Publications

4676

Rapid and accurate diagnosis of ‘Candidatus Phytoplasma’ is critical for disease management but remains challenging due to the pathogen's unculturable nature, low titers, uneven distribution and genetic complexity. While current official guidelines provide a highly adaptable taxonomic foundation for species assignment, translating these broad recommendations into standardized routine diagnostics remains difficult. This study implements the established species-assignment guidelines by developing a structured, proof-of-concept diagnostic framework utilizing a Multi-Locus Sequence Analysis (MLSA) workflow coupled with Oxford Nanopore Technologies (ONT) sequencing. The developed workflow targets four barcodes — 16S, secA, rplV and tufB — and replaces fixed demarcation criteria with a best-match principle for non-16S barcodes. Furthermore, the study offers guidance for barcode selection and challenges the use of 16S rRNA sequencing as a sole screening assay. Moreover, a read-based quantification system incorporating control thresholds was implemented to provide an objective and reliable assessment of the results. The workflow was validated using spiked dilution series and naturally infected samples, demonstrating that the ONT workflow achieves sensitivity and accuracy comparable to, or exceeding, that of traditional Sanger sequencing. Although PCR multiplexing was found to reduce sensitivity, limiting the immediate throughput advantages over Sanger sequencing, the proposed framework offers significant value through flexible species assignment and stringent contamination controls. Consequently, this method serves as a robust, high-resolution tool for resolving complex phytoplasma infections.

Plant-pathogenic bacteria threaten crop productivity and quality, yet chemical options remain limited compared with those for fungal and oomycete diseases. Current management relies mainly on copper bactericides, limited antibiotics, induced-resistance agents, biocontrol and resistant cultivars. However, copper and streptomycin resistance, efflux-mediated multidrug tolerance and rapid pathogen adaptation have weakened these strategies. Target-oriented research provides a framework for exploring agricultural antibacterials, anti-virulence agents, anti-colonization strategies, resistance sensitizers and host-resistance interventions, but many of these approaches remain conceptual, model-system, greenhouse or medical-bacteriology-derived rather than proven field solutions. This review classifies bacterial control targets into two interconnected groups: growth-essential targets, including peptidoglycan biosynthesis, membrane/envelope systems, nucleic-acid processes, protein synthesis, metabolism, nutrient transport and cell division; and anti-virulence/anti-adaptation targets, including secretion systems, quorum sensing, biofilms, motility, adhesion, cell-wall-degrading enzymes, tolerance systems, oxidative-stress responses and host susceptibility factors. Using “Candidatus Liberibacter asiaticus” (CLas) as a case study, genome annotation and infection-stage transcript-abundance data prioritized Sec-dependent secretion, outer-membrane/surface proteins, Bam assembly, nutrient transporters, Clp proteostasis, redox adaptation and core cellular processes as candidate target classes. Envelope-associated, secretion/anti-virulence, nutrient-acquisition and stress-sensitization modules may represent potential directions for downstream validation, but CLas candidates remain hypothesis-generating priorities requiring validation for essentiality, conservation, druggability, delivery feasibility, crop safety and field performance.

ABSTRACT We present the complete de novo assembly of a Candidatus Walczuchella monophlebidarum genome (286,606 bp), a flavobacterial endosymbiont of the giant-scale insect Icerya purchasi . The genome was assembled from metagenomic short read Illumina sequences obtained from DNA of citrus leaves collected in Carinthia, Austria in November 2024.

Abstract A new distribution map is provided for Candidatus Phytoplasma rubi Malembic-Maher et al. Mollicutes: Acholeplasmatales: Acholeplasmataceae. Hosts: Rubus spp.

Abstract Krum.hol.zi.bac.te.ri.o'ta. N.L. neut. n. Krumholzibacterium , type genus of the type class of the phylum; L. neut. pl. n. suff. ‐ota , ending to denote a phylum; N.L. neut. pl. n. Krumholzibacteriota , the phylum of the class Krumholzibacteria. The phylum Candidatus Krumholzibacteriota was proposed to accommodate genomic assemblies of yet‐uncultured microorganisms. The type species, of the genus, Candidatus Krumholzibacterium zodletonense Zgenome0171 T was recovered from an anaerobic, sulfide‐ and sulfur‐rich spring in Southwestern Oklahoma, USA. The phylum comprises one described class, order, family, genus, and species. However, currently available genomes ( n  = 90) in GTDB (release 08‐RS214) are accommodated in 1 class, 6 orders, 8 families, 30 genera, and 58 species. Phylogenetically, the phylum is closely related to Gemmatimonadota and multiple phyla with no cultured representatives ( Ca . Edwardsbacteria, Ca . Eisenbacteria, Ca . Latescibacterota, Ca . Zixibacteria, 4572‐55, ARS69, GCA‐2686955, JAAXHH01, JABDJQ01, JAGLTZ01, JAGLYR01, TA06, TA06_A, and VGIX01); all of which are part of the FCB ( Fibrobacteres , Chlorobia , Bacteroidetes ) superphylum in the Gracilicutes clade. Candidatus Krumholzibacteriota genomes have mainly been recovered from marine, freshwater, engineered, and host‐associated ecosystems. The prevailing in‐situ conditions in sampled habitats, gene content analysis, and the use of predictive machine learning‐based approaches suggest a phylum‐wide preference for anaerobic conditions, mesophilic temperatures, circumneutral pH, and low salinities. Metabolically, the type species of the type genus of the phylum is predicted to be a fermentative anaerobe with preference for amino acids over sugars as substrates. Later studies indicate that some members of the phylum harbor respiratory capacities (dissimilatory sulfate reduction, dissimilatory sulfite reduction, iron‐reduction, and high‐affinity oxygen respiration with a few members possessing the potential for low‐affinity oxygen respiration), while others possess the machinery for anaerobic degradation of aromatic hydrocarbons. Finally, although all members of Candidatus Krumholzibacteriota remain uncultured, a representative of the phylum was enriched in a methane‐ and iron‐fed bioreactor inoculated with Bothnian Sea sediments. Type genus : Candidatus Krumholzibacterium Youssef et al. 2019, CL2.

Abstract A new distribution map is provided for Candidatus Phytoplasma ziziphi Jung et al. Mollicutes: Acholeplasmatales: Acholeplasmataceae. Main host: jujube (Ziziphus jujuba), also other stone fruits.

Abstract He.pat.in'co.la. Gr. neut. n. hêpar (gen. hêpatos) , liver; L. masc./fem. n. incola , inhabitant, dweller; N.L. masc. n. Hepatincola , a dweller of the liver. Pseudomonadota / Alphaproteobacteria / Rhodospirilalles / Candidatus Hepatincolaceae / Candidatus Hepatincola Candidatus Hepatincola is a genus of uncultured, obligately host‐associated bacteria originally discovered in the lumen of the midgut glands (hepatopancreas) of terrestrial isopods. In this niche, the bacteria exhibit a characteristic morphology: elongated rods with a stalk‐like appendage at one pole, which can be inserted between the microvilli of the midgut epithelium. All known members of the genus belong to two species ( Ca . Hepatincola porcellionum and a yet to be named Ca . Hepatincola sp.) associated with terrestrial isopods. However, closely related genera from the same family have been observed in numerous aquatic and terrestrial members of the Ecdysozoa (molting invertebrates). Ca . Hepatincola and closely related genera all have reduced genomes of about 1.3 Mb with low DNA G + C content ( c . 30%) and extremely limited biosynthetic capacities but rich in transporters to import essential nutrients and precursors from the environment. In line with its ecological niche (the host gut lumen, a nutrient‐rich environment) and genomic characteristics, Ca . Hepatincola is rather a nutrient scavenger than a nutrient provider for the host. Ca . Hepatincola was initially classified within the order Rickettsiales and later within the Holosporales , but recent phylogenomic analyses support its phylogenetic placement within the order Rhodospirillales . DNA G + C content (mol % ) : 30. Type species : Candidatus Hepatincola porcellionum Wang et al. (2004), CL1.