Zulma M. Cucunubá

Countering the Zika epidemic in Latin America

Epidemic dynamics are key and data gaps must be addressed As evidence grew for a causal link between Zika infection and microcephaly and other serious congenital anomalies (1), the World Health Organization (WHO) declared the Latin American Zika epidemic a public health emergency of international concern in February 2016 (2). The speed of spread [see the figure, top, and the supplementary materials (SM)] has made effective public health responses challenging. Immediate responses have included vector control (3) and advice to delay pregnancy in a few countries (4), followed by an extended recommendation to all affected countries by WHO in June 2016. These have merits but are likely to have limited effectiveness (5) and may interact antagonistically. Fuller understanding of dynamics and drivers of the epidemic is needed to assess longer-term risks to prioritize interventions.

Countering Zika in Latin America

Epidemic dynamics are key and data gaps must be addressed As evidence grew for a causal link between Zika infection and microcephaly and other serious congenital anomalies (1), the World Health Organization (WHO) declared the Latin American Zika epidemic a public health emergency of international concern in February 2016 (2). The speed of spread [see the figure, top, and the supplementary materials (SM)] has made effective public health responses challenging. Immediate responses have included vector control (3) and advice to delay pregnancy in a few countries (4), followed by an extended recommendation to all affected countries by WHO in June 2016. These have merits but are likely to have limited effectiveness (5) and may interact antagonistically. Fuller understanding of dynamics and drivers of the epidemic is needed to assess longer-term risks to prioritize interventions.

Contemporary cryptic sexuality in Trypanosoma cruzi

Clonal propagation is considered to be the predominant mode of reproduction among many parasitic protozoa. However, this assumption may overlook unorthodox, infrequent or cryptic sexuality. Trypanosoma cruzi, which causes Chagas disease, is known to undergo non‐Mendelian genetic exchange in the laboratory. In the field, evidence of extant genetic exchange is limited. In this study, we undertook intensive sampling of T. cruzi Discrete Typing Unit I in endemic eastern Colombia. Using Fluorescence‐activated cell sorting, we generated 269 biological clones from 67 strains. Each clone was genotyped across 24 microsatellite loci. Subsequently, 100 representative clones were typed using 10 mitochondrial sequence targets (3.76 Kbp total). Clonal diversity among humans, reservoir hosts and vectors suggested complex patterns of superinfection and/or coinfection in oral and vector‐borne Chagas disease cases. Clonal diversity between mother and foetus in a congenital case demonstrates that domestic TcI genotypes are infective in utero. Importantly, gross incongruence between nuclear and mitochondrial markers is strong evidence for widespread genetic exchange throughout the data set. Furthermore, a confirmed mosaic maxicircle sequence suggests intermolecular recombination between individuals as a further mechanism of genetic reassortment. Finally, robust dating based on mitochondrial DNA indicates that the emergence of a widespread domestic TcI clade that we now name TcIDOM (formerly TcIa/VENDom) occurred 23 000 ± 12 000 years ago and was followed by population expansion, broadly corresponding with the earliest human migration into the Americas.

Molecular epidemiology of human oral Chagas disease outbreaks in Colombia.

Background Trypanosoma cruzi, the causative agent of Chagas disease, displays significant genetic variability revealed by six Discrete Typing Units (TcI-TcVI). In this pathology, oral transmission represents an emerging epidemiological scenario where different outbreaks associated to food/beverages consumption have been reported in Argentina, Bolivia, Brazil, Ecuador and Venezuela. In Colombia, six human oral outbreaks have been reported corroborating the importance of this transmission route. Molecular epidemiology of oral outbreaks is barely known observing the incrimination of TcI, TcII, TcIV and TcV genotypes. Methodology and Principal Findings High-throughput molecular characterization was conducted performing MLMT (Multilocus Microsatellite Typing) and mtMLST (mitochondrial Multilocus Sequence Typing) strategies on 50 clones from ten isolates. Results allowed observing the occurrence of TcI, TcIV and mixed infection of distinct TcI genotypes. Thus, a majority of specific mitochondrial haplotypes and allelic multilocus genotypes associated to the sylvatic cycle of transmission were detected in the dataset with the foreseen presence of mitochondrial haplotypes and allelic multilocus genotypes associated to the domestic cycle of transmission. Conclusions These findings suggest the incrimination of sylvatic genotypes in the oral outbreaks occurred in Colombia. We observed patterns of super-infection and/or co-infection with a tailored association with the severe forms of myocarditis in the acute phase of the disease. The transmission dynamics of this infection route based on molecular epidemiology evidence was unraveled and the clinical and biological implications are discussed.

Asymptomatic Plasmodium spp. infection in Tierralta, Colombia

With the aim of determining the prevalence of asymptomatic Plasmodium spp. infection by thick smear and PCR and its association with demographic and epidemiological characteristics in the village of Nuevo Tay, Tierralta, Córdoba, Colombia, a cross-sectional population study was carried out, using random probabilistic sampling. Venous blood samples were taken from 212 people on day 0 for thick smear and PCR. Clinical follow-up and thick smears were carried out on days 14 and 28. The prevalence of Plasmodium spp. infection was 17.9% (38/212; 95% CI: 12.5-23.3%) and the prevalence of asymptomatic Plasmodiumspp. infection was 14.6% (31/212; 95% CI: 9.6-19.6%). Plasmodium vivax was found more frequently (20/31; 64.5%) than Plasmodium falciparum (9/31; 29%) and mixed infections (2/31; 6.5%). A significantly higher prevalence of asymptomatic infection was found in men (19.30%) than in women (9.18%) (prevalence ratio: 2.10; 95% CI: 1.01-4.34%; p = 0.02). People who developed symptoms had a significantly higher parasitemia on day 0 than those who remained asymptomatic, of 1,881.5 +/- 3,759 versus 79 +/- 106.9 (p = 0.008). PCR detected 50% more infections than the thick smears. The presence of asymptomatic Plasmodium spp. infection highlights the importance of carrying out active searches amongst asymptomatic populations residing in endemic areas.

Increased CD4+/CD8+ Double-Positive T Cells in Chronic Chagasic Patients

Background CD4+/CD8+ double positive (DP) T cells have been described in healthy individuals as well as in patients with autoimmune and chronic infectious diseases. In chronic viral infections, this cell subset has effector memory phenotype and displays antigen specificity. No previous studies of double positive T cells in parasite infections have been carried out. Methodology/Principal Findings Seventeen chronic chagasic patients (7 asymptomatic and 10 symptomatic) and 24 non-infected donors, including 12 healthy and 12 with non-chagasic cardiomyopathy donors were analyzed. Peripheral blood was stained for CD3, CD4, CD8, HLA-DR and CD38, and lymphocytes for intracellular perforin. Antigen specificity was assessed using HLA*A2 tetramers loaded with T. cruzi K1 or influenza virus epitopes. Surface expression of CD107 and intracellular IFN-γ production were determined in K1-specific DP T cells from 11 chagasic donors. Heart tissue from a chronic chagasic patient was stained for both CD8 and CD4 by immunochemistry. Chagasic patients showed higher frequencies of DP T cells (2.1%±0.9) compared with healthy (1.1%±0.5) and non-chagasic cardiomyopathy (1.2%±0.4) donors. DP T cells from Chagasic patients also expressed more HLA-DR, CD38 and perforin and had higher frequencies of T. cruzi K1-specific cells. IFN-γ production in K1-specific cells was higher in asymptomatic patients after polyclonal stimulation, while these cells tended to degranulate more in symptomatic donors. Immunochemistry revealed that double positive T cells infiltrate the cardiac tissue of a chagasic donor. Conclusions Chagasic patients have higher percentages of circulating double positive T cells expressing activation markers, potential effector molecules and greater class I antigenic specificity against T. cruzi. Although K1 tetramer positive DP T cell produced little IFN-γ, they displayed degranulation activity that was increased in symptomatic patients. Moreover, K1-specific DP T cells can migrate to the heart tissue.

Enfermedad de Chagas aguda en Colombia, una entidad poco sospechada. Informe de 10 casos presentados en el periodo 2002 a 2005

Introduction. In Colombia, reported cases of acute Chagas disease are sporadic. Objective. Ten cases were described that had been reported to the Parasitology Laboratory of the Colombian National Health Institute between December 2002 and November 2005. Materials and methods. Information from clinical records, epidemiological report forms, laboratory and blood tests was collated. In addition the following data were compiled: demographic variables, clinical findings, results of laboratory tests and other exams (such as peripheral blood smears), IFAT for IgG antibodies, isolation in culture medium, inoculation in mice, polymerase chain reaction tests and isoenzyme eletrophoresis. Results. All the cases presented in known endemic areas for Chagas disease transmission in Colombia. Three cases were from Putumayo Province, two each from the provinces of Arauca, Casanare, Norte de Santander and one from Santander Province. The probable mode of transmission was vector-borne. Seven cases presented in adults aged 18 to 50, three in children aged 6 months to 2 years. Seven were male and three were female. The most frequent symptom was fever in nine cases. Signs of portal of entry were rare; only one patient presented a possible Romana´s sign. Three patients presented myocarditis, two acute cardiac failure and one cardiac tamponade. Parasitemia was evident in nine cases; five had positive IgG serological tests; five cases were confirmed through parasite isolation; isoenzyme electrophoresis showed Trypanosoma cruzi group I. Conclusions. Clinical variability prevailed. In none of the cases was a clinical diagnosis suspected. The diagnosis was made and confirmed through laboratory tests alone. The results highlight the importance of including this disease in the differential diagnosis of febrile syndrome in endemic regions due to its good response to etiological treatment and thereby preventing its progression to the chronic phase.

First Report of Human Trypanosoma cruzi Infection Attributed to TcBat Genotype

Chagas disease is an endemic disease of the American continent caused by Trypanosoma cruzi and divided into six discrete typing units (TcI – TcVI). Nearly 10 million people harbour the infection representing a serious issue in public health. Epidemiological surveillance allowed us to detect a bat‐related T. cruzi genotype (henceforth named TcBat) in a 5‐year‐old female living in a forest area in northwestern Colombia. Molecular tools determined a mixed infection of T. cruzi I and TcBat genotypes. This represents the first report of TcBat infection in humans; the epidemiological consequences of this finding are discussed herein.

Natural and emergent Trypanosoma cruzi I genotypes revealed by mitochondrial (Cytb) and nuclear (SSU rDNA) genetic markers

Chagas disease is a tropical and systemic disease caused by the parasite Trypanosoma cruzi. This parasite has been divided into six Discrete Typing Units (DTUs) due to its high genetic diversity. T. cruzi I (TcI) is the most prevalent DTU in Colombia and recently associated to cardiomyopathies. The aim of this study was to unravel the genetic variability among a set of 70 cell-single TcI clones from different geographical regions and hosts using the sequences of Cytb and SSU rDNA. The results showed two genotypes associated to transmission cycles of Chagas disease in Colombia and supports the previous descriptions using SL-IR. Phylogenetic networks were developed detecting recombination events within TcI. We also tested the phylogenetic relationships beneath TcI clones and TcIII/TcIV sequences observing the high relatedness of TcI clones from sylvatic cycle with TcIII/TcIV. We corroborate the high genetic diversity displayed by TcI, the plausible recombination within this DTU supporting the previous model of genetic exchange proposed in T. cruzi populations. We conclude inquiring the need to pursue new studies to elucidate the genetic structure of TcI across Chagas disease endemic countries.

Comparison of asymptomatic Plasmodium spp. infection in two malaria-endemic Colombian locations

BACKGROUND Asymptomatic infection by Plasmodium spp. could help explain the dynamics of malaria transmission in low-transmission regions. We therefore compared the prevalence of asymptomatic infection by Plasmodium spp. in two Colombian locations, Tierralta and Tumaco, with different transmission patterns, different ecological and epidemiological characteristics and potentially different associated factors. METHOD Two cross-sectional studies were carried out between September 2006 and November 2007. Infection by Plasmodium spp. was determined using polymerase chain reaction (PCR) and microscopy. RESULTS Prevalence of asymptomatic infection at day 0 in Tierralta was 11.3% (95% CI 7.2-16.8) by microscopy and 16.5% (95% CI 11.5-22.9) by PCR, while in Tumaco, values were 2.4% (95% CI 0.7-5.5) and 5.8% (95% CI 2.3-9.2) respectively. General prevalence of asymptomatic plasmodium infection after 28 days of follow-up was 5% (95% CI 3.2-7.7), 6.6% (95% CI 3.8-10.6%) in Tierralta and 3.4% (95% CI 1.5-6.6%) in Tumaco. Plasmodium vivax was predominant in Tierralta, P. falciparum in Tumaco. The main associated factors were male sex (aOR 2.5; 95% CI 1.0-6.7) and two to five previous malaria episodes (aOR 3.0; 95% CI 1.0-9.4). PCR detected 61% more infections than microscopy. CONCLUSION Measurement of the prevalence of asymptomatic Plasmodium spp. infection is suggested as part of the epidemiologic evaluation of malaria in low-transmission areas such as Colombia.