An international study was performed by 26 experienced PCR laboratories from 14 countries to assess the performance of duplex quantitative real-time PCR (qPCR) strategies on the basis of TaqMan probes for detection and quantification of parasitic loads in peripheral blood samples from Chagas disease patients. Two methods were studied: Satellite DNA (SatDNA) qPCR and kinetoplastid DNA (kDNA) qPCR. Both methods included an internal amplification control. Reportable range, analytical sensitivity, limits of detection and quantification, and precision were estimated according to international guidelines. In addition, inclusivity and exclusivity were estimated with DNA from stocks representing the different Trypanosoma cruzi discrete typing units and Trypanosoma rangeli and Leishmania spp. Both methods were challenged against 156 blood samples provided by the participant laboratories, including samples from acute and chronic patients with varied clinical findings, infected by oral route or vectorial transmission. kDNA qPCR showed better analytical sensitivity than SatDNA qPCR with limits of detection of 0.23 and 0.70 parasite equivalents/mL, respectively. Analyses of clinical samples revealed a high concordance in terms of sensitivity and parasitic loads determined by both SatDNA and kDNA qPCRs. This effort is a major step toward international validation of qPCR methods for the quantification of T. cruzi DNA in human blood samples, aiming to provide an accurate surrogate biomarker for diagnosis and treatment monitoring for patients with Chagas disease.

In recent years Venezuela has faced a severe economic crisis precipitated by political instability and a significant reduction in oil revenue. Public health provision has suffered particularly. Long-term shortages of medicines and medical supplies and an exodus of trained personnel have occurred against the backdrop of a surge in vector-borne parasitic and arboviral infections. Herein, we aim to assess comprehensively the impact of Venezuela's healthcare crisis on vectorborne diseases and the spillover to neighbouring countries. Methods Alongside the ongoing challenges affecting the healthcare system, health-indicator statistics have become increasingly scarce. Official data from the Ministry of Health, for example, are no longer available. To provide and update on vector-borne disease in Venezuela, this study used individualized data from nongovernmental organizations, academic institutions and professional colleges, various local health authorities and epidemiological surveillance programs from neighbouring countries, as well as data available through international agencies. Findings Between 2000-2015 Venezuela witnessed a 365% increase malaria cases followed by a 68% increase (319,765 cases) in late 2017. Neighbouring countries such as Brazil have reported an escalating trend of imported cases from Venezuelan from 1,538 (2014) to 3,129 (2017). Active Chagas disease transmission is reported with seroprevalence in children (<10 years) as high as 12.5% in one community tested (N=64). There has been a nine-fold rise in the mean incidence of dengue between 1990 to 2016. Estimated rates of chikungunya and Zika are 6,975 and 2,057 cases per 100,000 population, respectively, during their epidemic peaks. Interpretation The re-emergence of many arthropod-borne endemic diseases has set in place an epidemic of unprecedented proportions, not only in Venezuela but in the region. Data presented here demonstrates the complex determinants of this situation. National, regional and global authorities must take action to address these worsening epidemics and prevent their expansion beyond Venezuelan borders.

BackgroundMolecular epidemiology at the community level has an important guiding role in zoonotic disease control programmes where genetic markers are suitably variable to unravel the dynamics of local transmission. We evaluated the molecular diversity of Trypanosoma cruzi, the etiological agent of Chagas disease, in southern Ecuador (Loja Province). This kinetoplastid parasite has traditionally been a paradigm for clonal population structure in pathogenic organisms. However, the presence of naturally occurring hybrids, mitochondrial introgression, and evidence of genetic exchange in the laboratory question this dogma.Methodology/Principal FindingsEighty-one parasite isolates from domiciliary, peridomiciliary, and sylvatic triatomines and mammals were genotyped across 10 variable microsatellite loci. Two discrete parasite populations were defined: one predominantly composed of isolates from domestic and peridomestic foci, and another predominantly composed of isolates from sylvatic foci. Spatial genetic variation was absent from the former, suggesting rapid parasite dispersal across our study area. Furthermore, linkage equilibrium between loci, Hardy-Weinberg allele frequencies at individual loci, and a lack of repeated genotypes are indicative of frequent genetic exchange among individuals in the domestic/peridomestic population.Conclusions/SignificanceThese data represent novel population-level evidence of an extant capacity for sex among natural cycles of T. cruzi transmission. As such they have dramatic implications for our understanding of the fundamental genetics of this parasite. Our data also elucidate local disease transmission, whereby passive anthropogenic domestic mammal and triatomine dispersal across our study area is likely to account for the rapid domestic/peridomestic spread of the parasite. Finally we discuss how this, and the observed subdivision between sympatric sylvatic and domestic/peridomestic foci, can inform efforts at Chagas disease control in Ecuador.

Preventing the transmission of SARS-CoV-2 (causative agent for COVID-19) requires implementing contact and respiratory precautions. Modifying human behavior is challenging and requires understanding knowledge, attitudes, and practices (KAPs) regarding health threats. This study explored KAPs among people in Ecuador. A cross-sectional, internet-based questionnaire was used to assess knowledge about COVID-19, attitudes toward ability to control COVID-19, self-reported practices related to COVID-19, and demographics. A total of 2399 individuals participated. Participants had moderate to high levels of knowledge. Participants expressed mixed attitudes about the eventual control of COVID-19 in Ecuador. Participants reported high levels of adoption of preventive practices. Binomial regression analysis suggests unemployed individuals, househusbands/housewives, or manual laborers, as well as those with an elementary school education, have lower levels of knowledge. Women, people over 50 years of age, and those with higher levels of schooling were the most optimistic. Men, individuals 18-29, single, and unemployed people took the riskiest behaviors. Generally, knowledge was not associated with optimism or with practices. Our findings indicate knowledge about COVID-19 is insufficient to prompt behavioral change among Ecuadorians. Since current COVID-19 control campaigns seek to educate the public, these efforts' impacts are likely to be limited. Given attitudes determine people's actions, further investigation into the factors underlying the lack of confidence in the ability of the world, and of Ecuador, to overcome COVID-19, is warranted. Edu-communicational campaigns should be accompanied by efforts to provide economically disadvantaged populations resources to facilitate adherence to recommendations to prevent the spread of the virus.

Genetic exchange enables parasites to rapidly transform disease phenotypes and exploit new host populations. Trypanosoma cruzi , the parasitic agent of Chagas disease and a public health concern throughout Latin America, has for decades been presumed to exchange genetic material rarely and without classic meiotic sex. We present compelling evidence from 45 genomes sequenced from southern Ecuador that T. cruzi in fact maintains truly sexual, panmictic groups that can occur alongside others that remain highly clonal after past hybridization events. These groups with divergent reproductive strategies appear genetically isolated despite possible co-occurrence in vectors and hosts. We propose biological explanations for the fine-scale disconnectivity we observe and discuss the epidemiological consequences of flexible reproductive modes. Our study reinvigorates the hunt for the site of genetic exchange in the T. cruzi life cycle, provides tools to define the genetic determinants of parasite virulence, and reforms longstanding theory on clonality in trypanosomatid parasites.

BackgroundChagas disease, caused by infection with the protozoan Trypanosoma cruzi, remains a serious public health issue in Latin America. Genetically diverse, the species is sub-divided into six lineages, known as TcI–TcVI, which have disparate geographical and ecological distributions. TcII, TcV, and TcVI are associated with severe human disease in the Southern Cone countries, whereas TcI is associated with cardiomyopathy north of the Amazon. T. cruzi persists as a chronic infection, with cardiac and/or gastrointestinal symptoms developing years or decades after initial infection. Identifying an individual's history of T. cruzi lineage infection directly by genotyping of the parasite is complicated by the low parasitaemia and sequestration in the host tissues.Methodology/Principal FindingsWe have applied here serology against lineage-specific epitopes of the T. cruzi surface antigen TSSA, as an indirect approach to allow identification of infecting lineage. Chagasic sera from chronic patients from a range of endemic countries were tested by ELISA against synthetic peptides representing lineage-specific TSSA epitopes bound to avidin-coated ELISA plates via a biotin labelled polyethylene glycol-glycine spacer to increase rotation and ensure each amino acid side chain could freely interact with their antibodies. 79/113 (70%) of samples from Brazil, Bolivia, and Argentina recognised the TSSA epitope common to lineages TcII/TcV/TcVI. Comparison with clinical information showed that a higher proportion of Brazilian TSSApep-II/V/VI responders had ECG abnormalities than non-responders (38% vs 17%; p<0.0001). Among northern chagasic sera 4/20 (20%) from Ecuador reacted with this peptide; 1/12 Venezuelan and 1/34 Colombian samples reacted with TSSApep-IV. In addition, a proposed TcI-specific epitope, described elsewhere, was demonstrated here to be highly conserved across lineages and therefore not applicable to lineage-specific serology.Conclusions/SignificanceThese results demonstrate the considerable potential for synthetic peptide serology to investigate the infection history of individuals, geographical and clinical associations of T. cruzi lineages.

Background: The requirements for growth and survival of the intracellular pathogen Trypanosoma cruzi within mammalian host cells are poorly understood. Transcriptional profiling of the host cell response to infection serves as a rapid read-out for perturbation of host physiology that, in part, reflects adaptation to the infective process. Using Affymetrix oligonucleotide array analysis we identified common and disparate host cell responses triggered by T. cruzi infection of phenotypically diverse human cell types.

Abstract. Trypanosoma cruzi infection in the Ecuadorian Amazon region has recently been reported. A seroepidemiologic survey conducted in four provinces in this region indicates a seroprevalence rate of 2.4% among the 6,866 samples collected in 162 communities. Among children Յ 10 years of age, 1.2% were seropositive. Risk factors for T. cruzi seropositivity were having been born and remaining in the Ecuadorian Amazon provinces, age, living in a house with a thatch roof and open or mixed wall construction, recognizing the vector insects, and reporting being bitten by a triatomine bug. These data suggest active transmission of Chagas' disease in the Ecuadorian Amazon region is associated with poor housing conditions, and highlight the need for further studies aimed at understanding the biology of the insect vectors, reservoir species, and the clinical impact of T. cruzi infection as the basis for future educational and control programs in this region.