Publications

4474

ABSTRACT The functional roles of bacterial symbionts associated with microalgae remain understudied despite the importance of microalgae in biotechnology and environmental microbiology. 16S rRNA gene sequencing was conducted to analyze bacterial communities associated with two microalgae optimized for growth with flue gas containing 5%–10% CO 2 . Two dominant bacteria with no taxonomic classification beyond the class level (Paceibacteria) were discovered repeatedly in the most productive algal cultures. Long-read metagenomic sequencing was conducted to yield high-quality metagenomes, from which two novel species were discovered under the Seqcode (seqco.de/r:ywe1blo2), Phycocordibacter aenigmaticus gen. nov. sp. nov. and Minusculum obligatum gen. nov. sp. nov. The genus Phycocordibacter gen. nov. was proposed as the nomenclatural type of the family Phycocordibacteraceae fam. nov. and the order Phycocordibacterales ord. nov. Both bacteria possessed features typical of Patescibacteria such as reduced genomes (<800 kbp), lack of complete glycolysis and tricarboxylic acid (TCA) cycle pathways, and inability to synthesize amino acids. Instead, they rely on the reductive pentose phosphate pathway (Calvin cycle) for essential biosynthesis and redox balance. P. aenigmaticus may also rely on elemental sulfur oxidation ( sdo ), partial nitrite reduction ( nirK ), and sulfur-related amino acid metabolism (SAMe → SAH). Both bacteria were found in high relative abundance in cultures of Tetradesmus obliquus HTB1 (freshwater) and Nannochloropsis oceanica IMET1 (marine), suggesting a tight association with microalgae in various environments. The absence of full metabolic pathways for energy production suggests extreme metabolic limitations and obligate symbiosis, most likely with other bacteria associated with the microalgae. IMPORTANCE To our knowledge, this is the first report of Patescibacteria as dominant bacteria associated with microalgae or within a biologically mediated carbon capture system. Two novel Patescibacteria were found in two ecologically distinct microalgal cultures (one freshwater strain and one marine) regardless of whether the cultures were bubbled with air, 5% CO 2 , or 10% CO 2 . This unexpected and unprecedented dominance led to long-read sequencing and the assembly of high-quality metagenomes for both Patescibacteria, as well as five other bacteria in the system. The discovery of two novel species belonging to two novel genera, one novel family, and one novel order has enabled us to fill in gaps of a major, uncharacterized branch within the bacterial tree of life. Additionally, the extreme gene loss found in both Patescibacteria, Phycocordibacter aenigmaticus and Minusculum obligatum , contributes knowledge to a rapidly advancing body of research on the scavenging metabolic nature of this enigmatic and largely unclassified phylum.

Citrus greening or Huanglongbing (HLB) is a rapidly spreading disease caused by Candidatus Liberibacter species, which inhabit the phloem of citrus plants. It is primarily transmitted by psyllid insects. Managing HLB is challenging due to the bacterium's phloem-restricted nature, making diagnosis and treatment with antimicrobials difficult. Here in the present study, the bacterial consensus sequences were used for the primer design with In silico approaches and validated by PCR. The study was carried out in the Malwa region of Madhya Pradesh, India, which is known for citrus cultivations and remains affected by the HLB disease. Phylogenetic analysis of 100 Candidatus Liberibacter 16S rRNA sequences from NCBI was conducted using MEGA 6, with Malwa-specific refinement via MAFFT alignment. Eight previously deposited sequences formed the basis for the study. A consensus sequence was derived and used to design region-specific primers with GeneFisher. PCR validation was carried out with standard thermal cycling and gel electrophoresis. Phylogenetic analysis of 100 Candidatus Liberibacter 16S rRNA sequences from NCBI was conducted using Phylogenetic analysis showed high genetic variation among Liberibacter strains, supporting the need for region-specific detection. Eight sequences from the Malwa region were aligned using BioEdit, confirming conserved primer sites. Consensus primers were designed using GeneFisher and optimized for 16S rRNA targets. PCR validation identified 22 positives out of 100 samples, with primer set 2 detecting all target species effectively. The study concludes that primer set 2 enables rapid and accurate detection of HLB-causing bacteria in the Malwa region.

Cyanobacteria biodiversity remains underexplored despite their ecological importance and potential applications. To address this, we investigated two Leptolyngbya-like strains, LEGE 07170 and LEGE 191244, collected from marine tide pools in Portugal and Morocco using polyphasic approach. Phylogenetic analyses indicates that the strains form a distinct clade with strong statistical support in the Oculatellales order. The 16S rRNA gene identity matrix shows that the maximum shared values with the phylogenetically closest genera Gansulinema, Kaiparowitsia, Shahulinema, Aerofilum and Thermoleptolyngbya is consistently below 94.5%. Morphologically, LEGE 07170 and LEGE 191244 are indistinguishable to each other. Both optical and TEM analyses showed vacuole-like structures at the cells cross-walls and this character morphologically distinguishes these strains from their phylogenetically related genera. The 16S–23S ITS secondary structures also differed the LEGE-CC strains from their closest related genera.

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 Background Rare species, especially of the marine sedimentary biosphere, have long been overlooked owing to the complexity of sediment microbial communities, their sporadic temporal and patchy spatial abundance, and challenges in cultivating environmental microorganisms. In this study, we combined enrichments, targeted metagenomic sequencing, and extensive data mining to uncover uncultivated members of the archaeal rare biosphere in marine sediments. Results In protein-amended enrichments, we detected the ecologically and metabolically uncharacterized class Candidatus Penumbrarchaeia within the phylum Thermoplasmatota. By screening more than 8000 metagenomic runs and 11,479 published genome assemblies, we expanded the phylogeny of Ca. Penumbrarchaeia by 3 novel orders. All six identified families of this class show low abundance in environmental samples characteristic of rare biosphere members. Members of the class Ca. Penumbrarchaeia were predicted to be involved in organic matter degradation in anoxic, carbon-rich habitats. All Ca. Penumbrarchaeia families contain high numbers of taxon-specific orthologous genes, highlighting their environmental adaptations and habitat specificity. Besides, members of this group exhibit the highest proportion of unknown genes within the entire phylum Thermoplasmatota, suggesting a high degree of functional novelty in this class. Conclusions In this study, we emphasize the necessity of targeted, data-integrative approaches to deepen our understanding of the rare biosphere and uncover the functions and metabolic potential hidden within these understudied taxa.

AbstractBACKGROUNDAlthough it is known that Candidatus Liberibacter asiaticus (CLas), the agent of citrus Huanglongbing, circulates and multiplies within the insect vector Diaphorina citri, the specific factors enabling CLas transmission remain unclear. Previous studies have shown that ATPSyn‐β facilitates phytoplasma movement in vector insects, and functions as a transport protein in D. citri. In this study, the role of ATPSyn‐β was expected to be unveiled in CLas transmission in D. citri.RESULTSThe interaction between ATPSyn‐β of D. citri and the outer membrane protein A (OmpA) of CLas was identified using glutathione S‐transferase pull‐down and yeast two‐hybrid assays. No interactions existed between OmpA and vitellogenin (Vg). Bioinformatics analysis revealed that ATPSyn‐β contained two conserved domains, ATP‐synt_ab_N and ATP‐synt_ab. Notably, ATP‐synt_ab_N interacted with both OmpA and Vg, indicating a competitive relationship between Vg and OmpA for binding to ATPSyn‐β. After silencing ATPSyn‐β expression using RNAi, the feeding behavior of D. citri decreased, resulting in a lower acquisition of CLas. Subsequently, this inhibited CLas movement within D. citri and reduced the inoculation of CLas to healthy citrus plants.CONCLUSIONATPSyn‐β was involved in the acquisition, movement in vivo, and inoculation of CLas by interacting with OmpA, thus offering a novel target to prevent the spread of citrus Huanglongbing. © 2025 Society of Chemical Industry.