Publications

4373

Abstract Insects engage in manifold interactions with bacteria that can shift along the parasitism–mutualism continuum. However, only a small number of bacterial taxa managed to successfully colonize a wide diversity of insects, by evolving mechanisms for host-cell entry, immune evasion, germline tropism, reproductive manipulation, and/or by providing benefits to the host that stabilize the symbiotic association. Here, we report on the discovery of an Enterobacterales endosymbiont (Symbiodolus, type species Symbiodolus clandestinus) that is widespread across at least six insect orders and occurs at high prevalence within host populations. Fluorescence in situ hybridization in several Coleopteran and one Dipteran species revealed Symbiodolus’ intracellular presence in all host life stages and across tissues, with a high abundance in female ovaries, indicating transovarial vertical transmission. Symbiont genome sequencing across 16 host taxa revealed a high degree of functional conservation in the eroding and transposon-rich genomes. All sequenced Symbiodolus genomes encode for multiple secretion systems, alongside effectors and toxin-antitoxin systems, which likely facilitate host-cell entry and interactions with the host. However, Symbiodolus-infected insects show no obvious signs of disease, and biosynthetic pathways for several amino acids and cofactors encoded by the bacterial genomes suggest that the symbionts may also be able to provide benefits to the hosts. A lack of host-symbiont cospeciation provides evidence for occasional horizontal transmission, so Symbiodolus’ success is likely based on a mixed transmission mode. Our findings uncover a hitherto undescribed and widespread insect endosymbiont that may present valuable opportunities to unravel the molecular underpinnings of symbiosis establishment and maintenance.

Abstract Ammonia-oxidizing Nitrososphaeria are among the most abundant archaea on Earth and have profound impacts on the biogeochemical cycles of carbon and nitrogen. In contrast to these well-studied ammonia-oxidizing archaea (AOA), deep-branching non-AOA within this class remain poorly characterized because of a low number of genome representatives. Here, we reconstructed 128 Nitrososphaeria metagenome-assembled genomes from acid mine drainage and hot spring sediment metagenomes. Comparative genomics revealed that extant non-AOA are functionally diverse, with capacity for carbon fixation, carbon monoxide oxidation, methanogenesis, and respiratory pathways including oxygen, nitrate, sulfur, or sulfate, as potential terminal electron acceptors. Despite their diverse anaerobic pathways, evolutionary history inference suggested that the common ancestor of Nitrososphaeria was likely an aerobic thermophile. We further surmise that the functional differentiation of Nitrososphaeria was primarily shaped by oxygen, pH, and temperature, with the acquisition of pathways for carbon, nitrogen, and sulfur metabolism. Our study provides a more holistic and less biased understanding of the diversity, ecology, and deep evolution of the globally abundant Nitrososphaeria.

Abstract Acidimicrobiia are widely distributed in nature and suggested to be autotrophic via the Calvin–Benson–Bassham (CBB) cycle. However, direct evidence of chemolithoautotrophy in Acidimicrobiia is lacking. Here, we report a chemolithoautotrophic enrichment from a saline lake, and the subsequent isolation and characterization of a chemolithoautotroph, Salinilacustristhrix flava EGI L10123T, which belongs to a new Acidimicrobiia family. Although strain EGI L10123T is autotrophic, neither its genome nor Acidimicrobiia metagenome-assembled genomes from the enrichment culture encode genes necessary for the CBB cycle. Instead, genomic, transcriptomic, enzymatic, and stable-isotope probing data hinted at the activity of the reversed oxidative TCA (roTCA) coupled with the oxidation of sulfide as the electron donor. Phylogenetic analysis and ancestral character reconstructions of Acidimicrobiia suggested that the essential CBB gene rbcL was acquired through multiple horizontal gene transfer events from diverse microbial taxa. In contrast, genes responsible for sulfide- or hydrogen-dependent roTCA carbon fixation were already present in the last common ancestor of extant Acidimicrobiia. These findings imply the possibility of roTCA carbon fixation in Acidimicrobiia and the ecological importance of Acidimicrobiia. Further research in the future is necessary to confirm whether these characteristics are truly widespread across the clade.

Abstract Nitrate leaching from agricultural soils is increasingly found in groundwater, a primary source of drinking water worldwide. This nitrate influx can potentially stimulate the biological oxidation of iron in anoxic groundwater reservoirs. Nitrate-dependent iron-oxidizing (NDFO) bacteria have been extensively studied in laboratory settings, yet their ecophysiology in natural environments remains largely unknown. To this end, we established a pilot-scale filter on nitrate-rich groundwater to elucidate the structure and metabolism of nitrate-reducing iron-oxidizing microbiomes under oligotrophic conditions mimicking natural groundwaters. The enriched community stoichiometrically removed iron and nitrate consistently with the NDFO metabolism. Genome-resolved metagenomics revealed the underlying metabolic network between the dominant iron-dependent denitrifying autotrophs and the less abundant organoheterotrophs. The most abundant genome belonged to a new Candidate order, named Siderophiliales. This new species, “Candidatus Siderophilus nitratireducens,” carries genes central genes to iron oxidation (cytochrome c cyc2), carbon fixation (rbc), and for the sole periplasmic nitrate reductase (nap). Using thermodynamics, we demonstrate that iron oxidation coupled to nap based dissimilatory reduction of nitrate to nitrite is energetically favorable under realistic Fe3+/Fe2+ and NO3−/NO2− concentration ratios. Ultimately, by bridging the gap between laboratory investigations and nitrate real-world conditions, this study provides insights into the intricate interplay between nitrate and iron in groundwater ecosystems, and expands our understanding of NDFOs taxonomic diversity and ecological role.

Abstract SAR86 is one of the most abundant groups of bacteria in the global surface ocean. However, since its discovery over 30 years ago, it has remained recalcitrant to isolation and many details regarding this group are still unknown. Here, we report the cellular characteristics from the first SAR86 isolate brought into culture, Magnimaribacter mokuoloeensis strain HIMB1674, and use its closed genome in concert with over 700 environmental genomes to assess the phylogenomic and functional characteristics of this order-level lineage of marine Gammaproteobacteria. The SAR86 order Magnimaribacterales invests significant genomic resources into the capacity for $\beta$-oxidation, which is present in most genomes with high gene copy numbers. This cyclical set of reactions appears to be fed by components of cell membranes that include lipids such as phosphatidylcholine, phosphatidylethanolamine, glycolipids, and sulfolipids. In addition to the widespread capacity to degrade the side chain of steroidal compounds via $\beta$-oxidation, several SAR86 sublineages also appear able to fully degrade the steroid polycyclic ring structure as well as other aromatic, polycyclic, and heterocyclic molecules. Read recruitment from publicly available metagenomes reveals that the Magnimaribacterales compose up to 6% of the global surface ocean microbial community. Only a subset of genera drives these high relative abundances, with some more globally dominant and others restricted to specific oceanic regions. This study provides an unprecedented foundation through which to understand this highly abundant yet poorly understood lineage of marine bacteria and charts a path to bring more representatives of this order into laboratory culture.

Abstract The basal metazoan phylum Porifera (sponges) is increasingly used as a model to investigate ecological and evolutionary features of microbe–animal symbioses. However, sponges often host complex microbiomes, which has hampered our understanding of their interactions with their microbial symbionts. Here, we describe the discovery and characterization of the simplest sponge holobiont reported to date, consisting of the deep-sea glass sponge Aphrocallistes beatrix and two newly-described microbial symbionts: an autotrophic ammonia-oxidizing archaeon and a bacterial heterotroph. Omics analyses and metabolic modeling revealed the dependency of the ammonia-oxidizing archaea on sponge-derived ammonia to drive primary production, which in turn supports the bacterium’s growth by providing the dicarboxylate fumarate. Furthermore, virus-mediated archaeal lysis appears crucial to overcome the bacterium’s vitamin B12 auxotrophy. These findings reveal that the exchanges of vitamin B12 and dicarboxylate may be evolutionarily conserved features of symbiosis as they can also be found in interactions between free-living marine bacteria, and between microbes and plants or diatoms.

Abstract Anthropogenic influences have drastically increased nutrient concentrations in many estuaries globally, and microbial communities have adapted to the resulting hypereutrophic ecosystems. However, our knowledge of the dominant microbial taxa and their potential functions in these ecosystems has remained sparse. Here, we study prokaryotic community dynamics in a temporal–spatial dataset, from a subtropical hypereutrophic estuary. Screening 54 water samples across brackish to marine sites revealed that nutrient concentrations and salinity best explained spatial community variations, whereas temperature and dissolved oxygen likely drive seasonal shifts. By combining short and long read sequencing data, we recovered 2,459 metagenome-assembled genomes, proposed new taxon names for previously uncharacterised lineages, and created an extensive, habitat specific genome reference database. Community profiling based on this genome reference database revealed a diverse prokaryotic community comprising 61 bacterial and 18 archaeal phyla, and resulted in an improved taxonomic resolution at lower ranks down to genus level. We found that the vast majority (61 out of 73) of abundant genera (>1% average) represented unnamed and novel lineages, and that all genera could be clearly separated into brackish and marine ecotypes with inferred habitat specific functions. Applying supervised machine learning and metabolic reconstruction, we identified several microbial indicator taxa responding directly or indirectly to elevated nitrate and total phosphorus concentrations. In conclusion, our analysis highlights the importance of improved taxonomic resolution, sheds light on the role of previously uncharacterised lineages in estuarine nutrient cycling, and identifies microbial indicators for nutrient levels crucial in estuary health assessments.

Abstract Ammonia-oxidizing archaea and nitrite-oxidizing bacteria are common members of marine sponge microbiomes. They derive energy for carbon fixation and growth from nitrification—the aerobic oxidation of ammonia to nitrite and further to nitrate—and are proposed to play essential roles in the carbon and nitrogen cycling of sponge holobionts. In this study, we characterize two novel nitrifying symbiont lineages, Candidatus Nitrosokoinonia and Candidatus Nitrosymbion in the marine sponge Coscinoderma matthewsi using a combination of molecular tools, in situ visualization, and physiological rate measurements. Both represent a new genus in the ammonia-oxidizing archaeal class Nitrososphaeria and the nitrite-oxidizing bacterial order Nitrospirales, respectively. Furthermore, we show that larvae of this viviparous sponge are densely colonized by representatives of Ca. Nitrosokoinonia and Ca. Nitrosymbion indicating vertical transmission. In adults, the representatives of both symbiont genera are located extracellularly in the mesohyl. Comparative metagenome analyses and physiological data suggest that ammonia-oxidizing archaeal symbionts of the genus Ca. Nitrosokoinonia strongly rely on endogenously produced nitrogenous compounds (i.e. ammonium, urea, nitriles/cyanides, and creatinine) rather than on exogenous ammonium sources taken up by the sponge. Additionally, the nitrite-oxidizing bacterial symbionts of the genus Ca. Nitrosymbion may reciprocally support the ammonia-oxidizers with ammonia via the utilization of sponge-derived urea and cyanate. Comparative analyses of published environmental 16S rRNA gene amplicon data revealed that Ca. Nitrosokoinonia and Ca. Nitrosymbion are widely distributed and predominantly associated with marine sponges and corals, suggesting a broad relevance of our findings.

Abstract The evolutionary trajectory of Methylophilaceae includes habitat transitions from freshwater sediments to freshwater and marine pelagial that resulted in genome reduction (genome-streamlining) of the pelagic taxa. However, the extent of genetic similarities in the genomic structure and microdiversity of the two genome-streamlined pelagic lineages (freshwater “Ca. Methylopumilus” and the marine OM43 lineage) has so far never been compared. Here, we analyzed complete genomes of 91 “Ca. Methylopumilus” strains isolated from 14 lakes in Central Europe and 12 coastal marine OM43 strains. The two lineages showed a remarkable niche differentiation with clear species-specific differences in habitat preference and seasonal distribution. On the other hand, we observed a synteny preservation in their genomes by having similar locations and types of flexible genomic islands (fGIs). Three main fGIs were identified: a replacement fGI acting as phage defense, an additive fGI harboring metabolic and resistance-related functions, and a tycheposon containing nitrogen-, thiamine-, and heme-related functions. The fGIs differed in relative abundances in metagenomic datasets suggesting different levels of variability ranging from strain-specific to population-level adaptations. Moreover, variations in one gene seemed to be responsible for different growth at low substrate concentrations and a potential biogeographic separation within one species. Our study provides a first insight into genomic microdiversity of closely related taxa within the family Methylophilaceae and revealed remarkably similar dynamics involving mobile genetic elements and recombination between freshwater and marine family members.

Abstract Chlamydiae are ubiquitous intracellular bacteria and infect a wide diversity of eukaryotes, including mammals. However, chlamydiae have never been reported to infect photosynthetic organisms. Here, we describe a novel chlamydial genus and species, Candidatus Algichlamydia australiensis, capable of infecting the photosynthetic dinoflagellate Cladocopium sp. (originally isolated from a scleractinian coral). Algichlamydia australiensis was confirmed to be intracellular by fluorescence in situ hybridization and confocal laser scanning microscopy and temporally stable at the population level by monitoring its relative abundance across four weeks of host growth. Using a combination of short- and long-read sequencing, we recovered a high-quality (completeness 91.73% and contamination 0.27%) metagenome-assembled genome of A. australiensis. Phylogenetic analyses show that this chlamydial taxon represents a new genus and species within the Simkaniaceae family. Algichlamydia australiensis possesses all the hallmark genes for chlamydiae–host interactions, including a complete type III secretion system. In addition, a type IV secretion system is encoded on a plasmid and has previously been observed for only three other chlamydial species. Twenty orthologous groups of genes are unique to A. australiensis, one of which is structurally similar to a protein known from Cyanobacteria and Archaeplastida involved in thylakoid biogenesis and maintenance, hinting at potential chlamydiae interactions with the chloroplasts of Cladocopium cells. Our study shows that chlamydiae infect dinoflagellate symbionts of cnidarians, the first photosynthetic organism reported to harbor chlamydiae, thereby expanding the breadth of chlamydial hosts and providing a new contribution to the discussion around the role of chlamydiae in the establishment of the primary plastid.