Discovery of circulating miRNAs as biomarkers of chronic Chagas heart disease via a small RNA-Seq approach.

Chagas disease affects approximately 7 million people worldwide in Latin America and is a neglected tropical disease. Twenty to thirty percent of chronically infected patients develop chronic Chagas cardiomyopathy decades after acute infection. Identifying biomarkers of Chagas disease progression is necessary to develop better therapeutic and preventive strategies. Circulating microRNAs are increasingly reliable biomarkers of disease and therapeutic targets. To identify new circulating microRNAs for Chagas disease, we performed exploratory small RNA sequencing from the plasma of patients and performed de novo miRNA prediction, identifying potential new microRNAs. The levels of the new microRNAs temporarily named miR-Contig-1519 and miR-Contig-3244 and microRNAs that are biomarkers for nonchagasic cardiomyopathies, such as miR-148a-3p and miR-224-5p, were validated by quantitative reverse transcription. We found a specific circulating microRNA signature defined by low miR-Contig-3244, miR-Contig-1519, and miR-148a-3 levels but high miR-224-5p levels for patients with chronic Chagas disease. Finally, we predicted in silico that these altered circulating microRNAs could affect the expression of target genes involved in different cellular pathways and biological processes, which we will explore in the future.

Autoantibodies against the immunodominant sCha epitope discriminate the risk of sudden death in chronic Chagas cardiomyopathy.

In Chagas disease (ChD) caused by Trypanosoma cruzi, new biomarkers to predict chronic cardiac pathology are urgently needed. Previous studies in chagasic patients with mild symptomatology showed that antibodies against the immunodominant R3 epitope of sCha, a fragment of the human basic helix-loop-helix transcription factor like 5, correlated with cardiac pathology. To validate sCha as a biomarker and to understand the origin of anti-sCha antibodies, we conducted a multicenter study with several cohorts of chagasic patients with severe cardiac symptomatology. We found that levels of antibodies against sCha discriminated the high risk of sudden death, indicating they could be useful for ChD prognosis. We investigated the origin of the antibodies and performed an alanine scan of the R3 epitope. We identified a minimal epitope MRQLD, and a BLAST search retrieved several T. cruzi antigens. Five of the hits had known or putative functions, of which phosphonopyruvate decarboxylase showed the highest cross-reactivity with sCha, confirming the role of molecular mimicry in the development of anti-sCha antibodies. Altogether, we demonstrate that the development of antibodies against sCha, which originated by molecular mimicry with T. cruzi antigens, could discriminate electrocardiographic alterations associated with a high risk of sudden death.

Distribution of Trypanosoma cruzi discrete typing units in Bolivian migrants in Spain.

Chagas disease is caused by the protozoan Trypanosoma cruzi. This parasite is transmitted to humans mainly through the faeces of infected triatomine "kissing" bugs, by blood transfusions or organ donation from infected donors, and can be transmitted from mother to child. This disease is endemic in the Americas, where Bolivia has up to 28.8% prevalence in general population. Increased migration to Europe has made it emerge in countries where it was previously unknown, being Spain the second country in number of patients after the United States. T. cruzi is an organism with a rich genetic diversity, what has been grouped into six discrete typing units (DTUs). Some authors have linked these DTUs either to specific geographical distribution or to the different clinical presentations. Nevertheless little is known about its distribution in migrant populations. Our aim was to describe the T. cruzi strains isolated from a population of chronically infected Bolivian patients attending our clinic in Madrid. Thirty-three consecutive patients meeting this condition were selected for the study. Molecular characterization was performed by an algorithm that combines PCR of the intergenic region of the mini exon-gene, the 24Sα and 18S regions of rDNA and the variable region of the satellite DNA. A descriptive analysis was performed and associations between epidemiological/clinical data and the different DTUs were tested. Twenty-seven out of thirty-three patients had their DTU detected. Mean age was 36 years (IQR 31-43.3) and 23 were women (76.7%). The median time since arrival to Spain was 60 months (IQR 43-81). The most common DTU were TcV, TcIV and TcI. Four patients had cardiac involvement: 2 had TcV and 2 could not have their DTU determined. TcIII was not isolated from any patient. DTUs distribution in migrant population seems to be similar to that observed in the patients' countries of origin.

The Trypanosoma cruzi satellite DNA OligoC-TesT and Trypanosoma cruzi kinetoplast DNA OligoC-TesT for diagnosis of Chagas disease: a multi-cohort comparative evaluation study.

BACKGROUND: The Trypanosoma cruzi satellite DNA (satDNA) OligoC-TesT is a standardised PCR format for diagnosis of Chagas disease. The sensitivity of the test is lower for discrete typing unit (DTU) TcI than for TcII-VI and the test has not been evaluated in chronic Chagas disease patients. METHODOLOGY/PRINCIPAL FINDINGS: We developed a new prototype of the OligoC-TesT based on kinetoplast DNA (kDNA) detection. We evaluated the satDNA and kDNA OligoC-TesTs in a multi-cohort study with 187 chronic Chagas patients and 88 healthy endemic controls recruited in Argentina, Chile and Spain and 26 diseased non-endemic controls from D.R. Congo and Sudan. All specimens were tested in duplicate. The overall specificity in the controls was 99.1% (95% CI 95.2%-99.8%) for the satDNA OligoC-TesT and 97.4% (95% CI 92.6%-99.1%) for the kDNA OligoC-TesT. The overall sensitivity in the patients was 67.9% (95% CI 60.9%-74.2%) for the satDNA OligoC-TesT and 79.1% (95% CI 72.8%-84.4%) for the kDNA OligoC-Test. CONCLUSIONS/SIGNIFICANCE: Specificities of the two T. cruzi OligoC-TesT prototypes are high on non-endemic and endemic controls. Sensitivities are moderate but significantly (p = 0.0004) higher for the kDNA OligoC-TesT compared to the satDNA OligoC-TesT.

Clinicoepidemiological characteristics of HIV-infected immigrants attended at a tropical medicine referral unit.

BACKGROUND: Migration is a growing phenomenon with a well-known impact in infectious diseases epidemiology. Currently, immigrants represent almost 10% of the Spanish population. The majority come from countries where the prevalence of chronic viral illnesses is higher than in Spain. METHODS: To describe clinicoepidemiological features of human immunodeficiency virus (HIV)-infected immigrants attending our Unit and to compare differential characteristics depending on geographical origin, information from all new immigrants from January 1997 to December 2006 was collected. STUDY DESIGN: noninterventional retrospective chart review. RESULTS: We screened 1,609 patients of whom 77 (4.8%) were HIV antibody (Ab) positive. Of these, 80% were sub-Saharan Africans (SSAFR) and 20% were South-Central Americans (SCA). HIV prevalence was higher in SSAFR (5.6% vs 3.2%; p= 0.04). Overall, of those who were HIV Ab positive, 70% were male (median age 30 years), 59% heterosexuals, 9% hepatitis C virus coinfected, 8.6% hepatitis B virus coinfected, and 34% showed a positive tuberculin skin test. Median CD4 cell count was 263 cells/microL, median HIV-ribonucleic acid viral load 4.6 Log/mL, and 48% had a late diagnosis [acquired immunodeficiency syndrome (AIDS)-defining illness or <200 CD4 microL at the time of diagnosis]. Only 68% of patients for whom antiretroviral therapy was indicated actually started therapy and 22% were lost to follow-up just after diagnosis. SCA had lower CD4 cell counts (26 vs 168 cells/microL; p= 0.016), higher viral loads (5.3 vs 4.8 Log; p= 0.001), and were more likely to have an AIDS-defining illness (53% vs 21%; p= 0.04) compared to SSAFR. Tuberculin skin test reactivity was more common among SSAFR versus SCA [adjusted by CD4 count, odds ratio (OR) 6.3 and 95% confidence interval (CI): 0.65-60.5]. The main risk factor for late diagnosis was geographical origin: OR 4.6 (95% CI: 1.11-19.3) (SCA vs SSAFR; adjusted by the interval between the date of arrival in Spain and the date of HIV diagnosis). CONCLUSIONS: Almost half the HIV-infected immigrants were diagnosed in late stages. Patients were frequently lost to follow-up, and a significant minority did not start highly active antiretroviral therapy when indicated. SCA seem to have more severe immunosuppression at the time of diagnosis than SSAFR. Early voluntary routine HIV screening should be promoted.