Publications

3989

ABSTRACT Although the high-altitude limit for microbial survival in the Earth–atmosphere system has remained a scientific curiosity and topic of study, the ecological significance of long-distance microbial dispersal in the atmosphere has been perceived to have marginal relevance. Here, we report the characterization of novel plant pathogenic species of Curtobacterium that were isolated from samples collected at altitudes ranging from 1.5 to 29 km above sea level. Whole genome-based phylogenies of three strains, paired with plant challenge assays, indicate that each is a previously unrecognized species and causes disease on beans comparable to Curtobacterium flaccumfaciens . Isolates from the stratosphere (strain L6-1) and agricultural millet (G77) were identified to be the same species and designated as Curtobacterium aetherium sp. nov. C. aetherium displays high levels of tolerance to desiccation and UV radiation, which are stresses that increase in intensity with altitude. Back trajectory air mass analysis implied that the phytopathogens may have had an intercontinental source, but regional origins in the continental US cannot be excluded. The environmentally robust phytopathogens we have documented in the upper atmosphere provide new perspective on the role that high-altitude transport may play in microbial dispersal, gene flow, and the epidemiology of aerially dispersed plant disease. IMPORTANCE Enormous quantities and varieties of microorganisms are continually aerosolized and transported in the atmosphere, yet there is a limited understanding of the consequences to downwind ecosystems. While studying bacteria that survive extreme conditions at altitudes up to 29 km in the atmosphere, we discovered new species that have the capacity to cause disease in agriculturally relevant bean varieties. The hardiest isolate we characterized from the stratosphere is a member of the same species as an isolate from an agricultural source, which we have designated Curtobacterium aetherium . C. aetherium is a phytopathogen capable of enduring high-altitude and long-distance atmospheric transport while also possessing the potential to opportunistically infect crops in deposition locations.

Phytoplasmas cause infections in numerous plants in agricultural ecosystems, causing significant yield and quality losses in products. In recent years, it has been known that diseases caused by phytoplasmas cause economic losses in eggplant (Solanum melongena L.) cultivation. In Turkey, research on infections caused by phytoplasmas in eggplant growing areas is quite limited. This study was carried out to detect phytoplasma infections symptomatologically and molecularly in eggplant production areas in Şanlıurfa province. Fourteen samples were collected from eggplants exhibiting symptoms such as witches’ broom, flower abnormalities (virescence, phyllody), elongation of the pedicle, arising of new shoots from flower parts, yellowing and proliferation. Phytoplasma infection was detected in 8 symptomatic samples using 16S rRNA-specific primers, P1/P7 and R16F2n/R16R2, by direct and nested PCR. Sequence information of fragments obtained as a result of molecular studies was extracted and BLAST analyses were performed. According to nucleotide sequence similarity in the 16S rRNA gene region, it was determined that the genetic group of phytoplasma causing infection in eggplant was related to ‘Candidatus Phytoplasma solani’ (CaPsol) belonging to 16SrXII-A subgroup with 98% sequence identity. To our best knowledge, this study suggests comprehensive symptomatic diagnosis of CaPsol infecting eggplants in Türkiye.

Marine heterotrophic bacteria in coastal waters respond to the influx of carbon from natural and anthropogenic sources. We identified two nearly identical, (99.9% average nucleotide identity; 100% amino acid identity; same DNA G + C content of 52.3 mol%) high-quality (≥99% CheckM completeness and ≤ 1.3% contamination) draft metagenome-assembled genomes (MAGs; SJ0813 and SJ0972) from seawater microbiomes of a southern island of Singapore that is in a protected marine park. The MAGs were only assigned to the Cellvibrionaceae family according to Genome Taxonomy Database. Overall genome related indices to Pseudomaricurvus alkylphenolicus KU41GT as the closest phylogenetic relative revealed no more than 70.45% average nucleotide identity (ANIcutoff < 95%), below the 50% percentage of conserved proteins (POCPcutoff = 43.54%) for genera cutoff and low digital DNA–DNA hybridization values (DDH = 20.6 and 20.8%). The major respiratory quinone is predicted to be ubiquinone-9 from the annotation of 3-demethylubiquinone-9 3-methyltransferase (ubiG, K00568) involved in the last step of the ubiquinone biosynthesis pathway (M00117), which differed from the ubiquinone-8 utilized by known members of Cellvibrionaceae. Both MAGs contained a complete pathway for dissimilatory nitrate reduction to ammonia, which increases bioavailability of nitrogen in seawater. An identical choline dehydrogenase found in both MAGs have a low amino-acid identity (≤64.47%) compared to existing GMC family oxidoreductases, expanding on the diversity of this family of enzymes. The MAGs meet nearly all the minimum requirements but lack a 16S rRNA gene of sufficient length required for the proposed novel genus and species under SeqCode. Nevertheless, phylogenetic trees based on core-genome and RpoB as an alternative phylogenetic marker are congruent with the taxon standing as a monophyletic clade to other taxa of the order Cellvibrionales. Taken together, the MAGs (SJ0813 and SJ0972) represent an uncultured, undescribed genus and species in which we tentatively propose the name Candidatus Pelagadaptatus aseana gen. nov., sp. nov. and strain SJ0813TS (=BAABNI000000000.1TS) as type sequence. Phylogenetic inference from core-genome and RpoB phylogenetic trees placed Umboniibacter marinipuniceus KMM 3891T outside Cellvibrionaceae. We, therefore, propose the transfer of the genus Umboniibacter from the family Cellvibrionaceae to a new family Umboniibacteraceae according to the International Code of Nomenclature of Prokaryotes.

Surveys for exotic plant pests conducted during July and August of 2023 and 2024 across 20 vineyards in 12 counties throughout Minnesota, USA, revealed that less than 2% of the approximatively 3000 vines inspected (Vitis spp., hybrid grape varieties) exhibited symptoms suggestive of phytoplasma yellows disease. Observed symptoms included yellowing of leaf lamina, downward rolling of leaf margins, and necrosis of leaf margins. To investigate a potential association between these symptoms and phytoplasmas, two symptomatic plants were selected in August of 2024 from two distinct vineyards for further analysis. Petiole tissue DNA extracts were initially tested using a phytoplasma-specific real-time PCR assay (Hodgetts et al. 2009), which confirmed the presence of phytoplasma DNA in both symptomatic plants. Subsequently, the samples underwent semi-nested PCR amplification targeting the phytoplasma 16S rRNA gene, using primers P1/16S-SR followed by P1A/16S-SR (Deng and Hiruki 1991; Lee et al. 2004). The resulting amplicons were cloned and sequenced. No amplicon was produced from an asymptomatic grapevine DNA sample included as a test control. Analysis of the 16S rRNA gene sequences revealed that each plant was infected by one of two distinct phytoplasma strains, designated MN450 and MN466. Representative sequences obtained from the two samples exhibiting interoperon heterogeneity were deposited in the GenBank database under accession numbers: PQ889195 (rrnA) and PQ889196 (rrnB) for strain MN450, and PQ889197 (rrnA) and PQ889198 (rrnB), for strain MN466. Sequence analysis using the online phytoplasma classification tool iPhyClassifier (Zhao et al. 2009) showed that the 16S rRNA gene sequences from MN450 shared 1491/1495 bp (99.73%) and 1492/1495 bp (99.80%) identity (rrnA and rrnB, respectively) with that of the ‘'Ca. Phytoplasma pruni' rrnA’ reference strain (GenBank JQ044393), identifying MN450 as a ‘'Ca. Phytoplasma pruni' rrnA’-related strain and possibly representing a new subgroup within the 16Sr group III. For MN466, the 16S rRNA gene sequences shared 1471/1482 bp (99.26%) and 1469/1482 bp (99.12%) identity (rrnA and rrnB, respectively) with that of the ‘Ca. Phytoplasma asteris’ reference strain (GenBank M30790), and identifying the phytoplasma as a ‘Ca. P. asteris’-related strain belonging to subgroup 16SrI-A. To further characterize the two phytoplasma strains, the secY gene was PCR amplified from the DNA extracts as described by Lee et al. (2010). The obtained amplicons were cloned and sequenced, and representative secY gene sequences were deposited in GenBank under accession numbers PQ899621 (MN450) and PQ899622 (MN466). BLASTn searches against the NCBI core nucleotide database revealed that the secY gene sequence for MN450 exhibited 100% identity (1614/1614 bp) with ‘Ca. P. pruni’ (GenBank KM268860). Similarly, the sequence for MN466 showed 100% identity (1287/1287 bp) with ‘Ca. P. asteris’ (GenBank CP000061). In North America, grapevine yellows disease associated with strains related to 'Ca. P. asteris' and 'Ca. P. pruni' has previously been reported in various states and referred to as North American grapevine yellows (NAGY) by Davis et al. (2015). To our knowledge, this is the first report of NAGY in Minnesota, confirming that the two phytoplasma species detected are consistently associated with the disease in U.S. vineyards. While further research is needed to evaluate the potential impacts of NAGY in Minnesota, this study highlights the broader distribution of the disease across the United States.

AbstractThe severe Asiatic form of huanglongbing (HLB), caused by “Candidatus Liberibacter asiaticus” (CLas), threatens global citrus production via the citrus psyllid, Diaphorina citri. Culturing challenges of CLas necessitate reducing D. citri populations for disease management. CLas boosts the fecundity of CLas‐positive (CLas+) D. citri and fosters its own proliferation by modulating the insect host's juvenile hormone (JH), but the intricate endocrine regulatory mechanisms remain elusive. Here, it is reported that the D. citri ecdysis‐triggering hormone (DcETH) and its receptor DcETHR play pivotal roles in the reciprocal benefits between CLas and D. citri within the ovaries, influencing energy metabolism and reproductive development in host insects; miR‐210, negatively regulates DcETHR expression, contributing to this symbiotic interaction. CLas infection reduces 20‐hydroxyecdysone (20E) levels and stimulates DcETH release, elevating JH production via DcETHR, enhancing fecundity and CLas proliferation. Furthermore, circulating JH levels suppress 20E production in CLas+ ovaries. Collectively, the orchestrated functional interplay involving 20E, ETH, and JH increases energy metabolism and promotes the fecundity of CLas+ D. citri and CLas proliferation. These insights not only broaden the knowledge of how plant pathogens manipulate the reproductive behavior of insect hosts but also offer novel targets and strategies for combatting HLB and D. citri.