufactured by Eiken Chemic...
Memorias Do Instituto Oswaldo Cruz, Nov 1, 1997
By using improved pulsed field gel electrophoresis conditions, the molecular karyotype of the ref... more By using improved pulsed field gel electrophoresis conditions, the molecular karyotype of the reference clone CL Brener selected for Trypanosoma cruzi genome project was established. A total of 20 uniform chromosomal bands ranging in size from 0.45 to 3.5 Megabase pairs (Mbp) were resolved in a single run. The weighted sum of the chromosomal bands was approximately 87 Mbp. Chromoblots were hybridized with 39 different homologous probes, 13 of which identified single chromosomes. Several markers showed linkage and four different linkage groups were identified, each comprising two markers. Densitometric analysis suggests that most of the chromosomal bands contain two or more chromosomes representing either homologous chromosomes and/or heterologous chromosomes with similar sizes.
The Journal of Molecular Diagnostics, Apr 1, 2021
Vertical transmission of T. cruzi is the cause of congenital Chagas disease, a re-emerging infect... more Vertical transmission of T. cruzi is the cause of congenital Chagas disease, a re-emerging infectious disease that affects endemic and non-endemic regions alike. An early diagnosis is crucial because its prompt treatment achieves a high cure rate, precluding evolution to symptomatic chronic Chagas disease. However, early diagnosis involves low sensitive parasitological assays, making necessary serological confirmation after nine months of life. Aiming at implementing early diagnostic strategies suitable for minimally equipped laboratories, a T. cruzi-LAMP prototype was coupled to an automated DNA extraction device re-purposed from a 3D printer (PrintrLab). The whole process takes less than three hours to yield a result with an analytical sensitivity of 0.1 - 2 parasite equivalents per ml depending on the T. cruzi strain. Twenty five blood samples from neonates born to seropositive mothers were tested blindly. In comparison to qPCR, the PrintrLab - LAMP duo strategy showed high agreement, while both molecular-based methodologies yielded an optimal sensitivity and specificity with respect to congenital Chagas disease microscopy-based diagnosis. The PrintrLab-LAMP detected all ten congenitally transmitted T. cruzi infections, showing promise for point-of-care early diagnosis of congenital Chagas disease.
Nucleic Acids Research, 1992
International Journal of Infectious Diseases, Aug 1, 2018
Background: The hyperinfestation by Strongyloides stercolaris is a general syndrome with high mor... more Background: The hyperinfestation by Strongyloides stercolaris is a general syndrome with high morbidity and mortality, which usually occurs in patients with immunological compromise or in corticosteroid treatments. In critical patients with paralytic intestinal ileus, the decrease in the bioavailability of oral ivermectin leads to therapeutic failure. The use of Ivermectin parenteral veterinary use is an extreme resource in patients with paralytic ileus secondary to hyperinflation by Strongyloides stercoralis. (SHSS) Methods & Materials: Retrospective and observational study. Clinical records of patients diagnosed with hyperinfestation by Strongyloides stercolaris, from January 2012 to August 2017, were evaluated in a sanatorium in the City of Buenos Aires. Inclusion criteria: positive culture (fecal matter, sputum, BAL) for Strongyloides stercoralis and diagnosis of intestinal ileus. The following variables were analyzed: sex, median age, nationality, residence, immunological commitment, corticotherapy, clinical and laboratory manifestations, days of hospitalization, stay in the ICU, treatment (enteral and parenteral), clinical resolution and mortality. Results: Of the 5 patients with inclusion criteria, the following were determined: Median age of 49.2 years; 2 foreigners (Peru and Bolivia); 4 residents of the Province of Buenos Aires and one of CABA. 2 with immunological compromise (HIV, DBT and kidney transplant); all 5 received corticotherapy. Clinical manifestations: all patients presented paralytic intestinal ileus with respiratory compromise, one presented rash and 4 had eosinophilia. The treatment started with oral Ivermectin but in the face of failure all received subcutaneous ivermectin as a last option. The 5 patients had a stay in the ICU, 4 with satisfactory clinical resolution and one death. Conclusion: Strongyloid hyperinflation syndrome is always a serious condition that can be manifested by respiratory distress, sepsis, meningitis or intestinal ileus. In this last presentation, the impossibility of digestive absorption of drugs makes it the only therapeutic option to use parenteral Ivermectin in the face of failure of the oral or rectal route. Our patients were immunosuppressed or had received corticosteroids, but they did not come from endemic areas. We emphasize the importance of the diagnosis prior to immunosuppression and the performance of the culture as a screening.
Frontiers in Parasitology
Chagas disease (CD) caused by Trypanosoma cruzi remains a Neglected Tropical Disease with limited... more Chagas disease (CD) caused by Trypanosoma cruzi remains a Neglected Tropical Disease with limited access to diagnosis and treatment, particularly for chronically infected patients. Clinical trials are underway to improve treatment using new drugs or different regimens, and Real-Time PCR is used to assess the parasitological response as a surrogate biomarker. However, PCR-based strategies have limitations due to the complex nature of T. cruzi infection. The parasite exhibits asynchronous replication, different strains and clones, and diverse tissue tropism, making it challenging to determine optimal timeline points for monitoring treatment response. This mini-review explores factors that affect PCR-based monitoring and summarizes the endpoints used in clinical trials for detecting treatment failure. Serial sampling and cumulative PCR results may improve sensitivity in detecting parasitemia and treatment failure in these trials.
Clinical and Vaccine Immunology, Mar 1, 2004
Hepatitis C virus genotyping was assessed for 257 chronic hepatitis C patients with viral loads a... more Hepatitis C virus genotyping was assessed for 257 chronic hepatitis C patients with viral loads above 1,000 IU/ml. Twelve patients were coinfected with more than one genotype. Their median viral loads did not differ significantly from those observed for monoinfected patients, which in turn did not vary significantly among different genotypes.
The Journal of Infectious Diseases
Chagas disease in solid organ transplant recipients may present as a primary infection (PI). Earl... more Chagas disease in solid organ transplant recipients may present as a primary infection (PI). Early detection is crucial for timely treatment. This is the largest observational multicentre study evaluating qPCR for early diagnosis and treatment monitoring of PI in seronegative recipients of organs from seropositive donors. Of 34 patients admitted at 5 health centers, PI was detected by qPCR in 8 (23.5%) within a posttransplant period of 40 days (interquartile range [IQR], 31–50 days). No PI was detected by the Strout test or clinical symptoms/signs. All patients had favorable treatment outcome with negative qPCR 31 days (IQR, 18–35 days) after treatment, with no posttreatment relapse episodes.
Clinical Infectious Diseases
Background Chagas disease (CD) has significant global health impact, but safe, effective treatmen... more Background Chagas disease (CD) has significant global health impact, but safe, effective treatments remain elusive. The nitroimidazole fexinidazole is a potential treatment. Methods This double-blind, randomized, placebo-controlled, dose-finding, proof-of-concept study was conducted in Bolivia. Adults with serologically confirmed chronic indeterminate CD and positive PCR were randomly assigned to 1 of 6 fexinidazole regimens (1200 or 1800 mg/day for 2, 4, or 8 weeks) or placebo. Target recruitment was 20 patients/arm. The primary endpoint was sustained parasitological clearance by serial negative qPCR from end of treatment (EOT) until 6 months follow-up in the intention-to-treat (ITT) population. Follow-up was extended to 12 months. Results Enrollment was interrupted after 4/47 patients presented with transient asymptomatic grade 3 and 4 neutropenia. Treatment of ongoing patients was stopped in all patients administered >2 weeks. A total of 40 patients received treatment with fex...
Acta Tropica, 2021
Chagas disease, caused by the protozoan Trypanosoma cruzi, affects more than 6 million people wor... more Chagas disease, caused by the protozoan Trypanosoma cruzi, affects more than 6 million people worldwide. Following a mostly asymptomatic acute phase, the disease progresses to a long-lasting chronic phase throughout which life-threatening disorders to the heart and/or gastrointestinal tract will manifest in about 30% of those chronically infected. During the chronic phase, the parasitemia is low and intermittent, while a high level of anti-T. cruzi antibodies persist for years. These two features hamper post-chemotherapeutic follow-up of patients with the tools available. The lack of biomarkers for timely assessment of therapeutic response discourages a greater use of the two available anti-parasitic drugs, and complicates the evaluation of new drugs in clinical trials. Herein, we investigated in a blinded case-control study the serological reactivity over time of a group of parasite-derived antigens to potentially address follow up of T. cruzi chronically infected subjects after treatment. We tested PFR2, KMP11, HSP70, 3973, F29 and the InfYnity multiplexed antigenic array, by means of serological assays on a multi-national retrospective collection of samples. Some of the antigens exhibited promising results, underscoring the need for further studies to determine their potential role as treatment response biomarkers.
Infection, Genetics and Evolution, 2020
Kinetoplastids are a group of flagellated protozoa that infect a vast repertoire of mammals and i... more Kinetoplastids are a group of flagellated protozoa that infect a vast repertoire of mammals and insect vectors. From a zoonotic point of view, domestic animals are critical reservoirs for transmission of Kinetoplastidean parasites. Due to their proximity to humans, they assume substantial epidemiological importance in the context of these zoonoses and consequently in public health. Their reliable identification is relevant to understand their eco-epidemiological involvement in transmission cycles. This work aimed to develop an algorithm based on sequential Real-Time PCR (qPCR) assays targeted to different loci (24S alpha rDNA, ITS1 and Hsp70) allowing distinction among Trypanosoma cruzi, Trypanosoma rangeli, Trypanosoma evansi and Leishmania species in biological samples collected from mammalian reservoirs and triatomine vectors. The algorithm includes a first qPCR test targeted to endogenous genes conserved within mammals and within triatomine vectors as internal controls of DNA sample integrity and/or qPCR inhibition. This algorithm was evaluated in biological samples from domestic cattle (N = 14), dogs (N = 19) and triatomines (N = 19). Analytical sensitivity of 24S alpha rDNA for detection of T. rangeli was 10 fg of DNA, with a linear range between 10 fg and 10 ng. For T. cruzi it varied depending on the Discrete typing unit. The ITS1 qPCR showed an analytical sensitivity of 100 pg/reaction and 100 fg/reaction of Leishmania spp. and T. evansi DNAs. In mammal field samples, four T. cruzi 24S alpha rDNA sequences and fourteen ITS1 amplicons specific for T. evansi were detected. qPCR-HRM analysis directed to the Hsp70 gene diagnosed two dogs with Leishmania infantum infection. Among 19 triatomine field samples, T. cruzi was detected in five; T. rangeli in eight and one specimen showed a mixed infection. This diagnostic algorithm can provide more accurate records of kinetoplastidean infection burden in vectors and reservoirs, relevant to update current eco-epidemiological maps in co-endemic regions.
PLOS Neglected Tropical Diseases, 2019
PLOS Neglected Tropical Diseases, 2019
Several studies have proposed different genetic markers of susceptibility to develop chronic Chag... more Several studies have proposed different genetic markers of susceptibility to develop chronic Chagas cardiomyopathy (CCC). Many genes may be involved, each one making a small contribution. For this reason, an appropriate approach for this problematic is to study a large number of single nucleotide polymorphisms (SNPs) in individuals sharing a genetic background. Our aim was to analyze two CCR2 and seven CCR5 SNPs and their association to CCC in Argentina. A case-control study was carried out in 480 T. cruzi seropositive adults from Argentinean Gran Chaco endemic region (Wichi and Creole) and patients from Buenos Aires health centres. They were classified according to the Consensus on Chagas-Mazza Disease as non-demonstrated (non-DC group) or demonstrated (DC group) cardiomyopathy, i.e. asymptomatic or with CCC patients, respectively. Since, after allelic analysis, 2 out of 9 studied SNPs did not fit Hardy-Weinberg equilibrium in the unaffected non-DC group from Wichi patients, we analyzed them as a separate population. Only rs1800024T and rs41469351T in CCR5 gene showed significant differences within non-Wichi population (Creole + patients from Buenos Aires centres), being the former associated to protection, and the latter to risk of CCC. No evidence of association was observed between any of the analyzed CCR2-CCR5 gene polymorphisms and the development of CCC; however, the HHE haplotype was associated with protection in Wichi population. Our findings support the hypothesis that CCR2-CCR5 genes and their haplotypes are associated with CCC; however, depending on the population studied, different associations can be found. Therefore, the evolutionary context, in which the genes or haplotypes are associated with diseases, acquires special relevance.
Acta Tropica, 2015
Chagas disease is a major unsolved health issue in Latin America and an emerging threat worldwide... more Chagas disease is a major unsolved health issue in Latin America and an emerging threat worldwide. New drugs are urgently needed for chemotherapy as those available (benznidazole and nifurtimox) have variable efficacy and elevated toxicity. Efforts are actually oriented to improve tools and technologies (e.g. transgenic parasites, flow cytometry or image-based systems) for the screening of large numbers of candidate compounds for their activity against Trypanosoma cruzi (T. cruzi). Methods that test drug efficacy and selectivity in the same assay are suitable to accelerate the process of drug discovery. Here, we developed a GFP expressing T. cruzi from a moderate virulence stock and confirmed that the transgenic parasite retained the biological characteristics of the parental strain. With this tool, we established a flow cytometer-based method to simultaneously test drug activity against intracellular amastigotes and toxicity to the host cell. This one-step procedure allows determining the selectivity index of the tested compound in a sensitive and accurate manner even with low infection rates. This method can provide additional information on the interactions between drug, parasites and host cell and could be adapted to other trypanosomatids and protozoa with intracellular multiplication.
Brazilian Journal of Medical and Biological Research, 2012
Infection with Bartonella spp may cause cardiac arrhythmias, myocarditis and endocarditis in huma... more Infection with Bartonella spp may cause cardiac arrhythmias, myocarditis and endocarditis in humans. The aim of the present study was to evaluate a possible association between Bartonella spp bacteremia and endocarditis, arrhythmia and Chagas cardiomyopathy in patients from Brazil and Argentina. We screened for the presence of bacterial 16S rRNA in human blood by PCR using oligonucleotides to amplify a 185-bp bacterial DNA fragment. Blood samples were taken from four groups of subjects in Brazil and Argentina: i) control patients without clinical disease, ii) patients with negative blood-culture endocarditis, iii) patients with arrhythmias, and iv) patients with chronic Chagas cardiomyopathy. PCR products were analyzed on 1.5% agarose gel to visualize the 185-bp fragment and then sequenced to confirm the identity of DNA. Sixty of 148 patients (40.5%) with cardiac disease and 1 of 56 subjects (1.8%) from the control group presented positive PCR amplification for Bartonella spp, suggesting a positive association of the bacteria with these diseases. Separate analysis of the four groups showed that the risk of a Brazilian patient with endocarditis being infected with Bartonella was 22 times higher than in the controls. In arrhythmic patients, the prevalence of infection was 45 times higher when compared to the same controls and 40 times higher for patients with Chagas cardiomyopathy. To the best of our knowledge this is the first report of the association between Bartonella spp bacteremia and Chagas disease. The present data may be useful for epidemiological and prevention studies in Brazil and Argentina.
Parasitology, 2012
SUMMARYThe discrete typing units (DTUs) of Trypanosoma cruzi that infect domestic dogs and cats h... more SUMMARYThe discrete typing units (DTUs) of Trypanosoma cruzi that infect domestic dogs and cats have rarely been studied. With this purpose we conducted a cross-sectional xenodiagnostic survey of dog and cat populations residing in 2 infested rural villages in Pampa del Indio, in the humid Argentine Chaco. Parasites were isolated by culture from 44 dogs and 12 cats with a positive xenodiagnosis. DTUs were identified from parasite culture samples using a strategy based on multiple polymerase-chain reactions. TcVI was identified in 37 of 44 dogs and in 10 of 12 cats, whereas TcV was identified in 5 dogs and in 2 cats –a new finding for cats. No mixed infections were detected. The occurrence of 2 dogs infected with TcIII –classically found in armadillos– suggests a probable link with the local sylvatic transmission cycle involving Dasypus novemcinctus armadillos and a potential risk of human infection with TcIII. Our study reinforces the importance of dogs and cats as domestic reservoi...
Memórias do Instituto Oswaldo Cruz, 1997
Strategies to construct the physical map of the Trypanosoma cruzi nuclear genome have to capitali... more Strategies to construct the physical map of the Trypanosoma cruzi nuclear genome have to capitalize on three main advantages of the parasite genome, namely (a) its small size, (b) the fact that all chromosomes can be defined, and many of them can be isolated by pulse field gel electrophoresis, and (c) the fact that simple Southern blots of electrophoretic karyotypes can be used to map sequence tagged sites and expressed sequence tags to chromosomal bands. A major drawback to cope with is the complexity of T. cruzi genetics, that hinders the construction of a comprehensive genetic map. As a first step towards physical mapping, we report the construction and partial characterization of a T. cruzi CL-Brener genomic library in yeast artificial chromosomes (YACs) that consists of 2,770 individual YACs with a mean insert size of 365 kb encompassing around 10 genomic equivalents. Two libraries in bacterial artificial chromosomes (BACs) have been constructed, BACI and BACII. Both libraries represent about three genome equivalents. A third BAC library (BAC III) is being constructed. YACs and BACs are invaluable tools for physical mapping. More generally, they have to be considered as a common resource for research in Chagas disease.
Uploads
Papers by Alejandro Schijman