Abstract Climate change increasingly threatens potato production through intensified drought stress and emergence of vector-transmitted diseases. This study evaluated straw mulch application as a dual-purpose climate adaptation strategy to enhance potato performance under drought conditions and reduce the incidence of bacterial potato tuber wilt (BPW) caused by ‘ Candidatus Arsenophonus phytopathogenicus’ and ‘ Candidatus Phytoplasma solani’, transmitted by the planthoppers Pentastiridius leporinus and Hyalesthes obsoletus . Field trials conducted from 2022 to 2024 in Bingen am Rhein, Germany, tested five potato cultivars under mulched and unmulched conditions. Complementary greenhouse choice experiments investigated oviposition preferences of P. leporinus in response to soil type and mulch cover. The magnitude of mulch benefits correlated directly with drought severity, demonstrating greatest efficacy under high-stress conditions. Mulch application increased tuber yield by 33% overall, though effects varied significantly across years ( p = 0.003) and cultivars ( p = 0.006), with cultivar Darling showing exceptional drought responsiveness (119% yield increase under severe drought conditions). Mulch significantly reduced rubbery tuber incidence by 22% ( p = 0.019) and showed marginal reduction in wilting symptoms (29%, p = 0.072). In contrast, greenhouse experiments revealed mulch significantly increased vector oviposition ( p < 0.001), particularly at the straw surface layer. These findings demonstrate that mulch operates through complex ecological pathways that buffer drought stress and enhance disease tolerance despite increased vector reproduction. The net positive outcomes for both yield and disease management under drought conditions support strategic mulch deployment as a valuable climate adaptation tool, particularly when integrated with drought-responsive variety selection and precision management frameworks that balance enhanced vector habitat with improved plant resilience under combined abiotic and biotic stresses. This research provides the first comprehensive evaluation of cultivar-specific responses to mulch under combined drought and bacterial pathogen pressure, offering practical insights for sustainable potato production in changing climatic conditions.

Phytoplasmas are obligate intracellular parasitic bacteria that infect over 1000 plant species globally, causing devastating diseases characterized by yellowing, witches’ broom, phyllody, and significant yield losses in economically important crops. The unculturable nature of these pathogens has historically hindered their study; however, advances in molecular biology and genomics have substantially accelerated progress over the past two decades. This review provides a comprehensive overview of current knowledge on phytoplasma diseases and control technologies. In terms of taxonomy, phytoplasmas are currently classified into 37 16Sr groups with over 150 subgroups based on 16S rRNA gene analysis, and approximately 50 ‘Candidatus Phytoplasma’ species have been formally named. Genomic studies have revealed that phytoplasmas possess highly reduced genomes (530–1350 kb) lacking many essential metabolic pathways, reflecting their obligate parasitic lifestyle. Regarding pathogenesis, secreted effector proteins such as SAP (Secreted Aster Yellows Witches’ Broom Protein), TENGU (tengu-su inducer), and SWP (Secreted Wheat Blue Dwarf Protein) manipulate plant hormone signaling and developmental processes, leading to characteristic disease symptoms. Detection technologies have evolved from traditional microscopy to molecular methods, including nested PCR, real-time quantitative PCR, loop-mediated isothermal amplification (LAMP), and CRISPR/Cas-based systems (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated protein), with AI-based image recognition and remote sensing emerging as promising tools for large-scale field monitoring. Integrated management strategies encompassing agricultural practices, insect vector control, biological control agents, induced resistance, and breeding for resistance are discussed. Finally, future research directions, including functional genomics, microbiome-based approaches, and precision agriculture technologies, are highlighted. This review aims to provide researchers and practitioners with a systematic reference for understanding phytoplasma biology and developing effective disease management strategies.

ABSTRACT The planthopper Pentastiridius leporinus [Hemiptera: Cixiidae] is a vector of the two plant pathogenic bacteria ‘ Candidatus Arsenophonus phytopathogenicus’ and ‘ Candidatus Phytoplasma solani’ causing ‘bacterial potato tuber wilt’ in potato. An infection leads to symptoms such as rubbery and non‐processable tubers and consequently to substantial decreases in yield and tuber quality as well as to abandonment of seed potato production in affected areas. Up to date, resistant potato varieties are not yet known, and the tool box of integrated pest management to prevent the spread of P. leporinus as causal vector is quite limited. Mortality experiments designed as no‐choice trials on potato cultivar ‘Gala’ ( Solanum tuberosum ) and on its wild relative S. bulbocastanum revealed significant effects of the latter on the survival rate of the vector. LC/MS analysis of the insects and their intestines showed that C16:3‐solamine, a bioactive compound of S. bulbocastanum , was retrieved only in individuals from the S. bulbocastanum plants. Furthermore, choice‐tests were carried out to study the vector behaviour. A preference towards S. bulbocastanum vs. S. tuberosum ‘Gala’ was observed despite its lethal effects. Volatile organic compounds of the two different species were collected and analysed by GC/MS. The two profiles differed in 39 of 80 compounds. The wild potato species S. bulbocastanum and its compounds are discussed as potential starting points for further research on sustainable management of P. leporinus .

Coconut lethal yellowing disease (CLYD) caused by phytoplasma is a destructive disease affecting coconut and other palm species. The disease has been reported in Mozambique, Tanzania, and some West African countries, where millions of coconut trees have been affected. Although the disease was first reported in 1999 in Northern Coast of Kenya, no further studies have been conducted, leaving the giant coconut industry at risk of destruction. It is for this reason that a surveillance was undertaken to study the extent of the disease spread and the specific Candidatus phytoplasma strain causing the disease. A total of 125 coconut tree samples were collected in Kwale, Kilifi, Lamu, and Tana River counties by drilling the tree trunks aseptically. DNA extractions were done, followed by PCR process done using P1/P7 universal primers, gel electrophoresis and amplicon sequencing. For the coconut tree drillings results, the BLAST comparison with existing data in the NCBI database revealed it to be Candidatus phytoplasma palmae strain isolate Tanz08, under gene accession No. GU952107.1. Further phylogenetic analysis using the maximum likelihood method indicated that the strain is closely related to those reported in Tanzania. The CLYD positive samples were identified from Lamu and Kwale counties, with 6 cases from each County. Although there were no positive cases reported in Kilifi and Tana River counties, the presence of the vectors suggests that more studies need to be done on the disease. Further studies should be done on the alternative hosts, varieties prone to the disease, characterization and diversity of the vectors, the incubation period and the host pathogen relationship.

X-disease is an agricultural pathogen caused by the phytoplasma 'Candidatus Phytoplasma pruni' and is responsible for significant economic losses to cherry production across the Pacific Northwest. Euscelidius variegatus (Hemiptera: Cicadellidae) (Kirschbaum) is a key vector of the X-disease phytoplasma, and the most prevalent vector species in cherry orchards in The Dalles, OR. Little is known about the phenology of this important vector species. To determine the phenology of this species, E. variegatus adults were collected from commercial cherry orchards in The Dalles and Hood River, OR, and placed on potted barley plants in the laboratory. Development time and longevity of the F1 generation was recorded. Development time from egg to adult was 35.3 ± 2.5 d (n = 125). Adult leafhoppers lived an average of 52.6 d (SD = 32.9 d) (n = 34). We fit the survival data to a Gompertz distribution that produced a probability curve that estimated 50, 80, and 95% mortality at 41, 81, and 104 d, respectively. We also measured egg production and placement. Over 3 d females (n = 10) laid from 11 to 47 eggs with an average of 24.5 (SD = 10.6). The distribution of egg height produced a bell curve, with the highest number of eggs (n = 26: 10.6%) laid at 9 cm. Most eggs were found between 5 cm and 17 cm (n = 219: 89.4%). Our findings provide essential data for future degree-day models, enhancing understanding of population dynamics, and informing integrated pest management decisions.

Pear decline, caused by '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 its potential role in 'Ca. P. pyri' spreading under Chilean field conditions.

ABSTRACTPhytoplasma (‘Candidatus Phytoplasma’), a bacterial pathogen, is a significant plant health concern worldwide, resulting in substantial economic losses. In jujube (Ziziphus jujuba), it is often referred to as a cancer‐like disease that can destroy the whole plant and cause great economic loss, posing serious challenges to jujube's sustainable development. Here, the phytoplasma tolerance mechanism in jujube was revealed by the identification of a highly phytoplasma‐tolerant jujube genotype. Integrated transcriptomic and metabolomic analyses were conducted to compare a phytoplasma‐susceptible genotype Fuxiang with a tolerant genotype T13. The results revealed that the ZjWRKY40 gene was significantly upregulated in T13 about 3‐fold at the third development stage, suggesting its key role in mediating phytoplasma tolerance. A phytoplasma effector SJP4JWB was identified and shown to interact with ZjWRKY40 by yeast two‐hybrid and co‐immunoprecipitation methods. In addition, ZjWRKY40 was found to bind to the promoter of zju‐miR157, thereby regulating its expression. Moreover, zju‐miR157 targeted ZjSPL3 and negatively affected the phytoplasma tolerance by downregulating ZjSPL3 expression. Together, these findings outline a regulatory network involving SJP4JWB–ZjWRKY40–zju‐miR157, which provides important insights into the molecular mechanism of the phytoplasma tolerance in jujube and lays a foundation for developing tolerant genotypes through molecular breeding.

Occurrence of Candidatus Phytoplasma australasiaticum (16SrII-D) species infecting roses and weed plants in Andhra Pradesh, India

The X-disease phytoplasma (‘ Candidatus Phytoplasma pruni’) is an obligate pathogen that is capable of infection, persistence, and pathogenicity in both its major plant host ( Prunus spp.) and leafhopper vector ( Colladonus spp.) species. How ‘ Ca. P. pruni’ interacts with its plant and insect hosts, and how it alters its gene expression to do so, is unknown. Therefore, in this study, we conducted comparative RNA sequencing and differential gene expression analysis on ‘ Ca. P. pruni’-infected Prunus avium and Colladonus reductus samples. We found that the phytoplasma altered the expression of approximately 32% of its annotated protein-coding and pseudogenes, including intercellular transporters, proteolytic activity, and membrane structure, as well as upregulating genes associated with potential mobile units when in insect tissues versus in plant tissues. Most notably, differential expression was observed in genes that were identified by in silico analysis as being putative secreted effectors that may play a role in allowing phytoplasma infection and survival in these two very different host systems, as well as inducing X-disease in Prunus spp., which offers targets for control of this damaging pathogen by disrupting phytoplasma–host interactions. [Formula: see text] Copyright © 2026 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

The aim – to establish the phytopathogenic complex of tomato diseases in open soil conditions of the Left-Bank Forest-Steppe of Ukraine. Methods – general scientific, field, statistical. Results. Having analyzed the species composition of tomato disease pathogens in open soil conditions of the Left-Bank Forest-Steppe of Ukraine, it was found that plants are affected by fungal, bacterial and viral pathogens. The dominant role in the phytopathogenic complex of tomato belongs to diseases of fungal etiology – 63,0 %; pathogens of bacterial and viral etiology occupy 34,0 % and 3,0 %, respectively. Fungal pathogens caused early blight (Alternaria solani Ell et Mart.), fusarium wilt (Fusarium oxysporum Schltdl.), anthracnose (Colletotrichum coccodes (Wallz.) Hugres); bacterial diseases were represented by pathogens of soft bacterial rot (Erwinia carotovora subsp. carotovora), bacterial spot (Xantomonas vesicatoria Doidge), tomato stolbur (“Candidatus Phytoplasma”); viral - cucumber mosaic virus (CMV). According to the results of the study of the pathogenesis of tomato diseases, the following symptoms of the diseases were established: leaf spotting, wilting, fruit rot, disruption of plant reproductive functions, color change, growth inhibition. The most harmful tomato diseases in the fruit formation phase (BBCH 77) were early blight, the severity of of which reached 32,3 %; in the maturity of fruit phase (BBCH 89) – soft bacterial rot, the severity of disease was 11,1 %. Conclusions. The features of the formation of the phytopathogenic complex of tomato in open ground conditions were determined, which allowed us to determine the most harmful crop diseases in the conditions of the Forest-Steppe zone of Ukraine. The results obtained can be used for further research on the development of tomato plant protection systems to limit their development.

First report of 'Candidatus Phytoplasma citri'-related strain (16srII-C subgroup) associated with little leaf and witches’-broom disease of sunn hemp (Crotalaria juncea) in India

The Mexican yam bean (Pachyrhizus erosus L. Urban), a high-yielding leguminous root crop with good nutritional value and economic potential as a field crop, is widely cultivated in southern China (Han et al. 2023; Ravi et al. 2025). From September to November 2025, plants showing floral abnormalities, phyllody, proliferation of axillary shoots with shortened internodes, and stunting were found in a farmer’s field at Baihou District (24°17′50.489″N, 116°46′23.916″E) and Dapu County, Meizhou City, Guangdong Province, China. These disease symptoms, suspected to be induced by phytoplasma, had an infection rate of about 30% in the field, causing a yield loss of approximately 20%. For further investigation, total DNA was extracted from fresh leaves of ten symptomatic and four asymptomatic samples using a Plant Genomic DNA Purification Kit (DP305; Tiangen; Beijing; China). Conventional nested PCR was performed using primer pairs P1/P7 followed by R16mF2/mR1 (Lee et al. 1993) , and amplified a fragment of the expected size only from the symptomatic samples. The phytoplasma strain was designated as GDDPPE2502. The PCR products were sequenced, and a representative nucleotide sequence of 1,410 bp was deposited in GenBank under the accession number PX705183. A BLASTn similarity search revealed that the sequence shared 99.93% identity with that of the 16SrII-V subgroup reference strain ‘Candidatus Phytoplasma australasiaticum’ BAWM-TWN (GenBank accession no. GCA_030586915.1, 1,383/1,384 bp), and 99.86% with the SPLL subgroup reference strain ‘Ca. P. australasiaticum subsp. Ipomoeae’ o7C (GenBank accession no. GCA_024425275.1, 1,382/1,384 bp) (Rodrigues Jardim et al. 2023). Moreover, Virtual RFLP analysis using iPhyClassifier (Zhao et al. 2009) revealed that the phytoplasma strain was a member of the 16SrII-A subgroup with a 1.00 similarity coefficient to the reference phytoplasma strain (GenBank accession no. L33765). To validate the results, the translocase protein (secY) genes were amplified using primer pairs secYwbF1/secYwbR1 followed by secYwbF2/secYwbR2 (Wang et al. 2025), and the 0.8-kb band was sequenced and deposited in GenBank (GenBank accession no. PX715155). BLAST analysis showed that the secY sequence of GDDPPE2502 shared 100% of sequence identities to several ‘Ca. P. australasiaticum’ strains (GenBank accession no. CP133702.1, CP040925.1, CP097312.1 and CP171825.1). To further characterize the phytoplasma, phylogenetic analysis based on the 16S rRNA and secY genes, conducted by MEGA 7.0(Kumar et al. 2016), confirmed that the GDDPPE2502 consistently clustered within the same subclade as the 16SrII group ‘Ca. P. australasiaticum’ related strain NCHU2014 (16SrII-A), BAWM-TWN (16SrII-V) and o7C (SPLL). Taken together, according to the latest ‘Candidatus Phytoplasma’ species description guidelines (Bertaccini et al. 2022; Rodrigues Jardim et al. 2023; Wei and Zhao 2022), the phytoplasma GDDPPE2502 was related to ‘Ca. P. australasiaticum’ (16SrII-A). To our knowledge, the present study is the first to report a ‘Ca. P. australasiaticum’ -related strain associated with yam bean in Guangdong, China. These findings highlight the need for further research to identify the potential insect vectors and transmission pathways, which will be beneficial for the epidemiological monitoring and prevention of the related diseases.

Pear decline (PD), associated with 'Candidatus Phytoplasma pyri', is one of the most severe diseases affecting pear cultivation in Europe and the United States. Several psyllid species act as vectors of phytoplasmas belonging to the 16SrX group and play a key role in the epidemiology of the disease. This study aimed to characterize the epidemiology of pear decline in Sicily using integrated field, molecular, vector, and remote sensing approaches, four years after the first detection of PD in the region. Visual surveys and molecular analyses were conducted over two years in eight pear orchards. A total of 115 plant samples and 101 Cacopsylla spp. specimens, selected from 1435 collected individuals, were analysed, confirming the presence of 'Ca. P. pyri' in 69% of symptomatic plants and in 4.6% of C. pyri individuals. Genetic characterization revealed a high degree of similarity among the phytoplasma isolates analysed. Remote sensing analyses conducted since 2018, combined with vector population monitoring, confirmed the epidemic nature of PD and indicated the persistence of a risk of further pathogen spread within the region, supporting the use of remote sensing as a complementary tool for large-scale disease monitoring.

'Candidatus Phytoplasma solani' of the 16SrXII group is an emerging vector-borne pathogen in European crop production. The cixiid planthopper Hyalesthes obsoletus transmits 16SrXII-A stolbur phytoplasmas that are associated with diseases in grapevine, potato, and various weeds. While 16SrXII-P genomes transmitted by Pentastiridius leporinus are available, no genome of an H. obsoletus-transmissible 16SrXII-A phytoplasma has been reported from Germany. Here, we present insights into the phylogenetic position and pathogen-host interactions through the functional reconstruction of the complete 832,614 bp genome of the H. obsoletus transmissible 'Ca. P. solani' 16SrXII-A strain POT from a potato field. Phylogenetic analyses highlight the heterogeneity within the stolbur group using whole-genome alignment and a BUSCO-based core gene analysis approach. The POT chromosome shares highest average nucleotide identity with Italian bindweed-associated genomes and displays strong synteny with the c5 strain. Consistent with the typical phytoplasma architecture, the POT genome combines mobile-element-driven instability with a conserved core metabolism. Virulence factors include transposon-linked effectors but lack pathogenicity island organisation. POT further differs from other 16SrXII-group phytoplasmas through unique collagen-like proteins that could contribute to virulence. These findings provide a robust genomic framework that improves diagnostics, enables strain-level resolution and supports the assessment of breeding materials under stolbur phytoplasma pressure, thereby refining our understanding of stolbur phytoplasma diversity and highlighting the evolutionary divergence within the 16SrXII subgroup.

Little leaf and witches’ broom disease associated with phytoplasma infection has recently been reported in Tamil Nadu, across all growing seasons. A survey was conducted to assess the incidence of sunnhemp little leaf and witches’ broom disease in various sunnhemp growing regions of Tamil Nadu during 2022-2024, across all seasons. The highest incidence of little leaf and witches broom disease was observed during the Kharif 2023 in Pudukkottai district with an infection rate of 0.55 % to 88.67 %. Infected plants exhibited characteristic symptoms including phyllody (leaf-like structures replacing flowers), virescence (green pigmentation in floral parts), yellowing, reduced leaf size, stunted growth, witches’ broom (proliferation of shoots), leaf curling, bunchy appearance at stem ends, floral gigantism, twig die-back and unseasonal yellowing or reddening of leaves. The disease is caused by Candidatus phytoplasma and is transmitted by leafhoppers. However, the phytoplasma cannot be cultured under in vitro conditions. Therefore, in this study, nested PCR approach was employed as a rapid and reliable molecular diagnostic technique for detecting phytoplasma. Specific second-round primers were used to confirm the presence of phytoplasma in little leaf and witches’ broom infected samples. The infected plant material was analyzed using a 16S rRNA region-specific primer to identify the pathogen. Sequence analysis revealed that the phytoplasma infecting sunnhemp had over 99 % nucleotide similarity with other isolates of Candidatus: Phytoplasma aurantifolia from the NCBI database, confirming the association of Candidatus: Phytoplasma aurantifolia (16SrII group) with sunnhemp little leaf and witches’ broom disease.

End-point PCR, followed by restriction fragment length polymorphism (RFLP) or sequencing and sequence analysis of the 16S rRNA gene, is a cornerstone method for the universal detection and identification of phytoplasmas, including previously undescribed strains. This protocol describes the use of end-point PCR in direct, nested, or semi-nested systems, coupled with RFLP analysis using 17 restriction enzymes, to classify phytoplasmas into distinct 16Sr ribosomal groups and subgroups while addressing challenges such as inter-operon heterogeneity and mixed infections. Alternatively, sequencing of the PCR-amplified 16S rRNA gene, followed by sequence analysis (e.g., virtual RFLP or sequence homology), enables classification into either 16Sr groups/subgroups or 'Candidatus Phytoplasma' species. The method's sensitivity, cost-effectiveness, and compatibility with established classification frameworks make it invaluable for epidemiological studies, quarantine measures, and the delineation of 'Candidatus Phytoplasma' species. By providing a clear framework for the precise diagnosis of phytoplasma-associated diseases in diverse plant and insect hosts, this protocol supports rapid responses to outbreaks and helps mitigate the economic impact of phytoplasmas.

Imp is a gene coding for the immunodominant membrane protein Imp that is supposed to be involved in host-pathogen interactions. It was identified in some 'Candidatus Phytoplasma' species and showed a considerable sequence variability. For this characteristic it can be exploited to complement conventional phytoplasma classification based on the conserved 16S rRNA gene, enhancing the differentiation of closely related phytoplasma strains. This protocol describes a method for amplification and sequence analysis of the imp gene of Flavescence dorée (FD) related phytoplasmas, that can be useful for detecting and genotyping the strains involved, for example, in the complex FD epidemiology.

Since phytoplasmas cannot be cultured in vitro, it is impossible to study a protein of interest in vivo by manipulating its corresponding gene. It is therefore necessary to recombinantly express a protein of interest for use in an in vitro study. Although recombinant expression of proteins in E. coli is well established, there are some issues to consider when expressing recombinant phytoplasma proteins. Here, we describe a detailed method for the recombinant expression of an effector protein, SAP11, from 'Candidatus Phytoplasma mali' in E. coli. Some important points are highlighted.

ABSTRACT X‐disease, caused by strains in the 16SrIII‐A subgroup of ‘ Candidatus Phytoplasma pruni’, is a devastating disease of Prunus species (stone fruits). Multiple outbreaks of this disease have occurred across much of North America for more than a century, with the most recent one beginning around 2010 in the Pacific Northwest of the United States, causing severe damage to the stone fruit industry. Sensitive and specific detection of X‐disease is critical to prevent the propagation and spread of infected plant material; however, current PCR‐based detection workflows lack specificity to X‐disease and require one or more PCR assays followed by additional testing to confirm the species identity. In this study, two real‐time PCR assays, one targeting the secY gene and the other targeting the tuf gene, were developed for specific detection of X‐disease in stone fruits. The CPH‐TXD assay targeting the tuf gene provided high‐sensitivity target detection of as few as five target copies with significantly reduced cross‐reactivity compared to current methods. The CPH‐SXD assay targeting the secY gene detected a minimum of 30 target copies and showed no cross‐reaction with closely related strains. The repeatability and reproducibility of both assays were verified with acceptable results. Together, these assays provide efficient and high‐specificity detection methods to combat the growing threat of X‐disease in North America.

Flavescence dorée and Bois noir are two major grapevine yellows diseases affecting European viticulture. Accurate, early detection of the associated bacterial agents-Flavescence dorée phytoplasma (FDp) and 'Candidatus Phytoplasma solani'-in both grapevines and insect vectors is critical for effective disease monitoring and control. Quantitative PCR (qPCR) is a highly sensitive molecular tool widely used for phytoplasma detection, often implemented in multiplex formats for simultaneous identification of multiple targets. However, the integration of internal quality controls is essential to validate DNA extraction efficiency and rule out PCR inhibition. While species-specific endogenous controls are commonly used, their limited host range restricts assay flexibility. This protocol describes a novel triplex qPCR assay enabling the simultaneous detection of FDp, 'Ca. P. solani', and a universal endogenous control based on a conserved region of the eukaryotic 28S rRNA gene. This universal control allows broad applicability across diverse eukaryotic hosts, including both grapevine and key insect vectors, facilitating robust and flexible diagnostics in a single reaction setup.