Exploration and characterization of "Amoebozoa" diversity and investigation of their diversity patterns at regional and global scales
Authors:   , Blandenier Quentin 
Published: August 2023

The world eukaryotic diversity is dominated by (mostly) single-celled organisms referred to as protists. Among them, the Amoebozoa are one of the most numerous, diverse and characteristic groups in soil, thus playing important roles in ecosystem functioning. However, their study has been impeded by the difficulty in detecting them and the lack of stable morphological traits in most groups. Nevertheless, some amoebozoans such as the Hyalospheniformes (Arcellinida) are characterized by a self-constructed test (i.e. shell) which facilitates their identification, and are then considered as a suitable model group for investigating diversity patterns of repartition. The recent development of DNA barcoding has helped considerably taxonomic identification, whereas metabarcoding has allowed revealing microbial community composition without observational and cultivation biases. These methods have proved efficient for several microbial groups, but only few studies have been designed for Amoebozoa and available protocols are still rather scarce. The aims of my thesis were then to 1) improve and develop molecular methods to study the amoebozoan diversity and ecology, 2) estimate their taxonomic and functional diversity in the soil, 3) improve the taxonomic and phylogenetic frame for this diversity in order to build a sound basis for further research and 4) characterize the ecological drivers which are likely to influence microbial diversity at local, continental and global scales. We first identified a new molecular marker to survey arcellinids taxa, which proved to be efficient for discriminating closely-related taxa and simultaneously investigating deep relationships among distant taxa (Chapter 2). In addition, we also adapted a metabarcoding protocol with specific COI primers to survey the diversity within the genus Nebela at a fine taxonomical resolution (Chapter 6). Then, we isolated, cultivated and described the first member of a deep-branching environmental clade of Amoebozoa (Chapter 3). This amoeba, one of the smallest amoeboid species described, presents a unique life cycle with an alternation of phagotrophic active trophozoites and osmotrophic fungi-like ramifications. Its presence has been pervasively reported in many soil metabarcoding studies, but this organism had never been characterized. By contrast, Hyalospheniformes are known since the works of Ehrenberg in the 19th century. However, their diversity at the species level remains poorly characterized. In chapter 4, we showed that the iconic testate amoeba species Nebela militaris did not belong to genus Nebela but branched as a separate entity in the Hyalospheniformes tree. Therefore, we erected the new genus Alabasta for this species (Chapter 4). In addition, we demonstrated that Hyalospheniformes diversity had been greatly underestimated. Indeed, our morphological and molecular results have revealed the presence for several species within the genera Apodera, Alocodera and Padaungiella. This new diversity has implications on microbial biogeography as Apodera vas and Alocodera cockayni were previously considered as two non-cosmopolite species with very broad geographical ranges and large ecological tolerances. Furthermore, we showed that the situation was far more complex, suggesting the existence of narrow local endemisms and ecological specialists, similarly to genera Hyalosphenia and Nebela (Chapter 5). Finally, we explored the diversity patterns of the genus Nebela along an elevation gradient (Chapter 6). We observed a decrease of abundance and diversity in high elevation corresponding to a typical mid-domain effect. Our study also revealed several unknown phylotypes restricted to the higher elevation that seemed to present competitive exclusion with the generalist taxa from lower elevation. In conclusion, this thesis highlights that molecular methods associated to robust morphological observations are efficient to reveal and describe the diversity of Amoebozoa. Furthermore, these microbial organisms display biogeographical and macroecological patterns similarly to animals, plants and fungi, when all groups are studied at the same taxonomical rank, i.e. species level.