VESPA: an optimized protocol for accurate metabarcoding-based characterization of vertebrate eukaryotic endosymbiont and parasite assemblages.

Protocols for characterizing taxonomic assemblages by deep sequencing of short DNA barcode regions (metabarcoding) have revolutionized our understanding of microbial communities and are standardized for bacteria, archaea, and fungi. Unfortunately, comparable methods for host-associated eukaryotes have lagged due to technical challenges. Despite 54 published studies, issues remain with primer complementarity, off-target amplification, and lack of external validation. Here, we present VESPA (Vertebrate Eukaryotic endoSymbiont and Parasite Analysis) primers and optimized metabarcoding protocol for host-associated eukaryotic community analysis. Using in silico prediction, panel PCR, engineered mock community standards, and clinical samples, we demonstrate VESPA to be more effective at resolving host-associated eukaryotic assemblages than previously published methods and to minimize off-target amplification. When applied to human and non-human primate samples, VESPA enables reconstruction of host-associated eukaryotic endosymbiont communities more accurately and at finer taxonomic resolution than microscopy. VESPA has the potential to advance basic and translational science on vertebrate eukaryotic endosymbiont communities, similar to achievements made for bacterial, archaeal, and fungal microbiomes.

Comparative prevalence of Oxalobacter formigenes in three human populations.

There has been increasing interest in the human anaerobic colonic bacterium Oxalobacter formigenes because of its ability to metabolize oxalate, and its potential contribution to protection from calcium oxalate kidney stones. Prior studies examining the prevalence of this organism have focused on subjects in developed countries and on adults. Now using O. formigenes-specific PCR, we have compared the prevalence of these organisms among subjects in two remote areas in which modern medical practices have hardly been present with a USA group of mothers and their infants for the first three years of life. Among the Amerindians of the Yanomami-Sanema and Yekwana ethnic groups in Venezuela and the Hadza in Tanzania, O. formigenes was detected in 60-80% of the adult subjects, higher than found in adults from USA in this and prior studies. In young children, the prevalence was much lower in USA than in either tribal village. These data extend our understanding of the epidemiology of O. formigenes carriage, and are consistent with the hypothesis that the rising incidence of kidney stones is associated with the progressive loss of O. formigenes colonization in populations that have been highly impacted by modern medical practices.

Trachoma in 3 Amerindian Communities, Venezuelan Amazon, 2018.

Trachoma is among the most common infectious causes of blindness. During January-May 2018, a total of 4 trachoma cases were diagnosed among Amerindians of the Yanomami ethnic group in 3 communities of southern Venezuela. This country has social and environmental conditions conducive to the endemicity of this neglected tropical disease.

Publisher Correction: Complex virome in feces from Amerindian children in isolated Amazonian villages.

In the original version of this Article, the affiliation details for Eric Delwart were incorrectly given as 'Blood Systems Research Institute, San Francisco, CA 94118, USA and Amazonic Center for Research and Control of Tropical Diseases (CAICET), Puerto Ayacucho 7101, Venezuela'. The correct affiliations are 'Blood Systems Research Institute, San Francisco, CA 94118, USA and Department of Laboratory Medicine, University of California at San Francisco, San Francisco, CA 94118, USA'. This has been corrected in both the PDF and HTML versions of the Article.

Complex virome in feces from Amerindian children in isolated Amazonian villages.

The number of viruses circulating in small isolated human populations may be reduced by viral extinctions and rare introductions. Here we used viral metagenomics to characterize the eukaryotic virome in feces from healthy children from a large urban center and from three Amerindian villages with minimal outside contact. Numerous human enteric viruses, mainly from the Picornaviridae and Caliciviridae families, were sequenced from each of the sites. Multiple children from the same villages shed closely related viruses reflecting frequent transmission clusters. Feces of isolated villagers also contained multiple viral genomes of unknown cellular origin from the Picornavirales order and CRESS-DNA group and higher levels of nematode and protozoan DNA. Despite cultural and geographic isolation, the diversity of enteric human viruses was therefore not reduced in these Amazonian villages. Frequent viral introductions and/or increased susceptibility to enteric infections may account for the complex fecal virome of Amerindian children in isolated villages.

Changes in the Gut Microbiota of Urban Subjects during an Immersion in the Traditional Diet and Lifestyle of a Rainforest Village.

People living traditional lifestyles have higher gut microbiota diversity than urban subjects. We hypothesized that shifting lifestyles from an urban environment to a traditional rainforest village would lead to changes in the microbiota of visitors, which would become more similar to the microbiota of villagers. Here, we characterized at different time points the microbiota of 7 urban visitors (5 adults and 2 children) staying in a rainforest Amerindian village for 16 days and compared them with a reference collection of samples from age-matched local villagers. We performed a 16S rRNA gene survey of samples from multiple body sites (including fecal, oral, nasal, and skin samples) using Illumina MiSeq sequencing. The main factor segregating the microbiotas of each body site was the human group (i.e., visitors versus villagers), with the visitor microbiota tending to have lower alpha diversity; the lowered alpha diversity was statistically significant in the microbiota of skin and in the children's fecal and oral microbiota. During the rainforest period, all visitors experienced microbiota changes within their personal cloud of variation. For all body sites, the microbiota conformations in the visitor children better matched the microbiota conformations in villagers of the same age than did those of the visitor adults, which showed a lower "microbiota age" than the microbiota of the villagers. The results suggest higher stability in the adult microbiota, with the less resilient children's microbiota responding more to dietary changes.IMPORTANCE Despite the limitations of a small study, our results evidence higher resilience of the gut microbiota with respect to dietary manipulation in adults than in children and urge further studies to understand the extent of microbiota plasticity in response to dietary changes and the mechanisms underlying microbiota resilience. These studies are relevant to the potential of future human pre- and probiotics in preventing or curing microbiota-associated diseases.

Long-term comparative pharmacovigilance of orally transmitted Chagas disease: first report.

BACKGROUND: Two old drugs are the only choice against Trypanosoma cruzi and little is known about their secondary effects in the acute stage of oral-transmitted Chagas disease (ChD). METHODS: A cross-sectional analytical surveillance study was conducted in a sizable cohort of patients seen during the largest acute foodborne ChD microepidemic registered so far. Individuals were treated with benznidazole (BNZ) or nifurtimox (NFX). 'Common Terminology Criteria for Adverse Events' was assessed to categorize side effects according to severity. RESULTS: Out of 176 treatments applied, 79% had one or more adverse effects, which predominated in adults (97.8%) as compared to children (75.5%). Risk of side effects with NFX was significantly higher than BNZ. Four adults and a child treated with NFX had severe side effects (pulmonary infarction, facial paralysis, neutropenia, blurred vision, bone marrow hypoplasia) warranting hospitalization, and drug suspension. Adverse effects frequently reported with NFX were abdominal pain, hyporexia, weight loss, headache, nausea and lymphocytosis, whereas skin rash, neurosensory effects, hyporexia, fatigue, pyrosis, abdominal pain and eosinophilia were observed with BNZ. CONCLUSIONS: Frequency and severity of side effects during treatment of acute oral infection by T. cruzi demand direct supervision and close follow-up, even in those asymptomatic, to prevent life-threatening situations.

Evidence of suppression of onchocerciasis transmission in the Venezuelan Amazonian focus.

BACKGROUND: The World Health Organization (WHO) has set goals for onchocerciasis elimination in Latin America by 2015. Most of the six previously endemic countries are attaining this goal by implementing twice a year (and in some foci, quarterly) mass ivermectin (Mectizan®) distribution. Elimination of transmission has been verified in Colombia, Ecuador and Mexico. Challenges remain in the Amazonian focus straddling Venezuela and Brazil, where the disease affects the hard-to-reach Yanomami indigenous population. We provide evidence of suppression of Onchocerca volvulus transmission by Simulium guianense s.l. in 16 previously hyperendemic Yanomami communities in southern Venezuela after 15 years of 6-monthly and 5 years of 3-monthly mass ivermectin treatment. METHODS: Baseline and monitoring and evaluation parasitological, ophthalmological, entomological and serological surveys were conducted in selected sentinel and extra-sentinel communities of the focus throughout the implementation of the programme. RESULTS: From 2010 to 2012-2015, clinico-parasitological surveys indicate a substantial decrease in skin microfilarial prevalence and intensity of infection; accompanied by no evidence (or very low prevalence and intensity) of ocular microfilariae in the examined population. Of a total of 51,341 S. guianense flies tested by PCR none had L3 infection (heads only). Prevalence of infective flies and seasonal transmission potentials in 2012-2013 were, respectively, under 1% and 20 L3/person/transmission season. Serology in children aged 1-10 years demonstrated that although 26 out of 396 (7%) individuals still had Ov-16 antibodies, only 4/218 (2%) seropositives were aged 1-5 years. CONCLUSIONS: We report evidence of recent transmission and morbidity suppression in some communities of the focus representing 75% of the Yanomami population and 70% of all known communities. We conclude that onchocerciasis transmission could be feasibly interrupted in the Venezuelan Amazonian focus.

The microbiome of uncontacted Amerindians.

Most studies of the human microbiome have focused on westernized people with life-style practices that decrease microbial survival and transmission, or on traditional societies that are currently in transition to westernization. We characterize the fecal, oral, and skin bacterial microbiome and resistome of members of an isolated Yanomami Amerindian village with no documented previous contact with Western people. These Yanomami harbor a microbiome with the highest diversity of bacteria and genetic functions ever reported in a human group. Despite their isolation, presumably for >11,000 years since their ancestors arrived in South America, and no known exposure to antibiotics, they harbor bacteria that carry functional antibiotic resistance (AR) genes, including those that confer resistance to synthetic antibiotics and are syntenic with mobilization elements. These results suggest that westernization significantly affects human microbiome diversity and that functional AR genes appear to be a feature of the human microbiome even in the absence of exposure to commercial antibiotics. AR genes are likely poised for mobilization and enrichment upon exposure to pharmacological levels of antibiotics. Our findings emphasize the need for extensive characterization of the function of the microbiome and resistome in remote nonwesternized populations before globalization of modern practices affects potentially beneficial bacteria harbored in the human body.

Primer registro de triatóminos naturalmente infectados por Trypanosoma cruzi en el estado Bolívar, Venezuela / First record of naturally infected triatomines with trypanosoma cruzi, Bolivar state, Venezuela

La enfermedad de chagas o tripanosomiasis americana está adquiriendo cada vez mayor importancia en la región amazónica de Venezuela, en la cual hasta ahora se carecía de información de la presencia de trypanosoma cruzi en triatóminos, reservorios y humanos, particularmente para el estado Bolívar, el estado de mayor superficie del país, situado al sur del río Orinoco. Cuatro ejemplares de triatoma maculata fueron recolectados en el peridomicilio de la comunidad de Maniapure, en el municipio Cedeño. En dos de los ejemplares se determinó la infección natural por trypanosoma cruzi al examen al fresco de heces con presencia de tripomastigotos metacíclicos infectantes. Los parásitos fueron inoculados en modelos murinos y aislados en cultivos para su caracterización molecular. Se confirmó el diagnóstico de este parásito por pruebas parasitológicas y moleculares, caracterizándose los aislados como TcI. La importancia de estos hallazgos pioneros podría motivar el estudio de la transmisión vectorial de la enfermedad de chagas en este estado catalogado como no endémico.

American trypanosomiasis or chagas disease is becoming increasingly important in the Amazon region of Venezuela, in which up to now information was lacking of the presence of trypanosoma cruzi in triatomins, reservoirs and humans, particularly for Bolivar state, the state with the largest territory of the country, located in the south of the Orinoco river. Four specimens of triatoma maculata were collected in the peri-domiciliar area of the community of Maniapure, in the Cedeño municipality. Two of these individuals were found naturally infected with trypanosoma cruzi upon examination of fresh stool with presence of metacyclic trypomastigotes. Parasites were inoculated in murine models and isolated for molecular characterization. Parasite isolates were molecularly characterized as TcI. The importance of these pioneering findings should motivate the study of the vector or oral transmission of Chagas disease in this non endemic State of Venezuela.

Hidatidosis poliquística autóctona en dos pacientes Yanomami en el Alto Orinoco, Amazonas, Venezuela / Polycystic hydatid disease in two patients indigenous Yanomami in the Upper Orinoco, Amazonas, Venezuela

La infección por Echinococcus sp. es hipoendémica en Venezuela. Sólo cuatro casos de hidatidosis autóctona por E. vogeli han sido reportados, tres de ellos en la región de la Guayana venezolana. En Febrero del año 2009 se realizó el diagnóstico clínico-sero-epidemiológico de hidatidosis poliquística en una paciente femenina de la etnia Yanomami, procedente de Parima B, Alto Orinoco, en la Amazonía venezolana. Se resolvió con tratamiento médico y quirúrgico por laparoscopia y se evidenció en el quiste la presencia de ganchos rostelares compatibles con E. vogeli. En Abril del 2009 en una segunda paciente Yanomami de igual procedencia, se le diagnosticó hidatidosis por E. vogeli siendo operada exitosamente por cirugía laparoscópica asistida por robot. Dos casos humanos en una misma población y la presencia de factores de riesgo como la tenencia de perros domésticos y la comunicación por informantes indígenas del hallazgo de quistes en hígados de animales de cacería (Cuniculus paca o lapa y Dasyprocta sp. o picure), hacen pensar en transmisión activa en la cuenca del Alto Orinoco y en zonas selváticas de la Guayana venezolana. El presente, es el primer registro de casos de hidatidosis poliquística en indígenas de la etnia Yanomami.

Infection by Echinococcus sp. is hypoendemic in Venezuela. Only four cases of autochthonous E. vogeli hydatidosis have been reported, including three in the Venezuelan region of Guayana. In February 2009, based on epidemiological data, signs and symptoms and serological tests, a female patient of the Yanomami ethnic group, was diagnosed with a polycystic hydatid disease in Parima B, Alto Orinoco, in the Venezuelan Amazon. Rostellar hooks compatible with E. vogeli were found in the cyst. It was resolved with medical and surgical treatment by laparoscopy. A second Yanomami patient from the same location was diagnosed with E. vogeli hydatidosis in April 2009, being successfully operated with robot-assisted laparoscopy. Two human cases in the same population and the presence of risk factors such as domestic dog ownership and findings of cysts in livers of hunted animals (such as Cuniculus and Dasyprocta sp.) reported by indigenous informants, suggest active transmission in the Upper Orinoco basin and forested areas of the Venezuelan Guayana. These are the first reported cases of polycystic hydatid disease of the Yanomami ethnic group.