Publications

4560

Abstract Citrus Huanglongbing (HLB) is the most destructive disease in citriculture, mainly caused by Candidatus Liberibacter asiaticus (CLas). However, the immune response of citrus to CLas at the cellular level remains to be elucidated. In this study, the first single-cell atlas of rough lemon (Citrus jambhiri Lush.) root apexes were generated using single-nucleus RNA sequencing at 20 weeks post-inoculation with CLas. According to gene expression patterns, the single-cell atlas was partitioned into 20 transcriptionally distinct clusters, and five cell types were identified within these clusters. A significant number of defense-related genes were co-upregulated across the five cell types following CLas infection, whereas genes involved in signal transduction pathways, such as tubulin beta-6 chain (TUBB1) and the phospholipase D alpha 1 (PLD1), were concurrently downregulated. Based on pseudotime trajectory analysis, the key pathways and genes involved in the coordination of cell differentiation and resistance in citrus under CLas infection were characterized. Following CLas infection, the development of phloem cells was significantly delayed, and the differentiation of cambium cells into xylem cells was evident. The expression of genes associated with lignin synthesis was significantly upregulated in these cells. The reduction in phloem cell differentiation and the enhanced differentiation of cambium cells into defense-related xylem cells may represent the primary vascular immune mechanisms exhibited by citrus plants in response to CLas infection. Additionally, DNA-binding one zinc finger transcription factor DOF2.4 was found to potentially serve dual roles in regulating vascular cell development and inducing plant resistance against CLas. In conclusion, this study collectively provides insights into the cellular innate immunity responses of citrus to CLas infection. These findings hold significant implications for the sustainable development of citriculture amidst the ongoing global HLB epidemic, and offer novel insights into vascular immunity and plant defense responses.

Abstract Numerous cyanobacterial strains previously identified as Scytonema hyalinum were determined to be phylogenetically distant from the type species of Scytonema, S. hofmannii . Morphological and molecular evidence suggests this distinct clade necessitates placement in a new genus, and we have described Kalymmatonema gen. nov. herein. Kalymmatonema has been demonstrated to exhibit five ribosomal operons, all of which differed in both sequence and secondary structure of conserved helical domains in the 16S–23S internal transcribed spacer rRNA region. Four of these operon copies were highly similar in 16S and 23S rRNA gene sequences, whereas the divergent fifth copy is thought to represent a whole‐operon horizontal gene transfer event. Through in‐depth analysis, we were able to recognize seven species new to science, the type species K. desertorum sp. nov., K. arcangelii comb. nov., K. chimaera sp. nov., K. ethiopiense sp. nov., K. gypsitolerans sp. nov., K. mateoae sp. nov., and K. oahuense sp. nov. We also created the new combination, K. hyalinum comb. nov., in order to include the original Scytonema hyalinum in this new genus based upon the common morphological feature of a mucilaginous apical cap on the trichomes. Kalymmatonema displays a complex evolution of its ribosomal operons, with evidence not only of horizontal gene transfer but also of internal rearrangements and mobile genetic elements that have transposed the tRNA‐containing region of the ITS rRNA region among the four similar operons. Additional investigation of the evolutionary history of this interesting genus will likely lead to a better understanding of the processes shaping ribosomal evolution in cyanobacteria.

ABSTRACT Insects of the suborder Auchenorrhyncha harbour multiple ancient endosymbionts that jointly produce essential nutrients lacking from the host's diet. Compared to cicadas, leafhoppers, and spittlebugs, our understanding of the multipartite symbioses among planthoppers, an extremely diverse insect group, is still very limited. Herein, we assembled the genomes of the primary endosymbionts of two planthopper species from the Cixiidae family, Cixius wagneri and Pentastiridius leporinus , both vectors of phytopathogenic Arsenophonus in Europe. Each species harboured a different tripartite endosymbiont consortium: while P. leporinus carried the well‐known combination ‘ Candidatus Karelsulcia muelleri’, ‘ Ca . Vidania fulgoroideae’, and ‘ Ca . Purcelliella pentastirinorum’, C. wagneri harboured a yet unknown Gammaproteobacterium in addition to Karelsulcia and Vidania . This new endosymbiont ‘ Ca . Mirabilia symbiotica’ is likely much older than Purcelliella , considering its extremely reduced genome. In both species, Karelsulcia and Vidania jointly produce the 10 essential amino acids, whereas Purcelliella and Mirabilia provide the non‐essential amino acid cysteine and slightly different gene sets encoding B vitamins. Our findings confirm the functional stability of multipartite planthopper endosymbiont consortia despite changing partners over evolutionary time. In addition, we describe a new Rickettsia strain from the Meloidae group colonising P. leporinus , highlighting the diversity of bacterial endosymbionts associated with planthoppers.

ABSTRACTSesame phyllody, a destructive disease caused by phytoplasma infection, induces severe morphological abnormalities, including floral virescence, phyllody, witches' broom, leaf deformation, and stunted growth. This study aimed to characterize phytoplasma isolates from diverse regions of India, identifying them as Candidatus Phytoplasma asteris (16Sr‐I), Candidatus Phytoplasma citri (16Sr‐II), and Candidatus Phytoplasma australasia (16Sr‐II). Whole‐genome sequencing of Candidatus Phytoplasma asteris isolate SPGN revealed a genome size of 563,754 bp, encoding 542 proteins, including several genes associated with antibiotic resistance. Effector prediction analysis identified key virulence‐associated proteins, such as SAP50‐like, SAP34‐like, TENGU‐su inducer, and immunodominant membrane proteins, which manipulate host development and immune responses. Transcriptomic analysis of infected sesame plants revealed significant gene expression alterations, with upregulated genes linked to floral malformation, vascular tissue modifications, and stress responses, while the downregulated genes were associated with flavonoid metabolism and immune signaling. Phytoplasma infection disrupted hormonal pathways, leading to increased expression of auxin, cytokinin, and gibberellin‐related genes, suggesting hormonal dysregulation as a key factor in symptom development. Furthermore, immune suppression was evident through the downregulation of key defense‐related genes, including those involved in MAPK signaling and pathogenesis‐related protein families. These findings enhance our understanding of phytoplasma pathogenesis in sesame and provide potential targets for developing effective disease management strategies.

AbstractBACKGROUNDAutophagy is a conserved mechanism by which eukaryotic organisms defend against pathogen infection. However, the molecular mechanisms underlying the role of autophagy in the interactions of insect vectors with the phloem‐limited bacterial pathogen remain unclear. The citrus Huanglongbing (HLB)‐associated pathogen ‘Candidatus Liberibacter asiaticus’ (CLas) seriously endangers development of the citrus industry. It spreads via Diaphorina citri in a persistent and propagative manner.RESULTSIn this study, a total of 30 autophagy‐related genes (ATG) were identified in the D. citri genome, among which multiple genes were significantly regulated after CLas infection. Concurrently, CLas infection also leads to an increased number of autophagosomes and enhanced accumulation of ATG8‐II. Ultrastructural observations revealed the presence of bacterial‐like structures within autophagosomes in the midgut of CLas‐infected D. citri. Furthermore, both activation and inhibition of autophagy significantly influenced CLas titers. However, autophagy cannot completely eliminate CLas in D. citri. We identified a CLas effector, SDE4040 (CLIBASIA_04040), that interacts with DcATG8 and co‐localizes on autophagosomes in D. citri. Co‐expression of SDE4040 and DcATG8 induces autophagy in Spodoptera frugiperda (Sf9) cells.CONCLUSIONTaken together, these results indicate that CLas infection activates the autophagy pathway in D. citri, contributing to a reduction in bacterial titer. Our data also revealed that CLas may trigger complex interactions with the insect. © 2025 Society of Chemical Industry.

Abstract Citrus greening disease, caused by ‘Candidatus Liberibacter asiaticus’, severely impacts citrus production worldwide. The development of sustainable control strategies for this disease is restricted by the unavailability of the bacterium in pure culture. Herein, the metabolic profile of the waxworm larvae, Galleria mellonella, was compared to that of Diaphorina citri, the vector of ‘Ca. L. asiaticus’. Our findings showed that G. mellonella larvae possess the nutritional needs to host ‘Ca. L. asiaticus’, supporting its short-term persistence, and responds to infection with a visible immune reaction by producing melanin upon bacterial invasion. The inoculated larvae exhibit detectable bacterial titers for up to four days when inoculated with infected citrus phloem sap or D. citri haemolymph, after which bacterial titers decline, and infected larvae show reduced survival compared to mock-treated and ‘Ca. L. asiaticus’-free controls. Metabolic profiling of G. mellonella, D. citri, and honeybees (Apis mellifera) reveals distinct chemical compositions in their haemolymph. G. mellonella contains higher levels of amino acids, organic acids, nucleotides, and sugar-nucleotides, providing essential nutrients for ‘Ca. L. asiaticus’, while D. citri is enriched in monosaccharides and sugar-alcohols. Citric acid was detected exclusively in the haemolymph of G. mellonella. These findings suggest G. mellonella as a convenient model that can transiently host ‘Ca. L. asiaticus’ for short-term use, which would facilitate high-throughput screening of antimicrobial compounds against ‘Ca. L. asiaticus’, as well as exploring host-pathogen interactions. This model could accelerate the development of effective treatments against citrus greening and inform broader strategies for managing vector-borne plant diseases.