Plant Disease

Chokecherry ( Prunus virginiana L.) is a native shrub that is widely found across North America and is a host of ‘ Candidatus Phytoplasma pruni’ associated with X-disease of stone fruits. This pathogen is of major concern to U.S. cherry and peach growers, with the most recent and ongoing outbreak in the Pacific Northwest causing major economic harm. To investigate the role of chokecherry as a potential reservoir and source of orchard-associated strains, we surveyed wild chokecherry trees across 12 U.S. states and compared the strains present with those in commercially cultivated Prunus species. Approximately 47% of chokecherry sampled tested positive for a ‘ Ca. P. pruni’-related strain and phylogenetic analysis revealed high strain diversity, including six distinct clades and four newly identified lineages. In contrast, ‘ Ca. P. pruni’ strains isolated from cultivated cherry and peach trees exhibited lower diversity and were infected by known disease outbreak-associated strains such as Northwestern-X. These data suggest limited transmission between wild and cultivated hosts in the western United States, with patterns indicating spillover from orchards into nearby chokecherry populations rather than the reverse. However, in states east of the Rocky Mountains, chokecherry likely functions as a reservoir for orchard infections.

In Germany, maize (Zea mays L.) is cultivated on approximately 2.4 Mha, making it one of the most widely grown arable crops alongside wheat and barley (German Federal Statistical Office, 2025). Although maize has previously been identified as a host of Candidatus Phytoplasma solani (PHYPSO, 16SrXII-A), particularly in relation to maize redness (MR) disease detected in southeastern Europe (Jović et al., 2009), its role as a host of ‘Candidatus Arsenophonus phytopathogenicus’ (ARSEPH) has not been reported previously. The planthopper Pentastiridius leporinus (Hemiptera: Cixiidae) is the major vector for transmitting both pathogens in many crops in Germany (Rinklef et al., 2024). MR symptoms include patchy premature drying, reddish discoloration of leaves and midribs, poorly filled or aborted ears, and malformed kernels. In August and September 2025, maize plants showing MR-like symptoms were observed in different fields in Otzberg 49°48'25.3"N, 8°52'28.8"E (South Hesse), Bellheim 49°10′19.9N, 8°15′50.8E, Bockenheim 49°36'18.6"N, 8°11'30. E"O, Ibersheim 49°43′36.5″N, 8°24′34.6″E (Rhineland-Palatinate) and Ettling 48°48'50.0"N, 11°40'17.2"E (Baden-Württemberg). Samples from roots (25), leaves (10), ears (14) and kernels (9) of 14 selected symptomatic plants were collected for molecular analysis. DNA was extracted using a CTAB-based protocol. ARSEPH was detected by qPCR targeting the manA gene according to Zübert and Kube (2021). Positive results were obtained from ears of six plants from Ibersheim and Ettling, leaves of three plants from Bockenheim and Otzberg, kernels of two plants from Bellheim, and roots of one plant from Bockenheim. Four ARSEPH-positive samples (two ears and two kernels) were further analyzed using the Fra4/Fra5 PCR assay (Zreik et al. 1998). Sanger sequencing of the resulting amplicons followed by BLAST analysis confirmed all four sequences as ‘Candidatus Arsenophonus phytopathogenicus’ with 100% identity to reference sequence PP400342. The sequences have been deposited in GenBank under accession numbers PZ623476, PZ623478, PZ623483, and PZ626926. Furthermore, all samples were screened for PHYPSO using the qPCR assay described by Behrmann et al. (2022) with primers KL464/KL465. Seven ear samples from Ibersheim yielded positive qPCR signals. To further validate these results, DNA from qPCR-positive samples was subjected to nested PCR targeting the 16S rRNA gene using primer pairs P1/P7 (Schneider et al., 1995) and R16F2n/R16R2 (Gundersen and Lee, 1996), followed by Sanger sequencing and BLAST analysis against the NCBI GenBank database. However, repeated sequencing did not provide sufficient sequence identity to the reference strain STOL11 (GenBank accession number AF248959) to confirm the qPCR-positive samples as PHYPSO. To our knowledge, this is the first report of ARSEPH in maize in Germany. Its detection in maize extends the known host range of this phloem-limited bacterium. The high ARSEPH loads in ears and kernels detected in reproductive tissues are consistent with preferential accumulation in sink organs during grain filling. Whether maize contributes to pathogen persistence and transmission within the Pentastiridius leporinus or other potential vectors’ pathosystem remains to be investigated.

Early and accurate detection of ‘Candidatus Phytoplasma pruni’ is essential for the control of X-disease in Prunus species, but this is made difficult by pathogen biology and the ability to consistently recover nucleic acids, both of which increase the likelihood of false negatives. Here we examined the latter, using high- and low-titer infected sweet cherry (P. avium) samples to compare three tissue disruption methods, four nucleic acid extraction chemistries, and four PCR chemistries. Results showed that method selection had minimal impact on high-titer samples but significantly affected low-titer detection. Bead-beating disruption consistently outperformed rotary and manual grinding, yielding lower Cq values and more reliable amplification. Among extraction methods, CTAB-based protocols provided the highest nucleic acid recovery and detection sensitivity, particularly when paired with bead beating. Column- and magnetic bead–based kits performed less consistently, especially with low-titer samples, where false negatives were frequent. PCR chemistry also influenced results, although RT-qPCR did not significantly improve sensitivity over qPCR. Importantly, commonly used host internal controls failed to reliably indicate pathogen detection success, as high-abundance targets masked poor extraction efficiency. Cumulatively these data suggest the need for optimizing diagnostic protocols to ensure capture of low-concentration pathogens to avoid false negatives.

‘Candidatus Liberibacter solanacearum’ (Lso) haplotype D, transmitted by the carrot psyllid Bactericera trigonica, is a major threat to carrot production, particularly in the Mediterranean Basin and the Middle East. We investigated the impact of plant age at the time of inoculation on symptom development and yield under greenhouse and outdoor conditions. In greenhouse trials, 1- and 2-month-old plants were inoculated with psyllids harboring Lso. Both age groups showed reduced taproot weight compared with controls, but younger plants were more severely affected, exhibiting 90% yield reduction, while 2-month-old plants showed a 30% reduction. Witches’ broom symptoms appeared earlier in plants inoculated at 1 month (5 weeks postinoculation); however, within 2 weeks, plants inoculated at 2 months reached comparable levels of symptom incidence and severity. Additional greenhouse experiments using Lso-free psyllids confirmed that yield losses and disease symptoms were specifically due to Lso infection and not psyllid feeding. In outdoor trials, we tested the effects of inoculation at 7, 11, 15, and 19 weeks postsowing (wps) on disease incidence and yield. Significant yield losses occurred only in plants inoculated at 7 and 11 wps, with up to 45% reduction by harvest time. Symptoms began to appear at 20 wps in plants inoculated at 7 and 11 wps. At 23 wps, symptoms also appeared in plants inoculated at 15 wps, with a similar incidence to that observed in the earlier inoculation groups. Molecular detection of Lso at harvest (23 wps) confirmed that all plants inoculated at 19 wps or earlier were infected. These results demonstrate that carrots are highly susceptible to Lso-induced yield losses at the earlier stages of growth and highlight the importance of managing early season vector pressure to minimize economic damage.

Pear decline, associated to ‘Candidatus Phytoplasma pyri’, has emerged in Chilean pear orchards in recent years. While several Cacopsylla species are potential vectors of ‘Ca. P. pyri’, the disease’s full epidemiological cycle remains uncertain. Cacopsylla bidens, present in Chilean orchards, has recently been reported as a vector. This study conducted year-long surveys in two commercial pear orchards across different Chilean regions, capturing C. bidens in zones with ‘Ca. P. pyri’-infected pear trees. All developmental stages were collected, with peak abundances occurring in March and April. Outside the study zones, C. bidens were found in pine trees but not in adjacent cultivated areas. Two seasonal morphotypes, summer and winter forms, were identified. Molecular analysis detected ‘Ca. P. pyri’ in a high proportion of insects, with maximum infection rates in March and April. These findings advance our understanding of ‘Ca. P. pyri’ spatial and temporal dynamics and their potential role in ‘Ca. P. pyri’ spreading under Chilean field conditions.