First Report of a 'Candidatus Phytoplasma sacchari'-Related Strain Associated with Yellowing and Decline of Silver Bluestem in Texas, U.S.A.

Silver bluestem [Bothriochloa laguroides (DC.) Herter] is a warm-season grass native to Texas. This perennial grass plays a crucial role in maintaining ecological balance and supporting wildlife in the region. In September 2022, while investigating the ecological impact of invasive grass species on a grassland located near Pipe Creek (TX), B. laguroides plants were observed showing symptoms that included yellowing of the blades and occasionally brown discoloration of the midveins and stems (Fig. S1). Disease incidence was estimated as 2% of silver bluestem plants in the 2 hectares surveyed. To investigate the possibility of a phytoplasma association with the symptoms, four symptomatic and four asymptomatic leaf samples were collected for further study. Total DNA was extracted from leaf midribs using a DNeasy Plant Mini Kit (Qiagen). The DNA extracts were tested using a phytoplasma-specific quantitative PCR assay (Hodgetts et al. 2009), which identified two out of the four symptomatic B. laguroides samples as positive for phytoplasmas. A semi-nested PCR assay for amplification of the 16S rRNA gene fragment was then performed on these samples with primers P1/16S-SR followed by P1A/16S-SR (Deng, and Hiruki 1991; Lee et al. 2004), and two additional housekeeping genes (tuf and secA) were amplified as previously described (Makarova et al. 2012; Hodgetts et al. 2008; Bekele et al. 2011). All amplicons of the expected size, 1.5 kb (16S rRNA), 0.4 kb (tuf) and 0.6 kb (secA), were purified and bi-directionally sequenced using primers from each gene second round PCR amplification. Analysis of the sequences derived from the three gene fragments revealed no variation between the two plant samples and confirmed they originated from a phytoplasma, termed strain TXSB-2 (Texas Silver Bluestem). Sequences from a single B. laguroides plant DNA extract were deposited in GenBank with accession numbers OR711913 (16S rRNA), OR709687 (tuf) and OR709688 (secA). A BLAST search of the 16S rRNA gene sequence from TXSB-2 against the NCBI nucleotide database, showed 99.58% sequence identity with an unclassified phytoplasma clone 139-1 from a leafhopper collected in Australia (MW281491) (Fig. S2). The partial nucleotide sequence of the tuf and secA genes showed 90.60% and 89.78% similarity, respectively, to the corresponding genes in 'Ca. P. sacchari' strain SCWL1 (CP115156) associated with sugarcane in China. The iPhyClassifier, an interactive online tool for phytoplasma identification and classification (Zhao et al. 2009), was used to determine the 'Candidatus Phytoplasma' species affiliation and group/subgroup classification status of this phytoplasma strain. The result showed that the TXSB-2 16S rDNA shared 98.94% sequence identity with that of the 'Ca. P. sacchari' reference strain (GenBank accession: MN889545), indicating TXSB-2 is a 'Ca. P. sacchari'-related strain. The result from virtual restriction fragment length polymorphism (RFLP) analysis of the 16S rDNA F2nR2 fragment revealed that TXSB-2 possessed a collective RFLP pattern that is distinct from the reference patterns of all established phytoplasma ribosomal subgroups and is proposed as the representative strain of a new subgroup designated as 16SrXI-H. 'Candidatus Phytoplasma sacchari' has been reported associated with sugarcane grassy shoot disease, which is considered among the most damaging diseases of sugarcane across parts of Southeast Asia and India (Kirdat et al. 2021). The same phytoplasma was recently confirmed infecting sorghum in India (Nithya et al. 2024). To our knowledge, this is the first report of a 'Ca. P. sacchari'-related strain infecting B. laguroides in the United States. Moreover, B. laguroides is a new host for strains related to 'Ca. P. sacchari'. Further investigation is required to elucidate the prevalence of this disease in the area, its natural vectors, and the potential consequences arising from this novel phytoplasma strain within its ecosystem in Texas.

Fungal Endophyte Alternaria tenuissima Can Affect Growth and Selenium Accumulation in Its Hyperaccumulator Host Astragalus bisulcatus.

Endophytes can enhance plant stress tolerance by promoting growth and affecting elemental accumulation, which may be useful in phytoremediation. In earlier studies, up to 35% elemental selenium (Se0) was found in Se hyperaccumulator Astragalus bisulcatus. Since Se0 can be produced by microbes, the plant Se0 was hypothesized to be microbe-derived. Here we characterize a fungal endophyte of A. bisulcatus named A2. It is common in seeds from natural seleniferous habitat containing 1,000-10,000 mg kg-1 Se. We identified A2 as Alternaria tenuissima via 18S rRNA sequence analysis and morphological characterization. X-ray microprobe analysis of A. bisulcatus seeds that did or did not harbor Alternaria, showed that both contained >90% organic seleno-compounds with C-Se-C configuration, likely methylselenocysteine and glutamyl-methylselenocysteine. The seed Se was concentrated in the embryo, not the seed coat. X-ray microprobe analysis of A2 in pure culture showed the fungus produced Se0 when supplied with selenite, but accumulated mainly organic C-Se-C compounds when supplied with selenate. A2 was completely resistant to selenate up to 300 mg L-1, moderately resistant to selenite (50% inhibition at ∼50 mg Se L-1), but relatively sensitive to methylselenocysteine and to Se extracted from A. bisulcatus (50% inhibition at 25 mg Se L-1). Four-week old A. bisulcatus seedlings derived from surface-sterilized seeds containing endophytic Alternaria were up to threefold larger than seeds obtained from seeds not showing evidence of fungal colonization. When supplied with Se, the Alternaria-colonized seedlings had lower shoot Se and sulfur levels than seedlings from uncolonized seeds. In conclusion, A. tenuissima may contribute to the Se0 observed earlier in A. bisulcatus, and affect host growth and Se accumulation. A2 is sensitive to the Se levels found in its host's tissues, but may avoid Se toxicity by occupying low-Se areas (seed coat, apoplast) and converting plant Se to non-toxic Se0. These findings illustrate the potential for hyperaccumulator endophytes to affect plant properties relevant for phytoremediation. Facultative endophytes may also be applicable in bioremediation and biofortification, owing to their capacity to turn toxic inorganic forms of Se into non-toxic or even beneficial, organic forms with anticarcinogenic properties.

Selenium distribution and speciation in the hyperaccumulator Astragalus bisulcatus and associated ecological partners.

The goal of this study was to investigate how plant selenium (Se) hyperaccumulation may affect ecological interactions and whether associated partners may affect Se hyperaccumulation. The Se hyperaccumulator Astragalus bisulcatus was collected in its natural seleniferous habitat, and x-ray fluorescence mapping and x-ray absorption near-edge structure spectroscopy were used to characterize Se distribution and speciation in all organs as well as in encountered microbial symbionts and herbivores. Se was present at high levels (704-4,661 mg kg(-1) dry weight) in all organs, mainly as organic C-Se-C compounds (i.e. Se bonded to two carbon atoms, e.g. methylselenocysteine). In nodule, root, and stem, up to 34% of Se was found as elemental Se, which was potentially due to microbial activity. In addition to a nitrogen-fixing symbiont, the plants harbored an endophytic fungus that produced elemental Se. Furthermore, two Se-resistant herbivorous moths were discovered on A. bisulcatus, one of which was parasitized by a wasp. Adult moths, larvae, and wasps all accumulated predominantly C-Se-C compounds. In conclusion, hyperaccumulators live in association with a variety of Se-resistant ecological partners. Among these partners, microbial endosymbionts may affect Se speciation in hyperaccumulators. Hyperaccumulators have been shown earlier to negatively affect Se-sensitive ecological partners while apparently offering a niche for Se-resistant partners. Through their positive and negative effects on different ecological partners, hyperaccumulators may influence species composition and Se cycling in seleniferous ecosystems.

Selenium accumulation in plants--phytotechnological applications and ecological implications.

Selenium (Se) is an essential trace element for many organisms including humans, yet toxic at higher levels. Both Se deficiency and toxicity are problems worldwide. Since plants readily accumulate and volatilize Se, they may be used both as a source of dietary Se and for removing excess Se from the environment. Plant species differ in their capacity to metabolize and accumulate Se, from non-Se accumulators (< 100 mg Se/kg DW), to Se-accumulators (100-1000 mg Se/kg DW) to Se hyperaccumulators (> 1,000 mg Se/kg DW). Here we review plant mechanisms of Se metabolism in these various plant types. We also summarize results from genetic engineering that have led to enhanced plant Se accumulation, volatilization, and/or tolerance, including field studies. Before using Se-accumulating plants at a large scale we need to evaluate the ecological implications. Research so far indicates that plant Se accumulation significantly affects the plant's ecological interactions below and above ground. Selenium canprotect plants from fungal pathogens and from a variety of invertebrate and vertebrate herbivores, due to both deterrence and toxicity. However, specialist (Se-tolerant herbivores), detritivores and endophytes appear to utilize Se hyperaccumulator plants as a resource. These findings are relevant for managing phytoremediation of Se and similar elements.

White locoweed toxicity is facilitated by a fungal endophyte and nitrogen-fixing bacteria.

Mutualistic interactions with fungal endophytes and dinitrogen-fixing bacteria are known to exert key biological influences on the host plant. The influence of a fungal endophyte alkaloid on the toxicity of a plant has been documented in Oxytropis sericea. Oxytropis sericea is a perennial legume responsible for livestock poisoning in western North America. Livestock poisoning is attributed to the alkaloid swainsonine, which is synthesized inside the plant by the fungal endophyte Embellisia sp. In this study, the ability of Oxytropis sericea to form a dinitrogen-fixing symbiosis with Rhizobium and the effects of this symbiosis on the production of swainsonine by Embellisia sp. were evaluated in a greenhouse environment. Seeds of O. sericea were grown in plastic containers. Twenty-week-old O. sericea seedlings were inoculated with four strains of Rhizobium. Twenty weeks after inoculation, plant growth and root nodulation by Rhizobium were measured. Dinitrogen fixation was confirmed using an acetylene reduction assay (ARA) on excised root nodules. Dry leaves were analyzed for swainsonine content. A second set of plants was treated with fungicide to evaluate the effect of reduced fungal endophyte infection on plant growth and swainsonine production. All inoculated plants produced indeterminate nodules. The ARA indicated that 98% of the excised nodules were fixing dinitrogen. Rhizobium-treated plants had greater swainsonine concentrations than the non-inoculated controls. Plants that established from seeds treated with fungicide had lower biomass than non-fungicide-treated controls and plants treated with foliar fungicide. Plants treated with foliar fungicide and the controls had greater swainsonine concentrations than the plants that received seed fungicide. This greenhouse study is the first report of nodulation and dinitrogen fixation in O. sericea. It also demonstrates that dinitrogen fixation increases the production of swainsonine in O. sericea plants infected with Embellisia sp. Results from this study suggest that dinitrogen fixation affects swainsonine production and has the potential to support the symbiosis between Embellisia sp. and O. sericea when soil nitrogen is limited. Oxytropis sericea competitiveness appears to be facilitated by an ability to simultaneously associate with Rhizobium and a fungal symbiont.

Epidemiolgía de las enfermedades renales en niños en Venezuela / Epidemiology of the renal diseases in children in Venezuela

El presente estudio, multicéntrico, tiene como objetivos presentar datos epidemiológicos de las nefropatías más frecuentemente observadas en niños en Venezuela, describir las características más importantes de las mismas y realizar una evaluación general de la disponibilidad de facilidades nefropediátricas en el País. A través de un cuestionario estándar se obtuvo información de 15 centros hospitalarios, sobre la patología observada durante un año. Fueron reportados 4018 casos de nefropatías en pacientes evaluados ambulatoriamente u hospitalizados. El número de ingresos de nefrología pediátrica fue 6,3 por ciento del total de primeras consultas pediátricas/año. Aproximadamente 70 por ciento de los casos correspondieron a tres categorías diagnósticas: a) Infección urinaria (32 por ciento), con detección de malformaciones congénitas del tracto urinario en 25 por ciento de los pacientes; b) Trastornos metabólicos (28 por ciento), fundamentalmente hipercalciuria, asociados o no a urolitiasis; c. Glomerulopatías, (9,5 por ciento), con alta frecuencia de nefritis posinfecciosa, para el caso de las patologías agudas, y predominio de la glomeruloesclerosis segmentaria y focal para el caso de las lesiones renales complejas primarias. El otro 30 por ciento de los casos correspondió a: síndrome nefrótico, con 80 por ciento de casos corticosensibles y 4 por ciento SN del primer año de la vida, con predominio en este grupo del secundario a lúes congénita; la insuficiencia renal aguda fue secundaria a deshidratación en la mitad de los casos observados y un número significativo por combinación de deshidratación y nefrotoxicidad. La insuficiencia renal crónica mostró una incidencia de 14,6 casos/año/millón hab < 15 años, siendo las causas de la misma glomerulopatías en 36 por ciento, uropatías en 30 por ciento y enfermedades heredofamiliares 12 por ciento. Un 68 por ciento de los pacientes con uremia terminal son incluidos en programas de rehabilitación, fundamentalmente diálisis peritoneal ambulatoria continua, pero sólo 30 por ciento recibe trasplante renal. Otras patologías observadas fueron: acidosis tubular renal 6 por ciento, hematuria primaria 4 por ciento enuresis 2 por ciento y otras 3 por ciento (tumores renales, hipertensión arterial, enfermedades quísticas renales y nefritis túbulo intersticial)