Research Article: Use of shotgun metagenomics for the identification of protozoa in the gut microbiota of healthy individuals from worldwide populations with various industrialization levels

Date Published: February 6, 2019

Publisher: Public Library of Science

Author(s): Ana Lokmer, Amandine Cian, Alain Froment, Nausicaa Gantois, Eric Viscogliosi, Magali Chabé, Laure Ségurel, Tiffany L. Weir.


Protozoa have long been considered undesirable residents of the human gut, but recent findings suggest that some of them may positively affect the gut ecosystem. To better understand the role and ecological dynamics of these commensal and potentially beneficial protozoan symbionts, we need efficient methods to detect them, as well as accurate estimates of their prevalence across human populations. Metagenomics provides such an opportunity, allowing simultaneous detection of multiple symbionts in a single analytical procedure. In this study, we collected fecal samples of 68 individuals from three Cameroonian populations with different subsistence modes and compared metagenomics-based and targeted methods of detection for two common protozoan genera: Blastocystis and Entamoeba. In addition, we analyzed our data along with publicly available fecal metagenomes from various worldwide populations to explore the prevalence and association patterns of ten protozoan genera. Regarding the detection method, microscopy was much less sensitive than metagenomics for Entamoeba, whereas qPCR was at least as sensitive as metagenomics for Blastocystis sp. However, metagenomics was more likely to detect co-colonizations by multiple subtypes. Out of the ten examined genera in 127 individuals from Cameroon, Tanzania, Peru, Italy or USA, only three (Blastocystis, Entamoeba and Enteromonas) had an overall prevalence exceeding 10%. All three genera were more common in less industrialized populations and their prevalence differed between continents and subsistence modes, albeit not in a straightforward manner. The majority (72.5%) of colonized individuals carried at least two protozoan species, indicating that mixed-species colonizations are common. In addition, we detected only positive and no negative association patterns between different protozoa. Despite the pitfalls of the metagenomic approach, ranging from the availability of good-quality sequencing data to the lack of standard analytical procedures, we demonstrated its utility in simultaneous detection of multiple protozoan genera, and especially its ability to efficiently detect mixed-species colonizations. Our study corroborates and expands prevalence results previously obtained for Blastocystis sp. and provides novel data for Entamoeba spp. and several other protozoan genera. Furthermore, it indicates that multiple protozoa are common residents of the healthy human gut worldwide.

Partial Text

Gut protozoa have long been exclusively a topic of parasitological research, although they may actually be more often commensal than pathogenic [1, 2]. Recent findings even suggest that some protozoa, such as Blastocystis sp. and non-pathogenic Entamoeba spp., could benefit their hosts, as they are associated with an increased gut microbiome diversity and a higher frequency of potentially beneficial bacterial taxa [2–6]. It has thus been proposed that the virtual eradication of gut protozoa in industrialized countries might negatively influence human health due to the disruption of ecological interactions in the gut ecosystem [2, 6]. In addition, there is a number of other protozoan species that may affect the gut ecosystem, but whose pathogenicity and prevalence are even less known [2]. In order to study the ecological and clinical impact of gut protozoa, we first need reliable estimates of their prevalence.

Our study is the first to use a metagenomic approach to determine the prevalence of gut protozoa other than Blastocystis sp. Specifically, we used MG data to examine the prevalence of ten protozoan genera in eight populations across the globe. The particular approach we applied enabled us to detect high prevalence of mixed colonization in healthy individuals from non-industrialized populations, including both closely and distantly related protozoa. Our results support the hypothesis that protozoa could be common residents of the healthy human gut [2], but also highlight the pitfalls of MG-based diagnostics due to variety of strategies that can be applied to the analysis of MG data. The role of MG in diagnostics will likely become more important with the increasing number of good-quality genomes in the public databases and the development of well-defined diagnostic protocols.




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