Culture-free genome-wide locus sequence typing (GLST) provides new perspectives on Trypanosoma cruzi dispersal and infection complexity.

Analysis of genetic polymorphism is a powerful tool for epidemiological surveillance and research. Powerful inference from pathogen genetic variation, however, is often restrained by limited access to representative target DNA, especially in the study of obligate parasitic species for which ex vivo culture is resource-intensive or bias-prone. Modern sequence capture methods enable pathogen genetic variation to be analyzed directly from host/vector material but are often too complex and expensive for resource-poor settings where infectious diseases prevail. This study proposes a simple, cost-effective 'genome-wide locus sequence typing' (GLST) tool based on massive parallel amplification of information hotspots throughout the target pathogen genome. The multiplexed polymerase chain reaction amplifies hundreds of different, user-defined genetic targets in a single reaction tube, and subsequent agarose gel-based clean-up and barcoding completes library preparation at under 4 USD per sample. Our study generates a flexible GLST primer panel design workflow for Trypanosoma cruzi, the parasitic agent of Chagas disease. We successfully apply our 203-target GLST panel to direct, culture-free metagenomic extracts from triatomine vectors containing a minimum of 3.69 pg/µl T. cruzi DNA and further elaborate on method performance by sequencing GLST libraries from T. cruzi reference clones representing discrete typing units (DTUs) TcI, TcIII, TcIV, TcV and TcVI. The 780 SNP sites we identify in the sample set repeatably distinguish parasites infecting sympatric vectors and detect correlations between genetic and geographic distances at regional (< 150 km) as well as continental scales. The markers also clearly separate TcI, TcIII, TcIV and TcV + TcVI and appear to distinguish multiclonal infections within TcI. We discuss the advantages, limitations and prospects of our method across a spectrum of epidemiological research.

2b-RAD genotyping for population genomic studies of Chagas disease vectors: Rhodnius ecuadoriensis in Ecuador.

BACKGROUND: Rhodnius ecuadoriensis is the main triatomine vector of Chagas disease, American trypanosomiasis, in Southern Ecuador and Northern Peru. Genomic approaches and next generation sequencing technologies have become powerful tools for investigating population diversity and structure which is a key consideration for vector control. Here we assess the effectiveness of three different 2b restriction site-associated DNA (2b-RAD) genotyping strategies in R. ecuadoriensis to provide sufficient genomic resolution to tease apart microevolutionary processes and undertake some pilot population genomic analyses. METHODOLOGY/PRINCIPAL FINDINGS: The 2b-RAD protocol was carried out in-house at a non-specialized laboratory using 20 R. ecuadoriensis adults collected from the central coast and southern Andean region of Ecuador, from June 2006 to July 2013. 2b-RAD sequencing data was performed on an Illumina MiSeq instrument and analyzed with the STACKS de novo pipeline for loci assembly and Single Nucleotide Polymorphism (SNP) discovery. Preliminary population genomic analyses (global AMOVA and Bayesian clustering) were implemented. Our results showed that the 2b-RAD genotyping protocol is effective for R. ecuadoriensis and likely for other triatomine species. However, only BcgI and CspCI restriction enzymes provided a number of markers suitable for population genomic analysis at the read depth we generated. Our preliminary genomic analyses detected a signal of genetic structuring across the study area. CONCLUSIONS/SIGNIFICANCE: Our findings suggest that 2b-RAD genotyping is both a cost effective and methodologically simple approach for generating high resolution genomic data for Chagas disease vectors with the power to distinguish between different vector populations at epidemiologically relevant scales. As such, 2b-RAD represents a powerful tool in the hands of medical entomologists with limited access to specialized molecular biological equipment.

Factores sociales asociados con la utilización de los servicios de atención prenatal en Ecuador / Social factors associated with use of prenatal care in Ecuador

Objetivos. La atención prenatal es uno de los pilares de la salud pública y permite el acceso a intervenciones tales como la prevención de la transmisión materno-infantil del VIH y de la sífilis congénita. Este artículo tiene como objetivo describir los factores sociales asociados con la utilización de los servicios de atención prenatal en Ecuador. Métodos. Entre 2011 y 2012, se realizó un análisis de la información procedente de las historias clínicas y de la entrevista a las participantes, que integraron una muestra probabilística a nivel nacional de 5 998 mujeres atendidas por parto o aborto en 15 servicios sanitarios en Ecuador con el objetivo de estimar la prevalencia de VIH, sífilis, enfermedad de Chagas y la cobertura de atención prenatal. Resultados. El estudio mostró que 94,1% de las mujeres había acudido a algún control prenatal, pero la asistencia al menos a cuatro controles fue 73,1%. Se encontró que el menor nivel educativo, el mayor número de embarazos, la ocupación en el sector agrícola o ganadero y la pertenencia a los grupos étnicos indígena, afroecuatoriano u otros minoritarios fueron factores asociados con la falta de uso (ningún control prenatal) o al uso inadecuado de la atención prenatal (menos de cuatro controles o primer control después de las 20 semanas de gestación) en Ecuador. Conclusiones. Estos resultados apuntan a la persistencia de desigualdades marcadas en el acceso y en la utilización de servicios de atención prenatal atribuibles a factores socioeconómicos y a la necesidad de fortalecer las estrategias para su abordaje para alcanzar la meta de la cobertura universal de atención prenatal.

Objectives. Prenatal care is a pillar of public health, enabling access to interventions including prevention of mother-to-child transmission of HIV and congenital syphilis. This paper describes social factors related to use of prenatal care in Ecuador. Methods. In 2011 and 2012, participant clinical history and interview information was analyzed from a national probability sample of 5 998 women presenting for delivery or miscarriage services in 15 healthcare facilities in Ecuador, to estimate prevalence of HIV, syphilis, and Chagas disease, and prenatal care coverage. Results. The study found that 94.1% of women had attended at least one prenatal visit, but that attendance at no less than four visits was 73.1%. Furthermore, lower educational level, greater number of pregnancies, occupation in the agriculture or livestock sector, and membership in ethnic indigenous, Afro-Ecuadorian, or other minority groups were factors associated with lack of use (no prenatal visits) or insufficient use of prenatal care (fewer than four visits or first visit at >20 weeks gestation) in Ecuador. Conclusions. These results point to persistence of marked inequalities in access to and use of prenatal health services attributable to socioeconomic factors and to the need to strengthen strategies to address them, to reach the goal of universal prenatal care coverage.

Factores sociales asociados con la utilización de los servicios de atención prenatal en Ecuador / Social factors associated with use of prenatal care in Ecuador

RESUMEN Objetivos La atención prenatal es uno de los pilares de la salud pública y permite el acceso a intervenciones tales como la prevención de la transmisión materno-infantil del VIH y de la sífilis congénita. Este artículo tiene como objetivo describir los factores sociales asociados con la utilización de los servicios de atención prenatal en Ecuador. Métodos Entre 2011 y 2012, se realizó un análisis de la información procedente de las historias clínicas y de la entrevista a las participantes, que integraron una muestra probabilística a nivel nacional de 5 998 mujeres atendidas por parto o aborto en 15 servicios sanitarios en Ecuador con el objetivo de estimar la prevalencia de VIH, sífilis, enfermedad de Chagas y la cobertura de atención prenatal. Resultados El estudio mostró que 94,1% de las mujeres había acudido a algún control prenatal, pero la asistencia al menos a cuatro controles fue 73,1%. Se encontró que el menor nivel educativo, el mayor número de embarazos, la ocupación en el sector agrícola o ganadero y la pertenencia a los grupos étnicos indígena, afroecuatoriano u otros minoritarios fueron factores asociados con la falta de uso (ningún control prenatal) o al uso inadecuado de la atención prenatal (menos de cuatro controles o primer control después de las 20 semanas de gestación) en Ecuador. Conclusiones Estos resultados apuntan a la persistencia de desigualdades marcadas en el acceso y en la utilización de servicios de atención prenatal atribuibles a factores socioeconómicos y a la necesidad de fortalecer las estrategias para su abordaje para alcanzar la meta de la cobertura universal de atención prenatal.(AU)

ABSTRACT Objectives Prenatal care is a pillar of public health, enabling access to interventions including prevention of mother-to-child transmission of HIV and congenital syphilis. This paper describes social factors related to use of prenatal care in Ecuador. Methods In 2011 and 2012, participant clinical history and interview information was analyzed from a national probability sample of 5 998 women presenting for delivery or miscarriage services in 15 healthcare facilities in Ecuador, to estimate prevalence of HIV, syphilis, and Chagas disease, and prenatal care coverage. Results The study found that 94.1% of women had attended at least one prenatal visit, but that attendance at no less than four visits was 73.1%. Furthermore, lower educational level, greater number of pregnancies, occupation in the agriculture or livestock sector, and membership in ethnic indigenous, Afro-Ecuadorian, or other minority groups were factors associated with lack of use (no prenatal visits) or insufficient use of prenatal care (fewer than four visits or first visit at >20 weeks gestation) in Ecuador. Conclusions These results point to persistence of marked inequalities in access to and use of prenatal health services attributable to socioeconomic factors and to the need to strengthen strategies to address them, to reach the goal of universal prenatal care coverage.(AU)

[Social factors associated with use of prenatal care in Ecuador]. / Factores sociales asociados con la utilización de los servicios de atención prenatal en Ecuador.

OBJECTIVES: Prenatal care is a pillar of public health, enabling access to interventions including prevention of mother-to-child transmission of HIV and congenital syphilis. This paper describes social factors related to use of prenatal care in Ecuador. METHODS: In 2011 and 2012, participant clinical history and interview information was analyzed from a national probability sample of 5 998 women presenting for delivery or miscarriage services in 15 healthcare facilities in Ecuador, to estimate prevalence of HIV, syphilis, and Chagas disease, and prenatal care coverage. RESULTS: The study found that 94.1% of women had attended at least one prenatal visit, but that attendance at no less than four visits was 73.1%. Furthermore, lower educational level, greater number of pregnancies, occupation in the agriculture or livestock sector, and membership in ethnic indigenous, Afro-Ecuadorian, or other minority groups were factors associated with lack of use (no prenatal visits) or insufficient use of prenatal care (fewer than four visits or first visit at >20 weeks gestation) in Ecuador. CONCLUSIONS: These results point to persistence of marked inequalities in access to and use of prenatal health services attributable to socioeconomic factors and to the need to strengthen strategies to address them, to reach the goal of universal prenatal care coverage.