Publications

4413

ABSTRACT Candidatus Saccharimonadia is a class‐level lineage of ultrasmall bacteria within the phylum Minisyncoccota (formerly Candidate Phyla Radiation or Ca . Patescibacteria), commonly found in activated sludge processes treating municipal wastewater. In this study, we aimed to elucidate the metabolic potential of Ca . Saccharimonadia by using shotgun metagenomic sequencing combined with a filtration‐based size‐fractionation approach for activated sludge from five wastewater treatment plants. A total of 65 high‐quality metagenomic bins were recovered, belonging to four orders and 19 families of Ca . Saccharimonadia, including previously unreported lineages in activated sludge. These bins had small genomes (approximately 0.46–1.73 Mbp) with limited metabolic capabilities, indicating dependency on other microorganisms. Notably, the order Ca . Saccharimonadales retained a type IV secretion system and effector gene cluster for parasitic interactions with the hosts, suggesting that Ca . Saccharimonadales bacteria may exhibit a parasitic lifestyle. Co‐occurrence network analysis showed that members of the order Ca . Saccharimonadales were significantly correlated with multiple lineages, including Actinobacteriota , for which a parasitic relationship has been previously demonstrated. Our results shed light on the potential ecophysiology of the diverse members of Ca . Saccharimonadia, providing a comprehensive understanding of Ca . Saccharimonadia in activated sludge.

Abstract Pistachio ( Pistacia vera L.) is a vital crop for Türkiye's economy, occupying over 420 000 ha, primarily in south‐eastern Anatolia. In 2023, severe symptoms, such as curling, chlorosis and needle‐like leaf deformation, were observed on pistachio trees in Şanlıurfa Province. This prompted a molecular study of phytoplasma pathogens. A real‐time PCR targeting the 16SrDNA gene was used to analyse DNA from eight symptomatic and two asymptomatic trees. A phytoplasma was found in two symptomatic samples but not in asymptomatic controls. Positive samples were further analysed using nested PCR. One sequence with 99% identity to ‘ Candidatus Phytoplasma trifolii’ (16SrVI‐A subgroup) was found by bidirectional sequencing of PCR products. The identification of the phytoplasma was confirmed by a phylogenetic analysis showing the clustering of the UP 1 isolate with ‘ Ca. Phytoplasma trifolii’ reference strain and by a virtual RFLP analysis using 17 restrictions enzymes. This first report of ‘ Ca. Phytoplasma trifolii’ in pistachio in Türkiye highlights the presence of this phytoplasma in a key agricultural sector. Although no yield losses have been observed to date, broader surveys and monitoring of leafhopper vectors are necessary to assess the potential risk and implement timely management strategies.

Abstract Bactericera maculipennis (Crawford) and Bactericera cockerelli (Šulc) (Hemiptera: Triozidae) share hosts within the Solanaceae and Convolvulaceae (Solanales), and both are associated with “Candidatus Liberibacter solanacearum” (Lso). Lso, transmitted by B. cockerelli, causes diseases in solanaceous crops including zebra chip disease of potato. Up to 50% of B. maculipennis adults also harbor Lso, but transmission of Lso to plants by this psyllid has not been confirmed yet. The only documented field host of B. maculipennis in the Pacific Northwest is Convolvulus arvensis L. (Convolvulaceae) but diagnostic methods fail to detect Lso in leaves of this plant. It is therefore unclear how Lso persists within B. maculipennis populations. We surveyed species of Convolvulaceae and Solanaceae for B. maculipennis and report a widespread association between B. maculipennis and Lso throughout the western United States. Diagnostic polymerase chain reaction failed to detect Lso from leaves of C. arvensis yet readily detected Lso from stems where B. maculipennis nymphs tend to feed. Bactericera maculipennis transmitted Lso to species of Convolvulaceae in greenhouse experiments, confirming vector competency. We report high rates of Lso infection in populations of both B. maculipennis and B. cockerelli occurring on C. arvensis, but occurrence of B. cockerelli on C. arvensis was limited to autumn months only and with very low populations. Results suggest C. arvensis is a non-crop reservoir of Lso but do not suggest that B. maculipennis is a direct threat to solanaceous crops or that C. arvensis is a major source of Lso-infected B. cockerelli colonizing potato fields.

Mulberries (Morus spp.) are important crops valued for berry and silk production (Baciu et al., 2023). In March 2025, a mulberry tree in Dongshi, Taichung, Taiwan, was found bearing fruits showing phyllody symptoms. To determine whether the disease was associated with phytoplasma infection, symptomatic fruits were collected from five branches and tested. For the controls, asymptomatic berries were sampled from three symptomless mulberry trees in Wufeng, Taichung in April 2025. DNA was extracted from five symptomatic fruits collected from different branches and three symptomless berries obtained from separate asymptomatic trees. DNA was extracted with a Synergy 2.0 Plant DNA Extraction Kit (OPS Diagnostics). To ensure that DNA qualities were sufficient for PCR assays, the samples were tested with primers 28KJ and 28C targeting the plant’s 26S rDNA (Cullings 1992). PCR was conducted with the GoTaq Green Master Mix (Promega), and every sample produced the expected amplicon (~685 bp). Nested PCR tests were conducted using universal phytoplasma primer pairs P1/ P7 (outer; Schneider et al. 1995) and R16F2n/R16R2 (inner; Gundersen and Lee 1996). Four out of five symptomatic samples amplified the 1.25-kb target fragment in the nested assay, while none of the asymptomatic samples did. The 16S rDNA of a representative symptomatic sample was sequenced (GenBank/DDBJ accession no. LC897699) and subjected to analysis using iPhyClassifier (https://plantpathology.ba.ars.usda.gov/cgi-bin/resource/iphyclassifier.cgi). The detected phytoplasma was classified to the 16SrI-B subgroup with a similarity coefficient of 1.0 (compared to a reference pattern; ‘Candidatus Phytoplasma asteris’ OY; accession no. AP006628) and was found to share 99.68% 16S rDNA sequence similarity with a ‘Ca. Phytoplasma asteris’ reference strain (accession no. M30790). To validate this finding, the sample was also tested with other phytoplasma gene-specific nested primers targeting the elongation factor Tu (tuf) and ribosomal protein (rp) genes. For tuf, primer pairs fTuf1/rTuf1 and fTufAy/rTufAy were respectively used for the first and second round of PCR (Schneider and Gibb 1997). For rp, the outer and inner primer pairs were rpF1/rpR1 and rp(I)F1A/rp(I)R1A, respectively (Martini et al. 2007). Both target gene fragments were successfully amplified, and the inner amplicons were sequenced. BLASTn (core_nt) searches using quality-trimmed sequences showed that the obtained tuf sequence (accession no. LC896430) was most similar to that of ‘Ca. Phytoplasma asteris’ OY (accession no. AP006628; 99.9%; 830/831 bp) and those of various 16SrI strains. The detected rp sequence (accession no. LC896429) shared the highest identity with that of a ‘Ca. Phytoplasma asteris’ strain infecting Boehmeria pannosa (16SrI‑B; accession no. MT739521; 99.9%; 1,056/1,057 bp) and those of other 16SrI strains, and was very similar to that of ‘Ca. Phytoplasma asteris’ OY (accession no. AP006628; 99.6%; 1,053/1,057 bp). Accordingly, the phytoplasma detected in this study was identified as a ‘Ca. Phytoplasma asteris’-related strain belonging to subgroup 16SrI-B. In Taiwan, 16SrI phytoplasmas have been reported affecting plants such as Murraya exotica (Tseng et al. 2022). This is the first report of a 16SrI phytoplasma related with Morus symptoms in Taiwan. Since the disease directly impacts the commercial value of the berries, the pathogen should be carefully monitored in mulberry-producing areas.