On the non-oxygenic origins of thylakoids
Luc Cornet
Thylakoid membranes are the site of oxygenic photosynthesis, one of the most important biochemical processes on earth. The ancestral state of these membranes is represented today in Gloeobacterales, where they are lacking and photosynthesis instead takes place in the cytoplasmic membrane. The evolutionary transition from this ancestral state to the modern thylakoid membranes provided a major advantage, as it increased photosynthetic efficiency. However, how this significant transition occurred remains an understudied question. The biogenesis of modern thylakoid membranes relies on a highly synchronized process involving numerous assembly factors and showing important differences between the two photosystems. Together, these features suggest the existence of intermediate evolutionary states during the emergence of this compartment. Here, I propose a non-oxygenic origin of thylakoid membranes, where these intermediate states were initially dedicated to alternative electron flows. This hypothesis further addresses the paradox of cyanobacterial diversification in an euxinic environment, toxic to photosystem II.
https://www.nature.com/articles/s42003-025-09100-w
A BAC-guided haplotype assembly pipeline increases the resolution of the virus resistance locus CMD2 in cassava
Luc Cornet, Syed Shan-e-Ali Zaidi, Jia Li, Yvan Ngapout, Sara Shakir, Loic Meunier, Caroline Callot, William Marande, Marc Hanikenne, Stephane Rombauts, Yves Van de Peer & Hervé Vanderschuren
Cassava is an important crop for food security in the tropics where its production is jeopardized by several viral diseases, including the cassava mosaic disease (CMD) which is endemic in Sub-Saharan Africa and the Indian subcontinent. Resistance to CMD is linked to a single dominant locus, namely CMD2. The cassava genome contains highly repetitive regions making the accurate assembly of a reference genome challenging. In the present study, we generate BAC libraries of the CMD-susceptible cassava cultivar (cv.) 60444 and the CMD-resistant landrace TME3. We subsequently identify and sequence BACs belonging to the CMD2 region in both cultivars using high-accuracy long-read PacBio circular consensus sequencing (ccs) reads. We then sequence and assemble the complete genomes of cv. 60444 and TME3 using a combination of ONT ultra-long reads and optical mapping. Anchoring the assemblies on cassava genetic maps reveals discrepancies in our, as well as in previously released, CMD2 regions of the cv. 60444 and TME3 genomes. A BAC-guided approach to assess cassava genome assemblies significantly improves the synteny between the assembled CMD2 regions of cv. 60444 and TME3 and the CMD2 genetic maps. We then performed repeat-unmasked gene annotation on CMD2 assemblies and identify 81 stress resistance proteins present in the CMD2 region, among which 31 were previously not reported in publicly available CMD2 sequences. The BAC-assessed approach improved CMD2 region accuracy and revealed new sequences linked to virus resistance, advancing our understanding of cassava mosaic disease resistance.
https://link.springer.com/article/10.1186/s13059-025-03620-8
From Cytoplasmic Membrane to Thylakoids: Evolution of Membrane Biogenesis and Photosystem II assembly in early-diverging Cyanobacteria
Louise Hambücken, Denis Baurain, Luc Cornet
Thylakoid membranes (TM) in cyanobacteria and chloroplasts host the light-dependent reactions of oxygenic photosynthesis which involve a linear electron transfer (LET) chain composed of multi- subunit complexes, including notably Photosystem II (PSII). Gloeobacterales, the earliest-diverging cyanobacterial lineage, lack TM and perform photosynthesis within specialized regions of the cytoplasmic membrane (CM), thereby representing an ancestral state with respect to other cyanobacteria, all equipped with TM and known as Phycobacteria. The emergence of TM, which increased the membrane surface available for oxygenic photosynthesis, was a key innovation that likely contributed to the Great Oxidation Event. This evolutionary transition involved the formation of a distinct membrane compartment, followed by the relocation of LET components from the CM to TM. Here, we present a phylogenomic analysis identifying three candidate proteins associated with membrane trafficking that may contribute to TM biogenesis, including the SPFH family member Slr1106, which we show was acquired via lateral gene transfer. Moreover, evolutionary analysis of 36 PSII assembly factors indicates key modifications in late-stage PSII assembly, notably in manganese homeostasis, and highlights structural changes in the early-acting YidC translocase that may have facilitated the relocation of LET components from the CM to TM. Altogether, our phylogenetic and functional prediction analyses of proteins involved in membrane dynamics and PSII assembly factors bring new insights into the molecular innovations that led to the emergence of TM.
https://doi.org/10.1101/2025.11.06.686923
A FAIR Amplicon Sequencing Workflow for Long-term Environmental Monitoring
Marie Harmel, Benoit Durieu, Valentina Savaglia, Bjorn Tytgat, Denis Baurain, Annick Wilmotte, Elie Verleyen, Luc Cornet
Antarctica represents one of the last pristine environments on Earth, providing a unique opportunity to to study the effects of climate change and anthropogenic activities. Ice-free areas, such as the inland nunataks of the Sør Rondane Mountains (SRM), host unique terrestrial and lacustrine ecosystems, of which the simplified food webs rely almost exclusively on microbial primary production. Because of their small size, low productivity and hence low biomass, these microbial communities are fragile. Seven SRM sites were selected to be part of the Antarctic Specially Protected Area (ASPA) 179. The MonASPA project has established an environmental monitoring program to evaluate the effectiveness of the management plan approved by the Antarctic Treaty System for these areas. A key component of MonASPA is the long-term monitoring of microbial biodiversity using 16S rRNA amplicon sequencing. To ensure consistent operation over decades, we developed the Reproducible Amplicon Sequencing Pipeline for Antarctic Monitoring (RASPAM), which is built on Apptainer containers and Nextflow workflows. RASPAM implements Amplicon Sequence Variants (ASVs) and Zero-radius Operational Taxonomic Units (ZOTUs) to provide high-resolution taxonomic affiliation. It incorporates, in addition to the SILVA database, a taxonomically curated 16S rRNA database for cyanobacteria and enables comparisons against NCBI databases to facilitate the identification of rare prokaryotic strains in environmental samples. RASPAM is Findable, Accessible, Interoperable, and Reproducible (FAIR) and represents a robust tool for long-term monitoring of microbial communities in Antarctic and other extreme environments.
https://doi.org/10.64898/2025.12.04.692289
Early-Diverging SQR Enzyme in Antarctic Gloeobacterales Indicates Sulfide Tolerance in Thylakoid-Lacking Cyanobacteria
Louise Hambücken, Edi Sudianto, Jimmy H. Saw, Denis Baurain, Luc Cornet
Oxygenic photosynthesis, which converts solar energy into carbohydrates via a linear electron transport chain and two photosystems (PSII and PSI), first appeared in cyanobacteria approximately 3.3 Ga and drove the Great Oxidation Event around 2.4 Ga. During this period, euxinic conditions—characterized by sulfidic, anoxic oceans—posed a metabolic challenge to cyanobacteria, as sulfide inhibits PSII, the reaction center responsible for water splitting. Here, we report the presence of an early-diverging form of the sulfide quinone reductase (SQR) enzyme in Antarctic representatives of Gloeobacterales, the earliest-branching cyanobacterial lineage lacking thylakoids. Phylogenetic analyses reveal that these SQR sequences are the earliest-diverging cyanobacterial SQR known to date, predating the multiple lateral gene transfer events previously observed in the phylum. Additional searches in metagenomic datasets indicate that such sequences are restricted to cold environments. Our findings unveil possible adaptive strategies of early cyanobacteria to cope with sulfidic stress and point to Antarctic lakes as preserved natural laboratories for investigating cyanobacterial diversification and the evolution of oxygenic photosynthesis under euxinic conditions.
https://doi.org/10.1101/2025.10.24.684318
Horizontal Gene Transfers Underpin Ribose Heterotrophy and Central Carbon Metabolism Remodeling in Gloeobacteraceae
Edi Sudianto, Denis Baurain, Luc Cornet
Gloeobacterales has long been considered a “living fossil” cyanobacterial order, owing to its lack of thylakoid membranes and basal phylogenetic position. However, our study reveals that Gloeobacterales actively integrate horizontally transferred genes into their core metabolism. In Gloeobacteraceae—one of the two families within the order—these genes encode a complete ribose ATP synthase binding cassette (ABC) importer and downstream enzymes, enabling the heterotrophic uptake of external ribose and its assimilation into central carbon metabolism, along with photosynthesis, indicative of photomixotrophy. Beyond ribose utilization, their central carbon metabolism exhibits a mosaic architecture shaped by the integration of foreign genes into the Calvin-Benson-Bassham cycle, the pentose phosphate pathway, and the Embden-Meyerhof-Parnas pathway. Uniquely, these genes appear to have been acquired through multiple independent transfer events, as reflected by their dispersed genomic locations and diverse bacterial donors, including other cyanobacteria and Pseudomonadota. These findings contradict the long-standing view of Gloeobacterales as metabolically primitive relics. Instead, Gloeobacterales is likely a dynamic lineage that continues to adapt and evolve through metabolic innovation and the assimilation of foreign genes into its genomes.
https://doi.org/10.1101/2025.11.10.686926
Taxonomic Description of Uncultured Cyanobacteria from Extreme Habitats through Genome-Based Classification
Edi Sudianto, Maximillian D. Shlafstein, Benoit Durieu, Marie Harmel, Luc Cornet, Jimmy H. Saw
Cyanobacteria form a morphologically and phylogenetically diverse group of oxygenic phototrophic bacteria inhabiting a wide range of environments, including extreme habitats such as hot springs and volcanic steam vents. Many lineages, particularly those from these extreme environments, remain uncultured and are known only from metagenome-assembled genomes (MAGs), limiting their integration into formal taxonomy. Analysis of 46 steam vent associated samples from Hawai‘i using 16S rRNA amplicon sequencing revealed that cyanobacteria dominate these communities. Gloeobacter kilaueensis dominated pit-like environments with low-light conditions, while Leptolyngbyaceae and other families are more dominant in structured soil and wall communities. We further reconstructed 38 high-quality cyanobacterial MAGs and incorporated them into a phylogenomic analysis comprising 343 cyanobacterial genomes, followed by genome-based comparisons against 9,026 reference genomes. This revealed eight novel species and one novel genus spanning five orders: Chroococcidiopsidales, Leptolyngbyales, Nostocales, Oculatellales, and Oscillatoriales. Following SeqCode guidelines, we provide the first formal taxonomic descriptions of cyanobacterial MAGs and propose guidelines for integrating genome-based and cultivated material. These findings highlight Hawaiian steam vents as hotspots of previously uncharacterized cyanobacterial diversity and underscore the importance of genome-based nomenclature.
https://doi.org/10.64898/2026.01.02.697360
Out of Antarctica: new insights into Antarctic Subcluster 5.2 picocyanobacteria based on high-quality genomes
Benoit Durieu, Valentina Savaglia, Mick Van Vlierberghe, Valérian Lupo, Denis Baurain, Annick Wilmotte, Luc Cornet
Synechococcus-like cyanobacteria are cosmopolitan unicellular picocyanobacteria that have colonized diverse aquatic and terrestrial habitats. The so-called ‘subcluster 5.2’ represents a particularly diversified subgroup, including marine and freshwater organisms adapted to extreme conditions, notably polar environments. We increased the genomic representation of polar taxa in this subcluster by reconstructing new high-quality genomes from five Antarctic lacustrine strains and one Arctic freshwater isolate using a combination of small Illumina and long Nanopore reads. A maximum likelihood (ML) phylogenomic analysis of these new assemblies combined with all publicly available good quality assemblies of the subcluster 5.2 suggests evidence of a dispersal process from Antarctica. Indeed, the topology of the phylogenomic tree indicates one basal Antarctic lineage followed by the emergence of two lineages, one Antarctic and one non-Antarctic (Spain). This finding is further supported by a 16S rRNA ML phylogenetic and a pangenomic analysis. Although secondary colonization of Antarctica by cyanobacteria following the cooling of the continent 34 million years ago has been reported, this study is the first to support an ‘Out-of-Antarctica’ scenario inside subcluster 5.2.
https://doi.org/10.64898/2025.12.17.694815
Rapid Radiations Outweigh Reticulations During the Evolution of a 750-Million-Year-Old Lineage of Cyanobacteria
Carlos J Pardo-De la Hoz , Diane L Haughland , Darcie Thauvette , Sydney Toni , Spencer Goyette , William White , Ian D Medeiros , Luc Cornet , Petr Dvořák , Diego Garfias-Gallegos , Jolanta Miadlikowska , Nicolas Magain , François Lutzoni
Species are a fundamental unit of biodiversity. Yet, the existence of clear species boundaries among bacteria has long been a subject of debate. Here, we studied species boundaries in the context of the phylogenetic history of Nostoc, a widespread genus of photoautotrophic and nitrogen-fixing cyanobacteria that includes many lineages that form symbiotic associations with plants (e.g. cycads and bryophytes) and fungi (e.g. cyanolichens). We found that the evolution of Nostoc was characterized by eight rapid radiations, many of which were associated with major events in the evolution of plants. In addition, incomplete lineage sorting associated with these rapid radiations outweighed reticulations during Nostoc evolution. We then show that the pattern of diversification of Nostoc shapes the distribution of average nucleotide identities (ANIs) into a complex mosaic, wherein some closely related clades are clearly isolated from each other by gaps in genomic similarity, while others form a continuum where genomic species boundaries are expected. Nevertheless, recently diverged Nostoc lineages often form cohesive clades that are maintained by within-clade gene flow. Boundaries to homologous recombination between these cohesive clades persist even when the potential for gene flow is high, i.e. when closely related clades of Nostoc co-occur or are locally found in symbiotic associations with the same lichen-forming fungal species. Our results demonstrate that rapid radiations are major contributors to the complex speciation history of Nostoc. This underscores the need to consider evolutionary information beyond thresholds of genomic similarity to delimit biologically meaningful units of biodiversity for bacteria.
https://doi.org/10.1093/molbev/msaf244
Melissospora conviva gen. nov., sp. nov., a novel actinobacterial genus isolated from beehive through cross-feeding interactions
Déborah Tellatin, Luc Cornet, Valdes Snauwaert, Philippe Compère, Marc Ongena, Loïc Quinton, Nudzejma Stulanovic, Silvia Ribeiro Monteiro, Augustin Rigolet, Pierre Burguet, Petra Van Damme, Lorena Carro and Sébastien Rigali
Most micro-organisms remain unculturable under standard laboratory conditions, limiting our understanding of microbial diversity and ecological interactions. One major cause of this uncultivability is the loss of access to essential cross-fed metabolites when bacteria are removed from their natural communities. During a bioprospecting campaign targeting actinomycetes of an Apis mellifera beehive, we identified five isolates (DT32, DT45T, DT55, DT59 and DT194) that required co-cultivation for growth recovery, suggesting a dependence on microbial interactions in their native habitat. Whole-genome sequencing and phylogenetic analysis positioned these isolates within a distinct lineage of Micromonosporaceae, separate from the five officially recognized clades of the Micromonospora genus. A combination of microscopic, chemotaxonomic and physiological characterizations further supported their uniqueness. Notably, they exhibited high auxotrophy, being unable to use all carbon sources tested, likely due to genome reduction (4.6 Mbp) compared to other Micromonosporaceae. Pangenomic comparisons with their closest Micromonospora relatives revealed gene losses in key metabolic pathways, including the glyoxylate bypass and the Entner–Doudoroff pathway, which may explain their metabolic reliance. These findings reveal a highly specialized, ecologically adapted lineage with deep evolutionary divergence and further support microbial interdependence isolation strategies to explore the microbial dark matter. We propose Melissospora conviva as a novel genus and species within the Actinomycetota phylum, with isolate DT45T as the representative type species and type strain, which has been deposited in public collections under the accession numbers DSM 117791 and LMG 33580.
https://doi.org/10.1099/ijsem.0.006868