Scientific Reports

Abstract Phytoplasmas are wall-less plant pathogens characterized by highly reduced genomes and limited metabolic capabilities, leading to an obligate host dependency. Since phytoplasma axenic cultivation has not yet been achieved, genomic approaches are essential to unravelling their biological and pathogenic traits. However, such insights are currently hindered by the scarcity of complete genomes and the lack of data for several clades, as is the case for the quarantine-relevant ‘ Candidatus Phytoplasma phoenicium’ (group 16SrIX-B), the causal agent of the severe stone-fruit disease almond witches’ broom (AlmWB). In this study, we present the complete genome sequence of the Lebanese strain F1A, which consists of a 552,248 bp chromosome with a 24.29% GC content and encodes 450 protein-coding genes. Comparative analyses with 16SrIX group draft genomes revealed high conservation among Lebanese AlmWB strains; however, comparisons with related 16SrIX-C phytoplasmas were limited by their low assembly completeness. Beyond providing a complete reference genome for this phytoplasma group, the F1A chromosome revealed distinct metabolic features. Strain F1A lacks the upper glycolytic pathway, indicating a metabolism that relies on glycerol-3-phosphate uptake and carboxylic acid fermentation. Notably, in addition to the conserved malate-to-acetate pathway, F1A encodes a complete citrate lyase complex, suggesting the potential for citrate utilization. Phylogenetic analysis of the associated 2-hydrocarboxylate symporter revealed the widespread occurrence of two phylogenetically distinct variants in phytoplasmas. Apart from the deduced metabolic capacities, the predicted effector repertoire shows similarities with those of other phytoplasmas, including proteins associated with branch proliferation and witches’ broom symptoms. Overall, these findings link genome reduction to host-dependent carboxylic acid metabolism in a quarantine-relevant phytoplasma, establishing a framework for comparative and functional studies of 16SrIX phytoplasmas and providing a basis for future investigations of AlmWB ecology and pathogenicity.

Abstract A comprehensive polyphasic taxonomic strategy was applied to the systematic characterization of strain DNPG3 T , which was isolated from the river Ganges, Hooghly, West Bengal, India. The Gram-positive, halotolerant, heavy-metal-tolerant strain exhibited the ability to degrade p -nitrophenol (PNP). Cellular fatty acid analysis revealed that the predominant components were anteiso-C 15:0 (24.61%), C 11:0 (21.06%), iso-C 16:0 (11.89%), C 16:0 (11.58%), and anteiso-C 17:0 (11.24%). Notably, the presence of C 11:0 , C 10:0 2-OH as major fatty acids differentiate strain DNPG3 T from its closely related members of the genus Brachybacterium . The predominant respiratory quinone was identified as menaquinone-7 (MK-7). Analysis of 16S rRNA gene sequence indicated that B. zhongshanense strain JB T was the closest relative of DNPG3 T , sharing 97.08% sequence similarity. Genome-based ANI value calculated using the EzBioCloud server revealed that B. zhongshanense JCM 15471 T was the closest genomic relative (85.49%). These values were further substantiated by digital DNA–DNA hybridization (dDDH) estimates calculated using the GGDC server. Taxonomic assignment using the GTDB database further indicated that strain DNPG3 T constitutes a previously unrecognized species within the genus Brachybacterium . Genome analysis of strain DNPG3 T identified eleven genomic islands, along with a rich repertoire of 194 carbohydrate-active enzyme (CAZyme) families, comprising 95 glycoside hydrolases and 53 glycosyltransferases. In addition, five biosynthetic gene clusters were detected. Collectively, these genomic features indicate the involvement of horizontal gene transfer events and highlighted the pronounced metabolic versatility of the strain, underscoring its potential for industrial enzyme production and secondary metabolite biosynthesis. Pan-genome analysis further indicates that the Brachybacterium pan-genome is open, reflecting substantial genetic diversity and ongoing gene acquisition within the genus. Comprehensive biochemical, physiological, chemotaxonomic, and phylogenetic analyses supported the assignment of strain DNPG3 T to the genus Brachybacterium while clearly distinguishing it from all currently described species within the genus. Accordingly, strain DNPG3 T was proposed to represent a novel species, for which the name Brachybacterium netajii sp. nov. is suggested. The type strain was DNPG3 T (= MTCC13125 T ).

Abstract The presence of fastidious phytopathogenic bacteria ‘Candidatus Arsenophonus phytopathogenicus’ and ‘Candidatus Phytoplasma solani’ challenges sugar beet cultivation in Austria. Reptalus artemisiae (Becker, 1865) (referred before as Reptalus quinquecostatus sensu Holzinger et al. 2003) is the principal vector of ‘Ca. P. solani’ to sugar beet in Serbia, while Pentastiridius leporinus is the primary vector of ‘Ca. A. phytopathogenicus’ in western Europe and can also transmit ‘Ca. P. solani’. This study investigates the epidemiology of the associated diseases in Austrian sugar beet fields. An extensive survey in 2024 at 33 locations revealed significant spatial and temporal variability in cixiid populations, with R. artemisiae emerging as the predominant species. Transmission trials and field surveys confirmed R. artemisiae as a vector of ‘Ca. A. phytopathogenicus’ and ‘Ca. P. solani’ 16SrXII-P driving a disease outbreak in Nikitsch. The predominance of R. artemisiae in nearly all monitored fields suggests its significant role in the pathogens’ epidemiology on a broader scale in the Pannonian plain. Spatial and temporal variations in pathogen incidence in Austrian sugar beet mirror patterns in Serbia, driven by R. artemisiae. The co-occurrence of ‘Ca. A. phytopathogenicus’ and ‘Ca. P. solani’ 16SrXII-A and 16SrXII-P and their transmission by two vectors complicate disease management efforts.

Abstract The genus “Candidatus Kouleothrix” (Eikelboom type 1851) is the major filamentous bacterium responsible for activated sludge bulking in Japanese activated sludge plants, where it is present in many in unusually high abundances. Global surveys have shown this genus embraces several species, although the contribution each of these makes to bulking is not known. This study followed their population dynamics in a full-scale bulking sludge plant in Japan over a 12-month period, which required the development of species-specific FISH probes, used to measure their relative abundances. Six of the 9 recognised species of “Ca. Kouleothrix” were detected there, two of which (midas_s_3423 and midas_s_35412) were at consistently high abundances and responsible for increases in the sludge volume index (SVI). Their abundances were also statistically correlated with their filament lengths, which showed higher correlations with SVI increases than did species abundances. This study examined the impacts of both operational parameters and other communities on the abundances of these two species over the 12-month period and showed marked interspecies differences in responses to both. Together these findings stress the importance in studies of this kind to identify populations to species level to reveal possible important ecological differences not seen at the genus level.