Publications

4994

Candidatus Liberibacter spp. can infect most citrus plants and rely entirely on phloem sieve tube cells of the host plant for survival. Candidatus Liberibacter primarily contains Ca. L. asiaticus (CLas), Ca. L. africanus (CLaf), and Ca. L. americanus (CLam). Among these, CLas is the most harmful and widely distributed and is the primary pathogen of the devastating citrus disease Huanglongbing (HLB). Effectors are among the core weapons secreted by pathogens into plant cells to attack the plant immune system. In this study, we focused on CLas-specific effectors and those that are highly expressed during the infection stage to identify essential virulence effectors. Using secretion signal peptide prediction analysis, 40 candidate effectors with potential secretory capabilities were identified. Transient expression of these candidate effectors in Nicotiana benthamiana revealed their impact on pattern-triggered immunity, including INF-induced cell death and microbial pattern-induced reactive oxygen species (ROS) bursts, and the resistance of N. benthamiana to the bacterial pathogen Pst DC3000. 10 candidate effectors capable of suppressing plant immunity were identified. The stable expression of these candidate effectors in Arabidopsis showed that several candidate effectors enhanced plant susceptibility to Pst DC3000 and inhibited flg22-induced ROS production and MAPK activation. Among the three candidate effectors that significantly suppressed ROS burst, one effector, E3 (CLIBASIA_03085), interacts with the plant NADPH oxidase RbohD, a key enzyme responsible for ROS production. This suggests that E3 likely inhibits ROS accumulation by directly targeting RbohD. Here, we identified multiple candidate effectors capable of suppressing microbial pattern-triggered immunity that may be essential virulence factors for CLas infection, enhancing our understanding of CLas pathogenesis.

‘Candidatus Phytoplasma solani’ of the 16SrXII group is an emerging vector-borne pathogen in European crop production. The cixiid planthopper Hyalesthes obsoletus transmits 16SrXII-A stolbur phytoplasmas that are associated with diseases in grapevine, potato, and various weeds. While 16SrXII-P genomes transmitted by Pentastiridius leporinus are available, no genome of an H. obsoletus-transmissible 16SrXII-A phytoplasma has been reported from Germany. Here, we present insights into the phylogenetic position and pathogen–host interactions through the functional reconstruction of the complete 832,614 bp genome of the H. obsoletus transmissible ‘Ca. P. solani’ 16SrXII-A strain POT from a potato field. Phylogenetic analyses highlight the heterogeneity within the stolbur group using whole-genome alignment and a BUSCO-based core gene analysis approach. The POT chromosome shares highest average nucleotide identity with Italian bindweed-associated genomes and displays strong synteny with the c5 strain. Consistent with the typical phytoplasma architecture, the POT genome combines mobile-element-driven instability with a conserved core metabolism. Virulence factors include transposon-linked effectors but lack pathogenicity island organisation. POT further differs from other 16SrXII-group phytoplasmas through unique collagen-like proteins that could contribute to virulence. These findings provide a robust genomic framework that improves diagnostics, enables strain-level resolution and supports the assessment of breeding materials under stolbur phytoplasma pressure, thereby refining our understanding of stolbur phytoplasma diversity and highlighting the evolutionary divergence within the 16SrXII subgroup.

Abstract Methanogens are strictly anaerobic archaea capable of energy conservation by methane production, yet their presence in oxic and arid environments challenges existing paradigms. In this study, we enriched and genomically characterized seven methanogenic cultures from desert biocrusts, affiliated with the genera Methanobacterium, Methanosarcina, and Methanocella. Six of these new enrichment cultures represent new species. Nonetheless, phylogenomic analyses revealed close genetic relationships with organisms from anoxic environments, indicating the absence of an evolutionary distinction. Comparative genomics exposed diverse though non-unique repertories of antioxidant (e.g. catalase, superoxide dismutase and desulfoferrodoxin), and desiccation-resistance genes (including genes for maintaining osmotic pressure and repair of cell wall and membrane), with Methanobacterium spp possessing the lowest gene abundance and diversity for oxygen and desiccation tolerance. Nevertheless, the occurrence of a Class I methanogen such as Methanobacterium in arid soils challenges the notion that members of this class are less oxygen tolerant than Class II. Pangenome analysis further uncovered unique genes enriched in membrane-associated functions and potentially non-functional stress-related genes. Via a global metagenomic survey we find that methanogens are underdetected in dryland soils, likely due to sequencing depth limitations. Our findings highlight previously overlooked methanogen diversity and ecological plasticity in oxic and desiccated habitats, and emphasize the need for further studies to elucidate their survival strategies.

Huanglongbing (HLB), caused by Candidatus Liberibacter asiaticus (CLas), is the most destructive citrus disease worldwide, severely reducing yield and fruit quality. Although no naturally resistant cultivars are available, citrus plants overexpressing Arabidopsis NPR1 ( At NPR1) display enhanced tolerance, yet the molecular mechanisms underlying this phenotype remain insufficiently understood. To uncover early transcriptional events associated with tolerance, we performed comparative RNA-seq and phytohormone profiling of susceptible wild-type (WT) and tolerant At NPR1-overexpressing ( At NPR1-OE) Duncan grapefruit at 0, 6, and 24 hours post infection (hpi). Before infection, At NPR1 plants downregulated genes involved in cytoskeleton organization, cell wall biogenesis, and receptor signaling, suggesting a primed basal defense state. After CLas exposure, At NPR1 plants exhibited stronger and earlier transcriptional reprogramming, with substantially more differentially expressed genes at 6 hpi than WT. At 24 hpi, At NPR1 plants showed suppression of callose synthase genes and selective induction of β-1,3-glucanases, indicating more controlled phloem callose regulation. Concurrently, attenuated expression of respiratory burst oxidase homologs and ROS-associated genes suggested a moderated and less damaging oxidative burst. At NPR1 plants maintained stable levels of salicylic acid, and gibberellins while preventing the CLas-induced induction of abscisic acid observed in WT. Collectively, these findings reveal that At NPR1 overexpression enhances HLB tolerance by integrating early transcriptional reprogramming with balanced structural, oxidative, and hormonal responses. This study provides a mechanistic framework for understanding NPR1-mediated tolerance to CLas during the initial stages of infection.

Abstract Ferdinand Cohn observed abundant filamentous bacteria in drinking water wells in 1870 that he named Crenothrix polyspora. Subsequent research has revealed the methanotrophic metabolism of Crenothrix bacteria and their disproportionately high activity in stratified lakes compared to unicellular methanotrophs, yet laboratory cultivation has proven elusive, leaving the ecophysiology of Crenothrix bacteria largely unknown. Here we report the isolation of two methanotrophic strains of the “lacustrine Crenothrix” clade from an iron-rich wetland and reveal their unique cell biology and ecology. We demonstrate that the strains are microaerobic and grow as filaments of cells, which are connected by unidirectionally oriented structures. The strains have broad genomic repertoires for addressing O2 limitation that are uniquely associated with lacustrine Crenothrix compared to related clades based on genome data. Aligning with laboratory observations, we identify lacustrine Crenothrix bacteria along potential redox gradients in the wetland at iron-rich snow sites, and we also detect such bacteria in diverse global ecosystems based on public metagenome searches. Together, our data strongly point to an ecophysiology of lacustrine Crenothrix bacteria that is tightly linked to O2 limitation, and we propose that the strains uniquely store or share metabolic intermediates between cells in filaments to thrive under such conditions. Our cultivation-based findings for these strains, which we name Allocrenothrix methanica, provide new insights into the diversity, evolution, and ecology of filamentous methanotrophs, connecting over 150 years of microbiology research and opening vast new opportunities to investigate bacteria contributing to the global methane cycle under O2 limitation.

Abstract Members of the bacterial family Endozoicomonadaceae are ubiquitous marine symbionts associated with diverse hosts, including corals, sponges, molluscs, and fishes. Despite their ecological pervasiveness, inhabiting hosts in deep-sea vents and sunlit coral reefs, their taxonomy remains inconsistent, largely due to reliance on 16S rRNA gene phylogenies that fail to fully resolve evolutionary relationships. Here, we integrate phylogenomics, comparative genomics, and overall genome relatedness indices across 63 high-quality genomes to re-define genus boundaries within the Endozoicomonadaceae family. Among the metrics, the core-proteome average amino acid identity proved the most robust for genus-level resolution, with a proposed identity cutoff of 76%, aligning with phylogenomic structure and maintaining taxonomic robustness across the family. Phylogenomic reconstructions further revealed that the taxon Endozoicomonas is paraphyletic, forming two well-supported but distinct clades. This prompts a taxonomic revision, dividing Endozoicomonas into Endozoicomonas sensu stricto and Neoendozoicomonas gen. nov., with Neoendozoicomonas montiporae designated as the type species. Furthermore, the sister clade to Endozoicomonas sensu stricto—comprising the genera Endonucleibacter and Sororendozoicomonas—is characterized by genomic streamlining, including reduced genome size, lower GC content, reduced number of orthogroups, and potential functional divergences consistent with host specialization. Functional annotations highlighted secretion and conjugation systems as key differentiators between genera, emphasizing potential distinct host-interaction strategies. This genome-based framework refines the taxonomy of the Endozoicomonadaceae, provides additional criteria for genus delimitation, and strengthens the evolutionary and ecological interpretation of these widespread marine bacterial symbionts.

Abstract Dermatophilaceae polyphosphate-accumulating organisms (PAOs), formerly classified as Tetrasphaera PAOs, play pivotal roles in enhanced biological phosphorus removal (EBPR). However, their phylogenetic diversity, ecological preferences, and metabolic traits remain poorly characterized, and a robust marker gene for their classification is lacking. Here, we performed an extensive phylogenomic and metabolic analysis of Dermatophilaceae PAOs utilizing 46 newly recovered metagenome-assembled genomes from a laboratory-scale EBPR reactor treating high-strength wastewater and full-scale wastewater treatment plants. These analyses revealed a previously uncharacterized PAO genus, named here as Candidatus Dermatophostum, which shows specific preference for high-phosphorus environments. Its representative species, Ca. Dermatophostum ammonifactor, was enriched in the EBPR reactor and its PAO phenotype was confirmed by polyphosphate staining and fluorescence in situ hybridization. Integrative meta-omics combining genomic, transcriptomic, and protein structure analyses revealed its specialized metabolic capabilities for phosphate metabolism, glycogen synthesis, and dissimilatory nitrate reduction to ammonium. Moreover, Ca. Dermatophostum was found to be widely distributed across wastewater treatment plants worldwide, underscoring both its diverse metabolic capabilities and potential engineering implications for mitigating nitrous oxide (N2O) emissions for EBPR system. Finally, we propose a ppk1-based classification framework that resolves Dermatophilaceae PAOs into six distinct clades, consistent with whole-genome phylogeny, and demonstrates that ppk1 can serve as a reliable marker gene for tracking these populations. Together, these findings expand the ecological and functional understanding of Dermatophilaceae PAOs and highlight their promise for advancing sustainable wastewater treatment and resource recovery.

Abstract Sandy sediments of the coastal ocean are generally not considered a habitat for highly active sulfate-reducing microorganisms (SRM) due to fluctuations in oxygen concentrations and carbon availability. This study investigated the taxonomic composition and gene expression patterns of sulfate-reducing communities in both sandy, organic-poor sediments and muddy, organic-rich sediments in the mesohaline southern Baltic Sea. Our results revealed that a novel order-level lineage within the class Thermodesulfovibrionia (phylum Nitrospirota), named Arenidesulfomonadales, contributed significantly to sulfate reduction in the sandy sediments and supported sulfate reduction rates similar to those in the organic-rich, muddy sediments. In the sands, Thermodesulfovibrionia effectively competed with Desulfobacterales, which strongly dominated the sulfate-reducing communities in the organic-rich sediments. Metatranscriptomic data revealed that Desulfobacterales functioned as typical secondary fermenters in both sediment types and expressed nearly complete sets of genes for the catabolism of fermentation products such as lactate, propionate, and butyrate. In contrast, the gene expression profile of Thermodesulfovibrionia suggests that hydrogenotrophic sulfate reduction, coupled with carbon fixation via the Wood–Ljungdahl pathway and a reduced capacity to oxidize organic compounds, was a competitive metabolic strategy enabling sulfate reduction to proceed independently of fermentation-derived carbon. Overall, our results suggest that overlooked lineages of SRM may contribute to higher levels of sulfate reduction than previously assumed in the extensive areas of permeable sediments of coastal and shelf seas.